M3受体亚型在NSCLC细胞增殖、侵袭和迁移中的作用及机制研究  

Studies on proliferation and migration effects and mechanisms of M3 mAChR in NSCLC cells

作　　者：王士奇[1] 魏晓莉[1] 闫海涛[1] WANG Shi qi;WEI Xiao li;YAN Hai tao(Institute of Pharmacology and Toxicology, Academy of Military Medical Science, Beijing100850, China)

机构地区：[1]军事医学科学院毒物药物研究所,北京100850

出　　处：《中国药理学通报》2017年第12期1661-1668,共8页Chinese Pharmacological Bulletin

基　　金：国家自然科学基金资助项目(No 81472187)

摘　　要：目的探讨M3R在NSCLC细胞增殖、侵袭和迁移中的作用及信号机制。方法 CCK-8法、划痕法和Transwell细胞侵袭实验检测M3R拮抗剂对NSCLC细胞增殖、迁移和侵袭的影响;钙成像观察M3R拮抗剂对卡巴胆碱诱导的内钙增加的影响;Western blot验证M3R拮抗剂对细胞增殖、迁移和细胞周期相关信号分子蛋白表达的影响。结果M3R拮抗剂1.25~80μmol·L^(-1)作用48 h后,能明显抑制NSCLC细胞增殖,抑制效果为R2-8018>达非那新(Darifenacin),H1299>H460;p-Akt、p-GSK3β、cyclin D1蛋白表达水平均随M3R拮抗剂作用时间延长明显下降,p21蛋白表达水平明显上调;R2-8018处理后,H1299细胞迁移和侵袭能力下降,MMP-2蛋白表达水平明显下降。结论拮抗M3R抑制Akt/GSK3β信号通路,抑制cyclin D1活性并上调p21,阻滞细胞周期进程,进而抑制细胞增殖;并通过下调MMP-2抑制H1299细胞的迁移和侵袭。AimTo investigate the effect of M3mAChR on the proliferation and migration of NSCLC and the molecular mechanisms.MethodsCCK8assay,Wound healing assay and Transwell invasion assaywere used to determine the cell proliferation,migration and invasion.Calcium imaging was used to identify the M3mAChR subtype mediating carbachol induced increase in intracellular calcium.Western blot was used to determine the protein level of proliferation,migration and cell cycle related signaling molecules.ResultsFollowing the treatment with M3R antagonists125~80μmol·L-1for48h,the proliferation of NSCLC cells was inhibited,the inhibitory effect:R28018>Darifenacin,H1299>H460.After the treatment with R28018,the migration and invasion of H1299significantly declined.Western blot showed that the protein level of p Akt,p GSK3β,cyclinD1declined significantly with the increase of time and that the protein level of p21increased.ConclusionM3R antagonists induce cell cycle arrest by suppressing the activation of Akt,down regulating GSK3βand cyclinD1,and up regulating p21,then inhibiting the proliferation of NSCLC cells.It also inhibits the invasion and migration by down regulating MMP2.

关 键 词：非小细胞肺癌 M3受体 增殖 侵袭 迁移 基质金属蛋白酶2 

分 类 号：R329.24[医药卫生—人体解剖和组织胚胎学] R348.1[医药卫生—基础医学]