国际肿瘤学杂志

国际肿瘤学杂志 ›› 2020, Vol. 47 ›› Issue (2): 65-69.doi: 10.3760/cma.j.issn.1673-422X.2020.02.001

• 论著 •    下一篇

GOLPH3通过PI3K/AKT/GSK3β信号调控子宫内膜癌细胞的增殖与凋亡

罗春翠1, 袁超燕1, 陈庆芬2()   

  1. 1 湖北民族大学附属民大医院妇科,恩施 445000
    2 陕西省第四人民医院妇产科,西安 710043
  • 收稿日期:2019-09-09 修回日期:2019-11-30 出版日期:2020-02-08 发布日期:2020-05-27
  • 通讯作者: 陈庆芬 E-mail:cqf369369@126.com

GOLPH3 regulates proliferation and apoptosis of endometrial carcinoma cells through PI3K/AKT/GSK3β signal

Luo Chuncui1, Yuan Chaoyan1, Chen Qingfen2()   

  1. 1 Department of Gynecology, Minda Hospital Affiliated to Hubei Minzu University, Enshi 445000, China
    2 Obstetrics and Gynecology of the Fourth People's Hospital of Shaanxi Province, Xi'an 710043, China
  • Received:2019-09-09 Revised:2019-11-30 Online:2020-02-08 Published:2020-05-27
  • Contact: Chen Qingfen E-mail:cqf369369@126.com

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摘要：

目的 探究高尔基体磷蛋白3(GOLPH3)通过PI3K/AKT/GSK3β信号调控子宫内膜癌(EC)细胞增殖与凋亡的机制。方法 将人子宫内膜腺癌HEC-1-B细胞分为对照组、GOLPH3组和GOLPH3+GDC-0941组。通过转染GOLPH3 pcDNA质粒过表达GOLPH3,PI3K抑制剂GDC-0941用于阻断PI3K/AKT/GSK3β通路。分别通过CCK-8法、克隆形成实验和流式细胞术检测细胞活力、细胞增殖能力和细胞凋亡。通过Western blotting检测PI3K/AKT/GSK3β通路中蛋白磷酸化水平。结果 转染实验成功,且PI3K抑制剂GDC-0941不会影响GOLPH3 mRNA和蛋白的表达。对照组、GOLPH3组、GOLPH3+GDC-0941组的相对细胞活力分别为(100.00±4.63)%、(131.56±7.85)%、(97.85±7.36)%,3组间差异有统计学意义(F=7.437,P<0.001);GOLPH3组高于对照组(P<0.001),GOLPH3+GDC-0941组低于GOLPH3组(P<0.001)。3组的细胞克隆形成数目分别为(46.86±3.56)个、(89.32±4.78)个、(46.48±4.05)个,差异有统计学意义(F=20.437,P<0.001);GOLPH3组多于对照组(P<0.001),GOLPH3+GDC-0941组少于GOLPH3组(P<0.001)。3组的细胞凋亡率分别为(5.17±0.61)%、(2.34±0.56)%、(6.85±0.53)%,差异有统计学意义(F=11.643,P<0.001);GOLPH3组低于对照组(P<0.001),GOLPH3+GDC-0941组高于GOLPH3组(P<0.001)。3组磷酸化PI3K/PI3K水平分别为1.00±0.09、3.32±0.19、0.93±0.06,磷酸化AKT/AKT水平分别为1.00±0.11、4.63±0.63、1.15±0.16,磷酸化GSK3β/GSK3β水平分别为1.00±0.08、4.06±0.57、1.04±0.14,差异均有统计学意义(F=12.532,P<0.001;F=16.792,P<0.001;F=15.311,P<0.001);各蛋白磷酸化水平GOLPH3组均高于对照组(均P<0.001),GOLPH3+GDC-0941组均低于GOLPH3组(均P<0.001)。结论 在EC细胞中,过表达GOLPH3可通过激活PI3K/AKT/GSK3β通路促进细胞增殖并抑制细胞凋亡,提示GOLPH3参与EC的发生和发展。

关键词: 子宫内膜肿瘤, 细胞增殖, 细胞凋亡, 高尔基体磷蛋白3, PI3K/AKT

Abstract:

Objective To explore the mechanism of Golgi phosphoprotein 3 (GOLPH3) regulating the proliferation and apoptosis of endometrial cancer (EC) cells through PI3K/AKT/GSK3β signal. Methods Human endometrial adenocarcinoma HEC-1-B cells were divided into control group, GOLPH3 group and GOLPH3 + GDC-0941 group. GOLPH3 was over-expressed by transfection of the GOLPH3 pcDNA. The PI3K inhibitor GDC-0941 was used to block the PI3K/AKT/GSK3β pathway. Cell viability, cell proliferation and apoptosis were measured by CCK-8 method, clone formation experiment and flow cytometry, respectively. The protein phosphorylation level in PI3K/AKT/GSK3β pathway was detected by Western blotting. Results Transfection experiments were successful, and the PI3K inhibitor GDC-0941 did not affect GOLPH3 mRNA and protein expression. The relative cell viability of the control group, the GOLPH3 group and the GOLPH3 + GDC-0941 group were (100.00±4.63)%, (131.56±7.85)% and (97.85±7.36)%, and the difference among the three groups were significant (F=7.437, P<0.001). The cell viability of the GOLPH3 group was higher than that of the control group (P<0.001). The cell viability of the GOLPH3 + GDC-0941 group was lower than that of the GOLPH3 group (P<0.001). The numbers of cell clones in the three groups were 46.86±3.56, 89.32±4.78 and 46.48±4.05, and the difference was significant (F=20.437, P<0.001). GOLPH3 group had more clones than control group (P<0.001). The number of clones formed in the GOLPH3 + GDC-0941 group was less than that in the GOLPH3 group (P<0.001). The apoptosis rates of the three groups were (5.17±0.61)%, (2.34±0.56)% and (6.85±0.53)%, and the difference was significant (F=11.643, P<0.001). The apoptosis rate of the GOLPH3 group was lower than that of the control group (P<0.001), and the apoptosis rate of the GOLPH3 + GDC-0941 group was higher than that of the GOLPH3 group (P<0.001). The phosphorylated PI3K/PI3K levels of the three groups were 1.00±0.09, 3.32±0.19 and 0.93±0.06, respectively; phosphorylated AKT/AKT levels were 1.00±0.11, 4.63±0.63 and 1.15±0.16, respectively; phosphorylated GSK3β/GSK3β levels were 1.00±0.08, 4.06±0.57 and 1.04±0.14, respectively. The differences were statistically significant (F=12.532, P<0.001; F=16.792, P<0.001; F=15.311, P<0.001). The phosphorylation levels of each protein in the GOLPH3 group were higher than those in the control group (all P<0.001), and the GOLPH3 + GDC-0941 group were lower than the GOLPH3 group (all P<0.001). Conclusion In EC cells, over-expression of GOLPH3 can promote cell proliferation and inhibit apoptosis by activating the PI3K/AKT/GSK3β pathway, suggesting that GOLPH3 is involved in the occurrence and development of EC.

Key words: Endometrial neoplasms, Cell proliferation, Apoptosis, Golgi phosphoprotein 3, PI3K/AKT