Publicação
Advancing cell-based therapies towards the treatment of myocardial infarction
| Resumo: | Cardiovascular diseases remain the leading cause of death worldwide, with current available therapies failing to prevent or revert cardiac dysfunction. Thus, efforts have been made towards the development of new approaches with the potential to promote heart regeneration. Growing evidence suggests that the beneficial effects of transplanted cells are conducted by cells’ secreted factors, such as extracellular vesicles (EVs). In this context, EV-based products are emerging as promising therapeutic strategies to repair cardiac damage after myocardial infarction. Since the clinical translation of human adipose tissue-derived MSC (hAT-MSC) and hAT-MSC-derived EVs is currently limited by their scalability, in this work, two different strategies for manufacturing MSC-derived EVs with increased cardiac regenerative potential while maximizing EV secretion yields were explored. hAT-MSC were (i) transiently preconditioned, through glucose starvation and transfec-tion with miR-145-5p inhibitor, known to impact their angiogenic potential and (ii) hAT-MSC were genetically modified with a lentiviral vector co-expressing apelin and FGF-2, two angiogenic and cardioprotective factors. hAT-MSC were expanded using a microcarrier-based culture system in stirred-tank bioreactors under hypoxic conditions. hAT-MSC-derived EVs were isolated from the conditioned medium by tangential flow filtration followed by size exclusion chromatography. The different strategies implemented resulted in increased EV productivities (1.3-1.8-fold increase). We have observed that decreasing glucose concentration below 1 mM can not only impact the ability of MSC to secrete EVs, but also enhance their bioactivity towards cardiac regeneration, shedding light on the importance of standardizing cell culture conditions to develop more robust platforms for EV production. Additionally, we have shown that chemically-based non-viral gene delivery of miR-145-5p inhibitor can constitute a suitable approach to target the therapeutic potential of hAT-MSC-derived EVs. Finally, we have observed that genetically modified hAT-MSC can also secrete EVs with cardiac regeneration potential. |
|---|---|
| Autores principais: | Sousa, Carolina Drumonde |
| Assunto: | Extracellular vesicles Mesenchymal stem/stromal cells Stirred-tank bioreactors Preconditioning strategies Genetically modified cells Cardiac regenerative medicine |
| Ano: | 2022 |
| País: | Portugal |
| Tipo de documento: | dissertação de mestrado |
| Tipo de acesso: | acesso aberto |
| Instituição associada: | Universidade Nova de Lisboa |
| Idioma: | inglês |
| Origem: | Repositório Institucional da UNL |
| Resumo: | Cardiovascular diseases remain the leading cause of death worldwide, with current available therapies failing to prevent or revert cardiac dysfunction. Thus, efforts have been made towards the development of new approaches with the potential to promote heart regeneration. Growing evidence suggests that the beneficial effects of transplanted cells are conducted by cells’ secreted factors, such as extracellular vesicles (EVs). In this context, EV-based products are emerging as promising therapeutic strategies to repair cardiac damage after myocardial infarction. Since the clinical translation of human adipose tissue-derived MSC (hAT-MSC) and hAT-MSC-derived EVs is currently limited by their scalability, in this work, two different strategies for manufacturing MSC-derived EVs with increased cardiac regenerative potential while maximizing EV secretion yields were explored. hAT-MSC were (i) transiently preconditioned, through glucose starvation and transfec-tion with miR-145-5p inhibitor, known to impact their angiogenic potential and (ii) hAT-MSC were genetically modified with a lentiviral vector co-expressing apelin and FGF-2, two angiogenic and cardioprotective factors. hAT-MSC were expanded using a microcarrier-based culture system in stirred-tank bioreactors under hypoxic conditions. hAT-MSC-derived EVs were isolated from the conditioned medium by tangential flow filtration followed by size exclusion chromatography. The different strategies implemented resulted in increased EV productivities (1.3-1.8-fold increase). We have observed that decreasing glucose concentration below 1 mM can not only impact the ability of MSC to secrete EVs, but also enhance their bioactivity towards cardiac regeneration, shedding light on the importance of standardizing cell culture conditions to develop more robust platforms for EV production. Additionally, we have shown that chemically-based non-viral gene delivery of miR-145-5p inhibitor can constitute a suitable approach to target the therapeutic potential of hAT-MSC-derived EVs. Finally, we have observed that genetically modified hAT-MSC can also secrete EVs with cardiac regeneration potential. |
|---|