BPS2026 – The effect of the maturation of cardiomyocytes on angiogenesis through paracrine secretion

The regeneration of mammalian hearts after myocardial infarction(MI) is extremely inefficient. Preliminary data indicated that loss of regenerative and angiogenic capacity of the heart after birth is mechanistically linked. We hypothesized that mature cardiomyocytes inhibit angiogenesis through a paracrine effect. To test this hypothesis, we purified extracellular vesicles (EVs) from human induced pluripotent stem-cell-derived cardiomyocytes (iPSC-CM) at different stages of maturation. We assessed their effect on iPS-endothelial cells (iPS-ECs). Also, to identify the RNAs affecting induction of cardiac-angiogenesis and arterialization, RNA content of EVs derived from iPS-CMs at different stages of maturation were analyzed. EVs were collected from three different stages of CM maturation. In early cardiomyocyte stage (early-CM), compared to late stages (early-late-CM) and the end of the late stage (late-mature-CM), tube-formation-assay demonstrated that CM-differentiation and maturation suppressed angiogenesis. However, early-CM-EVs and early-late-CM-EVs have a more suppressive effect compared to late-CM-EVs on angiogenesis over long periods. Early-late and late-CM-EVs have also caused a decrease in the mRNA expression level of endothelial cell markers CD31, CD105, CD144, and CD309. These results suggested that the CM-maturation phase has an inhibitory effect on endothelial cell differentiation, development, and angiogenesis. To assess the effect of CM-derived EVs on membrane fluidity, we have measured the general polarization (GP) level of ECs via confocal microscopy. We observed that the membrane fluidity of ECs dramatically decreased upon the late-CM-EV incubation compared to earlier stages of CM-EV groups. Hence, early and early-late CM-EV incubation also caused a reduction in membrane packing, suggesting that the paracrine effect of CM-derived EVs in the modulation of angiogenesis varies depending on the time point of maturation. Overall, ten different mRNAs have a role in the inhibition of angiogenesis and endothelial cell proliferation, helping innovative RNA-therapies to simultaneously induce cardiomyocyte proliferation and angiogenesis (TUBITAK-223N181).