Leading Edge
Review
Hallmarks of aging: An expanding universe
Carlos Lo´pez-Otı´n,1,2,3,* Maria A. Blasco,4 Linda Partridge,5,6 Manuel Serrano,7,8,9 and Guido Kroemer10,11,12,*
1Departamento de Bioquı´mica y Biologı´a Molecular, Instituto Universitario de Oncologı´a (IUOPA), Universidad de Oviedo, Oviedo, Spain
2Instituto de Investigacio´n Sanitaria del Principado de Asturias (ISPA), Oviedo, Spain
3Centro de Investigacio´n Biome´dica en Red de Ca´ncer (CIBERONC), Madrid, Spain
4Telomeres and Telomerase Group, Molecular Oncology Program, Spanish National Cancer Centre (CNIO), Madrid, Spain
5Department of Genetics, Evolution and Environment, Institute of Healthy Ageing, University College London, London, UK
6Max Planck Institute for Biology of Ageing, Cologne, Germany
7Institute for Research in Biomedicine (IRB Barcelona), Barcelona Institute of Science and Technology (BIST), Barcelona, Spain
8Catalan Institution for Research and Advanced Studies (ICREA), Barcelona, Spain
9Altos Labs, Cambridge, UK
10Centre de Recherche des Cordeliers, Equipe labellise´e par la Ligue contre le cancer, Universite´ de Paris, Sorbonne Universite´, INSERM
U1138, Institut Universitaire de France, Paris, France
11Metabolomics and Cell Biology Platforms, Gustave Roussy, Villejuif, France
12Institut du Cancer Paris CARPEM, Department of Biology, Hoˆpital Europe´en Georges Pompidou, AP-HP, Paris, France
*Correspondence: clo@uniovi.es (C.L.-O.), kroemer@orange.fr (G.K.)
https://doi.org/10.1016/j.cell.2022.11.001
llSUMMARYAging is driven by hallmarks fulfilling the following three premises: (1) their age-associated manifestation, (2)
the acceleration of aging by experimentally accentuating them, and (3) the opportunity to decelerate, stop, or
reverse aging by therapeutic interventions on them. We propose the following twelve hallmarks of aging:
genomic instability, telomere attrition, epigenetic alterations, loss of proteostasis, disabled macroautoph-
agy, deregulated nutrient-sensing, mitochondrial dysfunction, cellular senescence, stem cell exhaustion,
altered intercellular communication, chronic inflammation, and dysbiosis. These hallmarks are intercon-
nected among each other, as well as to the recently proposed hallmarks of health, which include organiza-
tional features of spatial compartmentalization, maintenance of homeostasis, and adequate responses to
stress.INTRODUCTION
Aging research explores the decline in function of organisms
during adulthood. Since 2013, when the first edition of the hall-
marks of aging was published in Cell,1 close to 300,000 articles
dealing with this subject have been published, which is as many
as during the preceding century. Hence, time has become ripe
for a new edition of the hallmarks of aging incorporating the
main knowledge obtained a decade on.
The distinction among ‘‘hallmarks’’ is intrinsically diffuse, since
they interact and are not independent of each other. Therefore,
their classification is inevitably arbitrary, but we proposed three
criteria that must apply for each hallmark of aging: (1) the time-
dependent manifestation of alterations accompanying the aging
process, (2) the possibility to accelerate aging by experimentally
accentuating the hallmark, and—most decisively—(3) the oppor-
tunity to decelerate, halt, or reverse aging by therapeutic inter-
ventions on the hallmark. Rather than elaborating a compendium
of age-associated alterations, we shall focus on the molecular,
cellular, and systemic processes mechanistically accounting
for their manifestation. That said, both in laboratory animals
and in human medicine, objective quantification of morpholog-
ical and functional decline affecting the aging organism is essen-
tial to measure biological aging. Indeed, disparity betweenbiological and chronological age can reflect the efficacy of
age-accelerating or -decelerating manipulations that evaluate
the contribution of a given hallmark to the aging process. For
this reason, standardized physiological measurements (e.g.,
respirometry to measure basal and maximal energy expendi-
ture), functional tests (e.g., at the sensory, psychomotor, and
cognitive levels), and ever more sophisticated ‘‘omics’’ technol-
ogies (e.g., genomics, epigenomics, transcriptomics, prote-
omics, and metabolomics), often applied at the single-cell level,
are instrumental for evaluating the spatiotemporal patterns of
health degradation and the (in)efficacy of anti-aging strategies.
In 2013, we suggested nine molecular, cellular, and systemic
hallmarks of aging: DNA instability, telomere attrition, epigenetic
alterations, loss of proteostasis, deregulated nutrient-sensing,
mitochondrial dysfunction, cellular senescence, stem cell
exhaustion, and altered intercellular communication.1 Recent
research has confirmed and extended the importance of all
these hallmarks. They have withstood scrutiny by tens of
thousands of aging researchers, but they require an update to
deal with the discoveries of the last decade. For example, in
2013, much of the evidence on anti-aging interventions was
limited to non-mammalian model organisms including yeast,
nematodes, and fruit flies. Fortunately, experiments involving
mice (and in some cases, non-human primates) have nowCell 186, January 19, 2023 ª 2022 Elsevier Inc. 243
Figure 1. The hallmarks of aging
The scheme compiles the 12 hallmarks of aging
proposed in this work: genomic instability, telo-
mere attrition, epigenetic alterations, loss of
proteostasis, disabled macroautophagy, deregu-
lated nutrient-sensing, mitochondrial dysfunction,
cellular senescence, stem cell exhaustion, altered
intercellular communication, chronic inflamma-
tion, and dysbiosis. These hallmarks are grouped
into three categories: primary, antagonistic, and
integrative.
ll
Reviewcorroborated the validity of most of these hallmarks in mammals.
Of note, human age-related diseases have statistically higher
chances to co-occur and to share genomic characteristics
when they are causally linked to the same hallmark rather than
to different hallmarks,2 clinically validating the approach that
we have chosen.
Besides the necessary update of the previous hallmarks,
we have also introduced some reorganizations and included
the following three additional hallmarks of aging: disabled mac-
roautophagy, chronic inflammation, and dysbiosis. Disabled
macroautophagy was initially treated as a special case of loss
of proteostasis. However, macroautophagy does not only affect
proteins but can target entire organelles and non-proteinaceous
macromolecules, justifying its discussion as a separate entity.
Moreover, we considered that the final hallmark that we listed
in 2013, altered intercellular communication, was too vast,
requiring a separate discussion of chronic inflammation and
age-associated dysbiosis (Figure 1).
The interdependence of aging hallmarks means that the
experimental accentuation or attenuation of one specific hallmark
usually affects other hallmarks as well. This underscores the fact
that aging is a complex process that has to be conceived as a
whole. Accordingly, each of the hallmarks should be considered
as a point-of-entry for future exploration of the aging process, as
well as for the development of new anti-aging medicines.244 Cell 186, January 19, 2023GENOMIC INSTABILITY
Genome integrity and stability are perva-
sively threatened by exogenous chemi-
cal, physical, and biological agents,
as well as by endogenous challenges
such as DNA replication errors, chromo-
some segregation defects, oxidative pro-
cesses, andspontaneoushydrolytic reac-
tions. The wide range of genetic lesions
causedby theseextrinsic or intrinsic sour-
ces of damage include point mutations,
deletions, translocations, telomere short-
ening, single- and double-strand breaks,
chromosomal rearrangements, defects
in nuclear architecture, and gene disrup-
tion caused by the integration of viruses
or transposons. All these molecular alter-
ations and the resulting genomic mosai-
cism may contribute to both normal and
pathological aging.3 Accordingly, organ-isms have evolved a complex array of DNA repair and mainte-
nance mechanisms to deal with the damage inflicted to nuclear
and mitochondrial DNA (mtDNA) and to ensure the appropriate
chromosomal architecture and stability. These DNA repair net-
works lose efficiencywith age, which accentuates the accumula-
tion of genomic damage and the ectopic accumulation of DNA in
the cytosol4 (Figure 2A).
Nuclear DNA
Cells from aged humans and model organisms accumulate so-
matic mutations at nuclear DNA.5 Other forms of damage,
such as chromosomal aneuploidy and copy-number variations,
are also associated with aging. All these DNA alterations may
affect essential genes and transcriptional pathways, resulting
in dysfunctional cells that may finally compromise tissue and
organismal homeostasis. This is especially relevant when DNA
damage impacts on stem cells, hampering their role in tissue
renewal or leading to their exhaustion, which in turn promotes
aging and increases susceptibility to age-related pathologies.6,7
The mutational burden in histologically normal human tissues is
remarkable. For example, normal esophageal epithelium cells
from young individuals already display hundreds of mutations
and may carry more than 2,000 mutations per cell by middle
age.8 The accumulation of DNA mutations throughout life is
likely tolerated because of the excessive energetic cost of the
Figure 2. Loss of cellular integrity caused by genomic instability,
telomere attrition, and epigenetic alterations
(A) Endogenous or exogenous agents cause a variety of DNA lesions that
contribute to both normal and pathological aging. Such lesions can by repaired
by a variety of mechanisms that lose efficiency with age. Excessive DNA
damage, insufficient DNA repair, alterations in nuclear architecture, and
telomere attrition favor the aging process. BER, base excision repair; HR,
homologous recombination; NER, nucleotide excision repair; NHEJ, non-ho-
mologous end joining; MMR, mismatch repair; SAC, spindle assembly
checkpoint; TERT, telomerase reverse transcriptase; TLS, trans-lesion syn-
thesis.
(B) Changes in the acetylation andmethylation of DNA or histones, as well as in
levels or activity of chromatin-associated proteins or of non-coding RNAs
(ncRNAs) induce epigenetic changes that contribute to the aging process. The
red portions of the hourglasses indicate age-associated alterations and the
blue portions strategies for avoiding them.
ll
Reviewcomplete repair of all genomic damages caused by exogenous
and endogenous challenges. Consequently, cells favor survival
over genomic integrity.9 These data also suggest that similar to
carcinogenesis, driver mutations alone may not be sufficient toaccelerate aging because they require a permissive microenvi-
ronment created by non-mutagenic promoting factors to
become penetrant.10
Comparative analysis of the mutational landscape across
mammalian species has shown that species-specific somatic
mutation rate is inversely correlated with lifespan.11 To date,
there is no clear evidence that the normal rate of mutation fixa-
tion is responsible for aging, but numerous studies have shown
that DNA repair deficiencies have the potential to cause aging.
Thus, alterations in DNA repair mechanisms accelerate
aging in mice and underlie several human progeroid syn-
dromes.12 Conversely, transgenic mice overexpressing the
mitotic checkpoint kinase BubR1 exhibit an extended healthy
lifespan13 (Table 1). Moreover, studies in humans and other
long-lived species have shown that enhanced DNA repair mech-
anisms coevolve with increased longevity.14 Sirtuin-6 (SIRT6)
may play a major role in this differential reparative efficiency
across species. Overexpression of SIRT6 in mice reduces
genomic instability, improves double-strand break repair, and
extends lifespan15 (Table 1), although other explanations, such
as improved glucose metabolism and restoration of energy
homeostasis, have been proposed to explain the prolongevity
effects of SIRT6.16 Notably, recent work has shown that
small-molecule activation of 8-oxoguanine DNA glycosylase 1
increases oxidative DNA damage repair andmay have therapeu-
tic applications in the context of aging and other processes
linked to excessive oxidative damage.17 These findings suggest
that interventions aimed at reducing the mutational load of nu-
clear DNA or at enhancing or rerouting its repair mechanisms
may delay aging and the onset of age-related diseases, but
further causal evidence in this regard is still missing.
Mitochondrial DNA
Genomic instability affecting mtDNA may contribute to aging and
age-related pathologies.96 mtDNA is strongly impacted by ag-
ing-associated mutations and deletions due to its high replicative
index, the limited efficiency of its repair mechanisms, its oxidative
microenvironment, and the lack of protective histones embracing
this small DNA molecule. Somatic mtDNA alterations increase
across human tissues during aging, but it remains unclearwhether
this increase truly impacts theagingprocessat the functional level.
The causal implication of mtDNA mutations in driving aging has
been difficult to assess because of ‘‘heteroplasmy,’’ which implies
the co-existence of mutated and wild-type genomes within the
same cell. However, deep-sequencing of aged cells revealed
that their mtDNA mutational load may substantially increase
through clonal expansion events.97 The accelerated expansion
of mitochondrial mutations with age has also been observed in
both primate oocytes and somatic tissues,98 as well as in lympho-
blasts from patients with neurodegenerative diseases.99 Of note,
ultra-sensitive sequencing indicates that most mtDNA mutations
in aged cells arise from replication errors caused by mtDNA poly-
merase g rather than from oxidative stress.96
Preliminary evidence that mtDNA mutations might be directly
involved in aging andage-relatedpathologieswasprovidedbyhu-
man disorders that are caused by mtDNA damage and partially
phenocopy aging.100 Further causative evidence has arisen from
studies on mice deficient in DNA polymerase g that exhibitCell 186, January 19, 2023 245
Table 1. Examples of anti-aging effects of hallmark-targeted interventions in mammals
Hallmark Species/model Intervention Outcome Ref
Genomic
instability
mouse BubR1 overexpression increased longevity North et al.13
mouse SIRT6 overexpression increased longevity Tian et al.15
HGPS mouse
HGPS human
farnesyl transferase inhibitors healthspan and lifespan extension Gordon et al.18
Telomere
attrition
mouse telomere lengthening lifespan extension Toma´s-Loba et al.19
telomerase-
null mouse
telomerase reactivation lifespan extension Jaskelioff et al.20
mouse pharmacological or genetic
activation of telomerase
delayed aging Bernardes de
Jesus et al.21
mouse hyperlong telomeres increased lifespan; metabolic health
improvement
Mun˜oz-Lorente et al.22
mouse telomere maintenance in
adult neurons
preservation of neuron survival and
cognitive function
Shim et al.23
mouse telomerase activation by
gene therapy strategy
improvement in models of
pulmonary fibrosis and aplastic
anemia
Povedano et al.24
and Ba¨r et al.25
Epigenetic
alterations
human a-ketoglutarate delayed epigenetic clock Demidenko et al.26
human thymus regeneration by
GH +DHEA+ metformin
delayed epigenetic clock Fahy et al.27
mouse Kat7 inactivation lifespan extension Wang et al.28
human stem
cells
KAT7 inactivation decreased H3 acetylation, reduced
cell senescence
Wang et al.28
mouse Sirt1 overexpression improved genomic stability and
metabolism
Bhatt and Tiwari29
mouse Sirt3 overexpression reversed regenerative capacity
of HSC
Bhatt and Tiwari29
mouse miR-188 depletion alleviated age-related vascular
problems
He et al.30
mouse miR-455-3p overexpression improved mitochondrial and
cognitive function; extended
lifespan
Kumar et al.31
aged or Sirt6
/

mouse
nucleoside reverse-transcriptase
inhibitors
improved healthspan and lifespan Simon et al.32
Loss of
proteostasis
mouse transgenic expression of LAMP2a
in hepatocytes or HSC
improved hepatocyte viability in
aged mice. Improved function and
metabolic properties of HSC
Dong et al.33
mouse pharmacological induction of CMA improved Alzheimer’s pathology
and arteriosclerosis in disease
models
Madrigal-Matute
et al.34
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Table 1. Continued
Hallmark Species/model Intervention Outcome Ref
mouse intranasal administration of
recombinant human HSP70
enhanced lifespan (m), improved
cognitive functions, and
proteasome activity; reduced brain
lipofuscin
Bobkova et al.35
mouse 4-phenylbutyrate administered
to aged mice
reduced ER stress in cortex and
hippocampus and improved
cognition
Hafycz et al.36
human guanabenz in patients with ALS
(FDA approved)
reduced progression to bulbar stage Dalla Bella et al.37
Disabled
macroautophagy
mouse transgenic overexpression of Atg5 improved longevity, metabolic
health and motor function
Pyo et al.38
mouse mutation of beclin 1
(Becn1F121A/F121A) to reduce its
inhibition by Bcl-2
extended longevity of C57BL/6 mice
and progeroid klotho-ko mice.
Prolonged neurogenesis
Ferna´ndez et al.39
and Wang et al.40
mouse spermidine in drinking water extended longevity, reduced
cardiac aging and oxidative stress,
sinusoidal dilation in liver
Eisenberg et al.41
mouse salicylates (salicylate,
acetylsalicylate)
EP300 inhibition; autophagy-
dependent hepatoprotection;
improved cancer
immunosurveillance
Castoldi et al.42
mouse nordihydroguairaretic acid longevity extension (m) with EP300
inhibition
Tezil et al.43
human oral NMN in prediabetic women
(phase III trial)
increased insulin sensitivity of
skeleton muscle
Yoshino et al.44
human oral NR in Parkinson disease
patients (phase 1 trial)
clinical improvement, reduced
inflammatory cytokines in serum
and cerebrospinal fluid
Brakedal et al.45
human NAM in patients with 2 non-
melanoma skin cancers in the
preceding 5 years (phase 3 trial)
reduced rates of new non-
melanoma skin cancers and
keratosis
Chen et al.46
human urolithin A to middle-aged adults
(randomized phase 2)
improved aerobic endurance and
physical performance; reduced
plasma CRP
Singh et al.47
Deregulated
nutrient-sensing
mouse inducible GH receptor knockout
at 6 months
longevity and enhanced insulin
sensitivity; less neoplasms (m)
Duran-Ortiz et al.48
mouse caloric restriction by 30% of male
C57BL/6J mice
lifespan extension of 10%–30% Acosta-Rodrı´guez
et al.49
human caloric restriction by 14% for 2 years
(phase 2)
improved thymopoiesis and anti-
inflammatory effects on adipose
tissues. Reduced PLA2G7
Spadaro et al.50
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Table 1. Continued
Hallmark Species/model Intervention Outcome Ref
expression (gene knockout in mice
ameliorates metabolic health)
mouse ß-hydroxybutyrate in drinking water higher energy expenditure,
improved motor fitness and memory
an et al.51
Mitochondrial
dysfunction
mouse TPP-thiazole (inhibitor of respiratory
chain complex IV)
improved mitochondrial
metabolism, reduced visceral fat
and higher glucose tolerance in
old mice
avallaie et al.52
mouse CRMP that preferentially acts to
uncouple hepatocyte mitochondria
administration to obese old mice
reduces hepatosteatosis and liver
insulin resistance
oedeke et al.53
mouse elamipretide to inhibit mitochondrial
permeability transition
improved mitochondrial function
and avoidance of diastolic heart
dysfunction, especially if combined
with NMN
hang et al.54
primates spontaneous or diet-induced
obesity in cynomolgus and rhesus
macaques, treated with CRMP
enhanced hepatic mitochondrial fat
oxidation, improved insulin
tolerance, reduced hepatic and
plasma triglycerides, reduced
cholesterol
oedeke et al.55
human clinical trial evaluating elamipretide
on Barth syndrome (randomized
phase 2/3 trial)
improved walking test, muscle and
cardiac parameters, overall
improvement
eid Thompson et al.56
human clinical trial with L-carnitine
supplementation to older man
(phase 2 trial)
increased muscle carnitine content
and fatty acid oxidation during
exercise
hee et al.57
Cellular
senescence
mouse genetic ablation of p16-
expressing cells
increased health- and lifespan.
Treatment starting at 1 year age
aker et al.58
mouse senolytic treatment with dasatinib +
quercetin
increased health- and lifespan.
Treatment starting at 2 years age
u et al.59
mouse senolytic treatment with fisetin increased health- and lifespan.
Treatment starting at 1.6 years age
ousefzadeh et al.60
human senolytic treatment with dasatinib +
quercetin of patients with pulmonary
fibrosis (phase 1 trial)
improved physical performance;
reduction in pro-inflammatory and
pro-fibrotic factors in serum;
elevation of aKlotho in urine
ustice et al.61
human senolytic treatment with dasatinib +
quercetin of patients with diabetic
kidney disease (phase 1 trial)
reduction of senescent cells and
macrophages in adipose tissue, and
of pro-inflammatory factors in serum
ickson et al.62
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Hallmark Species/model Intervention Outcome Ref
Stem cell
exhaustion
mouse transgenic expression of OSKM longevity extension of HGPS
progeroid mice. Preservation of
hippocampal neuro- genesis and
function in normal mice. Improved
tissue repair immediately after injury
or in a subsequent injury
Wang et al.,28
Ocampo et al.,63
Browder et al.,64
Chen et al.,65
Hishida et al.,66
Rodrı´guez-Matella´n
et al.,67 Gao et al.,68
and Doeser et al.69
mouse AAV2-driven expression of OSK
in the eye
restoration of visual acuity to old
mice and to mice with glaucoma.
Improved repair of crushed
optical nerve
Lu et al.70
Altered intercellular
communication
mouse dilution of blood from old mice
with saline/albumin
rejuvenation in multiple tissues Mehdipour et al.71
mouse blood transfusion improved muscle repair; reduced
liver steatosis and fibrosis
Rebo et al.72
mouse human umbilical cord plasma improved hippocampal
neurogenesis
Castellano et al.73
mouse heterochronic parabiosis rejuvenation in multiple tissues Ma et al.74 and Pa´lovics et al.75
mouse CCL3/MIP1a administration HSC rejuvenation Ma et al.74
mouse TIMP2 i.v. administration hippocampus rejuvenation Castellano et al.73
mouse IL-37 injection into old mice improved metabolism and
endurance exercise
Ballak et al.76
mouse GDF11 i.v. administration rejuvenation of brain, muscle and
pancreas, but pro-fibrotic effects
Frohlich and Vinciguerra77
mouse transgenic overexpression of VEGF improved health- and lifespan,
enhanced liver and muscle repair
Grunewald et al.78
mouse human cells YAP expression rejuvenation of old cells, prevention
of emergence of aging features
Sladitschek-Martens et al.79
human cells ECM from young fibroblasts rejuvenation of aged senescent cells Choi et al.80
mouse chondroitin 6-sulfo-transferase
overexpression
improved memory in old mice Yang et al.81
Chronic
inflammation
mouse blockade of TNF-a with etanercept
from 16 to 18 months of age in
C57BL/6 mice
prevention of sarcopenia and
increased lifespan (f)
Desdin-Mico et al.82; Sciorati et al.83
rat blockade of TNF-a with etanercept
from 24 to 26 months of age in male
Wistar albino rats
prevention of cognitive deficits,
endothelial dysfunction, peripheral
and neuro-inflammation
Gocmez et al.84
mouse knockout of prostaglandin E2
receptor EP2 in myeloid cells or
treatment of C57BL/6 mice with
EP300 inhibitors
improved cognition and reduced
age-associated inflammation
Minhas et al.85
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Table 1. Continued
Hallmark Species/model Intervention Outcome Ref
mouse knockout of NLRP3 in C57BL/6mice improved glucose tolerance,
cognition, motor performance and
female fertility due to reduced
ovarian aging
Marı´n-Aguilar et al.86
human treatment of patients with a history
of myocardial infarction and high
hsCRP with canakinumab (phase
3 trial)
reduced incidence of hypertension
and diabetes; reduced frequency of
recurrent myocardial infarction and
non-small cell lung cancer
Ridker et al.87
Dysbiosis HGPS mouse fecal microbiota transplantation
from WT mice; Akkermansia
muciniphila administration
enhanced healthspan and lifespan Ba´rcena et al.88
SAMP8 mouse Lactobacillus plantarum GKM3 longevity promotion and alleviation
of age-related cognitive impairment
Lin et al.89
mouse microbiota transplantation from
young mice to aged host
improved maintenance of brain
health and immunity
Boehme et al.90
mouse fecal microbiota transplantation improved ovarian function in
aged mice
Xu et al.91
mouse fecal microbiota transplantation improved germinal centre reactions
in lymph nodes
Stebegg et al.92
mouse indole metabolites reduction of inflammation
during aging
Krishnan et al.93
mouse short-chain fatty acids restored microglial function in
aged mice
Cryan et al.94
human oral administration of Akkermansia
muciniphila (randomized phase
1/2 trial)
improved metabolic parameters in
obese or diabetic patients
Depommier et al.95
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Reviewaccelerated aging and reduced lifespan associated with deletions
rather than point mutations in mtDNA101 (Table 1). Overall, these
data suggest that the avoidance, attenuation, or correction of
mtDNAmutations might contribute to extend healthspan and life-
span. Nevertheless, as in the case of nuclear DNA mutations,
experimental evidence demonstrating deceleration of aging
by gain of function in mtDNA repair mechanisms is still largely
missing.
Nuclear architecture
Defects in the nuclear lamina, which constitutes a scaffold
for tethering chromatin and protein complexes, can generate
genome instability.102 Accelerated aging syndromes such as the
Hutchinson-Gilford and the Ne´stor-Guillermo progeria syndromes
(HGPSandNGPS, respectively) are causedbymutations in genes
LMNA and BANF1 encoding protein components of nuclear lam-
ina. Alterations of the nuclear lamina andproduction of an aberrant
prelamin A isoform called progerin are also characteristics of
normal human aging, and lamin B1 levels decline during cellular
senescence.18 Animal and cellular models have facilitated the
identification of the response mechanisms and stress pathways
elicited by nuclear lamina aberrations caused by aging and proge-
ria, including activation of tumor suppressor protein p53 (TP53),
deregulation of the somatotrophic axis, and attrition of adult
stem cells.18
The causal implication of nuclear lamina abnormalities in pre-
mature aging has been corroborated by the observation that
decreasing prelamin A or progerin levels delays the onset of pro-
geroid features and extends lifespan in mouse models of HGPS.
This can be achieved by systemic injection of antisense oligonu-
cleotides, farnesyltransferase inhibitors, a combination of statins
and aminobisphosphonates, restoration of the somatotrophic
axis, or blockade of NF-kB signaling.103 Some of these interven-
tions have been already approved for use in progeria patients.104
Moreover, gene editing strategies have been recently developed
to correct LMNA mutations in cells from HGPS patients and in
animal models of this disease.105,106 Hopefully, these ap-
proaches will be clinically implemented for the future treatment
of progeria, but to date, no evidence is available showing that
reducing progerin would delay normal aging.
TELOMERE ATTRITION
DNA damage at the end of chromosomes (telomeres) contrib-
utes to aging and age-linked diseases.107 Replicative DNA poly-
merases are unable to complete the copy of telomere regions of
eukaryotic DNA. Accordingly, after several rounds of cell divi-
sion, telomeres undergo a substantial shortening that induces
genomic instability and finally leads to either apoptosis or cell
senescence. These deleterious effects can be prevented
by the reverse-transcriptase activity of telomerase, an active
ribonucleoprotein that elongates telomeres to maintain their
adequate length.108,109 However, most mammalian somatic
cells do not express telomerase, which leads to the progressive
and cumulative erosion of telomere sequences from chromo-
some ends throughout life. There are several examples in which
telomere attrition attenuates carcinogenesis through limiting the
replicative lifespan of malignant cells. Hence, in contrast togenomic instability which unambiguously favors oncogenesis,
telomere attrition may antagonize malignancy. For this reason,
we consider telomere attrition as a hallmark of aging that is
separable from genomic instability.110
Telomerase deficiency in humans is associated with prema-
ture development of diseases such as pulmonary fibrosis, aplas-
tic anemia, and dyskeratosis congenita, all of which hamper the
regenerative capacity of the affected tissues.111 Telomere short-
ening is also observed during normal aging in many different
species, including humans and mice.112 The telomeric attrition
rate is influenced by age, genetic variants, lifestyle, and social
factors; depends on the proliferative activity of the affected cells;
and predicts lifespan in a wide variety of species.112 Telomere
uncapping can also result from deficiencies in shelterins, a group
of proteins that block the DNA damage response at chromo-
some ends and modulate telomere length. Several loss-of-func-
tion models for shelterin components indicate a decline of tissue
regenerative capacity and accelerated aging, even in the pres-
ence of telomeres with a normal length.113
Genetically modified animal models have revealed causal links
between telomere attrition, cellular senescence, and organismal
aging. Mice with shortened or lengthened telomeres exhibit
decreased or increased lifespan, respectively.19 Notably, the
premature aging of telomerase-deficient mice can be reverted
when telomerase is genetically reactivated20 (Table 1). More-
over, normal aging can be delayed in mice by pharmacological
activation or systemic viral transduction of telomerase,21
whereasmice with hyperlong telomeres show increased lifespan
and metabolic health improvement22 (Table 1). Likewise, mice
engineered to maintain physiological levels of telomerase in
adult neurons preserve the survival of these cells and maintain
cognitive function in Alzheimer’s disease models23 (Table 1).
