Analysis of AAV9 biodistribution, transduction efficiency and AAV-miR-935 cardio-specific overexpression in a murine model of myocardial infarction

Myocardial infarction (MI) leads to an irreversible loss of cardiac myocytes, which compromises cardiac function. The cellular and molecular mechanisms involved in the ischemic event remain under study, being the cardiac exosomes transfer of non-coding RNAs a key process in this pathology. In this c...

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Autor: García-Olloqui, P. (Paula)|||/items/33aebcca-bf96-4a81-a74d-9fcd60b275cc
Tipo de recurso: tesis doctoral
Fecha de publicación:2019
País:España
Institución:Universidad de Navarra
Repositorio:Dadun. Depósito Académico Digital de la Universidad de Navarra
Idioma:español
OAI Identifier:oai:dadun.unav.edu:10171/58884
Acceso en línea:https://hdl.handle.net/10171/58884
Access Level:acceso abierto
Palabra clave:Materias Investigacion::Ciencias de la Salud
Materias Investigacion::Ciencias de la Salud::Microbiología y biología molecular
Materias Investigacion::Ciencias de la vida::Citología, biología celular
Cultivo celular
Patología cardiovascular
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spelling Analysis of AAV9 biodistribution, transduction efficiency and AAV-miR-935 cardio-specific overexpression in a murine model of myocardial infarctionGarcía-Olloqui, P. (Paula)|||/items/33aebcca-bf96-4a81-a74d-9fcd60b275ccMaterias Investigacion::Ciencias de la SaludMaterias Investigacion::Ciencias de la Salud::Microbiología y biología molecularMaterias Investigacion::Ciencias de la vida::Citología, biología celularCultivo celularPatología cardiovascularMyocardial infarction (MI) leads to an irreversible loss of cardiac myocytes, which compromises cardiac function. The cellular and molecular mechanisms involved in the ischemic event remain under study, being the cardiac exosomes transfer of non-coding RNAs a key process in this pathology. In this context, a comparative analysis of the exosomal compartment of human cardiac progenitor cells (CPC) was performed in comparison with human bone marrow-mesenchymal stem cells (MSC) and human dermal fibroblasts (HDF). A total of 481 differentially expressed microRNAs (miR) were found, being miR-935 the most differentially expressed in CPC. Analysis of miR-935 in vitro overexpression promoted a positive trend to increment cardiomyocyte survival, in response to oxidative stress. Furthermore, when miR-935 regulation was analyzed in a mouse model of MI disease, miR downregulation was shown five days post-infarct in different cardiac subpopulations. In order to study the role of miR-935 in MI and to achieve an optimal miR-935 overexpression in our mouse model, we developed a robust viral-based method for cardiac-specific miR overexpression. Cardiotropic Serotype 9-Adeno-Associated Virus (AAV9) were used for this purpose and their biodistribution was first determined in mice. Ubiquitous EF1α or the cardiac-specific TnT promoters were combined with Luciferase (Luc) or GFP reporters and administered by intramyocardial or intravenous injection, either in healthy or myocardial-infarcted mice. High transgene expression levels were found in the heart, but not in the liver, of mice receiving AAV-TnT, which was significantly higher after intramyocardial injection regardless of ischemia-induction. On the contrary, high hepatic transgene expression levels were detected with the EF1α-promoter, independently of the administration route and heart damage. Luc expression increased with both promoters in a time-dependent manner, reaching a peak by day 3-7 that was stable for at least 60 days. Moreover, tissue-specific GFP expression was found in cardiomyocytes with the TnT-vector, while minimal cardiac expression was detected with the ubiquitous one. Interestingly, we found that MI greatly increased the transcriptional activity of AAV genomes. Thus, we found AAV9-TnT as a robust and stable vector for cardiac-specific delivery after intramyocardial injection. Finally, the therapeutic potential of this vector in combination with miR-935 was evaluated in a mouse model of myocardial ischemia. Intramyocardial injection of AAV9-TnT-miR935 vector did not significantly improve heart function 60 days post-infarct. However, a slight positive trend in the ejection fraction and a decreased adverse cardiac remodeling are observed, suggesting miR-935 putative cardioprotective role.Pelacho-Samper, B. (Beatriz)Dadun. Depósito Académico Digital Universidad de Navarra20202020-03-1620202020-03-1620202020-03-1620192019-06-27doctoral thesishttp://purl.org/coar/resource_type/c_db06info:eu-repo/semantics/doctoralThesisapplication/pdfhttps://hdl.handle.net/10171/58884reponame:Dadun. Depósito Académico Digital de la Universidad de Navarrainstname:Universidad de NavarraEspañolspaopen accesshttp://purl.org/coar/access_right/c_abf2info:eu-repo/semantics/openAccessoai:dadun.unav.edu:10171/588842026-06-21T12:47:57Z
dc.title.none.fl_str_mv Analysis of AAV9 biodistribution, transduction efficiency and AAV-miR-935 cardio-specific overexpression in a murine model of myocardial infarction
title Analysis of AAV9 biodistribution, transduction efficiency and AAV-miR-935 cardio-specific overexpression in a murine model of myocardial infarction
spellingShingle Analysis of AAV9 biodistribution, transduction efficiency and AAV-miR-935 cardio-specific overexpression in a murine model of myocardial infarction
García-Olloqui, P. (Paula)|||/items/33aebcca-bf96-4a81-a74d-9fcd60b275cc
Materias Investigacion::Ciencias de la Salud
Materias Investigacion::Ciencias de la Salud::Microbiología y biología molecular
Materias Investigacion::Ciencias de la vida::Citología, biología celular
