2022, Vol. 26 ›› Issue (31): 5008-5013

Effect and mechanism of conditioned medium from hypoxia preconditioned human Wharton’s Jelly derived mesenchymal stem cells on myocardial ischemia/reperfusion injury

Li Yan1, 2, Zhang Yan2, Zhang Ningkun2, Chen Yu1, 2   

Abstract: BACKGROUND: Mesenchymal stem cells improve tissue injury mainly through their paracrine action, which is enhanced by hypoxic preconditioning. However, the effect of conditioned medium from hypoxia preconditioned human Wharton’s Jelly derived mesenchymal stem cells on myocardial ischemia/reperfusion injury and its mechanism still need to be further studied.
OBJECTIVE: To investigate the effect and mechanism of conditioned medium from hypoxia preconditioned human Wharton’s Jelly derived mesenchymal stem cells on the apoptosis of H9C2 cells after ischemia/reperfusion injury.
METHODS: Human Wharton’s Jelly derived mesenchymal stem cells were treated with normoxia or hypoxia to obtain conditioned medium. The H9C2 cells were subjected to hypoxia/reoxygenation and serum-free culture for 6 hours to mimic the myocardial ischemia/reperfusion injury model in vitro. Lactate dehydrogenase activity in H9C2 conditioned medium was detected using lactate dehydrogenase assay kit. Rat myocardial H9C2 cells were divided into four groups: normoxic culture group, model group, normoxic conditioned medium group and hypoxic conditioned medium group. After treatment with normoxic or hypoxic conditioned medium for 24 hours after hypoxia, the apoptosis rate of H9C2 cells was detected by flow cytometry. Western blot assay was applied to detect the expression of apoptotic markers (Bax and Bcl-2) and autophagic markers (Beclin-1, P62 and LC3) in each group.
RESULTS AND CONCLUSION: (1) The morphology of H9C2 cells did not change significantly after treatment with ischemia/reperfusion or human Wharton’s Jelly derived mesenchymal stem cells-conditioned medium. (2) Compared with the normoxic culture group, lactate dehydrogenase activity in supernatant of H9C2 cells in the model group was significantly increased, but decreased in both conditioned medium groups. (3) Compared with the model group, the apoptosis rate of H9C2 and the expression of Bax, Beclin-1, and LC3 protein were downregulated and the expression of p62 protein was upregulated in H9C2 cells in the normoxic conditioned medium group and hypoxic conditioned medium group. The change was more significant in the hypoxic conditioned medium group than that in the normoxic conditioned medium group. (4) The results demonstrated that the conditioned medium from hypoxia preconditioned human Wharton’s Jelly derived mesenchymal stem cells could reduce the ischemia/reperfusion injury induced apoptosis of H9C2, and the protective effect may be related to the inhibition of autophagy.
Key words: Wharton’s Jelly derived mesenchymal stem cells, ischemia/reperfusion injury, autophage, hypoxic preconditioning, myocardial infarction