Thus, aging can be modulated by telomerase activation.
Telomerase activation to decelerate aging and treat
telomere diseases
In humans, many studies have provided evidence for causal
associations between short telomere length and age-related
diseases.114 In particular, generation ofmousemodels with short
telomeres has demonstrated that telomeric attrition is at the
origin of telomere syndromes115 and prevalent age-associated
diseases, such as pulmonary and kidney fibrosis.24,116 These
links between telomere dynamics and organismal aging
have resulted in the design of new interventions to delay
aging and age-related diseases. As an example, telomerase
activation using a gene therapy strategy has shown therapeutic
effects on mouse models of pulmonary fibrosis and aplastic
anemia.24,25
EPIGENETIC ALTERATIONS
The large variety of epigenetic changes that contribute to aging
include alterations in DNA methylation patterns, abnormal post-
translational modification of histones, aberrant chromatin
remodeling, and deregulated function of non-coding RNAs
(ncRNAs) (Figure 2B). These regulatory and often reversible
changes impact on gene expression and other cellular pro-
cesses, resulting in the development and progression of severalCell 186, January 19, 2023 251
ll
Reviewage-related human pathologies, such as cancer, neurodegener-
ation, metabolic syndrome, and bone diseases. A vast array of
enzymatic systems is involved in the generation and mainte-
nance of epigenetic patterns. These enzymes include DNA
methyltransferases, histone acetylases, deacetylases, methyl-
ases, and demethylases, as well as protein complexes
implicated in chromatin remodeling or in ncRNA synthesis and
maturation.
DNA methylation
The human DNA methylation landscape accumulates multiple
changes with the passage of time.117 Early studies described
an age-associated global hypomethylation, but further ana-
lyses revealed that specific loci, including those of several
tumor suppressor genes and Polycomb target genes, are hy-
permethylated with age. Cells from patients and mice with pro-
geroid syndromes also exhibit DNA methylation changes that
partially recapitulate those found in normal aging.118 The func-
tional consequences of most of these age-related epimutations
are uncertain, as the majority of changes affect introns and in-
tergenic regions.119
Epigenetic clocks based on DNA methylation status at
selected sites have been introduced to predict chronological
age and mortality risk as well as to evaluate interventions
that may extend human lifespan.119 This has been demon-
strated with protocols aimed at thymus regeneration, which
resulted in improved risk indices for many age-related dis-
eases and a mean epigenetic age approximately 1.5 years
less than baseline after 1 year of treatment. Moreover, predic-
tions of human morbidity and mortality showed a 2-year
decrease in epigenetic versus chronological age, which per-
sisted 6 months after discontinuing treatment.27 Likewise,
a-ketoglutarate supplementation for 7 months turned back
the epigenetic clock by 8 years.26 In summary, DNA methyl-
ation changes are associated with aging, but there is no defin-
itive evidence that they actually cause aging. Further studies
will be necessary to demonstrate that defective maintenance
of DNA methylation produces accelerated aging and that
improved fidelity in maintenance of DNA methylation patterns
extends longevity. It will be also necessary to identify the mo-
lecular drivers responsible for the modulation of changes
occurring in the aged human methylome.
Histone modifications
Global loss of histones and tissue-dependent changes in their
post-translational modifications are also closely linked to aging.
Increased histone expression extends lifespan in Drosophila,120
whereas increased histone H4K16 acetylation or H3K4 trime-
thylation and decreased levels of H3K9 or H3K27 trimethylation
are found in fibroblasts from aged individuals and progeroid
patients. These histone modifications can lead to transcrip-
tional changes, loss of cellular homeostasis, and age-associ-
ated metabolic decline.121 Of note, loss of heterochromatic
marks at telomeres has been shown to lead to telomere
lengthening.122
Histone demethylases modulate lifespan by targeting com-
ponents of key longevity routes such as the insulin/insulin
growth factor-1 (IGF-1) signaling pathway. Other histone-modi-252 Cell 186, January 19, 2023fying enzymes such as members of the SIRT family of protein
deacetylases and ADP-ribosyltransferases also contribute to
healthy aging.29 Transgenic overexpression of SIRT1 improves
genomic stability andmetabolic efficiency during aging in mice,
although without increasing longevity.29 Overexpression of
mitochondrial SIRT3 reverses the regenerative capacity lost in
aged hematopoietic stem cells (HSCs) and can mediate the
beneficial effects of dietary restriction in longevity.123 Similarly,
Sirt6 ablation in mice results in accelerated aging,124 whereas
Sirt6 overexpression extends lifespan.16 The underlying mech-
anisms derive from the fact that Sirt6 is a multitask protein with
ability to interconnect chromatin dynamics with metabolism
and DNA repair.125 Finally, Sirt7 deficiency induces global
genomic instability, metabolic dysfunctions, and premature ag-
ing.29 Together, these findings are consistent with the idea that
a decrease in deacetylase activity would result in chromatin
relaxation, increased exposure to DNA damaging agents, and
enhanced genomic instability.126 Conversely, genetic inactiva-
tion of the histone acetyltransferase KAT7 in human stem cells
decreases histone H3K14 acetylation and alleviates cell senes-
cence features.28 Moreover, intravenous injection of lentiviral
vectors encoding Cas9/sg-Kat7 ameliorates hepatocyte
senescence and liver aging and extends lifespan in both normal
and progeroid mice.28 Inhibitors of histone acetyltransferases
also ameliorate the premature aging phenotype and extend life-
span of progeroid mice, whereas histone deacetylase activa-
tors promote longevity in part via upregulation of SIRT1 activ-
ity.127 Together, these findings suggest that histone-modifiers
should be further explored as part of therapeutic strategies
against age-associated cognitive decline, although it is still un-
clear whether these interventions influence aging and longevity
through purely epigenetic mechanisms, by impinging on DNA
repair and genome stability or via transcriptional alterations
affecting metabolic or signaling pathways.Chromatin remodeling
Besides DNA- and histone-modifiers, several chromosomal
proteins and chromatin remodeling factors, such as the hetero-
chromatin protein 1a (HP1a) and Polycomb group proteins
which are implicated in genomic stability DNA repair, maymodu-
late aging.128 Alterations in these epigenetic factors result in
profound changes in chromatin architecture, including global
heterochromatin loss and redistribution, which are common
events in aged cells.
The causal relevance of these chromatin alterations in aging
has been largely studied in invertebrates in which loss-of-func-
tion mutations in HP1a decrease longevity, whereas its overex-
pression expands healthspan and lifespan129 (Table 1). Similar
studies in mammals are still limited, but most studies indicate
that heterochromatin relaxation contributes to aging and
aging-related pathologies, whereas maintenance of heterochro-
matin promotes longevity. For example, loss of PIN1—a
prolyl isomerase essential to preserve heterochromatin is asso-
ciated with premature aging and neurodegeneration in different
species from Drosophila to mammals130 (Table 1). Nevertheless,
experiments aimed at extending vertebrate longevity by gain of
function of chromatin remodeling factors are still missing.
ll
ReviewNon-coding RNAs
The large and growing universe of ncRNAs, including lncRNAs
(such as telomeric RNAs or TERRA), microRNAs (miRNAs), and
circular RNAs, has emerged as epigenetic factors with ability
to influence aging. ncRNAs modulate healthspan and lifespan
by post-transcriptional targeting of components of longevity
networks or by regulating stem cell behavior.131 A circular RNA
mediates the effect of the insulin/IGF-1 signaling pathway on
Drosophila lifespan,132 but most studies have focused on
miRNAs, and there is still debate on the extent to which other
ncRNAs may derive from transcriptional noise, with their regula-
tory roles in human physiology and pathology only circum-
scribed to few specific cases.133
Gain- and loss-of-function studies first confirmed the capacity
of several miRNAs to modulate longevity in invertebrates. Sub-
sequent studies in mice have provided causal evidence on the
functional relevance of miRNAs in aging (Table 1). For example,
miRNA-188-3p expression is upregulated in skeletal endothe-
lium during aging and contributes to vascular problems associ-
ated with the passage of time. Depletion of miR-188 in mice al-
leviates the age-related decline in beneficial bone capillary
subtypes, whereas endothelial-specific overexpression of this
miRNA decreases bone mass and delays bone regeneration.30
Conversely, depletion ofmiR-455-3p inmice exhibits deleterious
effects on mitochondrial dynamics, cognitive behavior, and
lifespan, whereas its overexpression preserves these functions
and extends lifespan.31 Overall, these findings suggest that
miRNAs may causally contribute to aging and aging-related pa-
thologies and represent potential therapeutic targets for delaying
or ameliorating these conditions.
Derepression of retrotransposons
Recent studies have unveiled the role of retrotransposons in ag-
ing of complex metazoans, including humans.134 These retro-
transposable elements are mobile genetic units that can move
from one genomic location to another, using a molecular mech-
anism that involves an RNA intermediate. Retrotransposons
consist of long interspersed nuclear elements (LINEs), which
encode the required proteins for retrotransposition, and SINEs,
which are short, non-coding RNAs that hijack the LINE protein
machinery. Retrotransposons are reactivated in senescent cells
and during lifetime and generate deleterious effects through ge-
netic and epigenetic changes or by activation of immune path-
ways triggered after identification of retrotransposon nucleic
acids as foreign DNA.134 Mechanistically, epigenetic derepres-
sion of LINE-1 RNA inhibits the epigenetic reader Suv39H1,2
resulting in global reduction of H3K9me3 and heterochoma-
tin,135 whereas reverse transcription of LINE-1 RNA results in
double-stranded cDNA that activates the cGAS/STING/inter-
feron pathway.136
Treatments with nucleoside reverse-transcriptase inhibitors
(NRTIs), which suppress or attenuate retrotransposition, extend
lifespan of Sirt6-null mice and improve healthspan, ameliorating
bone and muscle phenotypes (Table 1). Likewise, treatment of
aged wild-type mice with NRTIs reduces the levels of DNA dam-
age markers.32 Moreover, in vivo targeting of retrotransposons
with antisense oligonucleotides increases the lifespan of proge-
roid mice.135 Notably, a rare SIRT6 variant in centenarians is astronger suppressor of LINE1 retrotransposons, enhances
genome stability, and can more robustly kill cancer cells than
wild-type SIRT6.137 Collectively, these findings suggest that ret-
rotransposons causally contribute to the aging process and
that interventions that oppose retrotransposon activity might
improve healthy longevity. Further clinical studies in aged popu-
lations with drugs targeting the different functions of retrotrans-
posons may delineate novel intervention strategies on aging and
aging-related pathologies.
Gene expression changes
Themechanisms underlying the effects of all the above epigenetic
factors converge at the modulation of gene expression levels. Ag-
ing causes an increase of the transcriptional noise and an aberrant
production and maturation of many mRNAs.138,139 Microarray-
based comparisons of young and old tissues from human and
other species have identified age-related transcriptional signa-
tures that result from epigenetic changes occurring during aging.
Environmental exposures also cause alterations ingene regulation
via DNA methylation alterations and histone modifications and
promote aging-related epigenetic changes including the acceler-
ation of epigenetic clocks.140
Single-cell transcriptomic and plasma proteomics of multiple
cell types and organs at several ages across the entire mouse life-
span have unveiled remarkable gene expression shifts during
aging.138 These changes specially affect certain biological pro-
cesses, such as inflammation, protein folding, extracellular matrix
(ECM) regulation, and mitochondrial function, which are widely
deregulated in aging.141 The common expression patterns
observed during aging in different tissuesmay help to guide future
interventions aimed at improving healthspan and lifespan (Ta-
ble 1). Likewise, the observed decline in transcriptional and
post-transcriptional efficiency and fidelity in the course of aging,
and its negative consequences on the proteome health may
also open new opportunities for prolongevity strategies.139
LOSS OF PROTEOSTASIS
Aging and several age-related morbidities, such as amyotrophic
lateral sclerosis (ALS), Alzheimer’s disease, Parkinson’s disease,
and cataract, are associated with impaired protein homeostasis
or proteostasis, leading to the accumulation of misfolded,
oxidized, glycated, or ubiquitinylated proteins that often form ag-
gregates as intracellular inclusion bodies or extracellular amyloid
plaques.142
Proteostasis collapse
Intracellular proteostasis can be disrupted due to the enhanced
production of erroneously translated, misfolded or incomplete
proteins (Figure 3). Geneticmanipulation of the ribosomal protein
RPS23 to improve the accuracy of RNA-to-protein translation
extends lifespan in Schizosaccharomyces pombe,Caenorhabdi-
tis elegans, andDrosophilamelanogaster,143 whereas amutation
in RPS9 that favors error-prone translation causes premature
aging in mice.144 Another mechanism driving the collapse of
the proteostasis network resides in slowed translation elonga-
tion and cumulative oxidative damage of proteins, increasingly
distracting the chaperones from folding healthy proteinsCell 186, January 19, 2023 253
Figure 3. Loss of protein and organellar
turnover
Loss of proteostasis and disabled macro-
autophagy are characterized by a deviation from
the young equilibrium state in which an accumu-
lation of waste products results from a variety of
age-associated alterations and simultaneously
waste removal is compromised through a variety
of mechanisms. The functional consequences of
these alterations are listed. Some strategies for
reestablishing proteostasis and autophagy are
exemplified on the left and on the right.
ll
Reviewrequired for cellular fitness.145 In addition, numerous age-related
neurodegenerative diseases including ALS and Alzheimer can
be caused by mutations in proteins that render them intrinsically
prone to misfolding and aggregation, hence saturating the
mechanisms of protein repair, removal, and turnover that are
required for maintenance of the healthy state.146
The proteostasis network also collapses whenmechanisms as-
suring quality control fail, for instance, due to reduced function of
the unfolded protein response (UPR) in the endoplasmic reticulum
(ER),147 when stabilization of correctly folded proteins is compro-
mised or when mechanisms for the degradation of proteins by
the proteasome or the lysosome become insufficient (Figure 3).
Reduction of proteasome activity has been observed in aged or-
gans including the brain of the short-lived fish Nothobranchius
furzeri.148 Moreover, some mono-ubiquitinylated proteins accu-
mulate in aging tissues from flies, mice, monkeys, and humans,
as documented for histone 2A.149
The degradation of proteins by the lysosomecan be achieved in
a specific fashion, through chaperone-mediated autophagy
(CMA), wherein proteins exposing a pentapeptide motif resem-
bling KFERQ first bind to heat shock protein HSC70 and then to
lysosome-associated membrane protein type 2A (LAMP2A),
which facilitates the translocation of the client protein into the
lumen of the lysosome.150 Hepatic LAMP2A expression declines
withage inmice, and its transgenic re-expression reduces liver ag-
ing.151 Protein aggregates can also be removed bymacroautoph-254 Cell 186, January 19, 2023agy upon their inclusion in two-membrane
vesicles, the autophagosomes, for their
later fusionwith lysosomes.152 Since auto-
phagosomes can envelop non-proteina-
ceous structures, this process will be dis-
cussed separately from proteostasis in
the next hallmark section (disabledmacro-
autophagy). Nonetheless, stimulation of
autophagy constitutes a valid strategy for
the elimination of intracellular protein ag-
gregates.153
Proteostasis, aging, and longevity
Perturbation of general proteostasis ac-
celerates aging. For example, feeding
D. melanogaster with advanced glyca-
tion end products (AGEs) or lipofuscin
(an aggregate of covalently cross-linked
proteins, sugars, and lipids) causes theaccumulation of AGE-modified and carbonylated proteins
with a reduction of healthspan and lifespan that is further
accentuated upon knockdown of the lysosomal protease
cathepsin D.154 Loss of the protease ZMPSTE24 abolishes
the normal proteolytic maturation of prelamin A and causes a
progeroid syndrome in mice, phenocopying that observed in
humans with loss-of-function mutations of ZMPSTE24.18 In
mice, knockout of LAMP2A (essential for CMA) in neurons pro-
foundly affects the proteome, yielding similar changes as found
in Alzheimer patients. Indeed, inhibition of CMA in mice exacer-
bates experimental Alzheimer’s disease, whereas its stimula-
tion by a pharmacological CMA activator attenuates the pa-
thology.155
Experimental amelioration of proteostasis can retard the
aging process (Table 1). Intranasal application of recombinant
humanHSP70 protein tomice enhances proteasome activity, re-
duces brain lipofuscin levels, enhances cognitive functions, and
extends lifespan.35 Similarly, administration of the chemical
chaperone 4-phenylbutyrate to aged mice reduces ER stress
in the brain and improves cognition.36 In nematodes and flies,
transfection-enforced overexpression of isolated proteasome
subunits improves proteostasis and increases lifespan.156
In mice, stimulation of CMA by transgenic expression of
LAMP2a HSCs improve the survival of the targeted cell popula-
tions,33 in line with the observation that pharmacological
enhancement of CMA attenuates Alzheimer’s pathology and
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Reviewarteriosclerosis.155,34 Hence, activation of CMAmay constitute a
valid strategy for delaying the aging process.
A phase 3 clinical trial has revealed that in patients with recent
ALS diagnosis, administration of the antihypertensive guana-
benz inhibits progression to the life-threatening bulbar stage.37
Guanabenz may act to stimulate the phosphorylation (or to
inhibit the dephosphorylation) of eukaryotic translation initiation
factor 2a (eIF2a), which occurs in the context of the ‘‘integrated
stress response (ISR)’’ as part of the UPR,157 although it remains
under debate to what extent the actions of guanabenz are
mediated by the stimulation of the ISR.158 Importantly, eIF2a
phosphorylation causes a switch from 50 cap-dependent to 50
cap-independent RNA translation, knowing that the latter is
enhanced by several longevity-extending manipulations.159
Moreover, eIF2a phosphorylation is essential for the induction
of stress granules, which are required for longevity extension
by dietary restriction in worms.160 Finally, eIF2a phosphorylation
is indispensable for the induction of autophagy,161 which is ama-
jor anti-aging mechanism (see below), suggesting a crosstalk
between UPR and autophagy in prolongevity pathways. Future
studies must determine whether the capacity of guanabenz to
attenuate neurodegeneration is mediated via ISR stimulation or
alternative mechanisms. Indeed, it has been proposed that
inhibitors of ISRmight be also used for the treatment of neurode-
generative diseases.162
DISABLED MACROAUTOPHAGY
Macroautophagy (that we will refer to as ‘‘autophagy’’) involves
the sequestration of cytoplasmic material in two-membrane ves-
icles, the autophagosomes, which later fuse with lysosomes for
the digestion of luminal content.152 Thus, autophagy is not only
involved in proteostasis but also affects non-proteinaceous
macromolecules (such as ectopic cytosolic DNA, lipid vesicles,
and glycogen) and entire organelles (including dysfunctional
mitochondria targeted by ‘‘mitophagy,’’ and other organelles
leading to ‘‘lysophagy,’’ ‘‘reticulophagy,’’ or ‘‘pexophagy’’), as
well as invading pathogens (‘‘xenophagy’’).152 An age-related
decline in autophagy constitutes one of the most important
mechanisms of reduced organelle turnover, justifying its discus-
sion as a new hallmark of aging. As a note of caution, genes
and proteins that participate in the autophagic process are
also involved in alternative degradation processes such as
LC3-associated phagocytosis of extracellular material,163 and
the extrusion of intracellular waste (e.g., dysfunctional mitochon-
dria) in the form of exospheres for their subsequent removal by
macrophages.164 That said, there is strong evidence that the
core process of autophagy is relevant to aging (Figure 3).
Accelerated aging due to autophagy inhibition
In humans, the expression of autophagy-related genes, such as
ATG5, ATG7, and BECN1, declines with age.165 CD4+ T lympho-
cytes isolated from the offspring of parents with exceptional
longevity show enhanced autophagic activity compared with
age-matched controls.166 Decreased autophagy in circulating B
andT lymphocytes fromaging donors is accompanied bya reduc-
tion of the pro-autophagic metabolite spermidine.167,168 Similarly,
in rodents, a progressive deterioration of autophagy has beendescribed for some organs, pleading in favor of the idea that auto-
phagicflux iscompromisedwithage.Reductionofautophagicflux
may participate in the accumulation of protein aggregates and
dysfunctional organelles, reduced elimination of pathogens, and
enhanced inflammation because autophagy eliminates proteins
involved in inflammasome and their upstream triggers.169
Genetic inhibition of autophagy accelerates the aging process
in model organisms. This process is partially reversible, as illus-
trated in mice in which Atg5 is downregulated by a doxycycline-
inducible shRNA. Atg5 knockdown causes the premature
degeneration and senescence of multiple organ systems leading
to premature death.170 Upon withdrawal of doxycycline, auto-
phagy restoration is accompanied by attenuated systemic
inflammation and segmental reduction of aging. Of note, in this
model, the transient inhibition of autophagy is followed by a ma-
jor increase in the incidence of malignancies. Hence, autophagy
apparently acts as a tumor-suppressive mechanism, which may
involve cell-autonomous processes and cancer immunosurveil-
lance.153 In patients, loss-of-function mutations of genes that
regulate or execute autophagy have been causally linked to a
broad spectrum of cardiovascular, infectious, neurodegenera-
tive, metabolic, musculoskeletal, ocular, and pulmonary disor-
ders, many of which resemble to premature aging at the histo-
pathological and functional levels.152,153
Autophagy stimulation for decelerated aging
There is ample evidence that stimulation of autophagic flux in-
creases healthspan and lifespan in model organisms (Table 1).
For example, increasing autophagy solely in the enterocytes of
the intestine increases Drosophila lifespan.120 In mice, trans-
genic overexpression of Atg5 under the control of a ubiquitously
expressed promoter is sufficient to extend lifespan and to
improve metabolic health and motor function.38 Moreover,
knockin mutation of beclin 1 (Becn1F121A/F121A) to reduce its inhi-
bition by Bcl-2 causes an increase in autophagic flux, as well as
an extension of lifespan. This effect is coupled to a reduction of
age-associated pathologies and spontaneous tumorigenesis,39
as well as to increased neurogenesis.40
Oral supplementation of spermidine to mice induces auto-
phagy in multiple organs and extends longevity by up to 25%,
accompanied by reduced cardiac aging. This latter effect is
lost upon cardiomyocyte-specific knockout of Atg7, suggesting
that it relies on autophagy.41 Mechanistically, the pro-autopha-
gic effects of spermidine have been linked to an inhibition of
the acetyl transferase EP300 (resulting in reduced acetylation
of several core autophagy proteins)171 or to the hypusination of
eIF5A, which is essential for the synthesis of the autophagy tran-
scription factor TFEB.167 Among these factors, EP300 is the
target of the longevity-enhancing drugs nordihydroguairaretic
acid43 and salicylate.42 Pharmacological inhibition of EP300
with C646 mimics the stimulatory effects of spermidine on auto-
phagy and cancer immunosurveillance.172 When circulating B
lymphocytes or CD8+ T cells from aged human donors are
cultured in the presence of spermidine, the cells recover
juvenile levels of TFEB and eIF5A, coupled to a normalization
of autophagic flux.167,168 Moreover, in Drosophila, hypusination
deficiency due to a heterozygous mutation or knockdown of
deoxyhypusine synthase abolished lifespan extension byCell 186, January 19, 2023 255
ll
Reviewspermidine supplementation.173 Deoxyhypusine synthase defi-
ciency in murine T cells triggers severe intestinal inflammation
coupled to epigenetic remodeling and rewiring of the tricarbox-
ylic acid cycle,174 whereas spermidine treatment of wild-type
mice protects against colitis and colon carcinogenesis.175
Hence, both EP300 inhibition and eIF5A hypusination appear
plausible targets to explain the in vivo effects of spermidine.
Pharmacological agents that induce mitophagy and have a
positive impact on murine healthspan include NAD+ precursors
(such as nicotinamide, nicotinamide mononucleotide, and nico-
tinamide riboside)176 and urolithin A.177 Clinical trials have
demonstrated the efficacy of NAD+ precursors in the chemopre-
vention of non-melanoma skin cancer,46 in reversing insulin
resistance in prediabetic women,44 and in reducing neuroinflam-
mation in patients with Parkinson’s disease.45 Moreover, a
phase 3 trial has revealed the capacity of urolithin A to improve
muscle strength and to reduce C-reactive protein (CRP).47
DEREGULATED NUTRIENT-SENSING
The nutrient-sensing network is highly conserved in evolution. It
includes extracellular ligands, such as insulins and IGFs, the
receptor tyrosine kinases with which they interact, as well as
intracellular signaling cascades. These cascades involve the
PI3K-AKT and the Ras-MEK-ERK pathways, as well as tran-
scription factors, including FOXOs and E26 factors, which
transactivate genes involved in diverse cellular processes. The
mechanistic target of rapamycin (MTOR) complex-1 (MTORC1)
responds to nutrients, including glucose and amino acids, and
to stressors such as hypoxia and low energy to modulate the
activity of numerous proteins including transcription factors
such as SREBP and TFEB. This network is a central regulator
of cellular activity, including autophagy, mRNA and ribosome
biogenesis, protein synthesis, glucose, nucleotide and lipid
metabolism, mitochondrial biogenesis, and proteasomal activ-
ity. Network activity responds to nutrition and stress status by
activating anabolism if nutrients are present and stress is low
or by inducing cellular defense pathways in response to stress
and nutrient-shortage. There is extensive intracellular crosstalk
and feedback within the network, and between it and other intra-
cellular signaling pathways. Genetically reduced activity of com-
ponents of the nutrient-sensing network can increase lifespan
and healthspan in diverse animal models178,179 (Table 1). More-
over, genetic association studies in humans have implicated the
FOXO3 transcription factor180 and genetic variants encoding
components of the network in human longevity.178 Epigenetic
age is also associated with nutrient-sensing in human cells.181
In youth, activity of this signaling network thus functions to
promote beneficial anabolic processes, but during adulthood,
it acquires pro-aging properties (Figure 4).
The somatotrophic axis—the first one historically implicated in
the control of aging—is a growth-stimulatory cascade that, at its
apex, involves growth hormone (GH) produced by the hypophy-
sis. GH acts on the GH receptor of hepatocytes to stimulate the
secretion of IGFs, in particular IGF1, which promotes growth and
development via the IGF1R to stimulate trophic signals through
activation of PI3K-AKT and the MTORC1 network.182 In multiple
model organisms, spontaneous or engineered mutations of this256 Cell 186, January 19, 2023pathway enhance lifespan and retard facets of age-associated
deterioration (Table 1). Innate defects in the somatotrophic axis
cause dwarfism, but inhibition of this axis from early adulthood
has beneficial effects on organismal health (Figure 4).
Another signaling pathway involved in nutrient-sensing
relies on the receptor tyrosine kinase ALK (Figure 4), which, in
mice, is induced in the hypothalamus by feeding183 and re-
sponds to the ligands augmentor a and b (Auga and Augb).184
In Drosophila, knockdown of ALK decreases triglyceride levels
and the expression of several insulin-like peptides, whereas ge-
netic or pharmacological inhibition of ALK extends healthspan
and lifespan, mostly in females.183 In mice, body-wide or hypo-
thalamus-specific deletion of ALK, as well as double knockout of
Auga and Augb, promotes resistance against diet-induced
obesity, and in humans, a loss-of-function mutation of ALK is
associated with leanness.183,184 Hence, this pathway may offer
additional targets for interventions on metabolic aging.
Drugs targeting diseases such as cancer and metabolic dis-
ease often engage the nutrient-sensing network, thus such
drugs are candidates for repurposing as geroprotectors. Rapa-
mycin and rapalogs, which disrupt the MTORC1 complex,
have proved to extend lifespan in model organisms even with
treatment starting late in adulthood.185 In mice, rapamycin can
increase diverse aspects of health, although it exacerbates
some age-related traits such as cataract, and it is protective in
models of neurodegenerative and other age-related diseases.
Elderly humans are susceptible to viral respiratory infections.
Pre-treatment with MTORC1 inhibitors increased the immune
response of elderly volunteers to immunization against influ-
enza186 and reduced viral respiratory infections in the ensuing
winter,187 thus pointing to a potential strategy for reverting
age-related immunosenescence.