Cultivo celular
Patología cardiovascular
title_short Analysis of AAV9 biodistribution, transduction efficiency and AAV-miR-935 cardio-specific overexpression in a murine model of myocardial infarction
title_full Analysis of AAV9 biodistribution, transduction efficiency and AAV-miR-935 cardio-specific overexpression in a murine model of myocardial infarction
title_fullStr Analysis of AAV9 biodistribution, transduction efficiency and AAV-miR-935 cardio-specific overexpression in a murine model of myocardial infarction
title_full_unstemmed Analysis of AAV9 biodistribution, transduction efficiency and AAV-miR-935 cardio-specific overexpression in a murine model of myocardial infarction
title_sort Analysis of AAV9 biodistribution, transduction efficiency and AAV-miR-935 cardio-specific overexpression in a murine model of myocardial infarction
dc.creator.none.fl_str_mv García-Olloqui, P. (Paula)|||/items/33aebcca-bf96-4a81-a74d-9fcd60b275cc
author García-Olloqui, P. (Paula)|||/items/33aebcca-bf96-4a81-a74d-9fcd60b275cc
author_facet García-Olloqui, P. (Paula)|||/items/33aebcca-bf96-4a81-a74d-9fcd60b275cc
author_role author
dc.contributor.none.fl_str_mv Pelacho-Samper, B. (Beatriz)
Dadun. Depósito Académico Digital Universidad de Navarra
dc.subject.none.fl_str_mv Materias Investigacion::Ciencias de la Salud
Materias Investigacion::Ciencias de la Salud::Microbiología y biología molecular
Materias Investigacion::Ciencias de la vida::Citología, biología celular
Cultivo celular
Patología cardiovascular
topic Materias Investigacion::Ciencias de la Salud
Materias Investigacion::Ciencias de la Salud::Microbiología y biología molecular
Materias Investigacion::Ciencias de la vida::Citología, biología celular
Cultivo celular
Patología cardiovascular
description Myocardial infarction (MI) leads to an irreversible loss of cardiac myocytes, which compromises cardiac function. The cellular and molecular mechanisms involved in the ischemic event remain under study, being the cardiac exosomes transfer of non-coding RNAs a key process in this pathology. In this context, a comparative analysis of the exosomal compartment of human cardiac progenitor cells (CPC) was performed in comparison with human bone marrow-mesenchymal stem cells (MSC) and human dermal fibroblasts (HDF). A total of 481 differentially expressed microRNAs (miR) were found, being miR-935 the most differentially expressed in CPC. Analysis of miR-935 in vitro overexpression promoted a positive trend to increment cardiomyocyte survival, in response to oxidative stress. Furthermore, when miR-935 regulation was analyzed in a mouse model of MI disease, miR downregulation was shown five days post-infarct in different cardiac subpopulations. In order to study the role of miR-935 in MI and to achieve an optimal miR-935 overexpression in our mouse model, we developed a robust viral-based method for cardiac-specific miR overexpression. Cardiotropic Serotype 9-Adeno-Associated Virus (AAV9) were used for this purpose and their biodistribution was first determined in mice. Ubiquitous EF1α or the cardiac-specific TnT promoters were combined with Luciferase (Luc) or GFP reporters and administered by intramyocardial or intravenous injection, either in healthy or myocardial-infarcted mice. High transgene expression levels were found in the heart, but not in the liver, of mice receiving AAV-TnT, which was significantly higher after intramyocardial injection regardless of ischemia-induction. On the contrary, high hepatic transgene expression levels were detected with the EF1α-promoter, independently of the administration route and heart damage. Luc expression increased with both promoters in a time-dependent manner, reaching a peak by day 3-7 that was stable for at least 60 days. Moreover, tissue-specific GFP expression was found in cardiomyocytes with the TnT-vector, while minimal cardiac expression was detected with the ubiquitous one. Interestingly, we found that MI greatly increased the transcriptional activity of AAV genomes. Thus, we found AAV9-TnT as a robust and stable vector for cardiac-specific delivery after intramyocardial injection. Finally, the therapeutic potential of this vector in combination with miR-935 was evaluated in a mouse model of myocardial ischemia. Intramyocardial injection of AAV9-TnT-miR935 vector did not significantly improve heart function 60 days post-infarct. However, a slight positive trend in the ejection fraction and a decreased adverse cardiac remodeling are observed, suggesting miR-935 putative cardioprotective role.
publishDate 2019
dc.date.none.fl_str_mv 2019
2019-06-27
2020
2020-03-16
2020
2020-03-16
2020
2020-03-16
dc.type.none.fl_str_mv doctoral thesis
http://purl.org/coar/resource_type/c_db06
dc.type.openaire.fl_str_mv info:eu-repo/semantics/doctoralThesis
format doctoralThesis
dc.identifier.none.fl_str_mv https://hdl.handle.net/10171/58884
url https://hdl.handle.net/10171/58884
dc.language.none.fl_str_mv Español
spa
language_invalid_str_mv Español
language spa
dc.rights.none.fl_str_mv open access
http://purl.org/coar/access_right/c_abf2
dc.rights.openaire.fl_str_mv info:eu-repo/semantics/openAccess
rights_invalid_str_mv open access
http://purl.org/coar/access_right/c_abf2
eu_rights_str_mv openAccess
dc.format.none.fl_str_mv application/pdf
dc.source.none.fl_str_mv reponame:Dadun. Depósito Académico Digital de la Universidad de Navarra
instname:Universidad de Navarra
instname_str Universidad de Navarra
reponame_str Dadun. Depósito Académico Digital de la Universidad de Navarra
collection Dadun. Depósito Académico Digital de la Universidad de Navarra
repository.name.fl_str_mv
repository.mail.fl_str_mv
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