1-磷酸鞘氨醇在高糖诱导血管内皮细胞功能损伤中的作用及其机制探讨

Discussion on the role and mechanism of sphingosine 1-phosphate in the high glucose-induced vascular endothelial cell function injury

目的探讨1-磷酸鞘氨醇(S1P)在高糖诱导血管内皮细胞功能损伤中的作用及其机制。方法取人脐带静脉内皮细胞培养,取5代以内对数生长周期的细胞进行实验,检测各组24 h细胞活力水平、细胞凋亡水平,PLC(磷脂酶C)、Src激酶拮抗剂与S1P混合作用凋亡水平、Akt激活情况。结果正常对照组、甘露醇高渗对照组的细胞活力以及细胞凋亡差异无统计学意义(P>0.05)。在高糖处理下,细胞活力显著下降,细胞凋亡水平显著上升。模型组1μmol/L、10μmol/L、20μmol/L剂量条件下细胞活力高于高糖处理组、细胞凋亡率低于高糖处理组,呈剂量依赖,差异具有统计学意义(P<0.05)。在含有10μmol/L浓度S1P的培养液中分别加入PLC抑制剂U63122、Src激酶抑制剂PP2,则凋亡率逐渐上升,差异有统计学意义(P<0.05)。应用U73122阻断PLC后,Akt磷酸化无显著的变化,PTX、S1P、U73122(P>0.05),添加Src激酶抑制剂PP2后,Akt的磷酸化水平显著下降(P<0.05)。结论1-磷酸鞘氨醇在高糖诱导血管内皮细胞功能损伤中起到抗凋亡作用,可能与其调节Src/Akt通路功能有关。

Objective To investigate the role and mechanism of sphingosine 1-phosphate（S1P）in high glucose-induced vascular endothelial cell function injury. Methods The human umbilical cord vein endothelial cell was collected for culture. The cells in the logarithmic growth cycle within 5 generations were tested. The 24h cell viability level, cell apoptosis level, apopsotsis level of the combined effect of PLC（phospholipase C）, Src kinase antagonist and S1P and Akt activation were detected in each group. Results There was no statistically significant difference in cell viability and apoptosis between the normal control group and the mannitol hypertonic control group（P〉0.05）. Under high glucose treatment, cell viability was decreased significantly and apoptosis levels were increased significantly. The cell viability in the model group was higher than that in the high glucose treatment group and the apoptosis rate was lower than that in the high glucose treatment group at doses of 1 μumol/L, 10 μumol/L, and 20 μumol/L, showing a dose-dependent trend, and the difference was statistically significant（P〈0.05）. When the PLC inhibitor U63122 and Src kinase inhibitor PP2 were added to the culture medium containing 10 μmol/L S1P, the apoptosis rate was increased gradually, and the difference was statistically significant（P〈0.05）. After U73122 was used to block PLC, there was no significant change in Akt phosphorylation, PTX, S1P, U73122（P〉0.05）. After the Src kinase inhibitor PP2 was added, the phosphorylation level of Akt was significantly decreased（P〈0.05）. Conclusion Sphingosine 1-phosphate（S1P） plays an anti-apoptotic role in high glucose-induced vascular endothelial cell function injury, which may be related to its regulation of Src/Akt pathway.