摘要
目的探究PTEN通过磷酸肌醇-3激酶/蛋白激酶B/哺乳动物雷帕霉素靶蛋白(PI3K/Akt/mTOR)通路介导的自噬调控宫颈癌细胞顺铂耐药的机制。方法2024年3月通过大剂量冲击法诱导Hela细胞顺铂耐药。采用转染沉默PTEN,利用蛋白质印迹检测其对PI3K/Akt/mTOR通路和自噬的影响。将Hela细胞分为si-NC组、si-PTEN组、si-NC+3-甲基腺嘌呤(3-MA)组和si-PTEN+3-MA组。3-MA处理转染si-NC、si-PTEN的Hela细胞以抑制自噬,随后用顺铂处理,通过CCK-8法、克隆形成实验分析顺铂对各组细胞活力和增殖的影响。在动物实验中将裸鼠组1分为OE-NC组、OE-PTEN组、OE-NC+顺铂组和OE-PTEN+顺铂组,分别利用转染OE-NC、OE-PTEN的顺铂耐药的Hela细胞构建荷瘤裸鼠模型,然后用顺铂干预裸鼠。裸鼠组2分为si-NC组、si-NC+顺铂组、si-PTEN+顺铂组和si-PTEN+3-MA+顺铂组,分别利用转染si-NC、si-PTEN的Hela细胞构建荷瘤裸鼠模型,通过腹腔注射3-MA抑制自噬,再利用顺铂干预,分别检测肿瘤生长情况。结果与非耐药Hela细胞比较,顺铂耐药的Hela细胞PI3K、Akt、mTOR蛋白磷酸化水平,以及自噬水平均明显升高,差异均有统计学意义(P<0.05)。与顺铂处理的si-NC组比较,si-PTEN组Hela细胞活力、增殖能力均明显提高,si-NC+3-MA组细胞活力、增殖能力均进一步降低,且si-PTEN+3-MA组细胞活力、增殖能力均明显低于si-PTEN组,差异均有统计学意义(P<0.05);过表达PTEN明显提高了耐药肿瘤细胞对顺铂的敏感性,此外沉默PTEN减弱顺铂对宫颈癌肿瘤模型的抑制作用,而抑制自噬则逆转了PTEN沉默对顺铂耐药的诱导,差异均有统计学意义(P<0.05)。结论PTEN的降低通过激活PI3K/Akt/mTOR通路诱导自噬促进宫颈癌细胞顺铂耐药。
Objective To explore the mechanism by which PTEN regulates autophagy to mediate cisplatin resistance in cervical cancer cells through the phosphoinositide-3 kinase/protein kinase B/mammalian rapamycin target protein(PI3K/Akt/mTOR)pathway.Methods In March 2024,Hela cells were induced to develop cisplatin resistance using a high-dose shock method.PTEN was silenced by transfection,and the effects on the PI3K/Akt/mTOR pathway and autophagy were detected using Western blotting.Hela cells were divided into si-NC group,si-PTEN group,si-NC+3-MA group,and si-PTEN+3-MA group.3-MA was used to inhibit autophagy in Hela cells transfected with si-NC and si-PTEN,and then treated with cisplatin.The effects of cisplatin on the viability and proliferation of each group of cells were analyzed using CCK-8 assay and clone formation experiment.In the animal experiment,the nude mouse group 1 was divided into OE-NC group,OE-PTEN group,OE-NC+cisplatin group,and OE-PTEN+cisplatin group.Nude mice models with tumor formation were constructed using cisplatin-resistant Hela cells transfected with OE-NC and OE-PTEN,and then treated with cisplatin.The nude mouse group 2 was divided into si-NC group,si-NC+cisplatin group,si-PTEN+cisplatin group,and si-PTEN+3-MA+cisplatin group.Nude mice models with tumor formation were constructed using Hela cells transfected with si-NC and si-PTEN,and 3-MA was used to inhibit autophagy before cisplatin intervention.The tumor growth was detected in each group.Results Compared with non-resistant Hela cells,the phosphorylation levels of PI3K,Akt,and mTOR proteins and the autophagy level in cisplatin-resistant Hela cells were significantly increased,and the differences were statistically significant(P<0.05).Compared with the si-NC group treated with cisplatin,the viability and proliferation ability of Hela cells in the si-PTEN group were significantly increased,while the viability and proliferation ability of the si-NC+3-MA group were further decreased,and the viability and proliferation ability of the si-PTEN+3-MA group were significantly lower than those of the si-PTEN group,and the differences were statistically significant(P<0.05).Overexpression of PTEN significantly increased the sensitivity of resistant tumor cells to cisplatin.In addition,silencing PTEN weakened the inhibitory effect of cisplatin on cervical cancer tumor models,and inhibiting autophagy reversed the induction of cisplatin resistance by PTEN silencing.The differences were statistically significant(P<0.05).Conclusion The reduction of PTEN induces autophagy through activating the PI3K/Akt/mTOR pathway,promoting cisplatin resistance in cervical cancer cells.
作者
杜颖
高天
赵纯全
张玉林
DU Ying;GAO Tian;ZHAO Chunquan;ZHANG Yulin(Department of Gynecology,the First Affiliated Hospital of Chongqing Medical University,Chongqing 400016,China)
出处
《现代医药卫生》
2026年第2期253-259,共7页
Journal of Modern Medicine & Health
基金
重庆市教育委员会科学技术研究项目(KJQN202200444)。
关键词
宫颈癌
顺铂
耐药
自噬
PTEN
Cervical cancer
Cisplatin
Drug resistance
Autophagy
PTEN
