肝激酶B1对高糖条件下ARPE-19细胞增殖、迁移能力的影响及机制研究

Effect and mechanism of liver kinase B1 on ARPE-19 cell proliferation and migration under high glucose condition

目的观察肝激酶B1(LKB1)对高糖条件下人视网膜色素上皮细胞(ARPE-19)增殖、迁移的影响,并探讨其相关调控机制。方法LKB1真核表达载体pCMV-LKB1转染ARPE-19细胞,筛选稳定过表达LKB1的ARPE-19细胞株。将ARPE-19细胞分为4组:正常对照组、高糖组(HG组)、高糖+空白组(HG+B组)、高糖+LKB1组(HG+LKB1组)。用EdU法检测各组细胞增殖情况,Transwell小室观察各组细胞迁移能力,蛋白免疫印迹法检测各组细胞中相关蛋白的表达。结果EdU法检测结果显示,HG+LKB1组与HG组、HG+B组相比细胞增殖率显著降低(P<0.01);Transwell小室观察发现,HG+LKB1组与HG组、HG+B组相比细胞迁移能力显著降低(P<0.01)。蛋白免疫印迹法检测结果显示,HG+LKB1组与HG组、HG+B组相比基质金属蛋白酶-2(MMP2)和血管内皮生长因子(VEGF)的表达显著降低(P<0.01),磷酸化的AMP依赖的蛋白激酶(AMPK)显著增加(P<0.01),而磷酸化的哺乳类动物雷帕霉素靶蛋白(mTOR)则显著下降(P<0.01)。结论LKB1可下调MMP2和VEGF的表达,抑制高糖条件下ARPE-19细胞的增殖和迁移,该调控机制可能通过AMPK-mTOR信号通路发挥作用。

Objective To observe the effect of liver kinase B1(LKB1)on the proliferation and migration of ARPE-19 cells under high glucose condition and to explore its regulatory mechanism.Methods LKB1 eukary-otic expression vector pCMV-LKB1 transfected into ARPE-19 cells and screened stable overexpression LKB1 cell lines.ARPE-19 cells were divided into four groups:(1)normal control group(the control group),(2)high glucose group(HG group),(3)high glucose+blank group(HG+B group),(4)high glucose+LKB1 group(HG+LKB1 group).Cell proliferation in each group was detected by EdU method,Transwell observes cell mi-gration ability of each group,Western blotting detected the expression of associated proteins in various cells.Results EdU results showed that the cell proliferation rate of HG+LKB1 group was significantly lower than that of HG group and HG+B group(P<0.01);Transwell results showed a significant decrease in cell migra-tion force in the HG+LKB1 groups as compared with the HG and HG+B groups(P<0.01).Western blotting re-sults showed that the expression of vascular endothelial grovoth factor(VEGF)and MMP2 in HG+LKB1 group was significantly lower than that in HG group and HG+B group(P<0.01),the phosphorylated adenos-ine monophosphate-dependent kinase(AMPK)increased significantly(P<0.01),and the phosphorylated mTOR decreased significantly(P<0.01).Conclusion LKB1 can down regulate the expression of MMP2 and VEGF and inhibit the proliferation and migration ability of ARPE-19 cells under high sugar conditions,and this regulatory mechanism may be carried out through AMPK-mTOR signaling pathways.