PTEN/mTOR通路在高糖诱导的人绒毛膜滋养层细胞HTR-8/SVneo凋亡中的作用机制探讨 #br#

目的 探讨高糖诱导人绒毛膜滋养层细胞HTR-8/SVneo凋亡的相关机制。方法 体外培养人绒毛膜滋 养层细胞HTR-8/SVneo，分为空白对照组、高糖组、NC组（阴性对照组）、第10号染色体缺失的磷酸酶及张力蛋白同 源物基因（PTEN）-siRNA 组（抑制 PTEN 表达组）及哺乳动物雷帕霉素靶蛋白（mTOR）通路抑制剂（GDC-0349）组。 四甲基偶氮唑蓝（MTT）法检测HTR-8/SVneo细胞增殖能力；流式细胞仪检测细胞凋亡情况；蛋白免疫印迹法检测凋 亡相关蛋白及PTEN/mTOR通路相关蛋白表达情况。结果 与空白对照组比较，高糖组HTR-8/SVneo细胞增殖抑制 率、凋亡率及Bcl-2相关X蛋白（Bax）、活化的含半胱氨酸的天冬氨酸蛋白水解酶3（cleaved caspase-3）、PTEN蛋白表 达水平均显著升高（P＜0.05），B淋巴细胞瘤-2（Bcl-2）、磷酸化磷脂酰肌醇-3激酶（p-PI3K）/PI3K、磷酸化蛋白激酶B （p-AKT）/AKT、磷酸化 mTOR（p-mTOR）/mTOR 蛋白表达水平显著降低（P＜0.05）；与高糖组、NC 组比较，PTENsiRNA组HTR-8/SVneo细胞增殖抑制率、凋亡率及Bax、cleaved caspase-3、PTEN蛋白表达水平均显著降低，Bcl-2、pPI3K/PI3K、p-AKT/AKT、p-mTOR/mTOR 蛋白表达水平显著升高（P＜0.05）；与 PTEN-siRNA 组比较，GDC-0349 组 HTR-8/SVneo细胞增殖抑制率、凋亡率及Bax、cleaved caspase-3蛋白表达水平均显著升高，Bcl-2、p-PI3K/PI3K、pAKT/AKT、p-mTOR/mTOR 蛋白表达水平显著降低（P＜0.05）。结论 高糖可抑制人绒毛膜滋养层细胞 HTR-8/ SVneo增殖能力，诱导其凋亡，可能是通过上调PTEN表达、抑制PI3K/AKT/mTOR通路活化实现的。

Mechanism of PTEN/mTOR pathway in high glucose-induced apoptosis of human villous trophoblasts HTR-8/SVneo #br#

Objective To study the mechanism of high glucose-induced apoptosis of human villous trophoblasts HTR- 8/SVneo. Methods Human villous trophoblasts HTR-8/SVneo were cultured in vitro and divided into the blank control group, the high glucose group, the negative control group (NC), the phosphatase and tensin homology deleted on chromosome ten (PTEN) -siRNA group (inhibition of PTEN expression) and the mammalian target of rapamycin (mTOR) pathway inhibitor group (GDC-0349 group). Methyl thiazolyl tetrazolium (MTT) assay and flow cytometry were used to detect the proliferation of HTR-8/SVneo cells and the apoptosis of HTR-8/SVneo cells, respectively. The expression of apoptosis related proteins and proteins related to PTEN/mTOR pathway were detected by Western blot assay. Results Compared with the blank control group, HTR-8/SVneo cell proliferation inhibition rate, apoptosis rate and Bcl-2 related X protein (Bax), cleaved caspase-3 and PTEN protein expression levels were significantly increased in the high glucose group (P＜0.05), while the expression levels of B-cell lymphoma-2 (Bcl-2), phosphorylation phosphatidylinositol-3-kinase (p-PI3K)/PI3K, phosphorylation protein kinase B (p-AKT)/AKT and phosphorylation mTOR (p-mTOR)/mTOR protein were significantly decreased (P＜0.05). Compared with the high glucose group and the NC group, the HTR-8/SVneo cell proliferation inhibition rate, apoptosis rate, Bax, cleaved caspase-3 and PTEN protein expression levels were significantly decreased in the PTEN-siRNA group (P＜0.05), while the expression levels of Bcl-2, p-PI3K/PI3K, p-AKT/AKT and p-mTOR/mTOR protein were significantly increased (P＜0.05). Compared with the PTEN-siRNA group, the proliferation inhibition rate, apoptosis rate and the protein expression levels of Bax, cleaved caspase-3 were significantly increased in the GDC-0349 group (P＜0.05), while the expression levels of Bcl-2, p-PI3K/PI3K, p-AKT/AKT and p-mTOR/mTOR protein were significantly decreased (P＜0.05). Conclusion High glucose can inhibit the growth and induce apoptosis of human villous trophoblasts HTR-8/SVneo, which may be achieved by up-regulating the expression of PTEN and inhibiting the activation of PI3K/AKT/mTOR pathway.