摘要目的 探讨脂筏在肝细胞生长因子受体介导的信号跨膜转导中的作用.方法 采用甲基-β-环糊精(MβCD)处理HepG2细胞,以干扰脂筏的形成,然后加入人工重组肝细胞生长因子以活化其受体.采用Western blot技术及计算机扫描定量分析分别检测MβCD处理组和对照组细胞磷脂酶Cγ1(PLCγ1)/二脂酰甘油/蛋白激酶C信号通路、磷脂酰肌醇-3-激酶/磷脂酰肌醇依赖的蛋白激酶/蛋白激酶B信号通路和有丝分裂原激活的蛋白激酶(MAPK)信号通路活性的变化.结果 (1)用MβCD处理细胞后,细胞PLCγ1磷酸化程度降低,为对照组的65%(P=0.022);胞浆中PLCγ1含量增加,为对照组的1.75倍(P=0.017);膜结合的PLCγ1减少,为对照组的70%(P=0.037).(2)用MβCD处理细胞后,胞浆中蛋白激酶B含量减少,为对照组的62%(P=0.028),同时膜结合的蛋白激酶B磷酸化程度降低,为对照组的86%(P=0.041).用MβCD处理细胞同时可抑制磷脂酰肌醇依赖的蛋白激酶由胞浆向质膜的转移及活化.(3)MβCD处理对MAPK信号通路,包括细胞外信号调节蛋白激酶/MAPK信号通路、p38/MAPK信号通路和C-Jun N末端蛋白激酶/MAPK信号通路均无显著的影响.结论 抑制脂筏形成可下调肝细胞生长因子/c-Met对PLCγ1/二脂酰甘油/蛋白激酶C信号通路和磷脂酰肌醇-3-激酶/磷脂酰肌醇依赖的蛋白激酶/蛋白激酶B信号通路的活化.

abstractsObjective To study the effects of lipid rafts on cell signal transmembrane transduction mediated by c-Met. Methods After HepG2Cells were treated with MβCD to disrupt the lipid rafts and were treated with artificial recombination hepatocyte growth factor to activate c-Met, the activities of PLCγ1/PKC, P13K/Akt and MAPK signaling pathways in HepG2 cells were analyzed using Western blot. Results (1) After disruption of lipid rafts with MβCD, phosphorylation of PLCγ1 decreased by 35% (P=0.022); the content of PLCγ in the cytoplasm increased by 1.75 fold (P=0.017); PLCγ1 conjugated with membrane decreased by 30% (P=0.037). (2) The content of PKB in the cytosol decreased by 38% (P=0.028), and the phosphorylation level of PKB conjugated with membrane decreased by 14% (P=0.041). At the same time, PDK translocation from cytosol to the plasma membrane and its activation were inhibited by treatment with MβCD. (3) Treatment with MβCD had no significant effect on ErK/MAPK, p38/MAPK and JNK/MAPK signaling pathways. Conclusion Disruption of lipid rafts with MβCD inhibits the activation of PLCγ1/PKC and PI3K/PKB signaling pathways by HGF/cMet, but has no effect on MAPK signaling pathway.