Mechanisms
In humans, IGF1 peaks during the second decade of life but
declines with aging. Inhibition of the GH/IGF1 pathway in adult
or late life extends lifespan in model organisms, including
mice.48 Inhibition of cardiac IGF1R by expression of a dominant
negative p110a isoform of PI3K increases maximum lifespan of
male mice and improves heart function in aged mice.188 More-
over, enzymatic inhibition of IGF1R with tyrosine kinase
inhibitors improves anticancer immunosurveillance requiring
autophagy induction in malignant cells.189 Long-term administra-
tion of an anti-IGF1R antibody enhances the longevity of female
(but not male) mice, although reducing inflammation and tumor
development. These findings suggest that the IGF1/IGF1R
signaling axis may constitute a target for anti-aging interventions.
In favor for this conjecture, in elderly women (R95 years), as well
as in a mixed population of older adults (mean age 76 years), low
IGF1 levels correlatewith a lowprobability of cognitive impairment
and death.190 Moreover, in a large cohort from the UK Biobank,
significant positive correlations were noted between the hazard
associated with high IGF-1 and age for dementia, diabetes,
vascular disease, osteoporosis, and overall mortality.191 In cente-
narians, the concentrations of IGF1BP2 and IGFBP6 are
elevated.192 Future will tell whether yet-to-be-developed anti-
bodies or small molecules that selectively inhibit IGF1R signaling
without affecting other receptor tyrosine kinases (and in particular
Figure 4. Metabolic alterations
A simplified version of intertwined trophic pathways is shown for deregulated nutrient-sensingwith their possible countermeasures to restore nutrient-sensing. Of
note, the reduced activity of the nutrient-sensing network influences numerous processes beyond metabolism modulation during aging, including resistance to
diverse stressors, activation of repair mechanisms, autophagy stimulation, or inflammation control. Similarly, for mitochondrial dysfunction, a series of age-
associated alterations with their possible antidotes are listed. The functional consequences of age-associated metabolic alterations, some of which are relevant
to other hallmarks of aging, are exemplified in the lower part of the graph.
ll
Reviewthe insulin receptor) might be used for the pulsatile inhibition of the
somatotropic axis to achieve health benefits with acceptable side
effects.
Effects of nutrition
Diet is one of the most practical targets for interventions into hu-
man aging. Mechanistically, overnutrition: (1) triggers intracel-
lular nutrient sensors, such as MTORC1 (activated by leucine
and other amino acids), and the acetyltransferase EP300 (acti-
vated by acetyl coenzyme A); (2) inhibits sensors that detect
nutrient scarcity, such as AMP-activated kinase (AMPK) and
the deacetylases SIRT1 and SIRT3 (which respond to NAD+);
and (3) abolishes catabolic reactions (glycogenolysis, proteoly-
sis for gluconeogenesis, and lipolysis coupled to ketogenesis)
with consequent suppression of adaptive cellular stress re-
sponses, including autophagy, antioxidant defense, and DNA
repair. Conversely, fasting and dietary restriction inhibit
MTORC1 and EP300; activate AMPK, SIRT1, and SIRT3; and
stimulate adaptive cellular stress responses as they suppress
the somatotrophic axis and extend longevity in multiple model
organisms including primates.193
Nutrient sensors constitute targets for potential longevity
drugs (Figure 4), but health benefits and extended lifespan might
also be achieved by dietary restrictions. Mechanistically, this ispossible via reduction of overall caloric intake, manipulation of
the dietary composition,194,195 or time-restricted feeding.196 Di-
etary restriction regimens are particularly successful in extend-
ing lifespan in male C57BL/6Jmice, if the animals are completely
deprived from nutrients during daytime.49 However, dietary re-
striction regimens do not extend lifespan in all mouse strains,
supporting the contention that they must be adapted to the
genetic makeup of each individual.197 In humans, clinical assays
based on dietary restriction are complicated by poor compli-
ance, yet suggest positive effects on immunity and inflam-
mation.50
Intermittent fasting (e.g., 1 day without nutrients, followed by
1 day of ad libitum feeding) can avoid long-term weight loss
induced by caloric restriction, yet increases lifespan in mice195
and improves biomarkers of health in clinical trials.198,199 Life
time extension of a similar intermittent fasting regimen in flies
has been attributed to the nighttime-specific upregulation of
autophagy-stimulatory genes,200 but this has not yet been inves-
tigated in mammals. Rapamycin-induced longevity extension
(which in flies partially depends on autophagy induction) can
be obtained by constant-long term exposure, as well as by inter-
mittent regimens,201 suggesting that pulsatile inhibition of this
axis is sufficient to obtain the benefits of lifespan extension.
The optimal interval for such intermittent treatments has notCell 186, January 19, 2023 257
ll
Reviewyet been determined for clinical use, although partial caloric re-
striction for 4–7 days every 3–4 weeks may be sufficient to
improve metabolic syndrome and anticancer immunosur-
veillance.202
Another potentially beneficial regimen is ketogenic diet, which
is a low-carbohydrate, high-fat, and adequate protein diet. Both
fasting and ketogenic diet increase the production of ketone
bodies (in particular 3-hydroxybutyrate), which are synthesized
from acetyl coenzyme A in the liver in an autophagy-dependent
fashion, can reach millimolar concentrations in the plasma and
replace glucose as an essential fuel, for instance, for the mainte-
nance of brain function.203 Permanent but not cyclic administra-
tion of 3-hydroxybutyrate in the drinking water increases lifespan
and healthspan in mice.51 This strongly suggests that this ketone
body mediates some of the beneficial effects of ketogenic diet.
Mechanistically, 3-hydroxybutyrate induces vasodilatation and
activates immune responses acting on GTP protein coupled re-
ceptor 109A,203 whereas it directly inhibits the NLRP3 inflamma-
some,204 indicating a potential pleiotropic mode of action.
MITOCHONDRIAL DYSFUNCTION
Mitochondria are not only the powerhouses of the cell but also
constitute latent triggers of inflammation (when reactive oxygen
species [ROS] or mtDNA leak out of the organelle causing activa-
tion of inflammasomes or cytosolic DNA sensors, respectively)
and cell death (when activators of caspases, nucleases, or other
lethal enzymes are released from the intermembrane space).146
With aging, mitochondrial function deteriorates due to multiple in-
tertwined mechanisms including the accumulation of mtDNAmu-
tations, deficient proteostasis leading to the destabilization of res-
piratory chain complexes, reduced turnover of the organelle, and
changes in mitochondrial dynamics. This situation compromises
the contribution of mitochondria to cellular bioenergetics, en-
hances the production of ROS, andmay trigger accidental perme-
abilization ofmitochondrial membranes causing inflammation and
cell death.182 Logically, the function of mitochondria is primordial
for the maintenance of health, and its progressive deterioration
contributes to the aging phenotype (Figure 4).
Mitochondrial function and longevity
Healthspan-extending interventions can stimulate the function of
mitochondria. For instance, placebo-controlled trials have re-
vealed positive effects of L-carnitine supplementation on both
pre-frail subjects and elderly men57 (Table 1). The effect is
possibly mediated by counteracting age-related declining
L-carnitine levels which may limit fatty acid oxidation by mito-
chondria.205 Paradoxically, in model organisms, lifespan can
be improved by compromising mitochondrial function, which in-
duces a hormetic response (‘‘mitohormesis’’), provided that this
inhibition is partial and occurs early during development. In
C. elegans, partial inhibition of mitochondrial protein synthesis
or import enhances lifespan through a mechanism involving
the mitochondrial UPR (UPRmt).206 In Drosophila, muscle-spe-
cific knockdown of complex I subunit NDUFS1/ND75 extends
longevity in an UPRmt-dependent fashion.207 Mild inhibition of
mitochondrial ATP synthesis with TPP-thiazole can improve
metabolic health in aging mice, reducing visceral fat and258 Cell 186, January 19, 2023improving glucose tolerance, mitochondrial quality, and oxida-
tive metabolism.52 Partial uncoupling of hepatic mitochondria
by means of a controlled release mitochondrial protonophore
(CRMP) also reverses age-related metabolic syndrome in mice
with high-fat diet-induced obesity.53 In non-human primate
models including spontaneously obese rhesus macaques and
high-fat, high-fructose-fed cynomolgus macaques, CRMP re-
verses signs of metabolic syndrome and improves fatty acid
oxidation.55 These effects are coupled to a reduction of hepatic
acetyl-coenzyme A levels, a phenomenon known to stimulate
autophagy.208 Protonophores inducemitophagy,209 whichmight
explain their positive effects on metabolism as well. Metformin,
an antidiabetic considered as a weak complex I inhibitor, has
been discussed as a possible anti-aging drug.210 However,
thus far, there is no evidence that challenging mitochondria
can increase healthspan or lifespan in humans.
Increased mitochondrial membrane permeability (MMP)
due to the absence of serum/glucocorticoid regulated kinase-1
decreases lifespan, which is further compromised when
autophagy is enhanced but normalized when autophagy is in-
hibited by knockdown of essential autophagy-relevant genes in
C. elegans.211 Hence, MMPmay constitute a life-threatening con-
dition that is aggravated by autophagy. A modified tetrapeptide,
elamipretide, has been developed to target cardiolipin in the inner
mitochondrial membrane (IMM) and then turned out to bind to the
IMM protein adenine-nucleotide translocator-1 to inhibit the
mitochondria permeability transition, which is one particular
mechanism leading to MMP.54 Elamipretide has positive effects
on multiple aging-related phenotypes in mice and has yielded
positive results in a clinical trial on patients with Barth syndrome56
(Table 1). It will be important to understand whether elamipretide
can be advantageously combined with other lifespan-enhancing
drugs including autophagy enhancers. In addition to these works,
there are also several preclinical and clinical studies evaluating the
potential beneficial effects of the antioxidant lipophilic cations Mi-
toQ and SkQ1.212 Further researchwill define the utility of all these
compounds in the context of other interventions aimed at amelio-
rating age-associated mitochondrial dysfunctions.
Mitochondrial microproteins and aging
Plasma levels of the microprotein humanin, which is encoded by
mtDNA, decline with age. However, centenarians and their
offspring exhibit high levels of humanin.213 Notably, humanin
levels negatively correlate with IGF1 in humans and treatment
of patients with GH-insufficiency, with GH or IGF1, reduces
circulating humanin.214 Transgenic expression of humanin in
C. elegans extends longevity through autophagy induction, and
treatment of middle-aged mice with the humanin analog HNG
improvesmetabolic healthspan and reduces systemic inflamma-
tion.213 Another mtDNA-encoded microprotein, MOTS-c,
declineswith age but can be induced by exercise.215MOTS-c fa-
vors the production of the metabolite 5-aminoimidazole-4-
carboxamide-1-beta-4-ribofuranoside (AICAR), which acts as
an endogenous AMPK agonist, thereby preventing age-depen-
dent and high-fat-diet-induced insulin resistance, as well as
diet-induced obesity.215 Hence, mitochondrial microproteins
emerge as potential anti-aging factors that link organellar func-
tion to organismal homeostasis.
ll
ReviewCELLULAR SENESCENCE
Cellular senescence is a response elicited by acute or chronic
damage.216 In humans, senescent cells accumulate inmultiple tis-
sues at different rates, from 2- to 20-fold when comparing young
(<35years) to old (>65years) healthy donors,217mainly affecting fi-
broblasts, endothelial cells, and immune cells, although all cell
types can undergo senescence during aging,218 a process that is
triggered at least in part by telomere shortening with aging.109 In
fact, even post-mitotic or slowly proliferating tissues, such as the
brain or the heart,may harbor senescent cells.219 In addition, focal
or tissue-specific accumulation of senescent cells occurs inmany
diseases.220 The most compelling evidence for the causal role of
cellular senescence in aging is that continued genetic or pharma-
cological elimination of senescent cells extends the healthspan
and longevity of naturally agedmice.59 Also, genetic or pharmaco-
logical elimination of senescent cells is therapeutic in many dis-
eases modeled in mice,221 and at least 3 clinical trials have been
completed and 15 clinical trials are ongoing or planned to target
senescence for a variety of indications.222
The types of damage that trigger primary senescence include
oncogenic signaling, genotoxic damage, critically short telo-
meres, mitochondrial damage, viral or bacterial infection, oxida-
tive damage, nutrient imbalance, and mechanical stress.216 In
addition, secondary or paracrine senescence can be triggered
by extracellular mediators of inflammation and fibrosis including
CCL2, IL-1b, IL-6, IL-8, and TGF-b.223 There is evidence suggest-
ing that primary and secondary senescence differ in relevant bio-
logical aspects, but the molecular basis of this distinction remains
elusive. Historically, the most salient feature of cellular senes-
cence is a stable proliferative arrest mediated by the activation
of the tumor suppressors TP53 and CDKN2A/p16, and their
downstream effectors CDKN1A/p21 and retinoblastoma-1 (RB1)
family proteins, respectively. Together, these proteins inhibit cy-
clin-dependent kinases (CDKs) and transcriptional activators
(E2F family) that drive the cell cycle.216 Another important event
during senescence is the depletion of lamin B1 from the nuclear
envelope. This results in the loss of lamin-associated heterochro-
matin and de novo formation of heterochromatin rich inH3K9me3,
a process that can be visualized as HP1a foci or senescence-
associated heterochromatin foci (SAHFs).224 The net result is a
long-term and viable proliferative arrest with a low rate of sponta-
neous escape. Depending on their molecular makeup, cancer
cells exposed to genotoxic therapy may undergo a canonical
senescence response with a highly stable cell cycle arrest or
can undergo a senescence-like response with a highly reversible
arrest or can even completely bypass senescence.225 Of note,
senescence also plays a role during embryogenesis in the pro-
grammed elimination of specific cells and structures.226
Senescence and human diseases
Cellular senescence is implicated in multiple non-proliferative
diseases, including lung fibrosis, kidney diseases, liver steatosis,
obesity-associated metabolic syndrome, type I and II diabetes,
atherosclerosis, as well as Alzheimer’s and Parkinson’s dis-
eases.220 The pathogenic role of cellular senescence in these
diseases can be explained by the senescence-associated
secretory phenotype (SASP). SASP results from three featuresof senescent cells: (1) the transcriptional derepression of endog-
enous retroviruses, most notably LINE-1, which causes cytosolic
leakage of double-stranded DNA and activates the cGAS/STING
and TLR pathways;136 (2) the mitochondrial overproduction of
ROS; and (3) the perturbation of the autophagy-lysosomal sys-
tem leading to an expansion of lysosomal content that facilitates
the histochemical detection of lysosomal senescence-associ-
ated beta-galactosidase (SABG).227
SASP is highly heterogeneous, depending on the cell type-spe-
cific activation of innate immunity signaling pathways (cGAS/
STING, TLRs, and NLRPs), mTORC1, and transcription factors
(NF-kB, CBPs, GATA4, and others). SASP usually has simulta-
neousandpartially conflictingconsequenceson themicroenviron-
ment: (1) to recruit and activate immunecells through the secretion
of chemokines (CCL2, CXCL2, and CXCL3) and cytokines (IL-1b,
IL-2, IL-6, and IL-8); (2) to suppress the immune system through
the secretion of TGF-b; (3) to trigger fibroblast activation and
collagen deposition through pro-fibrotic factors (TGF-b, IL-11,
and PAI1); (4) to remodel the ECM through the secretion of matrix
metalloproteases; (5) to trigger the activation and proliferation of
progenitor cells through the secretion of growth factors (EGF and
PDGF); and (6) to triggerparacrine senescence inneighboringcells
(TGF-b, TNF-a, and IL-8). Inmany diseases, the net effect of SASP
is chronic inflammation and progressive fibrosis.228
Although there is not a single unequivocal marker of cellular
senescence, this process can be identified by the co-existence
of a combination of features that, together, are specific and pro-
vide a molecular definition to the phenomenon:216 (1) lysosomal
expansion, detectable by SABG; (2) upregulation of CDK inhibi-
tors, particularly p16 and/or p21; (3) loss of LMNB1 from the
nuclear envelope; (4) loss of the chromatin component
HMGB1 from the nucleus and its extracellular release as an alar-
min; (5) heterochromatic foci, visualized as HP1g nuclear foci or
SAHFs; (6) high levels of ROS; (7) exacerbated DNA damage,
visualized as gH2AX nuclear foci; and (8) high levels of SASP fac-
tors, notably IL-6, TGF-b, PAI1, and others.
Given the association between cellular senescence and multi-
ple pathologies, the question arises about the biological purpose
of such a cellular response. Cellular senescence is a potent tu-
mor suppressor mechanism, but mounting evidence has linked
cellular senescence to tissue repair processes in which senes-
cent cells promote localized fibrosis and the recruitment of im-
mune cells that then remove damaged and senescent cells. In
this regard, tissue repair can be considered a two-step process:
cellular senescence followed by immune recruitment and im-
mune clearance of senescence (Figure 5A). In this scenario,
senescence is a temporally restricted response that programs
its self-elimination with a beneficial outcome.229 The patholog-
ical consequences of senescence only become visible when
the second step of immune clearance is not achieved, and the
accumulation of senescent cells and the SASP effects on the tis-
sue microenvironment eventually result in fibrosis.
Senolytics
The strong association between cellular senescence and multiple
pathologieshasspurred thesearch for small chemical compounds
that selectively kill senescent cells and that are referred to as ‘‘se-
nolytics.230’’ Of note, senolysis (elimination of senescent cells) isCell 186, January 19, 2023 259
Figure 5. Cellular senescence and stem cell
exhaustion
(A) Cellular senescence usually promotes tissue
repair after injury and protects the organism from
oncogenic damage. This is achieved in two steps:
(1) establishment of senescence and (2) recruit-
ment of immune cells that will eliminate the se-
nescent cells, thereby promoting tissue repair. If
any of these steps fails, the organism is prone to
develop diseases.
(B) Stem cell exhaustion results from the loss of
cellular plasticity required for tissue repair. Tissue
repair requires a modified microenvironment
through the secretion of cytokines (in part due to
the senescence-associated secretory response),
growth factors and modulators of the extracellular
matrix (ECM) that favors the de-differentiation and
plasticity of cells from different tissue compart-
ments. These injury-induced plastic cells may
acquire multipotent progenitor function. Transient
expression of OSKM factors represses the tran-
scription of cell identity programs causing global
de-differentiation (OSKMon) and the acquisition of
plasticity. For rejuvenation, the process must be
interrupted at this point (OSKMoff) to allow cells to
re-differentiate and to restore their original cell
identities.
ll
Reviewvery different from the cancellation of the senescence response,
which can result, for example, frommutation of p16 or p21. Senol-
ysisdoesnotprevent theexecutionof senescencebut rather reca-
pitulates the natural immune clearance of senescent cells
(Figure 5A). In support of this, mice subjected to long-term ge-
netic-induced or pharmacologically induced senolysis present
extended longevity without increased cancer incidence or signs
of defective tissue repair.59,58
The number of senolytic therapies is still limited, but some have
been extensively used in preclinical models of disease, as exem-
plified by navitoclax, dual treatment with dasatinib and quercetin
(D/Q), fisetin, cardiac glycosides, and others.221 The survival and
apoptotic resistance of senescent cells strongly depends on the
BCL2 family of proteins, specially BCLXL, but also BCL2 and
BCLW. This renders senescent cells highly vulnerable to navito-
clax, which targets these three proteins.231 Navitoclax has been
evaluated in clinical trials for antitumor activity and it is expected
that this drug (or derivatives lacking toxicity on platelets) will enter
clinical trials for senescence-associated diseases.232 Other po-
tential senolytic treatments such as D/Q230 and fisetin60 are
approved for human use and are being tested in various clinical tri-
als for multiple indications. The mechanistic basis for their action
remains unclear. Dasatinib is a promiscuous kinase inhibitor, and260 Cell 186, January 19, 2023quercetin and fisetin are natural flavonoids
with multiple targets. D/Q has been tested
in clinical trials with promising results in
the case of lung and kidney fibrosis.62,61
Cardiac glycosides inhibit the plasma
membrane Na+/K+-ATPase present in all
cells causing a cationic imbalance and
lowering the intracellular pH.233 The
mechanism of senolysis by cardiac glyco-
sides is likely connected to the vulnera-
bility of senescent cells to low intracellularpH. Thus, chemical inhibition of glutaminase deprives cells of a
mechanism to counteract low pH and results in senolysis.234 All
the above-discussed senolytic compounds exert therapeutic ac-
tivity in a wide range of murine disease models associated with
senescence. Senolysis can also be achieved by immunological
approaches that target proteins appearing on the surface of se-
nescent cells. In particular, antibodies directed against the glyco-
protein NMB (GPNMB)235 and CAR T cells directed against the
receptor uPAR236 attenuate senescence-associated disease
models in mice.
In summary, cellular senescence is an important response to
stress and damage that, in normal physiology, is followed by im-
mune clearance, but that upon aging or chronic damage fails to
be eliminated by immune mechanisms and hence is pathogenic
due to the abundant secretion of pro-inflammatory and pro-
fibrotic factors. Therapeutic strategies aimed at killing senescent
cells have been extensively explored in animal models and are
now in clinical trials (Table 1).
STEM CELL EXHAUSTION
Aging is associated with reduced tissue renewal at steady state,
as well as with impaired tissue repair upon injury, with each
ll
Revieworgan having its own strategy for renewal and repair.237 For
example, in skeletal muscle, one single-cell type, the satellite
cell, is placed at the apex of a unipotent and unidirectional hier-
archy, both for renewal and repair. In skin epidermis, which is
characterized by high renewal and exposure to injury, there are
multiple stem cell niches, particularly in association to the hair
follicles, each one generating its progeny and territory. However,
upon injury, multiple cells can acquire stem cell properties and
subvert territorial boundaries. Other organs like liver, lung, or
pancreas exhibit rather low renewal rates under normal condi-
tions, contrasting with the acquisition of stem cell properties
including proliferation and multipotency by different cell types
(Figure 5B). Indeed, tissue repair is believed to rely to a large
extent on injury-induced cellular de-differentiation and plasticity.
For example, in the intestine, brain, and lung, injury induces de-
differentiation of non-stem cells, which reactivates normally si-
lent embryonic and stemness transcription programs, thus
acquiring the plasticity needed for tissue repair.238–240 Injury-
induced plasticity (and its progressive loss with aging) may be
more relevant for aging than the plasticity of resident stem
cells under normal homeostatic conditions. Stem and progenitor
cells are all subject to the same hallmarks of aging as are
cells without stem potential, and for this reason, we do not
discuss here the abundant literature about the impact of each
hallmark of aging on stem cell function. Instead, we will focus
on a general strategy to counter the decline of stem cell function
with aging based on the concept of ‘‘cellular reprogramming.’’
This process is thought to act in a cell-autonomous manner on
multiple cell types; however, its impact on stem and progenitor
cells is considered of higher relevance because of its long-term
impact.
Rejuvenation of tissue repair by reprogramming
Cellular reprogramming toward pluripotency consists in the con-
version of adult somatic cells into embryonic pluripotent cells
(known as induced pluripotent stem cells or iPSCs) by the
concomitant action of four externally transduced transcription
factors, namely, OCT4, SOX2, KLF4, and MYC (OSKM).241 The
process of reprogramming usually requires several weeks during
which cells first lose their differentiated phenotype by transcrip-
tional repression of cell identity genes and subsequently trans-
activate pluripotency genes.242 Full reprogramming not only im-
plies a change of cellular identity but also cellular rejuvenation,
characterized by a number of aging features that are reset to
the embryonic state, as indicated by p16 reduction,243 extension
of telomeres,244 and resetting of the DNA methylation clock.245
Interestingly, rejuvenation occurs in a progressive fashion start-
ing shortly after the initiation of de-differentiation.246 Indeed, it is
possible to initiate reprogramming with OSKM, interrupt the pro-
cess at an intermediate state, and allow cells to return to their
original identity. This transient cellular perturbation, variously
known as ‘‘partial,’’ ‘‘transient,’’ or ‘‘intermediate’’ reprogram-
ming, is able to rejuvenate cellular markers of aging such as
the DNA methylation clock, DNA damage, epigenetic patterns,
and aging-associated changes in the transcriptome, both
in vitro and in vivo.63,64,70,246,247 Therefore, it can be proposed
that the processes of de-differentiation and rejuvenation are
coupled. Specifically, de-differentiation implies the erasure ofepigenetic and transcriptional programs, and thismay also erase
aging-associated alternations. Upon interruption of partial re-
programming, cells re-stablish their original epigenetic and tran-
scriptional status in a process of re-differentiation that, interest-
ingly, does not re-stablish the erased aging-associated changes
and therefore resets the epigenome and transcriptome to a
younger state.
Transient reprogramming in mice confers repair capacity to
old tissues so that a subsequent damage is repaired as efficiently
as in young individuals. This increased repair capacity has been
shown for models of tissue damage in the endocrine pancreas,63
skeletalmuscle,63 nerve fibers,70 eye,70 skin,64 heart,65 and liver.66
Also, tissue dysfunctions characteristic of natural aging, such as
reduced visual acuity70 and the loss of adult neurogenesis in the
hippocampus and long-term memory,67 can be partially reversed
by transient reprogramming. There are a few instances in which
transient reprogramming is beneficial also during the process of
tissue repair (and not only prior to the injury). This is the case for
traumatic brain injury68 and skinwound healing.69 Finally, it should
bementioned that the lifespan of progeroidmice can be extended
by transient reprogramming,63 although extension of longevity by
OSKM has not yet been reported for wild-type mice.
Partial reprogramming recapitulates features of natural tissue
repair (Figure 5B). In both cases, cells undergo a transient pro-
cess of de-differentiation, acquisition of embryonic and progen-
itor features, and subsequent re-differentiation. Thus, de- and
re-differentiation could explain tissue rejuvenation, in line with
the observation that transient de-differentiation of myocytes,
followed by their re-differentiation, induces rejuvenation of
the transcriptome.248 The natural process of tissue repair may
imply some degree of cellular rejuvenation, in accord with the
finding that the epigenetic methylation clock accelerates soon
after tissue injury and partially reverses during tissue repair.249
Moreover, tissue damage reportedly creates a tissue microenvi-
ronment that is highly permissive for IL-6-driven reprogram-
ming.250 Finally, cyclic expression of the transcription
factor FOXM1 extends the longevity of progeroid mice and
wild-type mice.251 Although the detailed mechanism is still
unexplored, FOXM1 is induced in the kidney upon injury and
participates in triggering de-differentiation and proliferation
of tubular epithelial cells during the repair process.252
Thus, several features of natural tissue repair and artificial
reprogramming may converge, perhaps allowing refinement
of strategies for restoring repair capacity in aging tissues.
ALTERED INTERCELLULAR COMMUNICATION
Aging is coupled to progressive alterations in intercellular
communication that increase the noise in the system and
compromise homeostatic and hormetic regulation. Thus, aging
involves deficiencies in neural, neuroendocrine, and hormonal
signaling pathways, including the adrenergic, dopaminergic,
and insulin/IGF1-based and renin-angiotensin systems, as well
as sex hormones commensurate with the loss of reproductive
functions.182,253 Although the primary causes of such alterations
are cell intrinsic, as this is particularly well documented for the
SASP, these derangements in intercellular communication
ultimately sum up to a hallmark on its own that bridges theCell 186, January 19, 2023 261
ll
Reviewcell-intrinsic hallmarks to meta-cellular hallmarks including the
chronification of inflammatory reactions coupled to the decline
of immunosurveillance against pathogens and premalignant
cells, as well as the alterations in the bidirectional communica-
tion between human genome and microbiome, which finally re-
sults in dysbiosis. A number of studies in this regard have
focused on the search for blood-borne systemic factors with
pro-aging or prolongevity properties, the role of diverse commu-
nication systems between cells, and the evaluation of the
functional relevance of ECM disruption during aging.
Pro-aging blood-borne factors
A single transfusion of old blood induces features of aging in
young mice within a few days,72 and the simple dilution of the
blood of old mice with saline buffer containing 5% albumin in-
duces rejuvenation in multiple tissues,71 indicating
the existence of circulating factors that favor the aging process.
