摘要
目的探讨衰老关键蛋白3(fibulin 3,FBLN3)在人结直肠癌SW480细胞增殖及转移中的作用。方法人结直肠癌SW480细胞株分为空白组(培养液2mL)、空载组(FBLN3阴性慢病毒+培养液2mL)和FBLN3过表达组(FBLN3过表达慢病毒+培养液2 mL);采用MTT法观察FBLN3对SW480细胞增殖的影响;流式细胞技术检测FBLN3对SW480细胞周期的影响;Transwell实验观察FBLN3对SW480细胞侵袭、转移的影响;Western blot法检测FBLN3对相关蛋白表达的影响。结果 FBLN3过表达组转染后24h(0.386±0.008)、48h(0.535±0.011)、72h(0.467±0.012)吸光度值均低于空白组(0.460±0.011、0.787±0.009、1.031±0.005)和空载组(0.445±0.009、0.750±0.004、0.949±0.014)(P<0.05),空白组与空载组比较差异无统计学意义(P>0.05);转染48h后FBLN3过表达组G1期细胞比率[(65.02±1.05)%]高于空白组[(47.01±0.90)%]和空载组[(48.25±1.41)%],S期细胞比例[(15.41±1.70)%]低于空白组[(34.90±1.28)%]和空载组[(28.82±2.40)%](P<0.05);转染48h后FBLN3过表达组侵袭细胞计数(47±5)和迁移细胞计数(64±10)均低于空白组(106±7、144±9)及空载组(105±12、145±6)(P<0.05),空白组与空载组比较差异无统计学意义(P>0.05);FBLN3过表达组p-AKT、p-哺乳动物雷帕霉素靶蛋白(mammalian target of rapamycin,mTOR)、基质金属蛋白酶(matrix metalloproteinase,MMP)-2、MMP-9、Bcl-2及细胞周期蛋白D1(cyclinD1)蛋白表达水平均低于空白组和空载组,凋亡诱导分子Bcl-2相关X蛋白(Bcl-2associated X protein,Bax)和细胞周期蛋白依赖性激酶抑制剂P21蛋白表达均高于空白组和空载组(P<0.05)。结论 FBLN3过表达可抑制AKT/mTOR信号通路活化,抑制侵袭转移因子MMP-2、-9,凋亡抑制因子Bcl-2及细胞周期蛋白cyclinD1表达,促进凋亡诱导分子Bax和细胞周期蛋白依赖性激酶抑制剂P21蛋白的表达,导致肿瘤细胞周期G_1期阻滞,抑制SW480细胞的增殖、侵袭及转移,从而抑制结直肠癌发生、发展。
Objective To investigate the effects of fibulin 3(FBLN3)on the proliferation and metastasis of SW480 human colorectal cancer cells.Methods SW480 cell lines were divided into blank group(2mL culture medium),no-load group(FBLN3negative lentivirus+2 mL culture medium)and FBLN3 overexpression group(FBLN3overexpression lentivirus+2 mL culture medium).The effect of FBLN3 on the proliferation of SW480 cells was observed by MTT assay,and flow cytometry was used to detect the effect of FBLN3 on the SW480 cell cycle.Transwell experiment was used to observe the influence of overexpression of FBLN3 on the invasion and metastasis of SW480 cells.The influence of overexpression of FBLN3 on the related proteins was detected by Western blot.Results The optical density values in 24,48 and 72hafter transfection were significantly lower in FBLN3 overexpression group(0.386±0.008,0.535±0.011,0.467±0.012)than those in blank group(0.460±0.011,0.787±0.009,1.031±0.005)and no-load group(0.445±0.009,0.750±0.004,0.949±0.014)(P<0.05),and there were no significant differences between blank group and no-load group(P>0.05).The percentage of G1 phase cells in 48 hafter transfection was significantly higher in FBLN3 overexpression group((65.02±1.05)%)than that in blank group((47.01±0.90)%)and no-load group((48.25±1.41)%),the percentage of S phase cells was significantly lower in FBLN3 overexpression group((15.41±1.70)%)than that in blank group((34.90±1.28)%)and no-load group((28.82±2.40)%)(P<0.05).The number of invasive cells and migratory cell count were significantly lower in FBLN3 overexpression group(47±5,64±10)than those in blank group(106±7,144±9)and no-load group(105±12,145±6)in 48 hafter transfection(P<0.05),and there were no significant differences between blank group and no-load group(P>0.05).The expressions of p-AKT,p-mammalian target of rapamycin(p-mTOR),matrix metalloproteinase(MMP)-2,MMP-9,Bcl-2and cyclin D1 protein in 48 hafter transfection were significantly lower in FBLN3 overexpression group than those in bla
出处
《中华实用诊断与治疗杂志》
2017年第5期435-439,共5页
Journal of Chinese Practical Diagnosis and Therapy
基金
扬州市科技局项目(YZ2011083)
