miR-520a induce re-enter in S phase of pluripotent stem cell derived cardiomyocytes

NATALIA LUCÍA SANTÍN VELAZQUE; XIMENA GARATE; MARIA AGUSTINA SCARAFIA; ALAN MIQUEAS MÖBBS; CAROLINA COLLI; GUADALUPE AMIN; ARIEL WAISMAN; GUSTAVO EMILIO SEVLEVER; CARLOS DANIEL LUZZANI; ALEJANDRO DAMIÁN LA GRECA; SANTIAGO GABRIEL MIRIUKA

Los Angeles

Congreso; International Society for Stem Cell Research; 2019

ISSCR

Human heart has little regenerative capacity after myocardial damage due to the low proliferation potential of cardiomyocytes (CM). Hence, there is a growing interest in the production of human induced pluripotent stem cell-derived CM (hiPSC-CM) as a potentially promising strategy for regenerative therapies, and in identifying factors playing relevant roles in the regulation of the hiPSC-CMs cell cycle. Moreover, recent findings highlights the regenerative capacity of mesenchymal stem cells (MSC) and the many properties of MSC-conditioned medium and extracellular vesicles (EV) in cell culture. The aim of this study is to investigate the ability of the hiPSC-CM to re-enter cell cycle after exposure to conditioned medium from WJ-MSC. Human iPSC-CMs were obtained with a monolayer protocol. We established FUCCI-hiPSCs, a line constitutively expressing Fluorescent Ubiquitination-based Cell Cycle Indicator, which consists of system that employs red fluorescence for G1 state and green for S/G2/M. FUCCI-hiPSC-CMS were incubated with WJ-MSC-conditioned medium for 24, 48 and 72 hours. At the first 24hs, hiPSC-CM in S/G2/M increased from 14.7% to 28.7% with conditioned medium. Interestingly, while replication activity decrease over time with control medium (near 5% at 48 and 72hs), with MSC-conditioned medium maintain up to 20%. Enhanced DNA synthesis was confirmed by EdU assay and subsequent immunofluorescence. Finally, we used LC-MS/MS proteomic approach to identify proteins presents in the secretome from WJ-MSC that could explain the effect in the hiPSC-CM. Through Gene Ontology analysis we found 147 proteins involved in cell cycle. Particularly, some of them were related to the G2/M checkpoint and mTOR pathway. In conclusion, this study reveals that the exposure to MSC-contidionated medium induce proliferation of hiPSC-CMs. Our next goal is to study the differential expressed genes in the proliferative hiPSC-CM.