Among the pro-aging blood-borne factors, the chemokine
CCL11/eotaxin and the inflammation related protein b2-micro-
globulin reduce neurogenesis,254,255 IL-6 and TGF-b impair he-
matopoiesis,256 and the complement factor C1q compromises
muscle repair.257 Theoretically, the neutralization of these fac-
tors might have potent anti-aging effects. Indeed, several among
the aforementioned factors are secreted in the context of SASP
andmay be co-responsible for the phenomenon of ‘‘contagious’’
aging, which also involves extracellular vesicles.258 Thus, so-
called ‘‘senomorphics’’ might be used to repress SASP and
slow down aging.
Anti-aging blood-borne factors
Soluble factors present in the blood of young mice effectively
restore renewal and repair capacity in old mice259 (Table 1). Het-
erochronic parabiosis experiments followed by extensive single-
cell transcriptomics have confirmed the capacity of young blood
to rejuvenate multiple tissues74 and to restore age-associated
reduction in general gene expression, in particular that of mito-
chondrial genes involved in the electron transport chain.75 The
chemokine CCL3/MIP-1a acts as a rejuvenating factor for he-
matopoietic stem and progenitor cells;74 the metalloproteinase
inhibitor TIMP2 has been implicated in rejuvenating the hippo-
campus;73 the anti-inflammatory interleukin IL-37 (which de-
clines in monocytes from aged humans) improves increased
endurance exercise and ameliorates whole-body metabolism
in old mice;76 the cytokine GDF11 rejuvenates some tissues,
such as muscle, brain, and endocrine pancreas, although it im-
pairs the function and repair of other tissues due to its pro-
fibrotic side effects;77 and finally, mice with transgene-enforced
VEGF overexpression exhibit enhanced liver and muscle repair,
improved general health and an extension in average longevity
by 40%.78
Long-range and short-range communication systems
The central nervous system controls multiple facets of aging
affecting peripheral organs, explaining how brain-specific gene
manipulations like overexpression of SIRT1, UCP1, or knockout
of IKBKB and TRPV1 can enhance mouse longevity (Table 1).
The precise mechanisms of these long-range activities are yet
to be determined.260 Of note, intercellular communication also262 Cell 186, January 19, 2023involves the interaction among short-lived extracellular mole-
cules (such as ROS, nitric oxide, nucleic acids, prostaglandins,
and other lipophilic molecules), soluble factors that are released
from various tissues including white adipose tissue (adipokines),
brown adipose tissue (baptokines), heart (cardiokines), liver
(hepatokines) and skeletal muscles (myokines, including exer-
kines produced in response to exercise), cell-bound ligands,
and receptors on other cells (as exemplified by IL-1a that can
remain cell-bound), as well as direct cell-to-cell interactions
mediated by tight junctions or gap junctions. All these communi-
cation systemsmay be altered during aging and hence are being
scrutinized for their potential pro- and anti-aging properties.258
Extracellular matrix
Aging causes numerous damages in the long-lived protein com-
ponents of the ECM, including AGEs, carbonylation and carba-
mylation, elastin fragmentation, and collagen crosslinking,261
thus leading to tissue fibrosis (fibroaging).262 This deleterious
process is in part due to the excessive release of TGF-b and
other growth factors, and the nuclear translocation of TAZ and
YAP transcription factors, which act as mechanotransducers
and trigger the expression of pro-fibrotic genes such as
transglutaminase-2, lysyl oxidase (LOX), and LOX-like en-
zymes.262 ECM stiffness also affects the function of senescent
cells, which in turn secrete matrix metalloproteases that amplify
the damage of the ECM,263 and proteolytically generate dam-
age-associated molecular patterns to activate pro-senescent,
pro-fibrotic, and pro-inflammatory pathways.262 The increasing
stiffness of the aging matrix may also favor WNT signaling to
induce fibroblast activation and expression of pro-fibrotic
genes.264 This pathway exhibits extensive crosstalk with other
pro-fibrotic pathways, such as NOTCH, RAS, TGF-b/SMAD,
and hedgehog/GLI, thereby demonstrating the complexity and
interconnections of mechanisms underlying the development
of age-linked fibrosis.262 Of note, mechanical change caused
by matrix stiffness is sufficient to cause age-related loss of func-
tion of oligodendrocyte progenitor cells in a processmediated by
the mechanoresponsive ion channel PIEZO1.265
Several studies have provided causal evidence for the
contribution of ECM stiffness to aging and have also suggested
approaches for improving healthy aging (Table 1). In vivo inhibi-
tion of Piezo1 using AAV vectors results in rejuvenation of the
oligodendrocyte progenitors in the brain of old mice.265 Genetic
inactivation of YAP/TAZ in stromal cells causes accelerated
aging, although sustaining YAP function rejuvenates old
cells and prevents the emergence of aging features by control-
ling cGAS-STING signaling.79 Moreover, mice engineered to
produce collagenase-resistant type I collagen (Col1a1r/r)
exhibit vascular cell senescence, accelerated aging, and short-
ened lifespan.266 The importance of collagen for human
longevity has been reinforced by the discovery of rare variants
in COL25A1—encoding a brain-specific collagen—that may
have a protective role against Alzheimer’s disease.267 Moreover,
ECM prepared from young human fibroblasts induces a youthful
state in aged senescent cells.80 ECM compounds such as chon-
droitin sulfate and hyaluronic acid restore the age-related
decline of collagen and increase lifespan in nematodes.268
Conversely, ectopic expression of human hyaluronidase
Figure 6. Derangement of supracellular
functions
Altered intercellular communication bridges the
cell-intrinsic hallmarks to meta-cellular hallmarks
including the chronic inflammation, and the alter-
ations in the crosstalk between human genome
and microbiome, which finally result in dysbiosis.
(A) Chronic inflammation during aging occurs as a
consequence of multiple derangements that stem
from all the other hallmarks. Several representa-
tive examples of anti-inflammatory interventions
with positive effects on healthspan and lifespan
are shown in the right part of the figure.
(B) Dysbiosis contributes to multiple pathological
conditions associated with aging. The human gut
microbiota significantly changes during aging,
finally leading to a general decrease in ecological
diversity. The main features of the mechanisms
underlying these microbiota changes and some
examples of interventions on the gut microbiota
composition which can promote healthy aging are
shown in the lower part of the right panel. CVDs,
cardiovascular diseases; SCFAs, short-chain
fatty acids.
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ReviewTMEM2 promotes resistance to ER stress and extends lifespan
in C. elegans through changes in p38/ERK MAPK signaling.269
In mice, deletion of chondroitin 6-sulfotransferase results in an
abnormal ECM in the brain, early memory loss, and accelerated
brain aging, whereas overexpression of this enzyme improved
memory in old mice.81 Retrospective analyses indicate that
oral intake of glucosamine/chondroitin sulfate leads to a reduc-
tion in all-cause mortality in humans.270 However, there is no
prospective proof thus far that such a prolongevity effect would
be mediated through an amelioration of the ECM.CHRONIC INFLAMMATION
Inflammation increases during aging (‘‘in-
flammaging’’) with systemic manifesta-
tions, as well as with pathological local
phenotypes including arteriosclerosis,
neuroinflammation, osteoarthritis, and
intervertebral discal degeneration.
Accordingly, the circulating concentra-
tions of inflammatory cytokines and bio-
markers (such as CRP) increase with ag-
ing. Elevated IL-6 levels in plasma
constitute a predictive biomarker of all-
cause mortality in aging human popula-
tions.271 In association with enhanced
inflammation, immune function declines,
a phenomenon that can be captured by
high-dimensional monitoring of myeloid
and lymphoid cells in the blood from pa-
tients and from mouse tissues.272 For
example, a population of age-associated
T cells—termed Taa cells—is composed
of exhausted memory cells that mediate
pro-inflammatory effects via granzyme
K. Shifts in T cell populations entail the
hyperfunction of pro-inflammatory TH1and TH17 cells, defective immunosurveillance (with a negative
impact on the elimination of virus-infected, malignant or senes-
cent cells), loss of self-tolerance (with a consequent age-associ-
ated increase in autoimmune diseases), and reduced mainte-
nance and repair of biological barriers, altogether favoring
systemic inflammation273 (Figure 6A).
Links between inflammation and other aging hallmarks
Inflammaging occurs as a result of multiple derangements that
stem from all the other hallmarks. For example, inflammation isCell 186, January 19, 2023 263
ll
Reviewtriggered by the translocation of nuclear and mtDNA, into the
cytosol where it stimulates pro-inflammatory DNA sensors,
especially when autophagy is ineffective and hence unable to
intercept ectopic DNA.4 Genomic instability favors clonal
hematopoiesis of indeterminate potential (CHIP), with the
expansion of myeloid cells that often bear a pro-inflammatory
phenotype, driving for instance cardiovascular aging.274 Intrigu-
ingly, the most frequent CHIP-associated mutations affect
the epigenetic modifiers DNMT3 (which methylates cytosine
residues in DNA) and TET2 (which catalyzes the oxidation of
methylcytosine to 5-hydroxymethylcytosine). Mechanistically,
CHIP affecting TET2 enhances IL-1b and IL-6 production by
myeloid cells and stimulates cardiovascular disease (CVD),
which is attenuated among individuals bearing a loss-of-function
mutation in the IL-6 receptor or treated with an IL-1b neutralizing
antibody.275
Overexpression of pro-inflammatory proteins can be second-
ary to epigenetic dysregulation, deficient proteostasis, or
disabled autophagy. Excessive trophic signals resulting in
activation of the GH/IGF1/PI3K/AKT/mTORC1 axis trigger
inflammation. In addition, inflammation is favored by the SASP
secondary to the accrual of senescent cells, as well as by the
accumulation of extracellular debris and infectious pathogens,
which are not cleared due to senescence, and by exhaustion
of myeloid and lymphoid cells. This latter phenomenon involves
age-associated thymic involution, abrogating thymopoiesis with
the consequent rarefaction of the T cell repertoire and the
inability to mount efficient immune responses against novel anti-
gens.276 Of note, thymopoiesis is improved by CR in humans,
and a CR-downregulated gene coding for platelet activation fac-
tor acetyl hydrolase A2 group VII (PLA2G7) can be knocked out in
mice to combat thymic atrophy.50 Finally, inflammaging is also
exacerbated by perturbations of circadian rhythms and by
intestinal barrier dysfunction.277
Anti-inflammatory, anti-aging interventions
Although systemic inflammation is mechanistically linked to all
the aforementioned age-associated alterations, inflammation
constitutes a hallmark on its own. Indeed, specific manipulations
of the inflammatory and immune system can accelerate or decel-
erate the aging process across different organ systems. For
example, a T cell-specific defect in the mitochondrial transcrip-
tion factor A (TFAM) is sufficient to drive cardiovascular, cogni-
tive, metabolic, and physical aging coupled to an increase in
circulating cytokines. The TNF-a inhibitor etanercept partially
reversed this phenotype.82 Heterozygous deletion of the DNA
repair protein ERCC1 in hematopoietic cells from mice is suffi-
cient to induce immunosenescence and aging of non-lymphoid
organs, as well as numerous signs of organ damage coupled
to reduced lifespan. This phenotype was alleviated by the seno-
lytic fisetin.278 These results support the idea that aging of the
immune system may drive organismal aging. Of note, adoptive
transfer of TFAM-null T cells, young ERCC-deficient spleno-
cytes, or aged wild-type splenocytes into young mice induced
senescence, whereas the transfer of young immune cells into
ERCC-deficient mice attenuated senescence, pointing to the ca-
pacity of immune cells to modulate organismal aging in both
positive and negative terms.82,278264 Cell 186, January 19, 2023There aremultiple examples of broad healthspan and lifespan-
expanding effects of anti-inflammatory treatments (Figure 6A;
Table 1). Thus, blockade of TNF-a prevents sarcopenia in mice
and improves cognition in aging rats.83,84 Blockade of the com-
mon type 1 interferon receptor (IFNAR1) reverses the accumula-
tion of monocytes in the aging mouse lung.279 Knockout of the
prostaglandin E2 receptor EP2 in myeloid cells or treatment of
aged mice with pharmacological EP2 inhibitors ameliorates
cognition.85 Knockout of the inflammasome protein NLRP3
improves metabolic biomarkers, glucose tolerance, cognition,
and motor performance and extends mouse longevity.86 Phar-
macological inhibitors of NLRP3 or of its downstream enzyme
caspase-1 have encouraging preclinical effects on normal and
accelerated aging models.280 Most importantly, inhibition of
the caspase-1 product IL-1b with canakinumab exemplifies an
anti-aging treatment applicable to patients. The phase 3 clinical
trial CANTOS evaluated the capacity of canakinumab to prevent
recurrent CVD in patients with a history of myocardial infarction
and signs of pronounced inflammation. Beyond meeting the pri-
mary endpoint of the trial, canakinumab reduced the incidence
of diabetes and hypertension, as well as the incidence of, and
mortality from, lung cancer.87 Finally, although long-term use
of non-steroidal anti-inflammatory agents such as aspirin may
have positive effects on human health—in particular with respect
to the prevention of CVD and gastrointestinal cancers—a large
phase 3 clinical trial in which aspirin was administered to over
70-year-old subjects yielded negative results.281 Hence, further
studies will be necessary to explore the value of prophylactic
treatments with aspirin at a younger age to combine aspirin
with other medications or to replace aspirin by less toxic anti-in-
flammatory drugs.
DYSBIOSIS
Over recent years, the gut microbiome has emerged as a key
factor in multiple physiological processes such as nutrient diges-
tion and absorption, protection against pathogens, and produc-
tion of essential metabolites including vitamins, amino acid de-
rivatives, secondary bile acids, and short-chain fatty acids
(SCFAs). The intestinal microbiota also signals to the peripheral
and central nervous systems and other distant organs and
strongly impacts on the overall maintenance of host health.146
Disruption of this bacteria-host bidirectional communication re-
sults in dysbiosis and contributes to a variety of pathological
conditions, such as obesity, type 2 diabetes, ulcerative colitis,
neurological disorders, CVDs, and cancer.282 The progress in
this field has generated an enormous interest in exploring gut mi-
crobiota alterations in aging (Figure 6B).
Microbiota alterations in aging
The microbial community within the intestinal tract is highly var-
iable among individuals as a consequence of host genetic vari-
ants (ethnicity), dietary factors, and lifestyle habits (culture), as
well as environmental conditions (geography), whichmakes diffi-
cult to unveil the relationships between microbiota and pleio-
tropic age-associated disease manifestations. Nonetheless,
some meta-analyses have revealed microbiota-disease associ-
ations that have been validated across distinct pathologies283
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Reviewand countries.284,285 Studies in both humans and animal models
have provided valuable information on clinical, epidemiological,
sociological, and molecular aspects that underlie the complex
effects of an aged microbiome on human health and disease.286
Once bacterial diversity is established during childhood, it re-
mains relatively stable during adulthood. However, the architec-
ture and activity of this bacterial community undergoes gradual
changes during aging, finally leading to a general decrease in
ecological diversity. Thus, several studies conducted on cente-
narian populations showed a reduction in core abundant taxa,
such as Bacteroides and Roseburia, but also an increase in
several genera such as Bifidobacterium and Akkermansia, which
appear to have prolongevity effects.287
These studies have been extended by recent analysis of the
gut microbiome and phenotypic data from over 9,000 individuals
of three independent cohorts spanning 18–101 years of age.288
Of note, individual gut microbiomes become increasingly more
unique to each individual with age, and this uniqueness is asso-
ciated with well-known microbial metabolites involved in im-
mune regulation, inflammation, and aging. In older age, healthy
participants show continued drift toward a unique microbial
composition, whereas this drift is reduced or absent in individ-
uals in worse health. The identified microbiome pattern of
healthy aging is characterized by a depletion of core taxa,
such as Bacteroides, present across most humans. Moreover,
in individuals approaching extreme age, retention of highBacter-
oides levels and a low gut microbiome uniqueness measure are
significantly associated with decreased survival. However, find-
ings in microbiota from centenarians and supercentenarians are
not always concordant with those derived from elderly popula-
tions. The ELDERMET study reported an increased dominance
of the core genera Bacteroides, Alistipes, and Parabacteroides
in old individuals compared with younger controls. This study
also identified age-related shifts in gut microbiota composition
linked to frailty, cognition, depression, and inflammation.289
Another study revealed age-related trajectories of themicrobiota
shared across populations of different ethnicities, as well
as a common age-related decrease in sex-dependent differ-
ences in gut microbiota. Of note, older adults exhibit higher
abundances of several health-promoting bacterial species,
including Akkermansia.290 These results suggest that age-
related physiological changes, beyond dietary changes and life-
style of older adults, may have profound effects on the human
gut microbiota.
The heterogeneity of findings in all these studies indicates that
theremay bemultiple gutmicrobiome trajectories of aging. How-
ever, there is an interesting convergence in plasma concentra-
tions of microbiota-produced amino acid derivatives. These me-
tabolites include indoles derived from gut bacterial degradation
of tryptophan, and several fermentation products of phenylala-
nine/tyrosine, such as p-cresolsulfate, phenylacetylglutamine,
and p-cresol glucuronide. This finding is consistent with data
from the ELDERMET cohort showing that fecal concentrations
of p-cresol correlate with increased frailty and may contribute
to age-associated decline in this population. Conversely, plasma
concentrations of certain indole metabolites correlate with
improved fitness in older adults. Indole metabolites increase
healthspan and lifespan in mice, at least in part, by attenuationof inflammatory responses through binding of the arylhydrocar-
bon receptor.93
Further metabolomics and functional analysis of the gutmicro-
biome of centenarians have shown its enrichment in some
particular bacteria, such as Alistipes putredinis and Odoribacter
splanchnicus. Some of these bacterial species are capable of
generating unique secondary bile acids, including isoallo-litho-
cholic, which exerts potent antimicrobial effects against gram-
positive multidrug-resistant pathogens such as Clostridioides
difficile and Enterococcus faecium.291 Thus, specific bile
acid metabolism may be involved in reducing the risk of patho-
biont infection and contribute to intestinal homeostasis,
thereby decreasing the susceptibility to age-associated chronic
diseases.
Fecal microbiota transplantation and aging
Multiomics studies in pathological aging have revealed that two
different mouse models of progeria exhibit intestinal dysbiosis
mainly characterized by an increase in the abundance of Proteo-
bacteria and Cyanobacteria and a decrease in levels of Verruco-
microbia. Consistent with these findings, human progeria
patients with HGPS or NGPS also show intestinal dysbiosis,
whereas long-lived humans exhibit a substantial reduction in
Proteobacteria and a significant increase in Verrucomicrobia.88
The causal implications of these changes were demonstrated
in vivo by fecal microbiota transplantation (FMT). FMT from
wild type to progeroid mice recipients enhanced healthspan
and lifespan in both accelerated-aging models, whereas admin-
istration of the verrucomicrobium Akkermansia muciniphila was
also sufficient to obtain such effects. Conversely, FMT from
progeroid donors to wild-type recipients induced detrimental
metabolic alterations. Restoration of secondary bile acids and
other metabolites depleted in progeroid mice phenocopied the
beneficial effects of reestablishing a healthy microbiome88
(Table 1).
FMT also revealed the causative role of gut dysbiosis in the
chronic systemic inflammation and the decline in adaptive
immunity associated with aging and age-related diseases.
Transfer of the gut microbiota from old mice to young germ-
free mice triggered inflammatory responses characterized by
enhanced CD4+ T cell differentiation in spleen, upregulation of
inflammatory cytokines, and increased circulation of inflamma-
tory factors of bacterial origin.292 FMT also provided evidence
for the implication or the gut microbiota in the maintenance of
brain health and immunity during aging.90 Microbiota from young
mice donors reversed aging-associated differences in hippo-
campal metabolites and brain immunity and ameliorated age-
associated impairments in cognitive behavior when transplanted
into an aged host. These works open the possibility of manipu-
lating the gut microbiota with pre-, pro-, and post-biotics to reju-
venate the immune system and the aging brain. Heterochronic
fecal transfers confirmed the causal link between age-depen-
dent changes in microbial composition and a decline in the func-
tion of the host immune system.92 Indeed, the defective germinal
center reaction in Peyer’s patches of aged mice can be rescued
by FMT from younger animals without affecting germinal center
reactions in peripheral lymph nodes. Finally, FMT from young
donor mice improves ovarian function and fertility in agedCell 186, January 19, 2023 265
ll
Reviewmice. These beneficial effects are associated with an improve-
ment in the immune microenvironment of aged ovaries, with
decreased macrophages and macrophage-derived multinucle-
ated giant cells, reduced levels of pro-inflammatory IFNg, and
increased abundance of the anti-inflammatory cytokine IL-4.91
Other prolongevity interventions on gut microbiota
The probiotic Lactobacillus plantarumGKM3 promotes longevity
and alleviates age-related cognitive impairment in the SAMP8
mouse model of accelerated aging.89 Interventions on gut
microbiota composition also restored the age-linked decline in
microglial maturation and function which causes altered brain
plasticity and promotes neurodegeneration. Recolonization ex-
periments or administration of gut microbiota metabolites,
such as SCFAs, prevented the age-associated decline of bene-
ficial Bifidobacterium, increased Akkermansia abundance, and
restored microglial function in middle-aged mice.94 Moreover,
caloric restriction diets induce structural changes of the gut mi-
crobiome increasing the abundance of Lactobacillus and other
species that influence healthy aging. The gut microbiota-
induced inflammaging and the consequent increase in insulin
resistance can also be reversed by restoring abundance of bene-
ficial SCFA-producing bacteria, such as A. muciniphila, in aged
mice and macaques.293 Similarly, a randomized, double-blind,
placebo-controlled pilot study in overweight/obese insulin-resis-
tant volunteers showed that oral administration of pasteurized
A. muciniphila improved insulin sensitivity reduced insulinemia
and plasma total cholesterol levels.95 Collectively, these results
underscore the causal links between aging and dysbiosis and
suggest that interventions aimed at restoring a youthful micro-
biome may extend healthspan and lifespan.
INTEGRATION OF HALLMARKS
All the 12 hallmarks of aging are strongly related among each
other. For example, genomic instability (including that caused
by telomere shortening) crosstalks to epigenetic alterations
(e.g., through the loss-of-function mutation of epigenetic modi-
fiers such as TET2), loss of proteostasis (e.g., due to the produc-
tion of mutated, misfolded proteins), disabled macroautophagy
(e.g., through the capacity of autophagy to remove supernumer-
ary centrosomes, extranuclear chromatin, and cytosolic DNA),
deregulated nutrient-sensing (e.g., because SIRT6 is an NAD+
sensor involved in DNA repair but also responding to nutrient
scarcity), mitochondrial dysfunction (e.g., due to the mutation
ofmtDNA), cellular senescence (e.g., because DNAdamage trig-
gers senescence), altered intercellular communication (e.g., by
hampering activation of communication pathways), chronic
inflammation (e.g., because CHIP and leakage of DNA into the
cytosol induce inflammation), and dysbiosis (e.g., because
mutations in intestinal cells favors dysbiosis, whereas specific
microbial proteins and metabolites act as mutagens). Similar
functional relationships can be listed for most if not all hallmarks
of aging, illustrating their formidable interconnectivity.
This entanglement is also visible at the level of experimental
anti-aging interventions that often simultaneously target several
hallmarks. Thus, SIRT activators including NAD+ precursors
attenuate genomic instability (via DNA repair), epigenetic alter-266 Cell 186, January 19, 2023ations (via histone deacetylation), loss of proteostasis (via the
removal of protein aggregates), disabled macroautophagy (via
autophagy enhancement), deregulated nutrient-sensing (via
activation of nutrient scarcity sensors), and mitochondrial
dysfunction (via an increase in mitophagy-dependent quality
control).176 Spermidine complexes to DNA (hence counteracting
genomic instability), affects translation (avoiding loss of proteo-
stasis), stimulates macroautophagy, reverses lymphocyte
senescence, prevents the exhaustion of muscle stem cells,
maintains circadian rhythms, suppresses inflammation, stimu-
lates cancer immunosurveillance, and is produced by intestinal
bacteria.294Metformin has a pleiotropicmode of action including
induction of autophagy, activation of the nutrient scarcity sensor
AMPK, inhibition of mitochondrial respiration, alleviation of
adipocyte senescence, suppression of inflammation, and
favorable shifts in the gut microbiota.210 Similarly, maintenance
of eubiosis by oral supplementation of A. muciniphila stimulates
intestinal autophagy, reduces metabolic syndrome, dampens
inflammation, and enhances anticancer immune responses.295
Indeed, a notable feature of effective anti-aging interventions,
such as lowered insulin/IGF-1 signaling296 and disruption of
the TORC1 complex,296,297 is the diversity of mechanisms by
which they target different aging hallmarks in different tissues
to maintain healthspan of the whole organism.
Although each of the 12 hallmarks of aging can be targeted
one by one, yielding tangible benefits for healthspan and lifespan
(Table 1), there is some kind of hierarchy among them (Figure 1).
Thus, as we initially proposed,1 the primary hallmarks, which
reflect damages affecting the genome, telomeres, epigenome,
proteome, and organelles, progressively accumulate with time
and unambiguously contribute to the aging process.298 The
antagonistic hallmarks, which reflect responses to damage,
play a more nuanced role in the aging process. For example,
trophic signaling and anabolic reactions activated by nutrient-
sensing have beneficial actions in youth but are largely pro-age-
ing later on. Thus, in an archetypal case of antagonistic
pleiotropy, the nutrient-sensing network contributes to organ
development until young adulthood but plays a detrimental role
beyond this stage. Additionally, reversible and low-dose mito-
chondrial dysfunction can stimulate beneficial counterreactions
(via mitohormesis), whereas limited and spatially confined
levels of cellular senescence contribute to the suppression of
oncogenesis and improve wound healing. Finally, the integrative
hallmarks arise when the accumulated damage inflicted
by the primary and antagonistic hallmarks cannot be compen-
sated any more, resulting in stem cell exhaustion, intercellular
communication alterations including ECM damage, chronic
inflammation, and dysbiosis, which together dictate the pace
of aging.
Recently, we postulated the existence of eight hallmarks of
health,146 which include organizational features of spatial
compartmentalization (integrity of barriers and containment of
local perturbations), maintenance of homeostasis over time
(recycling and turnover, integration of circuitries, and rhythmic
oscillations), and an array of adequate responses to perturbation
(homeostatic resilience, hormetic regulation, and repair and
regeneration). Undoubtedly, aging is linked to progressive dete-
rioration of these eight hallmarks of health, implying a ramping
Figure 7. Integration of hallmarks
All the 12 hallmarks of aging proposed in this work are functionally related among each other. These determinants of aging are also interconnected to the eight
hallmarks of health, which include organizational features of spatial compartmentalization, maintenance of homeostasis over time, and an array of responses to
perturbation.146 Finally, the hallmarks of aging are interconnected to the eight proposed strata of organismal organization146 and create amultidimensional space
of interactions that may contribute to explain some important features of the aging process.
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Reviewincapacity to maintain spatial compartmentalization (with the
consequent loss of integrity of internal and external barriers, as
well as the incapacity to contain perturbations of such barriers
in space and time), to assure long-term homeostasis (with
reduced capacity of recycling and turnover, inefficient coordina-
tion among different systems via integrated circuitries, and de-
synchronization of ultradian, circadian, or infradian rhythms),
and to adequately respond to stress by complete repair and
regeneration, homeostatic resilience, and hormetic regulation
(Figure 7). This decline affects all eight strata of organismal orga-
nization, across different classes of molecules (such as
DNA, RNA, proteins, and metabolites), organelles (such as
nuclei, mitochondria, and lysosomes), cell types (such as paren-
chymatous, auxiliary/stromal, and inflammatory/immune cells),
supracellular units constituting the minimal functional entities
of organs, entire organs within their anatomical boundaries,
organ systems (such as the gastrointestinal, respiratory, and
urinary tracts), systemic circuitries (with their endocrine, neuro-
logical, lymphatic, and vascular connections), as well as the
meta-organism (that includes the microbiota). As a result, the
12 hallmarks of aging are interconnected to the eight hallmarks
of failing health and the eight strata of organismal organization
(Figure 7), creating a multidimensional space of interactions
that may explain some features of the aging process.
