Conclusion
HIF-1α was persistently overexpressed in Hyp-CMC-SMSC as compared to Hyp-CMC (except at 72 h). Hyp-CMC-MSC showed a weaker HIF-1α expression than Hyp-CMC-SMSC in most tested time points, except after 8 h. The Ki-67 expression showed the strongest upregulation in Hyp-CMC after 24 and 48 h incubation, then returned to baseline level, while a temporary increase in Ki-67 expression in Hyp-CMC-MSC at 4 and 8 h and at 48 h in Hyp-CMC-SMSC could be observed. RhoA was increased in normoxic MSCs and in Hyp-CMC-SMSC over time, but not in Hyp-CMC-MSC. A temporary increase in IL-18 protein expression was detected in Hyp-CMC-SMSC and Hyp-CMC. Our study demonstrates timely dynamic changes in expression of different ischemia and regeneration-related genes of CMCs, depending from the culture condition, with stronger expression of HIF-1α, RhoA and IL-18 if the hypoxic CMC were subjected to the secretome of MSCs.
Methods
In this study, the regenerative potential of hypoxic human cardiomyocytes (group Hyp-CMC) in vitro was evaluated when co-cultured with human bone-marrow derived MSC (group Hyp-CMC-MSC) or stimulated with the secretome of MSC (group Hyp-CMC-SMSC). The secretome of normoxic MSC and CMC, and the hypoxic CMC was analyzed with a cytokine panel. Gene expression changes of HIF-1α, proliferation marker Ki-67 and cytokinesis marker RhoA over different reoxygenation time periods of 4, 8, 24, 48, and 72 h were analyzed in comparison to normoxic CMC and MSC. Further, the proinflammatory cytokine IL-18 protein expression change, metabolic activity and proliferation was assessed in all experimental setups.