生长素对人脐静脉内皮细胞增殖和迁移的影响及机制

The Mechanism of Ghrelin Influencing Human Umbilical Endothelial Cell Proliferation and Migration

目的观察生长素对血管紧张素Ⅱ所致的脐静脉内皮细胞增殖和迁移的影响。方法采取不同浓度的血管紧张素Ⅱ（10-9~10-6mol/L）刺激脐静脉内皮细胞24 h,获取最佳浓度（10-6mol/L）后用此浓度的血管紧张素Ⅱ分别刺激脐静脉内皮细胞0、6、12和24 h,观察脐静脉内皮细胞的增殖和迁移;并观察生长素（10-9~10-6mol/L）预处理2 h后与10-6mol/L血管紧张素Ⅱ共同孵育24 h和10-6mol/L生长素预处理0、0.5、1和2 h后与10-6mol/L血管紧张素Ⅱ共同孵育24 h对脐静脉内皮细胞增殖和迁移的影响。加入丝裂原活化蛋白激酶/细胞外信号调节激酶信号通路阻滞剂PD98059（25μmol/L）、生长激素促分泌素受体1 a阻断剂（[Dys3]GHRP-6 25μmol/L）预处理人脐静脉内皮细胞,观察其对生长素影响血管紧张素Ⅱ诱导的内皮细胞增殖和迁移的作用。水溶性四氮唑8法测细胞增殖,Transwell小室测细胞迁移,免疫印迹法测细胞中细胞外信号调节激酶1/2、磷酸化细胞外信号调节激酶1/2蛋白表达。结果血管紧张素Ⅱ呈浓度和时间依赖性促进内皮细胞迁移和增殖。生长素抑制血管紧张素Ⅱ引起的内皮细胞迁移和增殖呈浓度和时间依赖性。PD98059（25μmol/L）预处理可以抑制血管紧张素Ⅱ促内皮细胞增殖作用,[Dys3]GHRP-6（25μmol/L）预处理阻断生长素对内皮细胞迁移和增殖的抑制作用。生长素可减少磷酸化细胞外信号调节激酶1/2的表达。结论生长素可抑制血管紧张素Ⅱ诱导的内皮细胞增殖和迁移,其机制涉及细胞外信号调节激酶1/2途径。

Aim To observe the effect of ghrelin on human umbilicus endothelial cell（HUVEC） proliferation and migration. Methods Different concerntration of angiotesionⅡ（AngⅡ）（10-9～10-6 mol/L） was adopted to excite HUVEC for 24 hours,and the best concerntration of 10-6 mol/L was acquired,and used to stimulate HUVEC for 0,6,12,24 hours respectively to observe human umbilicus endothelial cell proliferation and migration.（10-9～10-6 mol/L） ghrelin was pretreated for 2 hours and cultivated for 24 hours with HUVEC and 10-6 mol/L ghrelin was used to pretreat for 0,0.5,1,2 hour and cultivate HUVEC for 24 hours to observe the cells proliferation and migration.MAPK /ERK1/2 singal pathy inhibitor PD98059（25 μmol/L）,GHSR1a receptor inhibitor（GHRP-6 25 μmol/L） were added to pretreat HUVEC to observe ghrelin effect on AngⅡ induced endothelial cell injury.Cell proliferation was measured with WST-8,and cell migration with Transwell;ERK1/2 and p-ERK1/2 protein expression with western blot. Results AngⅡ promoted HUVEC proliferation and migration with dose and time dependent manner.Ghrelin inhibited AngⅡpromoting effect on cell migration and proliferation with dose and time dependent manner.Pretreatment with PD98059（25 μmol/L） can inhibit the promoting effect of HUVEC proliferation and migration;pretreatment with GHRP-6（25 μmol/L） can block ghrelin＇s inhibiting effect on cell migration and proliferation effect.Ghrelin can decrease the p-ERK1/2 expression. Conclusion Ghrelin can inhibit AngⅡinduced HUVEC proliferation and migration,and the mechanism was involved in ERK1/2 pathway.