Heterochronic parabiosis experiments, in which the vascular
systems of young and old mice are connected, may illustrate
best the importance of systemic regulatory factors (such as
hormones and circulating cells) on the aging process. This
phenomenon has been extensively characterized at the level of
single-cell transcriptomics, yielding a spatiotemporal map ofthe capacity of the young system to rejuvenate an older one
or, vice versa, the ability of pro-aging factors to precipitate the
senescence of young cells.74,75 This type of experiment
demonstrates that aging relies on the integration of cell-autono-
mous and non-cell-autonomous mechanisms that also have
been revealed in Drosophila (in which stimulation of autophagy
in the intestine is sufficient to extend lifespan of the entire
organism)120 and mice (in which injection of a few thousands of
senescent fibroblasts is sufficient to trigger invalidating osteoar-
thritis).299 Hence, pro-aging and anti-aging mechanisms can be
communicated among distinct cell types, perhaps explaining
that ‘‘normal’’ aging usually affects multiple organs in a close-
to-synchronous fashion, at difference with ‘‘pathological’’ aging
in which time-dependent diseases precociously manifest in spe-
cific locations, in the form of initially isolated cardiovascular,
oncological, or neurodegenerative disorders. However, the
distinction between normal and pathological aging is debat-
able,300 and some progeroid syndromes manifest signs of
incomplete or segmental aging, as exemplified by the absence
of a central nervous phenotype in HGPS.
In view of the spectacular progress of developing longevity
strategies in mammalian model organisms and initial clinical tri-
als (Table 1), it will be important to develop rational strategies for
intervening into human aging. The question arises to which
extent strategies for extending human healthspan should be
based on the avoidance of age-accelerating environmental fac-
tors (such as pollution, stress, inadequate physical activity, and
unhealthy diets, often unavoidable in a context of poverty, pre-
cariousness, and wartime), the adoption of health-promoting
lifestyle factors (such as diet, exercise, regular sleeping patterns,Cell 186, January 19, 2023 267
ll
Reviewand social activities), the administration of relatively non-spe-
cific, pleiotropic drugs (exemplified by NAD+ precursors, metfor-
min, spermidine, or MTORC1 inhibitors), or more specific medi-
cal interventions. Such specific treatments may involve
pharmacological agents—with the prospective of a broad imple-
mentation, genetic or cell-based therapies—with rather complex
logistics and elevated costs, or bioengineering methods for sur-
gical tissue replacement, which most likely will mainly remain in
the realm of experimentation. Given the multiplicity of hallmarks
offering therapeutic strategies for decelerating, halting, or
reversing aging, it will be interesting to evaluate combination reg-
imens with the scope of maximizing benefits andminimizing side
effects. The question remains open whether such healthspan
and lifespan extending prophylactic treatments will profit from
personalization based on individual patient characteristics
determined by the genetic, epigenetic, metabolomic, or pheno-
typic assessments of aging clocks.
Aging is not yet a recognized target for drug development or for
treatment. For this reason, the first clinical trials evaluating anti-
aging interventions must deal with the prevention or mitigation
of age-associated pathologies rather than aging itself. Although
such trials have been designed to target high-risk populations
(such as patients with myocardial infarction and laboratory signs
of inflammation in the CANTOS trial or patients with frailty or car-
diovascular events tobe enrolled in futuremetformin-related trials)
and to measure the manifestation of a second cardiovascular
event or aggravation of frailty, there is a risk that they are pro-
grammed too late, which is of significant concern. Indeed, at
this point, academic geroscience may raise or fall as the function
of the outcomeof the first randomized, double-blindedphase 3 tri-
als. The new directions of the hallmarks of aging may provide an
improved framework for the development of effective interven-
tions aimed at the extension of healthy longevity.ACKNOWLEDGMENTS
We apologize for omitting relevant works and citations due to space
constraints. We acknowledge all members of our laboratories for helpful
comments during the elaboration of this manuscript. We thank Jose´ M.P.
Freije for critical reading of the manuscript. C.L-O. is supported by grants
from the European Research Council (ERC Advanced Grant, DeAge), Minis-
terio de Ciencia e Innovacio´n, Instituto de Salud Carlos III, and La Caixa Foun-
dation (HR17-00221). The Instituto Universitario de Oncologı´a is supported by
Fundacio´n Bancaria Caja de Ahorros de Asturias. M.B. is funded by Agencia
Estatal de Investigacio´n (AEI/MCI/10.13039/501100011033, project RETOS
SAF2017-82623-R), cofunded by European Regional Development Fund, ‘‘A
way of making Europe’’; Comunidad de Madrid with the Sinergy Project
COVIDPREclinicalMODels-CM and the ERC under the European Union’s
Horizon 2020 research and innovation programme (grant 882385) through
the project ERC-AvG SHELTERINS. The CNIO, certified as Severo Ochoa
Centre of Excellence by AEI/MCI/10.13039/501100011033, is supported by
the Spanish Government through the Instituto de Salud Carlos III. L.P. is
supported by Horizon 2020 Framework Programme 741989, the Max Planck
Society, and the BBSRC.M.S. is funded by a core grant from the IRB, La Caixa
Foundation, the Milky Way Research Foundation, and Secretaria d’Universi-
tats i Recerca del Departament d’Empresa i Coneixement of Catalonia (Grup
de Recerca Consolidat 2017 SGR 282). G.K. is supported by the Ligue contre
le Cancer (e´quipe labellise´e); Agence National de la Recherche (ANR)–Projets
blancs; AMMICa US23/CNRS UMS3655; Association pour la recherche sur le
cancer (ARC); Cance´ropoˆle Ile-de-France; Fondation pour la Recherche
Me´dicale (FRM); a donation by Elior; Equipex Onco-Pheno-Screen; European268 Cell 186, January 19, 2023Joint Programme on Rare Diseases; the European Union Horizon 2020
Projects Oncobiome and Crimson; Fondation Carrefour; Institut National du
Cancer; Institut Universitaire de France; LabEx Immuno-Oncology (ANR-18-
IDEX-0001); a Cancer Research ASPIRE Award from the Mark Foundation;
the RHU Immunolife; Seerave Foundation; SIRIC Stratified Oncology Cell
DNA Repair and Tumor Immune Elimination (SOCRATE); and SIRIC Cancer
Research and Personalized Medicine (CARPEM). This study contributes to
the IdEx Universite´ de Paris ANR-18-IDEX-0001.
DECLARATION OF INTERESTS
M.A.B. is founder and shareholder of Life Length, SL, which commercializes
telomere length measurements for biomedical use. M.S. is shareholder and
advisor of Rejuveron Senescence Therapeutics, AG, and Altos Labs, Inc.;
and shareholder of Senolytic Therapeutics, Inc., and Life Biosciences, Inc.
G.K. has been holding research contracts with Daiichi Sankyo, Eleor, Kaleido,
Lytix Pharma, PharmaMar, Osasuna Therapeutics, Samsara Therapeutics,
Sanofi, Sotio, Tollys, Vascage, and Vasculox/Tioma; consulting for Reithera
and is on the Board of Directors of the Bristol Myers Squibb Foundation
France. G.K. is a scientific co-founder of everImmune, Osasuna Therapeutics,
Samsara Therapeutics, and Therafast Bio. G.K. is the inventor of patents
covering therapeutic targeting of aging, cancer, cystic fibrosis, and metabolic
disorders and has been holding research contracts with Daiichi Sankyo, Eleor,
Kaleido, Lytix Pharma, PharmaMar, Osasuna Therapeutics, Samsara Thera-
peutics, Sanofi, Tollys, and Vascage; has been consulting for Reithera; is on
the Board of Directors of the Bristol Myers Squibb Foundation France, and
is a scientific co-founder of everImmune, Osasuna Therapeutics, Samsara
Therapeutics, and Therafast Bio. G.K.’s wife, Laurence Zitvogel has held
research contracts with 9 Meters Biopharma, Daiichi Sankyo, Pilege, was on
the on the Board of Directors of Transgene, is a co-founder of everImmune,
and holds patents covering the treatment of cancer and the therapeutic manip-
ulation of the microbiota. G.K.’s brother, Romano Kroemer, was an employee
of Sanofi and has consulted for Boehringer-Ingelheim. None of the funders had
any role in study design, data collection and analysis, decision to publish, or
preparation of the manuscript.
REFERENCES
1. Lo´pez-Otı´n, C., Blasco, M.A., Partridge, L., Serrano, M., and Kroemer,
G. (2013). The hallmarks of aging. Cell 153, 1194–1217. https://doi.org/
10.1016/j.cell.2013.05.039.
2. Fraser, H.C., Kuan, V., Johnen, R., Zwierzyna, M., Hingorani, A.D., Be-
yer, A., and Partridge, L. (2022). Biological mechanisms of aging predict
age-related disease co-occurrence in patients. Aging Cell 21, e13524.
https://doi.org/10.1111/acel.13524.
3. Vijg, J., and Dong, X. (2020). Pathogenic mechanisms of somatic muta-
tion and genome mosaicism in aging. Cell 182, 12–23. https://doi.org/
10.1016/j.cell.2020.06.024.
4. Miller, K.N., Victorelli, S.G., Salmonowicz, H., Dasgupta, N., Liu, T., Pas-
sos, J.F., and Adams, P.D. (2021). Cytoplasmic DNA: sources, sensing,
and role in aging and disease. Cell 184, 5506–5526. https://doi.org/10.
1016/j.cell.2021.09.034.
5. Huang, Z., Sun, S., Lee, M., Maslov, A.Y., Shi, M., Waldman, S., Marsh,
A., Siddiqui, T., Dong, X., Peter, Y., et al. (2022). Single-cell analysis of
somatic mutations in human bronchial epithelial cells in relation to aging
and smoking. Nat. Genet. 54, 492–498. https://doi.org/10.1038/
s41588-022-01035-w.
6. Blokzijl, F., de Ligt, J., Jager, M., Sasselli, V., Roerink, S., Sasaki, N.,
Huch, M., Boymans, S., Kuijk, E., Prins, P., et al. (2016). Tissue-specific
mutation accumulation in human adult stem cells during life. Nature
538, 260–264. https://doi.org/10.1038/nature19768.
7. Miller, M.B., Huang, A.Y., Kim, J., Zhou, Z., Kirkham, S.L., Maury, E.A.,
Ziegenfuss, J.S., Reed, H.C., Neil, J.E., Rento, L., et al. (2022). Somatic
genomic changes in single Alzheimer’s disease neurons. Nature 604,
714–722. https://doi.org/10.1038/s41586-022-04640-1.
ll
Review8. Martincorena, I., Fowler, J.C., Wabik, A., Lawson, A.R.J., Abascal, F.,
Hall, M.W.J., Cagan, A., Murai, K., Mahbubani, K., Stratton, M.R.,
et al. (2018). Somatic mutant clones colonize the human esophagus
with age. Science 362, 911–917. https://doi.org/10.1126/science.
aau3879.
9. Nik-Zainal, S., and Hall, B.A. (2019). Cellular survival over genomic
perfection. Science 366, 802–803. https://doi.org/10.1126/science.
aax8046.
10. Balmain, A. (2020). The critical roles of somatic mutations and environ-
mental tumor-promoting agents in cancer risk. Nat. Genet. 52, 1139–
1143. https://doi.org/10.1038/s41588-020-00727-5.
11. Cagan, A., Baez-Ortega, A., Brzozowska, N., Abascal, F., Coorens,
T.H.H., Sanders, M.A., Lawson, A.R.J., Harvey, L.M.R., Bhosle, S.,
Jones, D., et al. (2022). Somatic mutation rates scale with lifespan across
mammals. Nature 604, 517–524. https://doi.org/10.1038/s41586-022-
04618-z.
12. Hennekam, R.C.M. (2020). Pathophysiology of premature aging charac-
teristics in Mendelian progeroid disorders. Eur. J. Med. Genet. 63,
104028. https://doi.org/10.1016/j.ejmg.2020.104028.
13. North, B.J., Rosenberg,M.A., Jeganathan, K.B., Hafner, A.V., Michan, S.,
Dai, J., Baker, D.J., Cen, Y., Wu, L.E., Sauve, A.A., et al. (2014). SIRT2 in-
duces the checkpoint kinase BubR1 to increase lifespan. EMBO J. 33,
1438–1453. https://doi.org/10.15252/embj.201386907.
14. Quesada, V., Freitas-Rodrı´guez, S., Miller, J., Pe´rez-Silva, J.G., Jiang,
Z.F., Tapia, W., Santiago-Ferna´ndez, O., Campos-Iglesias, D., Kuderna,
L.F.K., Quinzin, M., et al. (2019). Giant tortoise genomes provide insights
into longevity and age-related disease. Nat. Ecol. Evol. 3, 87–95. https://
doi.org/10.1038/s41559-018-0733-x.
15. Tian, X., Firsanov, D., Zhang, Z., Cheng, Y., Luo, L., Tombline, G., Tan, R.,
Simon, M., Henderson, S., Steffan, J., et al. (2019). SIRT6 is responsible
for more efficient DNA double-strand break repair in long-lived species.
Cell 177, 622–638.e22. https://doi.org/10.1016/j.cell.2019.03.043.
16. Roichman, A., Elhanati, S., Aon, M.A., Abramovich, I., Di Francesco, A.,
Shahar, Y., Avivi, M.Y., Shurgi, M., Rubinstein, A., Wiesner, Y., et al.
(2021). Restoration of energy homeostasis by SIRT6 extends healthy life-
span. Nat. Commun. 12, 3208. https://doi.org/10.1038/s41467-021-
23545-7.
17. Michel, M., Benı´tez-Buelga, C., Calvo, P.A., Hanna, B.M.F., Mortuse-
wicz, O., Masuyer, G., Davies, J., Wallner, O., Sanjiv, K., Albers, J.J.,
et al. (2022). Small-molecule activation of OGG1 increases oxidative
DNA damage repair by gaining a new function. Science 376, 1471–
1476. https://doi.org/10.1126/science.abf8980.
18. Gordon, L.B., Rothman, F.G., Lo´pez-Otı´n, C., and Misteli, T. (2014). Pro-
geria: a paradigm for translational medicine. Cell 156, 400–407. https://
doi.org/10.1016/j.cell.2013.12.028.
19. Toma´s-Loba, A., Flores, I., Ferna´ndez-Marcos, P.J., Cayuela, M.L.,
Maraver, A., Tejera, A., Borra´s, C., Matheu, A., Klatt, P., Flores, J.M.,
et al. (2008). Telomerase reverse transcriptase delays aging in cancer-
resistant mice. Cell 135, 609–622. https://doi.org/10.1016/j.cell.2008.
09.034.
20. Jaskelioff, M., Muller, F.L., Paik, J.-H., Thomas, E., Jiang, S., Adams,
A.C., Sahin, E., Kost-Alimova, M., Protopopov, A., Cadin˜anos, J., et al.
(2011). Telomerase reactivation reverses tissue degeneration in aged
telomerase-deficient mice. Nature 469, 102–106. https://doi.org/10.
1038/nature09603.
21. Bernardes de Jesus, B., Vera, E., Schneeberger, K., Tejera, A.M., Ayuso,
E., Bosch, F., and Blasco, M.A. (2012). Telomerase gene therapy in adult
and old mice delays aging and increases longevity without increasing
cancer. EMBO Mol. Med. 4, 691–704. https://doi.org/10.1002/emmm.
201200245.
22. Mun˜oz-Lorente, M.A., Cano-Martin, A.C., and Blasco, M.A. (2019). Mice
with hyper-long telomeres show less metabolic aging and longer life-
spans. Nat. Commun. 10, 4723. https://doi.org/10.1038/s41467-019-
12664-x.23. Shim, H.S., Horner, J.W., Wu, C.-J., Li, J., Lan, Z.D., Jiang, S., Xu, X.,
Hsu,W.-H., Zal, T., Flores, I.I., et al. (2021). Telomerase reverse transcrip-
tase preserves neuron survival and cognition in Alzheimer’s disease
models. Nat Aging 1, 1162–1174. https://doi.org/10.1038/s43587-021-
00146-z.
24. Povedano, J.M., Martinez, P., Serrano, R., Tejera, A´., Go´mez-Lo´pez, G.,
Bobadilla, M., Flores, J.M., Bosch, F., and Blasco, M.A. (2018). Thera-
peutic effects of telomerase in mice with pulmonary fibrosis induced by
damage to the lungs and short telomeres. eLife 7, e31299. https://doi.
org/10.7554/eLife.31299.
25. Ba¨r, C., Povedano, J.M., Serrano, R., Benitez-Buelga, C., Popkes, M.,
Formentini, I., Bobadilla, M., Bosch, F., and Blasco, M.A. (2016). Telome-
rase gene therapy rescues telomere length, bone marrow aplasia, and
survival in mice with aplastic anemia. Blood 127, 1770–1779. https://
doi.org/10.1182/blood-2015-08-667485.
26. Demidenko, O., Barardo, D., Budovskii, V., Finnemore, R., Palmer, F.R.,
Kennedy, B.K., and Budovskaya, Y.V. (2021). Rejuvant, a potential life-
extending compound formulation with alpha-ketoglutarate and vitamins,
conferred an average 8 year reduction in biological aging, after an
average of 7 months of use, in the TruAge DNA methylation test. Aging
(Albany, NY) 13, 24485–24499. https://doi.org/10.18632/aging.203736.
27. Fahy, G.M., Brooke, R.T., Watson, J.P., Good, Z., Vasanawala, S.S.,
Maecker, H., Leipold, M.D., Lin, D.T.S., Kobor, M.S., and Horvath, S.
(2019). Reversal of epigenetic aging and immunosenescent trends in hu-
mans. Aging Cell 18, e13028. https://doi.org/10.1111/acel.13028.
28. Wang, W., Zheng, Y., Sun, S., Li, W., Song, M., Ji, Q., Wu, Z., Liu, Z., Fan,
Y., Liu, F., et al. (2021). A genome-wide CRISPR-based screen identifies
KAT7 as a driver of cellular senescence. Sci. Transl. Med. 13, eabd2655.
https://doi.org/10.1126/scitranslmed.abd2655.
29. Bhatt V., Tiwari A.K. Sirtuins, a key regulator of ageing and age-related
neurodegenerative diseases. Int. J. Neurosci. 2022, Published online
May 13, 2022:1–26. https://doi.org/10.1080/00207454.2022.2057849.
30. He,W.-Z., Yang,M., Jiang, Y., He, C., Sun, Y.-C., Liu, L., Huang, M., Jiao,
Y.-R., Chen, K.-X., Hou, J., et al. (2022). miR-188-3p targets skeletal
endothelium coupling of angiogenesis and osteogenesis during ageing.
Cell Death Dis. 13, 494. https://doi.org/10.1038/s41419-022-04902-w.
31. Kumar, S., Morton, H., Sawant, N., Orlov, E., Bunquin, L.E., Pradeep-
kiran, J.A., Alvir, R., and Reddy, P.H. (2021). MicroRNA-455-3p improves
synaptic, cognitive functions and extends lifespan: relevance to Alz-
heimer’s disease. Redox Biol. 48, 102182. https://doi.org/10.1016/j.
redox.2021.102182.
32. Simon, M., Van Meter, M., Ablaeva, J., Ke, Z., Gonzalez, R.S., Taguchi,
T., De Cecco, M., Leonova, K.I., Kogan, V., Helfand, S.L., et al. (2019).
LINE1 derepression in aged wild-type and SIRT6-deficient mice drives
inflammation. Cell Metab. 29, 871–885.e5. https://doi.org/10.1016/j.
cmet.2019.02.014.
33. Dong, S., Wang, Q., Kao, Y.-R., Diaz, A., Tasset, I., Kaushik, S., Thiruthu-
vanathan, V., Zintiridou, A., Nieves, E., Dzieciatkowska, M., et al. (2021).
Chaperone-mediated autophagy sustains haematopoietic stem-cell
function. Nature 591, 117–123. https://doi.org/10.1038/s41586-020-
03129-z.
34. Madrigal-Matute, J., de Bruijn, J., van Kuijk, K., Riascos-Bernal, D.F.,
Diaz, A., Tasset, I., Martı´n-Segura, A., Gijbels, M.J.J., Sander, B.,
Kaushik, S., et al. (2022). Protective role of chaperone-mediated auto-
phagy against atherosclerosis. Proc. Natl. Acad. Sci. USA 119.
e2121133119. https://doi.org/10.1073/pnas.2121133119.
35. Bobkova, N.V., Evgen’ev, M., Garbuz, D.G., Kulikov, A.M., Morozov, A.,
Samokhin, A., Velmeshev, D., Medvinskaya, N., Nesterova, I., Pollock,
A., et al. (2015). Exogenous Hsp70 delays senescence and improves
cognitive function in aging mice. Proc. Natl. Acad. Sci. USA 112,
16006–16011. https://doi.org/10.1073/pnas.1516131112.
36. Hafycz, J.M., Strus, E., and Naidoo, N. (2022). Reducing ER stress with
chaperone therapy reverses sleep fragmentation and cognitive decline
in agedmice. Aging Cell 21, e13598. https://doi.org/10.1111/acel.13598.Cell 186, January 19, 2023 269
ll
Review37. Dalla Bella, E., Bersano, E., Antonini, G., Borghero, G., Capasso, M., Ca-
ponnetto, C., Chio`, A., Corbo, M., Filosto, M., Giannini, F., et al. (2021).
The unfolded protein response in amyotrophic later sclerosis: results of
a phase 2 trial. Brain 144, 2635–2647. https://doi.org/10.1093/brain/
awab167.
38. Pyo, J.-O., Yoo, S.-M., Ahn, H.-H., Nah, J., Hong, S.-H., Kam, T.-I., Jung,
S., and Jung, Y.-K. (2013). Overexpression of Atg5 in mice activates
autophagy and extends lifespan. Nat. Commun. 4, 2300. https://doi.
org/10.1038/ncomms3300.
39. Ferna´ndez, A´.F., Sebti, S., Wei, Y., Zou, Z., Shi, M., McMillan, K.L., He,
C., Ting, T., Liu, Y., Chiang, W.-C., et al. (2018). Disruption of the beclin
1–BCL2 autophagy regulatory complex promotes longevity in mice. Na-
ture 558, 136–140. https://doi.org/10.1038/s41586-018-0162-7.
40. Wang, C., Haas,M., Yeo, S.K., Sebti, S., Ferna´ndez, A´.F., Zou, Z., Levine,
B., and Guan, J.-L. (2022). Enhanced autophagy in Becn1F121A/F121A
knockin mice counteracts aging-related neural stem cell exhaustion
and dysfunction. Autophagy 18, 409–422. https://doi.org/10.1080/
15548627.2021.1936358.
41. Eisenberg, T., Abdellatif, M., Schroeder, S., Primessnig, U., Stekovic, S.,
Pendl, T., Harger, A., Schipke, J., Zimmermann, A., Schmidt, A., et al.
(2016). Cardioprotection and lifespan extension by the natural polyamine
spermidine. Nat. Med. 22, 1428–1438. https://doi.org/10.1038/nm.4222.
42. Castoldi, F., Humeau, J., Martins, I., Lachkar, S., Loew, D., Dingli, F., Du-
rand, S., Enot, D., Bossut, N., Chery, A., et al. (2020). Autophagy-medi-
ated metabolic effects of aspirin. Cell Death Discov. 6, 129. https://doi.
org/10.1038/s41420-020-00365-0.
43. Tezil, T., Chamoli, M., Ng, C.-P., Simon, R.P., Butler, V.J., Jung, M., An-
dersen, J., Kao, A.W., and Verdin, E. (2019). Lifespan-increasing drug
nordihydroguaiaretic acid inhibits p300 and activates autophagy. NPJ
Aging Mech. Dis. 5, 7. https://doi.org/10.1038/s41514-019-0037-7.
44. Yoshino, M., Yoshino, J., Kayser, B.D., Patti, G.J., Franczyk, M.P., Mills,
K.F., Sindelar, M., Pietka, T., Patterson, B.W., Imai, S.-I., et al. (2021).
Nicotinamide mononucleotide increases muscle insulin sensitivity in pre-
diabetic women. Science 372, 1224–1229. https://doi.org/10.1126/sci-
ence.abe9985.
45. Brakedal, B., Do¨lle, C., Riemer, F., Ma, Y., Nido, G.S., Skeie, G.O.,
Craven, A.R., Schwarzlmu¨ller, T., Brekke, N., Diab, J., et al. (2022). The
NADPARK study: A randomized phase I trial of nicotinamide riboside
supplementation in Parkinson’s disease. Cell Metab. 34, 396–407.e6.
https://doi.org/10.1016/j.cmet.2022.02.001.
46. Chen, A.C., Martin, A.J., Choy, B., Ferna´ndez-Pen˜as, P., Dalziell, R.A.,
McKenzie, C.A., Scolyer, R.A., Dhillon, H.M., Vardy, J.L., Kricker, A.,
et al. (2015). A phase 3 randomized trial of nicotinamide for skin-cancer
chemoprevention. N. Engl. J. Med. 373, 1618–1626. https://doi.org/10.
1056/NEJMoa1506197.
47. Singh, A., D’Amico, D., Andreux, P.A., Fouassier, A.M., Blanco-Bose,W.,
Evans, M., Aebischer, P., Auwerx, J., and Rinsch, C. (2022). Urolithin A
improves muscle strength, exercise performance, and biomarkers of
mitochondrial health in a randomized trial in middle-aged adults. Cell
Rep. Med. 3, 100633. https://doi.org/10.1016/j.xcrm.2022.100633.
48. Duran-Ortiz, S., List, E.O., Ikeno, Y., Young, J., Basu, R., Bell, S.,
McHugh, T., Funk, K., Mathes, S., Qian, Y., et al. (2021). Growth hormone
receptor gene disruption in mature-adult mice improves male insulin
sensitivity and extends female lifespan. Aging Cell 20, e13506. https://
doi.org/10.1111/acel.13506.
49. Acosta-Rodrı´guez, V., Rijo-Ferreira, F., Izumo, M., Xu, P., Wight-Carter,
M., Green, C.B., and Takahashi, J.S. (2022). Circadian alignment of early
onset caloric restriction promotes longevity in male C57BL/6J mice. Sci-
ence 376, 1192–1202. https://doi.org/10.1126/science.abk0297.
50. Spadaro, O., Youm, Y., Shchukina, I., Ryu, S., Sidorov, S., Ravussin, A.,
Nguyen, K., Aladyeva, E., Predeus, A.N., Smith, S.R., et al. (2022). Caloric
restriction in humans reveals immunometabolic regulators of health
span. Science 375, 671–677. https://doi.org/10.1126/science.abg7292.270 Cell 186, January 19, 202351. Fan, S.-Z., Lin, C.-S., Wei, Y.-W., Yeh, S.-R., Tsai, Y.-H., Lee, A.C., Lin,
W.-S., andWang, P.-Y. (2021). Dietary citrate supplementation enhances
longevity, metabolic health, and memory performance through promot-
ing ketogenesis. Aging Cell 20, e13510. https://doi.org/10.1111/
acel.13510.
52. Tavallaie, M., Voshtani, R., Deng, X., Qiao, Y., Jiang, F., Collman, J.P.,
and Fu, L. (2020). Moderation of mitochondrial respiration mitigates
metabolic syndrome of aging. Proc. Natl. Acad. Sci. USA 117, 9840–
9850. https://doi.org/10.1073/pnas.1917948117.
53. Goedeke, L., Murt, K.N., Di Francesco, A., Camporez, J.P., Nasiri, A.R.,
Wang, Y., Zhang, X.-M., Cline, G.W., de Cabo, R., and Shulman, G.I.
(2022). Sex- and strain-specific effects of mitochondrial uncoupling on
age-related metabolic diseases in high-fat diet-fed mice. Aging Cell 21,
e13539. https://doi.org/10.1111/acel.13539.
54. Zhang, H., Alder, N.N., Wang, W., Szeto, H., Marcinek, D.J., and Rabino-
vitch, P.S. (2020). Reduction of elevated proton leak rejuvenates mito-
chondria in the aged cardiomyocyte. eLife 9, e60827. https://doi.org/
10.7554/eLife.60827.
55. Goedeke, L., Peng, L., Montalvo-Romeral, V., Butrico, G.M., Dufour, S.,
Zhang, X.-M., Perry, R.J., Cline, G.W., Kievit, P., Chng, K., et al. (2019).
Controlled-releasemitochondrial protonophore (CRMP) reverses dyslipi-
demia and hepatic steatosis in dysmetabolic nonhuman primates. Sci.
Transl. Med. 11, eaay0284. https://doi.org/10.1126/scitranslmed.
aay0284.
