尼古丁对舌鳞状细胞癌Cal27细胞的生物学影响

目的:探讨吸烟对舌鳞癌患者Cal27细胞系的影响。方法:将舌鳞癌Cal27细胞传代培养,将对数生长期细胞分为空白对照组、尼古丁组及7型烟碱乙酰胆碱受体(7nAChR)抑制组。空白对照组不作任何处理,尼古丁组用尼古丁连续处理细胞1周,7nAChR抑制组给予尼古丁及7nAChR抑制剂α-银环蛇毒素(-BTX)处理细胞1周。采用电子显微镜观察空白对照组及尼古丁组细胞形态;采用CCK-8试剂盒检测空白对照组及尼古丁组细胞增殖力;细胞划痕实验检测空白对照组及尼古丁组细胞迁移能力;Transwell小室检测空白对照组及尼古丁组细胞侵袭能力;免疫印迹实验测定尼古丁组、空白对照组及7nAChR抑制组细胞Wnt信号通路蛋白相对表达水平。采用SPSS 20.0软件包对数据进行统计学分析。结果:Cal27细胞形态为不规则多角形,体积较大,周围可见伪足伸展,在细胞培养基上呈铺路石样排列。空白对照组、尼古丁组及7nAChR抑制组Cal27细胞形态无显著差异。孵育96 h后,尼古丁组细胞数显著大于空白对照组及7nAChR抑制组(P<0.05);相同时间内尼古丁组细胞划痕愈合程度显著大于空白对照组与7nAChR抑制组(P<0.05);相同时间内,尼古丁组穿过小室细胞数显著大于空白对照组及7nAChR抑制组(P<0.05);尼古丁组细胞β-catenin、c-Myc、p-GSK3β及Ror2等Wnt通路蛋白相对表达水平显著高于空白对照组及7nAChR抑制组细胞(P<0.05)。结论:尼古丁可通过激活Wnt信号通路,提高舌鳞癌Cal27细胞增殖、迁移及侵袭能力。

Biological effect of nicotine on Cal27 cell line in patients with tongue squamous cell carcinoma

PURPOSE: To investigate the effect of nicotine on Cal27 cell line in patients with tongue squamous cell carcinoma. METHODS: Tongue squamous cell carcinoma Cal27 cells were subcultured, and the logarithmic growth phase cells were selected and divided into blank control group, nicotine-treated group and α7 nicotinic acetylcholine receptor inhibition-treated group (α7nAChR inhibition group). Cells in the blank control group received no treatment; cells in nicotine group received nicotine treatment, and cells in α7nAChR inhibition group were treated nicotine combined with α-bungarotoxin (α-BTX). The treatments lasted for 1 week. Cell morphology of blank control group and nicotine group was observed under electron microscope, cell proliferation was detected by CCK-8 kit, cell migration ability was detected by scratch test, cell invasion ability of blank control group and nicotine group was detected by Transwell chamber. The relative expression levels of Wnt signaling pathway proteins in nicotine group, blank control group and α7nAChR inhibition group were determined by Western blot. The data were statistically analyzed using SPSS 20.0 software package. RESULTS: Microscope showed that the rapidly adherent cells were uniformed polygon shape with large nucleus, and there was no significant difference among 3 groups. After 96 hours of incubation, the number of cells in nicotine group was significantly higher than that in the blank control group and α7nAChR inhibition group (P<0.05). The degree of scratch healing in nicotine group was significantly higher than that in the blank control group and α7nAChR inhibition group (P<0.05). The number of cells passing through the chamber in nicotine group was significantly higher than that of the blank control group andα7nAChR inhibition group (P<0.05). The relative expression levels of Wnt pathway proteins including β-catenin, c-Myc, p-GSK3β and Ror2 were significantly higher in nicotine group than those in the blank control group andα7nAChR inhibition group (P<0.05). CONCLUSIONS: Nicotine can enhance proliferation, migration and invasion of tongue squamous cell carcinoma Cal27 cells by activating Wnt signaling pathway.