RESUMO
Objective To investigate the effect of hypoxic pretreatment on the angiogenesis of aged human bone marrow mesenchymal stem cells (hBMSCs), so to provide experimental support for more effective autologous stem cell transplantation therapy in aged patients with ischemic myocardial injury. Methods The aged hBMSCs were cultured in a hypoxic incubator, and then the cell morphology was observed under inverted phase-contrast microscope, the surface markers were detected by flow cytometry, and the differentiation potential was detected by osteogenic and adipogenic differentiation. Subsequently, the conditioned medium (CM) of young hBMSCs under normoxic culture (norCM), the conditioned media of aged hBMSCs under normoxic and hypoxic culture(hypoCM) were collected respectively. They were named as norCM-young, norCM-old and hypoCM-old. The equal volume of medium, which was not treated with stem cells, was set as control group. Human umbilical vein endothelial cells (HUVECs) were treated with 4 conditioned media, the cell survival rate was detected by CCK-8 assay, and the tube formation experiment was used to detect the angiogenesis ability in vitro. The BCA method was used to detect the total protein concentration of the conditioned medium of each group, and the Western blotting was used to detect the expression of hypoxia inducible factor-1α (HIF-1α) in the cells and the conditioned media. Results There was not significant effect of hypoxic pretreatment on the morphology, surface markers and differentiation ability of aged hBMSCs (P > 0. 05. n ≥ 3). Compared with the norCM-old group, hypoCM-old group significantly improved the survival of HUVECs under hypoxia-reoxygenation condition (P < 0. 05, n = 5), and the tube formation ability of it (P<0. 01, n = 5). The total protein concentration of hypoCM-old group was significantly higher than that of norCM-old group (P<0. 05, n = 3). The expression of HIF-1α in hBMSCs of hypo-old group was significantly higher than that of nor-old group (P<0. 05,n = 3), while the content of HIF-1α in conditioned medium of hypoCM-old group was significantly higher than that in norCM-old group (P<0. 01,n = 3). Conclusion The aged hBMSCs pretreated with hypoxia can promote the survival and tube formation of HUVECs through paracrine in vitro, which is HIF-1α-related.