波生坦对高糖诱导静脉内皮细胞血管表达的影响

Effects and its mechanism of bosentan on expression of vascular endothelial vein with high glucose-induced

目的研究波生坦对高糖诱导静脉内皮细胞血管表达的影响。方法提取健康剖腹产孕妇脐静脉内皮细胞,在体外人工培养、分化、制成细胞悬液并培养。将内皮细胞随机分为3组:正常组、模型组、实验组。正常组加入正常糖浓度5.5 mmol·L^(-1);模型组加入糖浓度30 mmol·L^(-1);实验组在模型组的基础上加入波生坦0.1 mmol·L^(-1),各组培养24 h。用荧光实时定量PCR法检测人体静脉内皮细胞血管的内皮生长因子(VEGF)、转录活化因子3(STAT3)、Toll样受体4(TLR4)水平,用免疫印迹法检测人体静脉内皮细胞的磷酸化细胞外信号调节蛋白激酶(pE RK1/2)表达水平。结果模型组加入高糖0,12,24 h后,VEGF水平分别为0.14±0.02,0.25±0.04,0.41±0.07,pE RK1/2水平分别为0.50±0.07,1.27±0.24,1.92±0.16,加糖24 h后与加糖前比较,差异均有统计学意义(均P<0.05)。加入药物24 h后,实验组、模型组和正常组VEGF水平分别为0.07±0.01,0.41±0.07,0.26±0.03;这3组的TLR4分别为0.70±0.07,1.94±0.32,1.28±0.12;这3组的p ERK1/2分别为0.66±0.05,1.92±0.16,0.92±0.02,模型组与正常组比较,差异均有统计学意义(均P<0.05);实验组与模型组比较,差异均有统计学意义(均P<0.05)。结论波生坦可以有效减少高糖诱导静脉内皮细胞血管VEGF的表达,其机制可能与波生坦抑制ERK信号通路有关。

Objective To study effect and its mechanism of bosentan on expression of vascular endothelial cells with high glucose induced.Methods Human umbilical vein endothelial cells from healthy caesarean section were extracted,cultured,differentiated and culture of cell suspension was cultured in vitro. The endothelial cells were randomly divided into three groups： normal group,model group,and experimental group. The normal group was was added with normal sugar concentration of 5. 5 mmol·L^-1. The model group was added with sugar concentration of 30 mmol·L^-1. The experimental group added 0. 1 mmol·L^-1 of bosentan on the basis of the model group. Each group was cultured for 24 hours.The vascular endothelial growth factor（ VEGF）,activator of transcription3（ STAT3） and Toll like receptor 4（ TLR4） in human vein endothelial cells were detected by real-time fluorescence quantitative PCR. The expression of phosphorylated extracellular signal regulated protein kinase（ p ERK1/2） in human vein endothelial cells was detected by Western blot. Results After the model group was treated with high glucose 0,12and 24 h,the levels of VEGF were 0. 14 ± 0. 02,0. 25 ± 0. 04 and 0. 41 ± 0. 07,p ERK1/2 were 0. 50 ± 0. 07,1. 27 ± 0. 24 and 1. 92 ± 0. 16; compared with that before,the differences after adding the sugar 24 h were statistically significant（ all P〈0. 05）. After addition the drugs of 24 h,the VEGF levels in the experimental group,the model group and the normal group were respectively 0. 07 ± 0. 01,0. 41 ± 0. 07 and 0. 26 ± 0. 03; TLR4 in the three groups were 0. 70 ± 0. 07,1. 94 ± 0. 32 and 1. 28 ± 0. 12; p ERK1/2 in the three groups were 0. 66 ± 0. 05,1. 92 ± 0. 16 and0. 92 ± 0. 02; Compared with the normal group and model group,the differences were statistically significant（ all P〈0. 05）. Compared with the model group and the experimental group,the differences were statistically significant（ all P〈0. 05）. Conclusion Bosentan can effectively reduce the expression of VEGF in vascular endothelial cells induced by high glucose,and it is suitable for extensive promotion. The mechanism may be related to the inhibition of ERK signaling pathway by bosentan.and 24 h,the levels of VEGF were 0. 14 ± 0. 02,0. 25 ± 0. 04 and 0. 41 ± 0. 07,p ERK1/2 were 0. 50 ± 0. 07,1. 27 ± 0. 24 and 1. 92 ± 0. 16; compared with that before,the differences after adding the sugar 24 h were statistically significant（ all P〈0. 05）. After addition the drugs of 24 h,the VEGF levels in the experimental group,the model group and the normal group were respectively 0. 07 ± 0. 01,0. 41 ± 0. 07 and 0. 26 ± 0. 03; TLR4 in the three groups were 0. 70 ± 0. 07,1. 94 ± 0. 32 and 1. 28 ± 0. 12; p ERK1/2 in the three groups were 0. 66 ± 0. 05,1. 92 ± 0. 16 and0. 92 ± 0. 02; Compared with the normal group and model group,the differences were statistically significant（ all P〈0. 05）. Compared with the model group and the experimental group,the differences were statistically significant（ all P〈0. 05）. Conclusion Bosentan can effectively reduce the expression of VEGF in vascular endothelial cells induced by high glucose,and it is suitable for extensive promotion. The mechanism may be related to the inhibition of ERK signaling pathway by bosentan.