56. Reid Thompson, W., Hornby, B., Manuel, R., Bradley, E., Laux, J., Carr,
J., and Vernon, H.J. (2021). A phase 2/3 randomized clinical trial followed
by an open-label extension to evaluate the effectiveness of elamipretide
in Barth syndrome, a genetic disorder of mitochondrial cardiolipin meta-
bolism. Genet. Med. 23, 471–478. https://doi.org/10.1038/s41436-020-
01006-8.
57. Chee, C., Shannon, C.E., Burns, A., Selby, A.L., Wilkinson, D., Smith, K.,
Greenhaff, P.L., and Stephens, F.B. (2021). Increasing skeletal muscle
carnitine content in older individuals increases whole-body fat oxidation
during moderate-intensity exercise. Aging Cell 20, e13303. https://doi.
org/10.1111/acel.13303.
58. Baker, D.J., Childs, B.G., Durik, M., Wijers, M.E., Sieben, C.J., Zhong, J.,
Saltness, R.A., Jeganathan, K.B., Verzosa, G.C., Pezeshki, A., et al.
(2016). Naturally occurring p16(Ink4a)-positive cells shorten healthy life-
span. Nature 530, 184–189. https://doi.org/10.1038/nature16932.
59. Xu, M., Pirtskhalava, T., Farr, J.N., Weigand, B.M., Palmer, A.K., Wei-
voda, M.M., Inman, C.L., Ogrodnik, M.B., Hachfeld, C.M., Fraser, D.G.,
et al. (2018). Senolytics improve physical function and increase lifespan
in old age. Nat. Med. 24, 1246–1256. https://doi.org/10.1038/s41591-
018-0092-9.
60. Yousefzadeh, M.J., Zhu, Y., McGowan, S.J., Angelini, L., Fuhrmann-
Stroissnigg, H., Xu, M., Ling, Y.Y., Melos, K.I., Pirtskhalava, T., Inman,
C.L., et al. (2018). Fisetin is a senotherapeutic that extends health and
lifespan. EBiomedicine 36, 18–28. https://doi.org/10.1016/j.ebiom.
2018.09.015.
61. Justice, J.N., Nambiar, A.M., Tchkonia, T., LeBrasseur, N.K., Pascual, R.,
Hashmi, S.K., Prata, L., Masternak, M.M., Kritchevsky, S.B., Musi, N.,
et al. (2019). Senolytics in idiopathic pulmonary fibrosis: results from a
first-in-human, open-label, pilot study. EBioMedicine 40, 554–563.
https://doi.org/10.1016/j.ebiom.2018.12.052.
62. Hickson, L.J., Langhi Prata, L.G.P., Bobart, S.A., Evans, T.K., Giorgadze,
N., Hashmi, S.K., Herrmann, S.M., Jensen, M.D., Jia, Q., Jordan, K.L.,
et al. (2019). Senolytics decrease senescent cells in humans: preliminary
report from a clinical trial of dasatinib plus quercetin in individuals with
diabetic kidney disease. EBioMedicine 47, 446–456. https://doi.org/10.
1016/j.ebiom.2019.08.069.
63. Ocampo, A., Reddy, P., Martinez-Redondo, P., Platero-Luengo, A., Ha-
tanaka, F., Hishida, T., Li, M., Lam, D., Kurita, M., Beyret, E., et al. (2016).
In vivo amelioration of age-associated hallmarks by partial
ll
Reviewreprogramming. Cell 167, 1719–1733.e12. https://doi.org/10.1016/j.cell.
2016.11.052.
64. Browder, K.C., Reddy, P., Yamamoto, M., Haghani, A., Guillen, I.G.,
Sahu, S., Wang, C., Luque, Y., Prieto, J., Shi, L., et al. (2022). In vivo par-
tial reprogramming alters age-associated molecular changes during
physiological aging in mice. Nat Aging 2, 243–253. https://doi.org/10.
1038/s43587-022-00183-2.
65. Chen, Y., Lu¨ttmann, F.F., Schoger, E., Scho¨ler, H.R., Zelaraya´n, L.C.,
Kim, K.-P., Haigh, J.J., Kim, J., and Braun, T. (2021). Reversible
reprogramming of cardiomyocytes to a fetal state drives heart regenera-
tion in mice. Science 373, 1537–1540. https://doi.org/10.1126/science.
abg5159.
66. Hishida, T., Yamamoto, M., Hishida-Nozaki, Y., Shao, C., Huang, L.,
Wang, C., Shojima, K., Xue, Y., Hang, Y., Shokhirev, M., et al. (2022).
In vivo partial cellular reprogramming enhances liver plasticity and regen-
eration. Cell Rep. 39, 110730. https://doi.org/10.1016/j.celrep.2022.
110730.
67. Rodrı´guez-Matella´n, A., Alcazar, N., Herna´ndez, F., Serrano, M., and
A´vila, J. (2020). In vivo reprogramming ameliorates aging features in den-
tate gyrus cells and improves memory in mice. Stem Cell Rep. 15, 1056–
1066. https://doi.org/10.1016/j.stemcr.2020.09.010.
68. Gao, X., Wang, X., Xiong, W., and Chen, J. (2016). In vivo reprogramming
reactive glia into iPSCs to produce new neurons in the cortex following
traumatic brain injury. Sci. Rep. 6, 22490. https://doi.org/10.1038/
srep22490.
69. Doeser, M.C., Scho¨ler, H.R., andWu, G. (2018). Reduction of fibrosis and
scar formation by partial reprogramming in vivo. Stem Cells 36, 1216–
1225. https://doi.org/10.1002/stem.2842.
70. Lu, Y., Brommer, B., Tian, X., Krishnan, A., Meer, M., Wang, C., Vera,
D.L., Zeng, Q., Yu, D., Bonkowski, M.S., et al. (2020). Reprogramming
to recover youthful epigenetic information and restore vision. Nature
588, 124–129. https://doi.org/10.1038/s41586-020-2975-4.
71. Mehdipour, M., Skinner, C., Wong, N., Lieb, M., Liu, C., Etienne, J., Kato,
C., Kiprov, D., Conboy, M.J., and Conboy, I.M. (2020). Rejuvenation of
three germ layers tissues by exchanging old blood plasma with saline-al-
bumin. Aging (Albany, NY) 12, 8790–8819. https://doi.org/10.18632/ag-
ing.103418.
72. Rebo, J., Mehdipour, M., Gathwala, R., Causey, K., Liu, Y., Conboy, M.J.,
and Conboy, I.M. (2016). A single heterochronic blood exchange reveals
rapid inhibition of multiple tissues by old blood. Nat. Commun. 7, 13363.
https://doi.org/10.1038/ncomms13363.
73. Castellano, J.M., Mosher, K.I., Abbey, R.J., McBride, A.A., James, M.L.,
Berdnik, D., Shen, J.C., Zou, B., Xie, X.S., Tingle, M., et al. (2017). Human
umbilical cord plasma proteins revitalize hippocampal function in aged
mice. Nature 544, 488–492. https://doi.org/10.1038/nature22067.
74. Ma, S., Wang, S., Ye, Y., Ren, J., Chen, R., Li, W., Li, J., Zhao, L., Zhao,
Q., Sun, G., et al. (2022). Heterochronic parabiosis induces stem cell revi-
talization and systemic rejuvenation across aged tissues. Cell Stem Cell
29, 990–1005.e10. https://doi.org/10.1016/j.stem.2022.04.017.
75. Pa´lovics, R., Keller, A., Schaum, N., Tan, W., Fehlmann, T., Borja, M.,
Kern, F., Bonanno, L., Calcuttawala, K., Webber, J., et al. (2022). Molec-
ular hallmarks of heterochronic parabiosis at single-cell resolution. Na-
ture 603, 309–314. https://doi.org/10.1038/s41586-022-04461-2.
76. Ballak, D.B., Brunt, V.E., Sapinsley, Z.J., Ziemba, B.P., Richey, J.J., Zi-
gler, M.C., Johnson, L.C., Gioscia-Ryan, R.A., Culp-Hill, R., Eisen-
messer, E.Z., et al. (2020). Short-term interleukin-37 treatment improves
vascular endothelial function, endurance exercise capacity, and whole-
body glucose metabolism in old mice. Aging Cell 19, e13074. https://
doi.org/10.1111/acel.13074.
77. Frohlich, J., and Vinciguerra, M. (2020). Candidate rejuvenating factor
GDF11 and tissue fibrosis: friend or foe? GeroScience 42, 1475–1498.
https://doi.org/10.1007/s11357-020-00279-w.78. Grunewald, M., Kumar, S., Sharife, H., Volinsky, E., Gileles-Hillel, A.,
Licht, T., Permyakova, A., Hinden, L., Azar, S., Friedmann, Y., et al.
(2021). Counteracting age-related VEGF signaling insufficiency pro-
motes healthy aging and extends life span. Science 373, eabc8479.
https://doi.org/10.1126/science.abc8479.
79. Sladitschek-Martens, H.L., Guarnieri, A., Brumana, G., Zanconato, F.,
Battilana, G., Xiccato, R.L., Panciera, T., Forcato, M., Bicciato, S., Guz-
zardo, V., et al. (2022). YAP/TAZ activity in stromal cells prevents ageing
by controlling cGAS-STING. Nature 607, 790–798. https://doi.org/10.
1038/s41586-022-04924-6.
80. Choi, H.R., Cho, K.A., Kang, H.T., Lee, J.B., Kaeberlein, M., Suh, Y.,
Chung, I.K., and Park, S.C. (2011). Restoration of senescent human
diploid fibroblasts by modulation of the extracellular matrix. Aging Cell
10, 148–157. https://doi.org/10.1111/j.1474-9726.2010.00654.x.
81. Yang, S., Gigout, S., Molinaro, A., Naito-Matsui, Y., Hilton, S., Foscarin,
S., Nieuwenhuis, B., Tan, C.L., Verhaagen, J., Pizzorusso, T., et al. (2021).
Chondroitin 6-sulphate is required for neuroplasticity and memory in
ageing. Mol. Psychiatry 26, 5658–5668. https://doi.org/10.1038/
s41380-021-01208-9.
82. Desdı´n-Mico´, G., Soto-Heredero, G., Aranda, J.F., Oller, J., Carrasco, E.,
Gabande´-Rodrı´guez, E., Blanco, E.M., Alfranca, A., Cusso´, L., Desco,M.,
et al. (2020). T cells with dysfunctional mitochondria induce multimorbid-
ity and premature senescence. Science 368, 1371–1376. https://doi.org/
10.1126/science.aax0860.
83. Sciorati, C., Gamberale, R., Monno, A., Citterio, L., Lanzani, C., De Lor-
enzo, R., Ramirez, G.A., Esposito, A., Manunta, P., Manfredi, A.A.,
et al. (2020). Pharmacological blockade of TNFa prevents sarcopenia
and prolongs survival in aging mice. Aging (Albany, NY) 12, 23497–
23508. https://doi.org/10.18632/aging.202200.
84. Gocmez, S.S., Yazir, Y., Gacar, G., Demirtasx Sxahin, T., Arkan, S., Karson,
A., andUtkan, T. (2020). Etanercept improves aging-induced cognitive def-
icits by reducing inflammation and vascular dysfunction in rats. Physiol.
Behav. 224, 113019. https://doi.org/10.1016/j.physbeh.2020.113019.
85. Minhas, P.S., Latif-Hernandez, A., McReynolds, M.R., Durairaj, A.S.,
Wang, Q., Rubin, A., Joshi, A.U., He, J.Q., Gauba, E., Liu, L., et al.
(2021). Restoring metabolism of myeloid cells reverses cognitive decline
in ageing. Nature 590, 122–128. https://doi.org/10.1038/s41586-020-
03160-0.
86. Marı´n-Aguilar, F., Lechuga-Vieco, A.V., Alcocer-Go´mez, E., Castejo´n-
Vega, B., Lucas, J., Garrido, C., Peralta-Garcia, A., Pe´rez-Pulido, A.J.,
Varela-Lo´pez, A., Quiles, J.L., et al. (2020). NLRP3 inflammasome sup-
pression improves longevity and prevents cardiac aging in male mice.
Aging Cell 19, e13050. https://doi.org/10.1111/acel.13050.
87. Ridker, P.M., MacFadyen, J.G., Thuren, T., Everett, B.M., Libby, P., and
Glynn, R.J.; CANTOS Trial Group (2017). Effect of interleukin-1b inhibition
with canakinumab on incident lung cancer in patients with atheroscle-
rosis: exploratory results from a randomised, double-blind, placebo-
controlled trial. Lancet 390, 1833–1842. https://doi.org/10.1016/S0140-
6736(17)32247-X.
88. Ba´rcena, C., Valde´s-Mas, R., Mayoral, P., Garabaya, C., Durand, S., Ro-
drı´guez, F., Ferna´ndez-Garcı´a, M.T., Salazar, N., Nogacka, A.M., Garata-
chea, N., et al. (2019). Healthspan and lifespan extension by fecal micro-
biota transplantation into progeroid mice. Nat. Med. 25, 1234–1242.
https://doi.org/10.1038/s41591-019-0504-5.
89. Lin, S.-W., Tsai, Y.-S., Chen, Y.-L., Wang, M.-F., Chen, C.-C., Lin, W.-H.,
and Fang, T.J. (2021). Lactobacillus plantarum GKM3 promotes
longevity, memory retention, and reduces brain oxidation stress in
SAMP8 mice. Nutrients 13, 2860. https://doi.org/10.3390/nu13082860.
90. Boehme, M., Guzzetta, K.E., Bastiaanssen, T.F.S., van de Wouw, M.,
Moloney, G.M., Gual-Grau, A., Spichak, S., Olavarrı´a-Ramı´rez, L., Fitz-
gerald, P., Morillas, E., et al. (2021). Microbiota from youngmice counter-
acts selective age-associated behavioral deficits. Nat Aging 1, 666–676.
https://doi.org/10.1038/s43587-021-00093-9.Cell 186, January 19, 2023 271
ll
Review91. Xu, L., Zhang, Q., Dou, X., Wang, Y., Wang, J., Zhou, Y., Liu, X., and Li, J.
(2022). Fecal microbiota transplantation from young donor mice im-
proves ovarian function in aged mice. J. Genet. Genomics. https://doi.
org/10.1016/j.jgg.2022.05.006.
92. Stebegg, M., Silva-Cayetano, A., Innocentin, S., Jenkins, T.P., Canta-
cessi, C., Gilbert, C., and Linterman, M.A. (2019). Heterochronic faecal
transplantation boosts gut germinal centres in agedmice. Nat. Commun.
10, 2443. https://doi.org/10.1038/s41467-019-10430-7.
93. Krishnan, S., Ding, Y., Saedi, N., Choi, M., Sridharan, G.V., Sherr, D.H.,
Yarmush, M.L., Alaniz, R.C., Jayaraman, A., and Lee, K. (2018). Gut mi-
crobiota-derived tryptophan metabolites modulate inflammatory
response in hepatocytes and macrophages. Cell Rep. 23, 1099–1111.
https://doi.org/10.1016/j.celrep.2018.03.109.
94. Cryan, J.F., O’Riordan, K.J., Cowan, C.S.M., Sandhu, K.V., Bastiaans-
sen, T.F.S., Boehme, M., Codagnone,M.G., Cussotto, S., Fulling, C., Go-
lubeva, A.V., et al. (2019). The microbiota-gut-brain axis. Physiol. Rev.
99, 1877–2013. https://doi.org/10.1152/physrev.00018.2018.
95. Depommier, C., Everard, A., Druart, C., Plovier, H., Van Hul, M., Vieira-
Silva, S., Falony, G., Raes, J., Maiter, D., Delzenne, N.M., et al. (2019).
Supplementation with Akkermansia muciniphila in overweight and obese
human volunteers: a proof-of-concept exploratory study. Nat. Med. 25,
1096–1103. https://doi.org/10.1038/s41591-019-0495-2.
96. Sanchez-Contreras, M., and Kennedy, S.R. (2022). The complicated na-
ture of somatic mtDNA mutations in aging. Front. Aging 2, 805126.
https://doi.org/10.3389/fragi.2021.805126.
97. Greaves, L.C., Nooteboom, M., Elson, J.L., Tuppen, H.A.L., Taylor, G.A.,
Commane, D.M., Arasaradnam, R.P., Khrapko, K., Taylor, R.W., Kirk-
wood, T.B.L., et al. (2014). Clonal expansion of early tomid-life mitochon-
drial DNA point mutations drives mitochondrial dysfunction during hu-
man ageing. PLoS Genet. 10, e1004620. https://doi.org/10.1371/
journal.pgen.1004620.
98. Arbeithuber, B., Cremona, M.A., Hester, J., Barrett, A., Higgins, B., An-
thony, K., Chiaromonte, F., Diaz, F.J., and Makova, K.D. (2022).
Advanced age increases frequencies of de novomitochondrial mutations
inmacaque oocytes and somatic tissues. Proc. Natl. Acad. Sci. USA 119.
e2118740119. https://doi.org/10.1073/pnas.2118740119.
99. Wang, Y., Guo, X., Ye, K., Orth,M., andGu, Z. (2021). Accelerated expan-
sion of pathogenic mitochondrial DNA heteroplasmies in Huntington’s
disease. Proc. Natl. Acad. Sci. USA 118. e2014610118. https://doi.org/
10.1073/pnas.2014610118.
100. Macken, W.L., Vandrovcova, J., Hanna, M.G., and Pitceathly, R.D.S.
(2021). Applying genomic and transcriptomic advances to mitochondrial
medicine. Nat. Rev. Neurol. 17, 215–230. https://doi.org/10.1038/
s41582-021-00455-2.
101. Lujan, S.A., Longley, M.J., Humble, M.H., Lavender, C.A., Burkholder, A.,
Blakely, E.L., Alston, C.L., Gorman, G.S., Turnbull, D.M., McFarland, R.,
et al. (2020). Ultrasensitive deletion detection links mitochondrial DNA
replication, disease, and aging. Genome Biol. 21, 248. https://doi.org/
10.1186/s13059-020-02138-5.
102. Shin, J.-Y., and Worman, H.J. (2022). Molecular pathology of laminopa-
thies. Annu. Rev. Pathol. 17, 159–180. https://doi.org/10.1146/annurev-
pathol-042220-034240.
103. Lai, W.-F., and Wong, W.-T. (2020). Progress and trends in the develop-
ment of therapies for Hutchinson-Gilford progeria syndrome. Aging Cell
19, e13175. https://doi.org/10.1111/acel.13175.
104. Dhillon, S. (2021). Lonafarnib: first approval. Drugs 81, 283–289. https://
doi.org/10.1007/s40265-020-01464-z.
105. Santiago-Ferna´ndez, O., Osorio, F.G., Quesada, V., Rodrı´guez, F.,
Basso, S., Maeso, D., Rolas, L., Barkaway, A., Nourshargh, S., Folgue-
ras, A.R., et al. (2019). Development of a CRISPR/Cas9-based therapy
for Hutchinson-Gilford progeria syndrome. Nat. Med. 25, 423–426.
https://doi.org/10.1038/s41591-018-0338-6.272 Cell 186, January 19, 2023106. Koblan, L.W., Erdos, M.R., Wilson, C., Cabral, W.A., Levy, J.M., Xiong,
Z.M., Tavarez, U.L., Davison, L.M., Gete, Y.G., Mao, X., et al. (2021).
In vivo base editing rescues Hutchinson-Gilford progeria syndrome in
mice. Nature 589, 608–614. https://doi.org/10.1038/s41586-020-
03086-7.
107. Blackburn, E.H., Epel, E.S., and Lin, J. (2015). Human telomere biology: A
contributory and interactive factor in aging, disease risks, and protection.
Science 350, 1193–1198. https://doi.org/10.1126/science.aab3389.
108. Chakravarti, D., LaBella, K.A., and DePinho, R.A. (2021). Telomeres: his-
tory, health, and hallmarks of aging. Cell 184, 306–322. https://doi.org/
10.1016/j.cell.2020.12.028.
109. Blasco, M.A. (2005). Telomeres and human disease: ageing, cancer and
beyond. Nat. Rev. Genet. 6, 611–622. https://doi.org/10.1038/nrg1656.
110. Lo´pez-Otı´n, C., Pietrocola, F., Roiz-Valle, D., Galluzzi, L., and Kroemer,
G. (2023). Meta-hallmarks of aging and cancer. Cell Metab. 35. https://
doi.org/10.1016/j.cmet.2022.11.001.
111. Alder, J.K., andArmanios,M. (2022). Telomere-mediated lungdisease. Phys-
iol. Rev. 102, 1703–1720. https://doi.org/10.1152/physrev.00046.2021.
112. Whittemore, K., Vera, E., Martı´nez-Nevado, E., Sanpera, C., and Blasco,
M.A. (2019). Telomere shortening rate predicts species life span. Proc.
Natl. Acad. Sci. USA 116, 15122–15127. https://doi.org/10.1073/pnas.
1902452116.
113. Lim, C.J., and Cech, T.R. (2021). Shaping human telomeres: from shel-
terin and CST complexes to telomeric chromatin organization. Nat.
Rev. Mol. Cell Biol. 22, 283–298. https://doi.org/10.1038/s41580-021-
00328-y.
114. Martı´nez, P., and Blasco, M.A. (2011). Telomeric and extra-telomeric
roles for telomerase and the telomere-binding proteins. Nat. Rev. Cancer
11, 161–176. https://doi.org/10.1038/nrc3025.
115. Rossiello, F., Jurk, D., Passos, J.F., and d’Adda di Fagagna, F. (2022).
Telomere dysfunction in ageing and age-related diseases. Nat. Cell
Biol. 24, 135–147. https://doi.org/10.1038/s41556-022-00842-x.
116. Saraswati, S., Martı´nez, P., Gran˜a-Castro, O., and Blasco, M.A. (2021).
Short and dysfunctional telomeres sensitize the kidneys to develop
fibrosis. Nat. Aging 1, 269–283. https://doi.org/10.1038/s43587-021-
00040-8.
117. Seale, K., Horvath, S., Teschendorff, A., Eynon, N., and Voisin, S. (2022).
Making sense of the ageing methylome. Nat. Rev. Genet. 23, 585–605.
https://doi.org/10.1038/s41576-022-00477-6.
118. Bejaoui, Y., Razzaq, A., Yousri, N.A., Oshima, J., Megarbane, A.,
Qannan, A., Potabattula, R., Alam, T., Martin, G.M., Horn, H.F., et al.
(2022). DNA methylation signatures in Blood DNA of Hutchinson-
Gilford progeria syndrome. Aging Cell 21, e13555. https://doi.org/10.
1111/acel.13555.
119. Horvath, S., and Raj, K. (2018). DNA methylation-based biomarkers and
the epigenetic clock theory of ageing. Nat. Rev. Genet. 19, 371–384.
https://doi.org/10.1038/s41576-018-0004-3.
120. Lu, Y.-X., Regan, J.C., Eßer, J., Drews, L.F., Weinseis, T., Stinn, J., Hahn,
O., Miller, R.A., Gro¨nke, S., and Partridge, L. (2021). A TORC1-histone
axis regulates chromatin organisation and non-canonical induction of
autophagy to ameliorate ageing. eLife 10, e62233. https://doi.org/10.
7554/eLife.62233.
121. Oh, E.S., and Petronis, A. (2021). Origins of human disease: the chrono-
epigenetic perspective. Nat. Rev. Genet. 22, 533–546. https://doi.org/10.
1038/s41576-021-00348-6.
122. Blasco, M.A. (2007). The epigenetic regulation of mammalian telomeres.
Nat. Rev. Genet. 8, 299–309. https://doi.org/10.1038/nrg2047.
123. Brown, K., Xie, S., Qiu, X., Mohrin, M., Shin, J., Liu, Y., Zhang, D., Scad-
den, D.T., and Chen, D. (2013). SIRT3 reverses aging-associated degen-
eration. Cell Rep. 3, 319–327. https://doi.org/10.1016/j.celrep.2013.
01.005.
124. Mostoslavsky, R., Chua, K.F., Lombard, D.B., Pang,W.W., Fischer, M.R.,
Gellon, L., Liu, P., Mostoslavsky, G., Franco, S., Murphy, M.M., et al.
ll
Review(2006). Genomic instability and aging-like phenotype in the absence of
mammalian SIRT6. Cell 124, 315–329. https://doi.org/10.1016/j.cell.
2005.11.044.
125. Chang, A.R., Ferrer, C.M., and Mostoslavsky, R. (2020). SIRT6, a
mammalian deacylase with multitasking abilities. Physiol. Rev. 100,
145–169. https://doi.org/10.1152/physrev.00030.2018.
126. Mohrin, M., Shin, J., Liu, Y., Brown, K., Luo, H., Xi, Y., Haynes, C.M., and
Chen, D. (2015). Stem cell aging. A mitochondrial UPR-mediated meta-
bolic checkpoint regulates hematopoietic stem cell aging. Science 347,
1374–1377. https://doi.org/10.1126/science.aaa2361.
127. Bonkowski, M.S., and Sinclair, D.A. (2016). Slowing ageing by design: the
rise of NAD+ and sirtuin-activating compounds. Nat. Rev. Mol. Cell Biol.
17, 679–690. https://doi.org/10.1038/nrm.2016.93.
128. Swer, P.B., and Sharma, R. (2021). ATP-dependent chromatin remodel-
ers in ageing and age-related disorders. Biogerontology 22, 1–17.
https://doi.org/10.1007/s10522-020-09899-3.
129. Larson, K., Yan, S.-J., Tsurumi, A., Liu, J., Zhou, J., Gaur, K., Guo, D.,
Eickbush, T.H., and Li,W.X. (2012). Heterochromatin formation promotes
longevity and represses ribosomal RNA synthesis. PLoS Genet. 8,
e1002473. https://doi.org/10.1371/journal.pgen.1002473.
130. Napoletano, F., Ferrari Bravo, G., Voto, I.A.P., Santin, A., Celora, L., Cam-
paner, E., Dezi, C., Bertossi, A., Valentino, E., Santorsola,M., et al. (2021).
The prolyl-isomerase PIN1 is essential for nuclear Lamin-B structure and
function and protects heterochromatin under mechanical stress. Cell
Rep. 36, 109694. https://doi.org/10.1016/j.celrep.2021.109694.
131. Jusic, A., Thomas, P.B., Wettinger, S.B., Dogan, S., Farrugia, R., Gae-
tano, C., Tuna, B.G., Pinet, F., Robinson, E.L., Tual-Chalot, S., et al.
(2022). Noncoding RNAs in age-related cardiovascular diseases. Ageing
Res. Rev. 77, 101610. https://doi.org/10.1016/j.arr.2022.101610.
132. Weigelt, C.M., Sehgal, R., Tain, L.S., Cheng, J., Eßer, J., Pahl, A., Dieter-
ich, C., Gro¨nke, S., and Partridge, L. (2020). An insulin-sensitive circular
RNA that regulates lifespan in Drosophila. Mol. Cell 79, 268–279.e5.
https://doi.org/10.1016/j.molcel.2020.06.011.
133. Ponting, C.P., and Haerty, W. (2022). Genome-wide analysis of human
long noncoding RNAs: a provocative review. Annu. Rev. Genomics
Hum. Genet. 23, 153–172. https://doi.org/10.1146/annurev-genom-
112921-123710.
134. Gorbunova, V., Seluanov, A., Mita, P., McKerrow, W., Fenyo¨, D., Boeke,
J.D., Linker, S.B., Gage, F.H., Kreiling, J.A., Petrashen, A.P., et al. (2021).
The role of retrotransposable elements in ageing and age-associated dis-
eases. Nature 596, 43–53. https://doi.org/10.1038/s41586-021-03542-y.
135. Della Valle, F., Reddy, P., Yamamoto, M., Liu, P., Saera-Vila, A., Bensad-
dek, D., Zhang, H., Prieto Martinez, J., Abassi, L., Celii, M., et al. (2022).
LINE-1 RNA causes heterochromatin erosion and is a target for amelio-
ration of senescent phenotypes in progeroid syndromes. Sci. Transl.
Med. 14, eabl6057. https://doi.org/10.1126/scitranslmed.abl6057.
136. De Cecco, M., Ito, T., Petrashen, A.P., Elias, A.E., Skvir, N.J., Criscione,
S.W., Caligiana, A., Brocculi, G., Adney, E.M., Boeke, J.D., et al. (2019).
L1 drives IFN in senescent cells and promotes age-associated inflamma-
tion. Nature 566, 73–78. https://doi.org/10.1038/s41586-018-0784-9.
