Effect of angiotensin II on biological characteristics of human umbilical cord mesenchymal stem cells

Abstract

Objective To investigate the effects of different concentrations of angiotensin Ⅱ on the apoptosis, proliferation and paracrine of human umbilical cord mesenchymal stem cells in vitro and select the appropriate concentration of angiotensin Ⅱ to pretreat mesenchymal stem cells. Methods According to the method of the author′s laboratory established for isolate, culture and identification of the human umbilical cord mesenchymal stem cells, the 3-8th human umbilical cord mesenchymal stem cells were cultured in plates. The human umbilical cord mesenchymal stem cells were randomly divided into 4 groups (3 in each group): control group, 100 ng/mL angiotensin Ⅱ group, 500 ng/mL angiotensin Ⅱ group and 1000 ng/mL angiotensin Ⅱ group. The culture media containing 100 ng/mL, 500 ng/mL and 1000 ng/mL angiotensin Ⅱ were used to treat experimental groups, respectively, the control group was cultured with normal medium without Ang-Ⅱ. When the human umbilical cord mesenchymal stem cells were cultured for 24, 48 and 72 h, the cell morphology and density were observed by inverted microscope. The apoptosis was assessed by acridineorange/ethidium bromide staining. The cell proliferation activity was measured by Cell Counting Kit-8, and the cell cycle (S period) was tested by flow cytometry after propidium iodide staining. The levels of vascular endothelial growth factor, basic fibroblast growth factor, and hepatocyte growth factor in each group were detected by enzyme linked immunosorbent assay. Results When the human umbilical cord mesenchymal stem cells was cultured for 24 h, 48 h, 72 h, cell morphology under inverted microscope in each group was long-shuttle shape. A few cells were polygonal. The fusion rate of 100 ng/mL angiotensin Ⅱ group were 40%-45%, 70%-80%, 90%-95%, the fusion rate was significantly increased. Acridineorange/ethidium bromide staining showed that no apoptosis cells were observed in group of 100 ng/mL angiotensin Ⅱ and the group of control. The results of Cell Counting Kit-8 also showed that the cells value-added of 100 ng/mL angiotensin Ⅱ group was significantly faster than that in control group; The results of flow cytometry showed that the ratio of the number of cells in 100 ng/mL angiotensin Ⅱ group was significantly increased. Enzyme linked immunosorbent assay analysis showed that the contents of the vascular endothelial growth factor, basic fibroblast growth factor, hepatocyte growth factor of 100 ng/mL angiotensin Ⅱ group were highest, and compared with the control group, the difference was statistically significant (P<0.05). Conclusion 100 ng/mL angiotensin Ⅱ can inhibit apoptosis and promote cell proliferation, which may be related to a large number of secretion of vascular endothelial growth factor, basic fibroblast growth factor, hepatocyte growth factor. Key words: Angiotensin Ⅱ; Umbilical cord; Mesenchymal stem cells; Cell proliferation; Apoptosis; Paracrine