高糖环境下P物质活化Akt通路促进间充质干细胞增殖和成血管内皮细胞分化的研究

An experimental study of substance P promotes the proliferation of bone marrow mesenchymalstem cells and the differentiation of vascular endothelial cells by activating Akt pathway inhigh glucose environment

目的揭示高糖环境下P物质(SP)对人骨髓间充质干细胞(BMSC)增殖、迁移及成血管内皮细胞分化的影响,为其应用于治疗糖尿病皮肤溃疡等并发症提供依据。方法细胞高糖模型分为高糖对照组、高糖SP组和高糖蛋白激酶B(Akt)抑制剂组;正常环境培养的BMSC为正常对照组。CCK-8法检测细胞增殖情况;Transwell细胞小室迁移实验检测BMSC迁移数目;各组BMSC进行成血管内皮细胞分化实验,酶联免疫吸附试验检测成血管内皮细胞分化后各组BMSC的CD31蛋白含量;基质胶网眼形成实验检测成血管形成能力。Western blot检测各组BMSC的Akt和磷酸化Akt(p-Akt)蛋白表达情况。结果与正常对照组比较,高糖对照组吸光度(A_(450))值、迁移数目、CD31蛋白含量、网眼数目、Akt和p-Akt蛋白相对表达量均降低;与高糖对照组比较,高糖SP组A_(450)值、迁移数目、CD31蛋白含量、网眼数目、Akt和p-Akt蛋白相对表达量均升高;与高糖SP组相比,高糖Akt抑制剂组上述指标均降低(P<0.01)。结论高糖环境下,SP可活化Akt通路促进BMSC增殖、迁移和血管内皮细胞分化。

Objective To investigate the effect and mechanism of substance P(SP)on the proliferation,migration and vessel endothelial cell differentiation of human bone marrow mesenchymal stem cells(BMSCs)in high glucose environment,and to lay a foundation for the treatment of diabetic complications such as skin ulcers.Methods The cell high glucose model was divided into the high glucose control group,the high glucose SP group and the high glucose Akt inhibitor group.And BMSCs cultured in normal environment were used as the normal control group.CCK-8 assay was used to detect the cell proliferation.Transwell cell migration assay was used to determine the cell migration number of BMSCs.In each group,the differentiation of vascular cells was induced,and ELISA was used to detect CD31 protein content of BMSCs after differentiation for each group.Matrix mesh formation test was used to detect the angiogenesis ability.Western blot assay was used to detect the expression levels of Akt and phosphorylated Akt(p-Akt)in each group of BMSCs.Results Compared with the control group and high glucose control group,the absorbance(A_(450))value,migration number,CD31 protein content,mesh number,relative expression levels of Akt and P-Akt protein were decreased in the high glucose SP group.Compared with the highglucose group,the A_(450) value,migration number,CD31 protein content,mesh number,relative expression levels of Akt and PAkt protein were significantly increased in the high-glucose SP group(P<0.01).Compared with the high-glucose SP group,these indexes were all reduced in the high glucose Akt inhibitor group(P<0.01).Conclusion In high glucose environment,SP promotes the proliferation,migration and angiogenic differentiation of BMSCs by activating Akt pathway.