137. Simon, M., Yang, J., Gigas, J., Earley, E.J., Hillpot, E., Zhang, L., Zagor-
ulya, M., Tombline, G., Gilbert, M., Yuen, S.L., et al. (2022). A rare human
centenarian variant of SIRT6 enhances genome stability and interaction
with Lamin A. EMBO J. 41, e110393. https://doi.org/10.15252/embj.
2021110393.
138. Hernando-Herraez, I., Evano, B., Stubbs, T., Commere, P.-H., Jan
Bonder, M., Clark, S., Andrews, S., Tajbakhsh, S., and Reik, W. (2019).
Ageing affects DNA methylation drift and transcriptional cell-to-cell vari-
ability in mouse muscle stem cells. Nat. Commun. 10, 4361. https://doi.
org/10.1038/s41467-019-12293-4.
139. Bhadra, M., Howell, P., Dutta, S., Heintz, C., and Mair, W.B. (2020). Alter-
native splicing in aging and longevity. Hum. Genet. 139, 357–369. https://
doi.org/10.1007/s00439-019-02094-6.140. Ijomone, O.M., Ijomone, O.K., Iroegbu, J.D., Ifenatuoha, C.W., Olung,
N.F., and Aschner, M. (2020). Epigenetic influence of environmentally
neurotoxic metals. Neurotoxicology 81, 51–65. https://doi.org/10.1016/
j.neuro.2020.08.005.
141. Tabula Muris Consortium (2020). A single-cell transcriptomic atlas char-
acterizes ageing tissues in the mouse. Nature 583, 590–595. https://doi.
org/10.1038/s41586-020-2496-1.
142. Hipp, M.S., Kasturi, P., and Hartl, F.U. (2019). The proteostasis network
and its decline in ageing. Nat. Rev. Mol. Cell Biol. 20, 421–435. https://
doi.org/10.1038/s41580-019-0101-y.
143. Martinez-Miguel, V.E., Lujan, C., Espie-Caullet, T., Martinez-Martinez, D.,
Moore, S., Backes, C., Gonzalez, S., Galimov, E.R., Brown, A.E.X., Halic,
M., et al. (2021). Increased fidelity of protein synthesis extends lifespan.
Cell Metab. 33, 2288–2300.e12. https://doi.org/10.1016/j.cmet.2021.
08.017.
144. Shcherbakov, D., Nigri, M., Akbergenov, R., Brilkova, M., Mantovani, M.,
Petit, P.I., Grimm, A., Karol, A.A., Teo, Y., Sancho´n, A.C., et al. (2022).
Premature aging in mice with error-prone protein synthesis. Sci. Adv.
8, eabl9051. https://doi.org/10.1126/sciadv.abl9051.
145. Gerashchenko, M.V., Peterfi, Z., Yim, S.H., and Gladyshev, V.N. (2021).
Translation elongation rate varies among organs and decreases with
age. Nucleic Acids Res. 49, e9. https://doi.org/10.1093/nar/gkaa1103.
146. Lo´pez-Otı´n, C., and Kroemer, G. (2021). Hallmarks of health. Cell 184,
33–63. https://doi.org/10.1016/j.cell.2020.11.034.
147. Hetz, C., Zhang, K., and Kaufman, R.J. (2020). Mechanisms, regulation
and functions of the unfolded protein response. Nat. Rev. Mol. Cell
Biol. 21, 421–438. https://doi.org/10.1038/s41580-020-0250-z.
148. Kelmer Sacramento, E., Kirkpatrick, J.M., Mazzetto, M., Baumgart, M.,
Bartolome, A., Di Sanzo, S., Caterino, C., Sanguanini, M., Papaevgeniou,
N., Lefaki, M., et al. (2020). Reduced proteasome activity in the aging
brain results in ribosome stoichiometry loss and aggregation. Mol.
Syst. Biol. 16, e9596. https://doi.org/10.15252/msb.20209596.
149. Yang, L., Ma, Z., Wang, H., Niu, K., Cao, Y., Sun, L., Geng, Y., Yang, B.,
Gao, F., Chen, Z., et al. (2019). Ubiquitylome study identifies increased
histone 2A ubiquitylation as an evolutionarily conserved aging biomarker.
Nat. Commun. 10, 2191. https://doi.org/10.1038/s41467-019-10136-w.
150. Kaushik, S., and Cuervo, A.M. (2018). The coming of age of chaperone-
mediated autophagy. Nat. Rev. Mol. Cell Biol. 19, 365–381. https://doi.
org/10.1038/s41580-018-0001-6.
151. Rodriguez-Navarro, J.A., Kaushik, S., Koga, H., Dall’Armi, C., Shui, G.,
Wenk, M.R., Di Paolo, G., and Cuervo, A.M. (2012). Inhibitory effect of di-
etary lipids on chaperone-mediated autophagy. Proc. Natl. Acad. Sci.
USA 109, E705–E714. https://doi.org/10.1073/pnas.1113036109.
152. Levine, B., and Kroemer, G. (2019). Biological functions of autophagy
genes: a disease perspective. Cell 176, 11–42. https://doi.org/10.1016/
j.cell.2018.09.048.
153. Klionsky, D.J., Petroni, G., Amaravadi, R.K., Baehrecke, E.H., Ballabio,
A., Boya, P., Bravo-San Pedro, J.M., Cadwell, K., Cecconi, F., Choi,
A.M.K., et al. (2021). Autophagy in major human diseases. EMBO J. 40,
e108863. https://doi.org/10.15252/embj.2021108863.
154. Tsakiri, E.N., Iliaki, K.K., Ho¨hn, A., Grimm, S., Papassideri, I.S., Grune, T.,
and Trougakos, I.P. (2013). Diet-derived advanced glycation end prod-
ucts or lipofuscin disrupts proteostasis and reduces life span in
Drosophila melanogaster. Free Radic. Biol. Med. 65, 1155–1163.
https://doi.org/10.1016/j.freeradbiomed.2013.08.186.
155. Bourdenx, M., Martı´n-Segura, A., Scrivo, A., Rodriguez-Navarro, J.A.,
Kaushik, S., Tasset, I., Diaz, A., Storm, N.J., Xin, Q., Juste, Y.R., et al.
(2021). Chaperone-mediated autophagy prevents collapse of the
neuronal metastable proteome. Cell 184, 2696–2714.e25. https://doi.
org/10.1016/j.cell.2021.03.048.
156. Munka´csy, E., Chocron, E.S., Quintanilla, L., Gendron, C.M., Pletcher,
S.D., and Pickering, A.M. (2019). Neuronal-specific proteasome
augmentation via Prosb5 overexpression extends lifespan and reducesCell 186, January 19, 2023 273
ll
Reviewage-related cognitive decline. Aging Cell 18, e13005. https://doi.org/10.
1111/acel.13005.
157. Derisbourg, M.J., Hartman, M.D., and Denzel, M.S. (2021). Perspective:
modulating the integrated stress response to slow aging and ameliorate
age-related pathology. Nat Aging 1, 760–768. https://doi.org/10.1038/
s43587-021-00112-9.
158. Marciniak, S.J., Chambers, J.E., and Ron, D. (2022). Pharmacological
targeting of endoplasmic reticulum stress in disease. Nat. Rev. Drug Dis-
cov. 21, 115–140. https://doi.org/10.1038/s41573-021-00320-3.
159. Shen, Z., Hinson, A., Miller, R.A., and Garcia, G.G. (2021). Cap-indepen-
dent translation: A shared mechanism for lifespan extension by rapamy-
cin, acarbose, and 17a-estradiol. Aging Cell 20, e13345. https://doi.org/
10.1111/acel.13345.
160. Kuo, C.-T., You, G.-T., Jian, Y.-J., Chen, T.-S., Siao, Y.-C., Hsu, A.-L.,
and Ching, T.-T. (2020). AMPK-mediated formation of stress granules
is required for dietary restriction-induced longevity in Caenorhabditis el-
egans. Aging Cell 19, e13157. https://doi.org/10.1111/acel.13157.
161. Humeau, J., Leduc, M., Cerrato, G., Loos, F., Kepp, O., and Kroemer, G.
(2020). Phosphorylation of eukaryotic initiation factor-2a (eIF2a) in auto-
phagy. Cell Death Dis. 11, 433. https://doi.org/10.1038/s41419-020-
2642-6.
162. Halliday, M., Hughes, D., and Mallucci, G.R. (2017). Fine-tuning PERK
signaling for neuroprotection. J. Neurochem. 142, 812–826. https://doi.
org/10.1111/jnc.14112.
163. Galluzzi, L., and Green, D.R. (2019). Autophagy-independent functions of
the autophagy machinery. Cell 177, 1682–1699. https://doi.org/10.1016/
j.cell.2019.05.026.
164. Nicola´s-A´vila, J.A., Lechuga-Vieco, A.V., Esteban-Martı´nez, L., Sa´nchez-
Dı´az, M., Dı´az-Garcı´a, E., Santiago, D.J., Rubio-Ponce, A., Li, J.L., Balac-
hander, A., Quintana, J.A., et al. (2020). A network of macrophages sup-
ports mitochondrial homeostasis in the heart. Cell 183, 94–109.e23.
https://doi.org/10.1016/j.cell.2020.08.031.
165. Lipinski, M.M., Zheng, B., Lu, T., Yan, Z., Py, B.F., Ng, A., Xavier, R.J., Li,
C., Yankner, B.A., Scherzer, C.R., et al. (2010). Genome-wide analysis re-
veals mechanisms modulating autophagy in normal brain aging and in
Alzheimer’s disease. Proc. Natl. Acad. Sci. USA 107, 14164–14169.
https://doi.org/10.1073/pnas.1009485107.
166. Raz, Y., Guerrero-Ros, I., Maier, A., Slagboom, P.E., Atzmon, G., Barzilai,
N., and Macian, F. (2017). Activation-induced autophagy is preserved in
CD4+ T-cells in familial longevity. J. Gerontol. A Biol. Sci. Med. Sci. 72,
1201–1206. https://doi.org/10.1093/gerona/glx020.
167. Zhang, H., Alsaleh, G., Feltham, J., Sun, Y., Napolitano, G., Riffelmacher,
T., Charles, P., Frau, L., Hublitz, P., Yu, Z., et al. (2019). Polyamines con-
trol eIF5A hypusination, TFEB translation, and autophagy to reverse B
cell senescence. Mol. Cell 76, 110–125.e9. https://doi.org/10.1016/j.
molcel.2019.08.005.
168. Alsaleh, G., Panse, I., Swadling, L., Zhang, H., Richter, F.C., Meyer, A.,
Lord, J., Barnes, E., Klenerman, P., Green, C., et al. (2020). Autophagy
in T cells from aged donors is maintained by spermidine and correlates
with function and vaccine responses. eLife 9, e57950. https://doi.org/
10.7554/eLife.57950.
169. Deretic, V., and Kroemer, G. (2022). Autophagy inmetabolism and quality
control: opposing, complementary or interlinked functions? Autophagy
18, 283–292. https://doi.org/10.1080/15548627.2021.1933742.
170. Cassidy, L.D., Young, A.R.J., Young, C.N.J., Soilleux, E.J., Fielder, E.,
Weigand, B.M., Lagnado, A., Brais, R., Ktistakis, N.T., Wiggins, K.A.,
et al. (2020). Temporal inhibition of autophagy reveals segmental reversal
of ageing with increased cancer risk. Nat. Commun. 11, 307. https://doi.
org/10.1038/s41467-019-14187-x.
171. Xu Y., Wan W. Acetylation in the regulation of autophagy. Autophagy
2022, Published online April 18, 2022:1–18. https://doi.org/10.1080/
15548627.2022.2062112.274 Cell 186, January 19, 2023172. Pietrocola, F., Pol, J., Vacchelli, E., Rao, S., Enot, D.P., Baracco, E.E.,
Levesque, S., Castoldi, F., Jacquelot, N., Yamazaki, T., et al. (2016).
Caloric restriction mimetics enhance anticancer immunosurveillance.
Cancer Cell 30, 147–160. https://doi.org/10.1016/j.ccell.2016.05.016.
173. Liang, Y., Piao, C., Beuschel, C.B., Toppe, D., Kollipara, L., Bogdanow,
B., Maglione, M., Lu¨tzkendorf, J., See, J.C.K., Huang, S., et al. (2021).
eIF5A hypusination, boosted by dietary spermidine, protects from pre-
mature brain aging and mitochondrial dysfunction. Cell Rep. 35,
108941. https://doi.org/10.1016/j.celrep.2021.108941.
174. Puleston, D.J., Baixauli, F., Sanin, D.E., Edwards-Hicks, J., Villa, M., Ka-
bat, A.M., Kami
nski, M.M., Stanckzak, M., Weiss, H.J., Grzes, K.M., et al.
(2021). Polyamine metabolism is a central determinant of helper T cell
lineage fidelity. Cell 184, 4186–4202.e20. https://doi.org/10.1016/j.cell.
2021.06.007.
175. Gobert, A.P., Latour, Y.L., Asim,M., Barry,D.P., Allaman,M.M., Finley, J.L.,
Smith, T.M., McNamara, K.M., Singh, K., Sierra, J.C., et al. (2022). Protec-
tive roleof spermidine incolitis andcoloncarcinogenesis.Gastroenterology
162, 813–827.e8. https://doi.org/10.1053/j.gastro.2021.11.005.
176. Katsyuba, E., Romani, M., Hofer, D., and Auwerx, J. (2020). NAD+ ho-
meostasis in health and disease. Nat. Metab. 2, 9–31. https://doi.org/
10.1038/s42255-019-0161-5.
177. D’Amico, D., Andreux, P.A., Valde´s, P., Singh, A., Rinsch, C., and Au-
werx, J. (2021). Impact of the natural compound urolithin a on health, dis-
ease, and aging. TrendsMol. Med. 27, 687–699. https://doi.org/10.1016/
j.molmed.2021.04.009.
178. Slack, C., Alic, N., Foley, A., Cabecinha, M., Hoddinott, M.P., and Par-
tridge, L. (2015). The Ras-Erk-ETS-signaling pathway is a drug target
for longevity. Cell 162, 72–83. https://doi.org/10.1016/j.cell.2015.06.023.
179. Singh, P.P., Demmitt, B.A., Nath, R.D., and Brunet, A. (2019). The ge-
netics of aging: A vertebrate perspective. Cell 177, 200–220. https://
doi.org/10.1016/j.cell.2019.02.038.
180. Ji, J.S., Liu, L., Shu, C., Yan, L.L., and Zeng, Y. (2021). Sex difference and
interaction of SIRT1 and FOXO3 candidate longevity genes on life expec-
tancy: a 10-year prospective longitudinal cohort study. J. Gerontol. A
Biol. Sci. Med. Sci. 77, glab378. https://doi.org/10.1093/gerona/
glab378.
181. Kabacik, S., Lowe, D., Fransen, L., Leonard, M., Ang, S.-L., Whiteman,
C., Corsi, S., Cohen, H., Felton, S., Bali, R., et al. (2022). The relationship
between epigenetic age and the hallmarks of aging in human cells. Nat
Aging 2, 484–493. https://doi.org/10.1038/s43587-022-00220-0.
182. Amorim, J.A., Coppotelli, G., Rolo, A.P., Palmeira, C.M., Ross, J.M., and
Sinclair, D.A. (2022). Mitochondrial and metabolic dysfunction in ageing
and age-related diseases. Nat. Rev. Endocrinol. 18, 243–258. https://
doi.org/10.1038/s41574-021-00626-7.
183. Orthofer, M., Valsesia, A., Ma¨gi, R., Wang, Q.-P., Kaczanowska, J., Ko-
zieradzki, I., Leopoldi, A., Cikes, D., Zopf, L.M., Tretiakov, E.O., et al.
(2020). Identification of ALK in thinness. Cell 181, 1246–1262.e22.
https://doi.org/10.1016/j.cell.2020.04.034.
184. Ahmed, M., Kaur, N., Cheng, Q., Shanabrough, M., Tretiakov, E.O., Har-
kany, T., Horvath, T.L., and Schlessinger, J. (2022). A hypothalamic
pathway for Augmentor a-controlled body weight regulation. Proc.
Natl. Acad. Sci. USA 119. e2200476119. https://doi.org/10.1073/pnas.
2200476119.
185. Partridge, L., Fuentealba, M., and Kennedy, B.K. (2020). The quest to
slow ageing through drug discovery. Nat. Rev. Drug Discov. 19, 513–
532. https://doi.org/10.1038/s41573-020-0067-7.
186. Mannick, J.B., Teo, G., Bernardo, P., Quinn, D., Russell, K., Klickstein, L.,
Marshall, W., and Shergill, S. (2021). Targeting the biology of ageing with
mTOR inhibitors to improve immune function in older adults: phase 2b
and phase 3 randomised trials. Lancet Healthy Longev. 2, e250–e262.
https://doi.org/10.1016/S2666-7568(21)00062-3.
187. Mannick, J.B., Morris, M., Hockey, H.-U.P., Roma, G., Beibel, M., Kulma-
tycki, K., Watkins, M., Shavlakadze, T., Zhou, W., Quinn, D., et al. (2018).
ll
ReviewTORC1 inhibition enhances immune function and reduces infections in
the elderly. Sci. Transl. Med. 10, eaaq1564. https://doi.org/10.1126/sci-
translmed.aaq1564.
188. Abdellatif, M., Trummer-Herbst, V., Martin Heberle, A., Humnig, A.,
Pendl, T., Durand, S., Cerrato, G., Hofer, S.J., Islam, M., Voglhuber, J.,
et al. (2022). Fine-tuning cardiac insulin/insulin-like growth factor 1 re-
ceptor signaling to promote health and longevity. Circulation 145,
1853–1866. https://doi.org/10.1161/CIRCULATIONAHA.122.059863.
189. Wu, Q., Tian, A.-L., Li, B., Leduc, M., Forveille, S., Hamley, P., Galloway,
W., Xie, W., Liu, P., Zhao, L., et al. (2021). IGF1 receptor inhibition am-
plifies the effects of cancer drugs by autophagy and immune-dependent
mechanisms. J. Immunother. Cancer 9, e002722. https://doi.org/10.
1136/jitc-2021-002722.
190. Zhang, W.B., Aleksic, S., Gao, T., Weiss, E.F., Demetriou, E., Verghese,
J., Holtzer, R., Barzilai, N., and Milman, S. (2020). Insulin-like growth
factor-1 and IGF binding proteins predict all-cause mortality and
morbidity in older adults. Cells 9, E1368. https://doi.org/10.3390/
cells9061368.
191. Zhang, W.B., Ye, K., Barzilai, N., and Milman, S. (2021). The antagonistic
pleiotropy of insulin-like growth factor 1. Aging Cell 20, e13443. https://
doi.org/10.1111/acel.13443.
192. Sebastiani, P., Federico, A., Morris, M., Gurinovich, A., Tanaka, T., Chan-
dler, K.B., Andersen, S.L., Denis, G., Costello, C.E., Ferrucci, L., et al.
(2021). Protein signatures of centenarians and their offspring suggest
centenarians age slower than other humans. Aging Cell 20, e13290.
https://doi.org/10.1111/acel.13290.
193. Mattison, J.A., Colman, R.J., Beasley, T.M., Allison, D.B., Kemnitz, J.W.,
Roth, G.S., Ingram, D.K., Weindruch, R., de Cabo, R., and Anderson,
R.M. (2017). Caloric restriction improves health and survival of rhesusmon-
keys. Nat. Commun. 8, 14063. https://doi.org/10.1038/ncomms14063.
194. Solon-Biet, S.M., McMahon, A.C., Ballard, J.W.O., Ruohonen, K., Wu,
L.E., Cogger, V.C., Warren, A., Huang, X., Pichaud, N., Melvin, R.G.,
et al. (2020). The ratio of macronutrients, not caloric intake, dictates car-
diometabolic health, aging, and longevity in ad libitum-fed mice. Cell
Metab. 31, 654. https://doi.org/10.1016/j.cmet.2020.01.010.
195. Pak, H.H., Haws, S.A., Green, C.L., Koller, M., Lavarias, M.T., Richard-
son, N.E., Yang, S.E., Dumas, S.N., Sonsalla, M., Bray, L., et al. (2021).
Fasting drives the metabolic, molecular and geroprotective effects of a
calorie-restricted diet in mice. Nat. Metab. 3, 1327–1341. https://doi.
org/10.1038/s42255-021-00466-9.
196. Mitchell, S.J., Bernier, M., Mattison, J.A., Aon, M.A., Kaiser, T.A., Anson,
R.M., Ikeno, Y., Anderson, R.M., Ingram, D.K., and de Cabo, R. (2019).
Daily fasting improves health and survival in male mice independent of
diet composition and calories. Cell Metab. 29, 221–228.e3. https://doi.
org/10.1016/j.cmet.2018.08.011.
197. Mitchell, S.J., Madrigal-Matute, J., Scheibye-Knudsen, M., Fang, E.,
Aon, M., Gonza´lez-Reyes, J.A., Cortassa, S., Kaushik, S., Gonzalez-
Freire, M., Patel, B., et al. (2016). Effects of sex, strain, and energy intake
on hallmarks of aging inmice. Cell Metab. 23, 1093–1112. https://doi.org/
10.1016/j.cmet.2016.05.027.
198. Mattson, M.P., Longo, V.D., and Harvie, M. (2017). Impact of intermittent
fasting on health and disease processes. Ageing Res. Rev. 39, 46–58.
https://doi.org/10.1016/j.arr.2016.10.005.
199. Stekovic, S., Hofer, S.J., Tripolt, N., Aon, M.A., Royer, P., Pein, L., Sta-
dler, J.T., Pendl, T., Prietl, B., Url, J., et al. (2019). Alternate day fasting
improves physiological and molecular markers of aging in healthy, non-
obese humans. Cell Metab. 30, 462–476.e6. https://doi.org/10.1016/j.
cmet.2019.07.016.
200. Ulgherait, M., Midoun, A.M., Park, S.J., Gatto, J.A., Tener, S.J., Siewert,
J., Klickstein, N., Canman, J.C., Ja, W.W., and Shirasu-Hiza, M. (2021).
Circadian autophagy drives iTRF-mediated longevity. Nature 598, 353–
358. https://doi.org/10.1038/s41586-021-03934-0.
201. Anisimov, V.N., Zabezhinski, M.A., Popovich, I.G., Piskunova, T.S.,
Semenchenko, A.V., Tyndyk, M.L., Yurova, M.N., Rosenfeld, S.V., andBlagosklonny, M.V. (2011). Rapamycin increases lifespan and inhibits
spontaneous tumorigenesis in inbred female mice. Cell Cycle 10,
4230–4236. https://doi.org/10.4161/cc.10.24.18486.
202. Salvadori, G., Zanardi, F., Iannelli, F., Lobefaro, R., Vernieri, C., and
Longo, V.D. (2021). Fasting-mimicking diet blocks triple-negative breast
cancer and cancer stem cell escape. Cell Metab. 33, 2247–2259.e6.
https://doi.org/10.1016/j.cmet.2021.10.008.
203. McCarthy, C.G., Chakraborty, S., Singh, G., Yeoh, B.S., Schrecken-
berger, Z.J., Singh, A., Mell, B., Bearss, N.R., Yang, T., Cheng, X.,
et al. (2021). Ketone body b-hydroxybutyrate is an autophagy-dependent
vasodilator. JCI Insight 6, e149037. https://doi.org/10.1172/jci.insight.
149037.
204. Youm, Y.-H., Nguyen, K.Y., Grant, R.W., Goldberg, E.L., Bodogai, M.,
Kim, D., D’Agostino, D., Planavsky, N., Lupfer, C., Kanneganti, T.D.,
et al. (2015). The ketone metabolite b-hydroxybutyrate blocks NLRP3 in-
flammasome-mediated inflammatory disease. Nat. Med. 21, 263–269.
https://doi.org/10.1038/nm.3804.
205. Rattray, N.J.W., Trivedi, D.K., Xu, Y., Chandola, T., Johnson, C.H.,
Marshall, A.D., Mekli, K., Rattray, Z., Tampubolon, G., Vanhoutte, B.,
et al. (2019). Metabolic dysregulation in vitamin E and carnitine shuttle en-
ergy mechanisms associate with human frailty. Nat. Commun. 10, 5027.
https://doi.org/10.1038/s41467-019-12716-2.
206. Lionaki, E., Gkikas, I., Daskalaki, I., Ioannidi, M.-K., Klapa, M.I., and Tav-
ernarakis, N. (2022). Mitochondrial protein import determines lifespan
through metabolic reprogramming and de novo serine biosynthesis.
Nat. Commun. 13, 651. https://doi.org/10.1038/s41467-022-28272-1.
207. Owusu-Ansah, E., Song, W., and Perrimon, N. (2013). Muscle mitohorm-
esis promotes longevity via systemic repression of insulin signaling. Cell
155, 699–712. https://doi.org/10.1016/j.cell.2013.09.021.
208. Pietrocola, F., Galluzzi, L., Bravo-San Pedro, J.M., Madeo, F., and
Kroemer, G. (2015). Acetyl coenzyme A: a central metabolite and second
messenger. Cell Metab. 21, 805–821. https://doi.org/10.1016/j.cmet.
2015.05.014.
209. Youle, R.J. (2019). Mitochondria-Striking a balance between host and
endosymbiont. Science 365, eaaw9855. https://doi.org/10.1126/sci-
ence.aaw9855.
210. Kulkarni, A.S., Gubbi, S., and Barzilai, N. (2020). Benefits of metformin in
attenuating the hallmarks of aging. Cell Metab. 32, 15–30. https://doi.org/
10.1016/j.cmet.2020.04.001.
211. Zhou, B., Kreuzer, J., Kumsta, C., Wu, L., Kamer, K.J., Cedillo, L., Zhang,
Y., Li, S., Kacergis, M.C., Webster, C.M., et al. (2019). Mitochondrial
permeability uncouples elevated autophagy and lifespan extension.
Cell 177, 299–314.e16. https://doi.org/10.1016/j.cell.2019.02.013.
212. Zinovkin, R.A., and Zamyatnin, A.A. (2019). Mitochondria-targeted drugs.
Curr. Mol. Pharmacol. 12, 202–214. https://doi.org/10.2174/1874467212
666181127151059.
213. Yen, K., Mehta, H.H., Kim, S.J., Lue, Y., Hoang, J., Guerrero, N., Port, J.,
Bi, Q., Navarrete, G., Brandhorst, S., et al. (2020). The mitochondrial
derived peptide humanin is a regulator of lifespan and healthspan. Aging
(Albany, NY) 12, 11185–11199. https://doi.org/10.18632/aging.103534.
214. Lee, C., Wan, J., Miyazaki, B., Fang, Y., Guevara-Aguirre, J., Yen, K.,
Longo, V., Bartke, A., and Cohen, P. (2014). IGF-I regulates the age-
dependent signaling peptide humanin. Aging Cell 13, 958–961. https://
doi.org/10.1111/acel.12243.
215. Reynolds, J.C., Lai, R.W., Woodhead, J.S.T., Joly, J.H., Mitchell, C.J.,
Cameron-Smith, D., Lu, R., Cohen, P., Graham, N.A., Benayoun, B.A.,
et al. (2021). MOTS-c is an exercise-induced mitochondrial-encoded
regulator of age-dependent physical decline and muscle homeostasis.
Nat. Commun. 12, 470. https://doi.org/10.1038/s41467-020-20790-0.
216. Gorgoulis, V., Adams, P.D., Alimonti, A., Bennett, D.C., Bischof, O.,
Bishop, C., Campisi, J., Collado, M., Evangelou, K., Ferbeyre, G., et al.
(2019). Cellular senescence: defining a path forward. Cell 179, 813–
827. https://doi.org/10.1016/j.cell.2019.10.005.Cell 186, January 19, 2023 275
ll
Review217. Tuttle, C.S.L., Waaijer, M.E.C., Slee-Valentijn, M.S., Stijnen, T., Westen-
dorp, R., and Maier, A.B. (2020). Cellular senescence and chronological
age in various human tissues: A systematic review and meta-analysis.
Aging Cell 19, e13083. https://doi.org/10.1111/acel.13083.
