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dc.contributor.advisorde la Pompa, Jose Luis 
dc.contributor.authorGrivas, Dimitrios 
dc.date.accessioned2020-02-05T08:03:54Z
dc.date.available2020-02-05T08:03:54Z
dc.date.issued2019-12-19
dc.identifier.urihttp://hdl.handle.net/20.500.12105/9029
dc.description.abstractZebrafish have the ability to regenerate cardiac tissue after injury in contrast to mammals. Cryoinjury of the heart triggers a regenerative program that includes inflammatory response, cell proliferation and formation of a transient scar that eventually lead to complete tissue regeneration. This thesis focused on understanding the molecular and mechanical properties governing zebrafish heart regeneration. In particular, we studied the role of midkine-a (mdka) and caveolin-1 (cav1), two genes that were upregulated in a microarray analysis of regenerating hearts performed in the laboratory. Additionally, we studied the epigenetic regulation of mdka and tested in vivo different mdka cis-regulatory elements with the goal to identify the minimum regulatory regions that drive mdka expression in development and after injury. Midkine-a (Mdka) is a neurite-growth factor that is involved in the formation of the media floor plate in zebrafish. Here, we show that mdka expression was strongly induced after heart injury, although it was not expressed in intact hearts. In injured hearts, the onset of mdka expression was one-day post cryoinjury (dpci) in all the epicardial layer, whereas by 7dpci the expression became restricted to the epicardial cells covering the injured area. To study the role of Mdka in heart regeneration, we generated mdka-knock out (KO) zebrafish strains. In injured heart, mdka depletion did not trigger upregulation of mdkb and ptn, the two other members of the midkine gene family, which might compensate for the loss of Mdka. Analysis of 90dpci hearts showed that mdka deletion resulted in impaired heart regeneration, with the injured area enriched in collagen deposition. However, the proliferation ratio of cardiomyocytes (CM) and epicardial cells was not affected. Analysis of extracellular matrix turnover, immune cells infiltration in the injured area and revascularization of the regenerating tissue will clarify the reasons for the impaired cardiac regeneration in mdka-KO hearts. During development, mdka is expressed in neural tissues such us neural tube and brain, and is not detected neither in developing hearts nor in adult intact hearts. However, mdka expression was activated upon heart injury, having dynamic expression pattern. To investigate such a change in the expression pattern, we used available ChIP-seq and ATAC-seq data to analyse the epigenetic landscape of mdka. In mdka locus, there are three intronic enhancer sequences and two additional enhancers located upstream of mdka. We tested these enhancers and the mdka promoter linked to GFP reporter and studied their expression patterns in our transgenic lines. We found that the promoter of mdka and the intronic enhancers were responsible for the spatio-temporal expression pattern of mdka during development, in adult tissues and after injury of the heart and the fin. Caveolin-1 (Cav1) is the structural protein of caveolae, small membrane invaginations that are involved in signal transduction and mechanoprotection. Our expression analysis showed that Cav1 was expressed in the endocardium, endothelium and epicardium of intact adult heart. After heart cryoinjury, Cav1 was strongly expressed in epicardial cells covering the injury, and in the endocardium invading the damaged tissue. We introduced mutations in cav1 sequence that resulted in loss of Cav1 and caveolae. Analysis of the regeneration process revealed that the caveolae-depleted hearts regenerated normally, although cav1-KO hearts showed a transient defect on CM proliferation. However, loss of Cav1 and caveolae affected heart function, since the ejection fraction and the heart rate were significantly decreased in intact hearts. Using atomic force microscopy, we found that epicardial cells and cortical cardiomyocytes lacking caveolae were stiffer than the control hearts explaining the decline of cardiac function.es_ES
dc.description.sponsorshipThis doctoral thesis was performed in the Intercellular Signaling in Cardiovascular Development and Disease laboratory directed by Dr. José Luis de la Pompa at Centro Nacional de Investigaciones Cardiovasculares (CNIC) in Madrid. This study was financed by different grants of Ministerio de Ciencia, Innovación y Universidades (SAF2013-45543-R, SAF2015-71863-REDT y SAF2016-78370-R), CIBER Cardiovascular. Instituto de Salud Carlos III. Ref.: CB16/11/00399, Red de Terapia Celular. Instituto de Salud Carlos III. Ref.: RD16/0011/0021, CardioNeT- International Training Network on the Cellular and Molecular Bases of Heart Homeostasis and Repair. Agency: European Union, ref. 28600, 2012-2015. Dimitrios Grivas holds a PhD fellowship linked to the grant [PITN-GA-2011-289600] (CardioNeT)es_ES
dc.language.isoenges_ES
dc.rights.urihttp://creativecommons.org/licenses/by-nc-sa/4.0/*
dc.titleStudy of Midkine-a and Caveolin-1 in zebrafish heart regenerationes_ES
dc.typeTesis doctorales_ES
dc.rights.licenseAtribución-NoComercial-CompartirIgual 4.0 Internacional*
dc.identifier.doi10.4321/repisalud.9029
dc.contributor.funderMinisterio de Ciencia, Innovación y Universidades (España)es_ES
dc.contributor.funderCentro de Investigación Biomedica en Red - CIBERes_ES
dc.contributor.funderInstituto de Salud Carlos III - ISCIIIes_ES
dc.contributor.funderEuropean Commissiones_ES
dc.embargo.terms2020-12-19es_ES
dc.repisalud.orgCNICCNIC::Grupos de investigación::Señalización Intercelular durante el Desarrollo y la Enfermedad Cardiovasculares_ES
dc.repisalud.institucionCNICes_ES
dc.relation.projectIDinfo:eu-repo/grantAgreement/EC/FP7/289600es_ES
dc.rights.accessRightsinfo:eu-repo/semantics/embargoedAccesses_ES


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