中国麻风皮肤病杂志 ›› 2023, Vol. 39 ›› Issue (7): 479-484.doi: 10.12144/zgmfskin202307479

• 论著 • 上一篇    下一篇

EGFR shRNA联合雷帕霉素对Colo-16细胞增殖、迁移及侵袭的影响

王慧1,杨艳1,李竹茜1,展晓红2,魏志平3   

  1. 1青岛市城阳区人民医院皮肤科，青岛，266109；2青岛大学附属医院病理科，青岛，266109；3徐州医科大学附属医院皮肤科，徐州，221002
  • 出版日期:2023-07-15 发布日期:2023-07-05

Effects of EGFR shRNA combined with rapamycin on proliferation, migration and invasion of Colo-16 cells

WANG Hui1, YANG Yan1, LI Zhuqian1, ZHAN Xiaohong2, WEI Zhiping3   

  1. 1 Department of Dermatology, Chengyang District People's Hospital, Qingdao 266109,China;2 Department of Pathology, Affiliated Hospital of Qingdao University, Qingdao 266109,China;3 Department of Dermatology, Affiliated Hospital of Xuzhou Medical University, Xuzhou 221002,China
  • Online:2023-07-15 Published:2023-07-05

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摘要： 目的：明确表皮生长因子受体（EGFR）shRNA联合雷帕霉素（RAPA）对皮肤鳞癌Colo-16细胞增殖、迁移和侵袭的影响，并对其可能机制进行初步探究。方法：Colo-16细胞分为5组：正常细胞组（Colo-16细胞+磷酸盐缓冲液PBS）、阴性对照组（转染shRNA-NC质粒的Colo-16细胞+PBS）、EGFR shRNA组（转染EGFR shRNA质粒的Colo-16细胞+PBS）、RAPA组（Colo-16细胞+RAPA）、联合组（转染EGFR shRNA质粒的Colo-16细胞＋RAPA）。采用细胞计数试剂盒(CCK-8)和克隆形成实验测定细胞增殖能力；划痕实验和Transwell法检测细胞迁移和侵袭情况，采用Western blot、RT-PCR检测增殖、侵袭相关基因Ki-67、MMP-2、MMP-9蛋白和mRNA水平表达。多组间比较采用单因素方差分析，组间两两多重比较采用SNK-q检验。结果：EGFR shRNA组、RAPA组及联合组Colo-16细胞吸光度A值、细胞克隆形成率、划痕愈合率、侵袭细胞数目、Ki-67、MMP-2、MM-P9等基因蛋白和mRNA表达显著低于正常细胞组与阴性对照组，差异有统计学意义（均P<0.05），且联合的效果更加明显。正常细胞组与阴性对照组差异均无统计学意义（均P>0.05）。结论：EGFR shRNA联合雷帕霉素在抑制Colo-16细胞增殖、迁移及侵袭方面有协同增效的作用，其机制可能与EGFR/PI3K/AKT/mTOR信号通路的双重抑制作用相关。

关键词: 皮肤鳞状细胞癌, 表皮生长因子, RNA干扰, 雷帕霉素, 细胞增殖, 细胞迁移, 细胞侵袭

Abstract: Objective: To investigate the effect of epidermal growth factor receptor (EGFR) shRNA combined with rapamycin (RAPA) on the proliferation, migration and invasion of skin squamous cell carcinoma Colo-16 cells, and to explore its possible mechanism. Methods: Colo-16 cells were divided into five groups, normal cell group (Colo-16+phosphate buffer saline PBS), negative control group (Colo-16 cells+PBS transfected with shRNA-NC plasmid), EGFR shRNA group (Color-16 cells+PBS transfected with EGFR shRNA plasmid), RAPA group (conventionally cultured Colo-16 cells+RAPA), and combination group (Colo-16 cells+RAPA transfected with EGFR shRNA plasmid). Cell proliferation ability was measured by cell counting kit (CCK-8) and clone formation test. Scratch test and Transwell method were used to detect cell migration and invasion. The expression of Ki-67, MMP-2, MMP-9 protein and mRNA related to proliferation and invasion were detected by Western blot and RT-PCR . Single factor analysis of variance was used for comparison among multiple groups, and SNK-q test was used for multiple comparison between groups. Results: Colo-16 cell absorbance A value, cell clone formation rate, scratch healing rate, number of invasive cells, Ki-67, MMP-2, MM-P9 gene protein and mRNA expression in EGFR shRNA group, RAPA group and combination group were significantly lower than those in normal cell group and negative control group, with significant differences (Ps<0.05), and the combination effect was more obvious. There was no statistically significant difference between normal cell group and negative control group (Ps>0.05). Conclusion: EGFR shRNA combined with rapamycin has synergistic effect in inhibiting the proliferation, migration and invasion of Colo-16 cells, and its mechanism may be related to the double inhibition of EGFR/PI3K/AKT/mTOR signal pathway.

Key words: skin squamous cell carcinoma, epidermal growth factor, RNA interference, rapamycin, cell proliferation, cell migration, cell invasion