218. Xu, P., Wang, M., Song, W.-M., Wang, Q., Yuan, G.-C., Sudmant, P.H.,
Zare, H., Tu, Z., Orr, M.E., and Zhang, B. (2022). The landscape of human
tissue and cell type specific expression and co-regulation of senescence
genes. Mol. Neurodegener. 17, 5. https://doi.org/10.1186/s13024-021-
00507-7.
219. Mehdizadeh, M., Aguilar, M., Thorin, E., Ferbeyre, G., and Nattel, S.
(2022). The role of cellular senescence in cardiac disease: basic biology
and clinical relevance. Nat. Rev. Cardiol. 19, 250–264. https://doi.org/10.
1038/s41569-021-00624-2.
220. Serrano, M., and Mun˜oz-Espı´n, D. (2022). Cellular Senescence in Dis-
ease (Elsevier) https://doi.org/10.1016/C2019-0-04661-4.
221. Robbins, P.D., Jurk, D., Khosla, S., Kirkland, J.L., LeBrasseur, N.K.,
Miller, J.D., Passos, J.F., Pignolo, R.J., Tchkonia, T., and Niedernhofer,
L.J. (2021). Senolytic drugs: reducing senescent cell viability to extend
health span. Annu. Rev. Pharmacol. Toxicol. 61, 779–803. https://doi.
org/10.1146/annurev-pharmtox-050120-105018.
222. Wissler Gerdes, E.O., Misra, A., Netto, J.M.E., Tchkonia, T., and Kirkland,
J.L. (2021). Strategies for late phase preclinical and early clinical trials of
senolytics. Mech. Ageing Dev. 200, 111591. https://doi.org/10.1016/j.
mad.2021.111591.
223. Birch, J., and Gil, J. (2020). Senescence and the SASP: many therapeutic
avenues. Genes Dev. 34, 1565–1576. https://doi.org/10.1101/gad.
343129.120.
224. Sati, S., Bonev, B., Szabo, Q., Jost, D., Bensadoun, P., Serra, F., Lou-
biere, V., Papadopoulos, G.L., Rivera-Mulia, J.-C., Fritsch, L., et al.
(2020). 4D genome rewiring during oncogene-induced and replicative
senescence. Mol. Cell 78, 522–538.e9. https://doi.org/10.1016/j.mol-
cel.2020.03.007.
225. Chakradeo, S., Elmore, L.W., and Gewirtz, D.A. (2016). Is senescence
reversible? Curr. Drug Targets 17, 460–466. https://doi.org/10.2174/
1389450116666150825113500.
226. Rhinn, M., Ritschka, B., and Keyes, W.M. (2019). Cellular senescence in
development, regeneration and disease. Development 146, dev151837.
https://doi.org/10.1242/dev.151837.
227. Young, A.R.J., Cassidy, L.D., and Narita, M. (2021). Autophagy and
senescence, converging roles in pathophysiology as seen through
mouse models. Adv. Cancer Res. 150, 113–145. https://doi.org/10.
1016/bs.acr.2021.02.001.
228. Faget, D.V., Ren, Q., and Stewart, S.A. (2019). Unmasking senescence:
context-dependent effects of SASP in cancer. Nat. Rev. Cancer 19,
439–453. https://doi.org/10.1038/s41568-019-0156-2.
229. Meyer, K., Lo´pez-Domı´nguez, J.A., Maus, M., Kovatcheva, M., and
Serrano, M. (2022). Senescence as a therapeutic target: current state
and future challenges. In Cellular Senescence in Disease, M. Serrano
and D. Mun˜oz-Espı´n, eds. (Academic Press), pp. 425–442. Chapter 16.
https://doi.org/10.1016/B978-0-12-822514-1.00014-6.
230. Zhu, Y., Tchkonia, T., Pirtskhalava, T., Gower, A.C., Ding, H., Giorgadze,
N., Palmer, A.K., Ikeno, Y., Hubbard, G.B., Lenburg, M., et al. (2015). The
Achilles’ heel of senescent cells: from transcriptome to senolytic drugs.
Aging Cell 14, 644–658. https://doi.org/10.1111/acel.12344.
231. Zhu, Y., Tchkonia, T., Fuhrmann-Stroissnigg, H., Dai, H.M., Ling, Y.Y.,
Stout, M.B., Pirtskhalava, T., Giorgadze, N., Johnson, K.O., Giles, C.B.,
et al. (2016). Identification of a novel senolytic agent, navitoclax, targeting
the Bcl-2 family of anti-apoptotic factors. Aging Cell 15, 428–435. https://
doi.org/10.1111/acel.12445.
232. He, Y., Zhang, X., Chang, J., Kim, H.-N., Zhang, P., Wang, Y., Khan, S.,
Liu, X., Zhang, X., Lv, D., et al. (2020). Using proteolysis-targeting
chimera technology to reduce navitoclax platelet toxicity and improve276 Cell 186, January 19, 2023its senolytic activity. Nat. Commun. 11, 1996. https://doi.org/10.1038/
s41467-020-15838-0.
233. Triana-Martı´nez, F., Picallos-Rabina, P., Da Silva-A´lvarez, S., Pietrocola,
F., Llanos, S., Rodilla, V., Soprano, E., Pedrosa, P., Ferreiro´s, A., Barra-
das, M., et al. (2019). Identification and characterization of cardiac glyco-
sides as senolytic compounds. Nat. Commun. 10, 4731. https://doi.org/
10.1038/s41467-019-12888-x.
234. Johmura, Y., Yamanaka, T., Omori, S., Wang, T.W., Sugiura, Y., Matsu-
moto, M., Suzuki, N., Kumamoto, S., Yamaguchi, K., Hatakeyama, S.,
et al. (2021). Senolysis by glutaminolysis inhibition ameliorates various
age-associated disorders. Science 371, 265–270. https://doi.org/10.
1126/science.abb5916.
235. Suda, M., Shimizu, I., Katsuumi, G., Yoshida, Y., Hayashi, Y., Ikegami, R.,
Matsumoto, N., Yoshida, Y., Mikawa, R., Katayama, A., et al. (2021).
Senolytic vaccination improves normal and pathological age-related
phenotypes and increases lifespan in progeroid mice. Nat Aging 1,
1117–1126. https://doi.org/10.1038/s43587-021-00151-2.
236. Amor, C., Feucht, J., Leibold, J., Ho, Y.-J., Zhu, C., Alonso-Curbelo, D.,
Mansilla-Soto, J., Boyer, J.A., Li, X., Giavridis, T., et al. (2020). Senolytic
CAR T cells reverse senescence-associated pathologies. Nature 583,
127–132. https://doi.org/10.1038/s41586-020-2403-9.
237. Clevers, H., and Watt, F.M. (2018). Defining adult stem cells by function,
not by phenotype. Annu. Rev. Biochem. 87, 1015–1027. https://doi.org/
10.1146/annurev-biochem-062917-012341.
238. Tata, P.R., and Rajagopal, J. (2017). Plasticity in the lung: making and
breaking cell identity. Development 144, 755–766. https://doi.org/10.
1242/dev.143784.
239. Lin, B., Coleman, J.H., Peterson, J.N., Zunitch, M.J., Jang, W., Herrick,
D.B., and Schwob, J.E. (2017). Injury induces endogenous reprogram-
ming and dedifferentiation of neuronal progenitors to multipotency. Cell
Stem Cell 21, 761–774.e5. https://doi.org/10.1016/j.stem.2017.09.008.
240. Murata, K., Jadhav, U., Madha, S., van Es, J., Dean, J., Cavazza, A., Wu-
cherpfennig, K., Michor, F., Clevers, H., and Shivdasani, R.A. (2020).
Ascl2-dependent cell dedifferentiation drives regeneration of ablated in-
testinal stem cells. Cell Stem Cell 26, 377–390.e6. https://doi.org/10.
1016/j.stem.2019.12.011.
241. Takahashi, K., and Yamanaka, S. (2006). Induction of pluripotent stem
cells frommouse embryonic and adult fibroblast cultures by defined fac-
tors. Cell 126, 663–676. https://doi.org/10.1016/j.cell.2006.07.024.
242. Deng, W., Jacobson, E.C., Collier, A.J., and Plath, K. (2021). The tran-
scription factor code in iPSC reprogramming. Curr. Opin. Genet. Dev.
70, 89–96. https://doi.org/10.1016/j.gde.2021.06.003.
243. Li, H., Collado, M., Villasante, A., Strati, K., Ortega, S., Can˜amero, M.,
Blasco, M.A., and Serrano, M. (2009). The Ink4/Arf locus is a barrier for
iPS cell reprogramming. Nature 460, 1136–1139. https://doi.org/10.
1038/nature08290.
244. Marion, R.M., Strati, K., Li, H., Tejera, A., Schoeftner, S., Ortega, S.,
Serrano, M., and Blasco, M.A. (2009). Telomeres acquire embryonic
stem cell characteristics in induced pluripotent stem cells. Cell Stem
Cell 4, 141–154. https://doi.org/10.1016/j.stem.2008.12.010.
245. Horvath, S. (2013). DNAmethylation age of human tissues and cell types.
Genome Biol. 14, R115. https://doi.org/10.1186/gb-2013-14-10-r115.
246. Gill, D., Parry, A., Santos, F., Okkenhaug, H., Todd, C.D., Hernando-Her-
raez, I., Stubbs, T.M., Milagre, I., and Reik, W. (2022). Multi-omic
rejuvenation of human cells by maturation phase transient reprogram-
ming. eLife 11, e71624. https://doi.org/10.7554/eLife.71624.
247. Chondronasiou, D., Gill, D., Mosteiro, L., Urdinguio, R.G., Berenguer-
Llergo, A., Aguilera, M., Durand, S., Aprahamian, F., Nirmalathasan, N.,
Abad, M., et al. (2022). Multi-omic rejuvenation of naturally aged tissues
by a single cycle of transient reprogramming. Aging Cell 21, e13578.
https://doi.org/10.1111/acel.13578.
248. Roux, A.E., Zhang, C., Paw, J., Zavala-Solorio, J., Malahias, E., Vijay, T.,
Kolumam, G., Kenyon, C., and Kimmel, J.C. (2022). Diverse partial
ll
Reviewreprogramming strategies restore youthful gene expression and tran-
siently suppress cell identity. Cell Syst. 13, 574–587.e11. S2405-
4712(22)00223-X. https://doi.org/10.1016/j.cels.2022.05.002.
249. Poganik, J.R., Zhang, B., Baht, G.S., Kerepesi, C., Yim, S.H., Lu, A.T.,
Haghani, A., Gong, T., Hedman, A.M., Andolf, E., et al. (2022). Biological
age is increased by stress and restored upon recovery https://doi.org/10.
1101/2022.05.04.490686.
250. Mosteiro, L., Pantoja, C., Alcazar, N., Mario´n, R.M., Chondronasiou, D.,
Rovira, M., Fernandez-Marcos, P.J., Mun˜oz-Martin, M., Blanco-Aparicio,
C., Pastor, J., et al. (2016). Tissue damage and senescence provide crit-
ical signals for cellular reprogramming in vivo. Science 354, aaf4445.
https://doi.org/10.1126/science.aaf4445.
251. Ribeiro, R., Macedo, J.C., Costa, M., Ustiyan, V., Shindyapina, A.V.,
Tyshkovskiy, A., Gomes, R.N., Castro, J.P., Kalin, T.V., Vasques-No´voa,
F., et al. (2022). In vivo cyclic induction of the FOXM1 transcription factor
delays natural and progeroid aging phenotypes and extends healthspan.
Nat Aging 2, 397–411. https://doi.org/10.1038/s43587-022-00209-9.
252. Chang-Panesso, M., Kadyrov, F.F., Lalli, M., Wu, H., Ikeda, S., Kefa-
loyianni, E., Abdelmageed, M.M., Herrlich, A., Kobayashi, A., and Hum-
phreys, B.D. (2019). FOXM1 drives proximal tubule proliferation during
repair from acute ischemic kidney injury. J. Clin. Invest. 129, 5501–
5517. https://doi.org/10.1172/JCI125519.
253. Miller, H.A., Dean, E.S., Pletcher, S.D., and Leiser, S.F. (2020). Cell non-
autonomous regulation of health and longevity. eLife 9, e62659. https://
doi.org/10.7554/eLife.62659.
254. Villeda, S.A., Luo, J., Mosher, K.I., Zou, B., Britschgi, M., Bieri, G., Stan,
T.M., Fainberg, N., Ding, Z., Eggel, A., et al. (2011). The ageing systemic
milieu negatively regulates neurogenesis and cognitive function. Nature
477, 90–94. https://doi.org/10.1038/nature10357.
255. Smith, L.K., He, Y., Park, J.-S., Bieri, G., Snethlage, C.E., Lin, K., Gontier,
G., Wabl, R., Plambeck, K.E., Udeochu, J., et al. (2015). b2-microglobulin
is a systemic pro-aging factor that impairs cognitive function and neuro-
genesis. Nat. Med. 21, 932–937. https://doi.org/10.1038/nm.3898.
256. Valletta, S., Thomas, A., Meng, Y., Ren, X., Drissen, R., Sengu¨l, H., Di
Genua, C., and Nerlov, C. (2020). Micro-environmental sensing by
bone marrow stroma identifies IL-6 and TGFb1 as regulators of hemato-
poietic ageing. Nat. Commun. 11, 4075. https://doi.org/10.1038/s41467-
020-17942-7.
257. Naito, A.T., Sumida, T., Nomura, S., Liu, M.-L., Higo, T., Nakagawa, A.,
Okada, K., Sakai, T., Hashimoto, A., Hara, Y., et al. (2012). Complement
C1q activates canonical Wnt signaling and promotes aging-related
phenotypes. Cell 149, 1298–1313. https://doi.org/10.1016/j.cell.2012.
03.047.
258. Fafia´n-Labora, J.A., and O’Loghlen, A. (2020). Classical and nonclassical
intercellular communication in senescence and ageing. Trends Cell Biol.
30, 628–639. https://doi.org/10.1016/j.tcb.2020.05.003.
259. Rando, T.A., and Jones, D.L. (2021). Regeneration, rejuvenation, and
replacement: turning back the clock on tissue aging. Cold Spring
Harb. Perspect. Biol. 13, a040907. https://doi.org/10.1101/cshper-
spect.a040907.
260. Folgueras, A.R., Freitas-Rodrı´guez, S., Velasco, G., and Lo´pez-Otı´n, C.
(2018). Mouse models to disentangle the hallmarks of human aging.
Circ. Res. 123, 905–924. https://doi.org/10.1161/CIRCRESAHA.118.
312204.
261. Fedintsev, A., and Moskalev, A. (2020). Stochastic non-enzymatic modi-
fication of long-lived macromolecules - A missing hallmark of aging.
Ageing Res. Rev. 62, 101097. https://doi.org/10.1016/j.arr.2020.101097.
262. Selman, M., and Pardo, A. (2021). Fibroageing: an ageing pathological
feature driven by dysregulated extracellular matrix-cell mechanobiology.
Ageing Res. Rev. 70, 101393. https://doi.org/10.1016/j.arr.2021.101393.
263. Levi, N., Papismadov, N., Solomonov, I., Sagi, I., and Krizhanovsky, V.
(2020). The ECM path of senescence in aging: components and modi-
fiers. FEBS J. 287, 2636–2646. https://doi.org/10.1111/febs.15282.264. Hu, H.-H., Cao, G., Wu, X.-Q., Vaziri, N.D., and Zhao, Y.-Y. (2020). Wnt
signaling pathway in aging-related tissue fibrosis and therapies. Ageing
Res. Rev. 60, 101063. https://doi.org/10.1016/j.arr.2020.101063.
265. Segel, M., Neumann, B., Hill, M.F.E., Weber, I.P., Viscomi, C., Zhao, C.,
Young, A., Agley, C.C., Thompson, A.J., Gonzalez, G.A., et al. (2019).
Niche stiffness underlies the ageing of central nervous system progenitor
cells. Nature 573, 130–134. https://doi.org/10.1038/s41586-019-1484-9.
266. Vafaie, F., Yin, H., O’Neil, C., Nong, Z., Watson, A., Arpino, J.-M., Chu,
M.W.A., Wayne Holdsworth, D., Gros, R., and Pickering, J.G. (2014).
Collagenase-resistant collagen promotes mouse aging and vascular
cell senescence. Aging Cell 13, 121–130. https://doi.org/10.1111/
acel.12155.
267. Erikson, G.A., Bodian, D.L., Rueda, M., Molparia, B., Scott, E.R., Scott-
Van Zeeland, A.A., Topol, S.E., Wineinger, N.E., Niederhuber, J.E., Topol,
E.J., et al. (2016). Whole-genome sequencing of a healthy aging cohort.
Cell 165, 1002–1011. https://doi.org/10.1016/j.cell.2016.03.022.
268. Statzer, C., Jongsma, E., Liu, S.X., Dakhovnik, A., Wandrey, F., Mozhar-
ovskyi, P., Zu¨lli, F., and Ewald, C.Y. (2021). Youthful and age-related ma-
treotypes predict drugs promoting longevity. Aging Cell 20, e13441.
https://doi.org/10.1111/acel.13441.
269. Schinzel, R.T., Higuchi-Sanabria, R., Shalem, O., Moehle, E.A., Webster,
B.M., Joe, L., Bar-Ziv, R., Frankino, P.A., Durieux, J., Pender, C., et al.
(2019). The hyaluronidase, TMEM2, promotes ER homeostasis and
longevity independent of the UPRER. Cell 179, 1306–1318.e18. https://
doi.org/10.1016/j.cell.2019.10.018.
270. King, D.E., and Xiang, J. (2020). Glucosamine/chondroitin and mortality
in a US NHANES cohort. J. Am. Board Fam. Med. 33, 842–847. https://
doi.org/10.3122/jabfm.2020.06.200110.
271. Hirata, T., Arai, Y., Yuasa, S., Abe, Y., Takayama, M., Sasaki, T., Kuni-
tomi, A., Inagaki, H., Endo, M., Morinaga, J., et al. (2020). Associations
of cardiovascular biomarkers and plasma albumin with exceptional sur-
vival to the highest ages. Nat. Commun. 11, 3820. https://doi.org/10.
1038/s41467-020-17636-0.
272. Mogilenko, D.A., Shpynov, O., Andhey, P.S., Arthur, L., Swain, A., Esau-
lova, E., Brioschi, S., Shchukina, I., Kerndl, M., Bambouskova, M., et al.
(2021). Comprehensive profiling of an aging immune system reveals
clonal GZMK+ CD8+ T cells as conserved hallmark of inflammaging. Im-
munity 54, 99–115.e12. https://doi.org/10.1016/j.immuni.2020.11.005.
273. Carrasco, E., Go´mez de Las Heras, M.M., Gabande´-Rodrı´guez, E., Des-
dı´n-Mico´, G., Aranda, J.F., and Mittelbrunn, M. (2022). The role of T cells
in age-related diseases. Nat. Rev. Immunol. 22, 97–111. https://doi.org/
10.1038/s41577-021-00557-4.
274. Jaiswal, S., Natarajan, P., Silver, A.J., Gibson, C.J., Bick, A.G., Shvartz, E.,
McConkey,M., Gupta,N., Gabriel, S., Ardissino, D., et al. (2017). Clonal he-
matopoiesis and risk of atherosclerotic cardiovascular disease. N. Engl. J.
Med. 377, 111–121. https://doi.org/10.1056/NEJMoa1701719.
275. Svensson, E.C., Madar, A., Campbell, C.D., He, Y., Sultan, M., Healey,
M.L., Xu, H., D’Aco, K., Fernandez, A., Wache-Mainier, C., et al. (2022).
TET2-driven clonal hematopoiesis and response to canakinumab: an
exploratory analysis of the CANTOS randomized clinical trial. JAMA Car-
diol. 7, 521–528. https://doi.org/10.1001/jamacardio.2022.0386.
276. Mittelbrunn, M., and Kroemer, G. (2021). Hallmarks of T cell aging. Nat.
Immunol. 22, 687–698. https://doi.org/10.1038/s41590-021-00927-z.
277. Routy, B., Gopalakrishnan, V., Daille`re, R., Zitvogel, L., Wargo, J.A., and
Kroemer, G. (2018). The gut microbiota influences anticancer immuno-
surveillance and general health. Nat. Rev. Clin. Oncol. 15, 382–396.
https://doi.org/10.1038/s41571-018-0006-2.
278. Yousefzadeh, M.J., Flores, R.R., Zhu, Y., Schmiechen, Z.C., Brooks,
R.W., Trussoni, C.E., Cui, Y., Angelini, L., Lee, K.-A., McGowan, S.J.,
et al. (2021). An aged immune system drives senescence and ageing of
solid organs. Nature 594, 100–105. https://doi.org/10.1038/s41586-
021-03547-7.Cell 186, January 19, 2023 277
ll
Review279. D’Souza, S.S., Zhang, Y., Bailey, J.T., Fung, I.T.H., Kuentzel, M.L., Chit-
tur, S.V., and Yang, Q. (2021). Type I interferon signaling controls the
accumulation and transcriptomes of monocytes in the aged lung. Aging
Cell 20, e13470. https://doi.org/10.1111/acel.13470.
280. Gonza´lez-Dominguez, A., Montan˜ez, R., Castejo´n-Vega, B., Nun˜ez-
Vasco, J., Lendines-Cordero, D., Wang, C., Mbalaviele, G., Navarro-
Pando, J.M., Alcocer-Go´mez, E., and Cordero, M.D. (2021). Inhibition
of the NLRP3 inflammasome improves lifespan in animal murine model
of Hutchinson-Gilford progeria. EMBO Mol. Med. 13, e14012. https://
doi.org/10.15252/emmm.202114012.
281. McNeil, J.J., Wolfe, R., Woods, R.L., Tonkin, A.M., Donnan, G.A., Nelson,
M.R., Reid, C.M., Lockery, J.E., Kirpach, B., Storey, E., et al. (2018). Ef-
fect of aspirin on cardiovascular events and bleeding in the healthy
elderly. N. Engl. J. Med. 379, 1509–1518. https://doi.org/10.1056/
NEJMoa1805819.
282. Zmora, N., Soffer, E., and Elinav, E. (2019). Transforming medicine with
the microbiome. Sci. Transl. Med. 11, eaaw1815. https://doi.org/10.
1126/scitranslmed.aaw1815.
283. Gacesa, R., Kurilshikov, A., Vich Vila, A., Sinha, T., Klaassen, M.A.Y.,
Bolte, L.A., Andreu-Sa´nchez, S., Chen, L., Collij, V., Hu, S., et al.
(2022). Environmental factors shaping the gut microbiome in a Dutch
population. Nature 604, 732–739. https://doi.org/10.1038/s41586-022-
04567-7.
284. Lee, K.A., Thomas, A.M., Bolte, L.A., Bjo¨rk, J.R., de Ruijter, L.K., Arma-
nini, F., Asnicar, F., Blanco-Miguez, A., Board, R., Calbet-Llopart, N.,
et al. (2022). Cross-cohort gut microbiome associations with immune
checkpoint inhibitor response in advanced melanoma. Nat. Med. 28,
535–544. https://doi.org/10.1038/s41591-022-01695-5.
285. McCulloch, J.A., Davar, D., Rodrigues, R.R., Badger, J.H., Fang, J.R.,
Cole, A.M., Balaji, A.K., Vetizou, M., Prescott, S.M., Fernandes, M.R.,
et al. (2022). Intestinal microbiota signatures of clinical response and im-
mune-related adverse events in melanoma patients treated with anti-PD-
1. Nat. Med. 28, 545–556. https://doi.org/10.1038/s41591-022-01698-2.
286. Ghosh, T.S., Shanahan, F., and O’Toole, P.W. (2022). The gut micro-
biome as a modulator of healthy ageing. Nat. Rev. Gastroenterol. Hepa-
tol. 19, 565–584. https://doi.org/10.1038/s41575-022-00605-x.
287. Biagi, E., Franceschi, C., Rampelli, S., Severgnini, M., Ostan, R., Turroni,
S., Consolandi, C., Quercia, S., Scurti, M., Monti, D., et al. (2016). Gut mi-
crobiota and extreme longevity. Curr. Biol. 26, 1480–1485. https://doi.
org/10.1016/j.cub.2016.04.016.
288. Wilmanski, T., Diener, C., Rappaport, N., Patwardhan, S., Wiedrick, J.,
Lapidus, J., Earls, J.C., Zimmer, A., Glusman, G., Robinson, M., et al.
(2021). Gut microbiome pattern reflects healthy ageing and predicts sur-
vival in humans. Nat. Metab. 3, 274–286. https://doi.org/10.1038/
s42255-021-00348-0.278 Cell 186, January 19, 2023289. Ghosh, T.S., Das,M., Jeffery, I.B., andO’Toole, P.W. (2020). Adjusting for
age improves identification of gut microbiome alterations in multiple dis-
eases. eLife 9, e50240. https://doi.org/10.7554/eLife.50240.
290. Zhang, X., Zhong, H., Li, Y., Shi, Z., Ren, H., Zhang, Z., Zhou, X., Tang, S.,
Han, X., Lin, Y., et al. (2021). Sex- and age-related trajectories of the adult
human gut microbiota shared across populations of different ethnicities.
Nat Aging 1, 87–100. https://doi.org/10.1038/s43587-020-00014-2.
291. Sato, Y., Atarashi, K., Plichta, D.R., Arai, Y., Sasajima, S., Kearney, S.M.,
Suda, W., Takeshita, K., Sasaki, T., Okamoto, S., et al. (2021). Novel
bile acid biosynthetic pathways are enriched in the microbiome of
centenarians. Nature 599, 458–464. https://doi.org/10.1038/s41586-
021-03832-5.
292. Fransen, F., van Beek, A.A., Borghuis, T., Aidy, S.E., Hugenholtz, F., van
der Gaast-de Jongh, C., Savelkoul, H.F.J., De Jonge, M.I., Boekschoten,
M.V., Smidt, H., et al. (2017). Aged gut microbiota contributes to system-
ical inflammaging after transfer to Germ-free mice. Front. Immunol. 8,
1385. https://doi.org/10.3389/fimmu.2017.01385.
293. Ragonnaud, E., and Biragyn, A. (2021). Gutmicrobiota as the key control-
lers of ‘‘healthy’’ aging of elderly people. Immun. Ageing 18, 2. https://doi.
org/10.1186/s12979-020-00213-w.
294. Madeo, F., Eisenberg, T., Pietrocola, F., and Kroemer, G. (2018). Spermi-
dine in health and disease. Science 359, eaan2788. https://doi.org/10.
1126/science.aan2788.
295. Cani, P.D., Depommier, C., Derrien, M., Everard, A., and de Vos, W.M.
(2022). Akkermansia muciniphila: paradigm for next-generation benefi-
cial microorganisms. Nat. Rev. Gastroenterol. Hepatol. 19, 625–637.
https://doi.org/10.1038/s41575-022-00631-9.
296. Liu, G.Y., and Sabatini, D.M. (2020). mTOR at the nexus of nutrition,
growth, ageing and disease. Nat. Rev. Mol. Cell Biol. 21, 183–203.
https://doi.org/10.1038/s41580-019-0199-y.
297. Selvarani, R., Mohammed, S., and Richardson, A. (2021). Effect of rapa-
mycin on aging and age-related diseases-past and future. GeroScience
43, 1135–1158. https://doi.org/10.1007/s11357-020-00274-1.
298. Gladyshev, V.N., Kritchevsky, S.B., Clarke, S.G., Cuervo, A.M., Fiehn, O.,
de Magalha˜es, J.P., Mau, T., Maes, M., Moritz, R.L., Niedernhofer, L.J.,
et al. (2021). Molecular damage in aging. Nat Aging 1, 1096–1106.
https://doi.org/10.1038/s43587-021-00150-3.
299. Xu, M., Bradley, E.W., Weivoda, M.M., Hwang, S.M., Pirtskhalava, T.,
Decklever, T., Curran, G.L., Ogrodnik, M., Jurk, D., Johnson, K.O.,
et al. (2017). Transplanted senescent cells induce an osteoarthritis-like
condition in mice. J. Gerontol. A Biol. Sci. Med. Sci. 72, 780–785.
https://doi.org/10.1093/gerona/glw154.
300. Gladyshev, T.V., and Gladyshev, V.N. (2016). A disease or not a disease?
Aging as a pathology. Trends Mol. Med. 22, 995–996. https://doi.org/10.
1016/j.molmed.2016.09.009.