摘要
目的观察ARHI基因对胰腺癌细胞增殖的影响,探讨其抑制细胞增殖的机制。方法 2009年3-9月人胰腺癌细胞株PANC-1选自北京协和医院,根据转入质粒将培养细胞分为Control组(无转染质粒的PANC-1细胞)、Vector组(稳定表达pIRES2-EGFP质粒的PANC-1细胞)、ARHI组(稳定表达pIRES2-EGFP-ARHI质粒的PANC-1细胞)。通过四甲基偶氮唑盐(MTT)法分析3组胰腺癌细胞在培养第0、1、2、3、4、5天增殖情况;应用流式细胞仪检测3组胰腺癌细胞周期;通过Western blotting法检测3组ARHI、磷酸化丝氨酸蛋白激酶(p-AKT)、MDM2、p53、p21^(WAF1)、丝氨酸蛋白激酶(AKT)表达;应用碘化丙啶(PI)-3K/AKT抑制剂LY294002干预ARHI组,检测ARHI、p-AKT、MDM2、p53、p21^(WAF1)、AKT表达。结果 3组培养第1、2、3、4、5天细胞增殖情况比较,差异均有统计学意义(P<0.05)。Vector组培养第4、5天细胞增殖情况与Control组比较,差异有统计学意义(P<0.05);ARHI组培养第1、2、3、4、5天细胞增殖情况与Control组和Vector组比较,差异均有统计学意义(P<0.05)。3组细胞周期比较,差异均有统计学意义(P<0.05)。Vector组G_0-G_1期、S期细胞所占比例与Control组比较,差异有统计学意义(P<0.05);ARHI组G_0-G_1期、S期细胞所占比例与Control组和Vector组比较,差异有统计学意义(P<0.05);ARHI组G_2-M期细胞所占比例与Control组比较,差异有统计学意义(P<0.05)。与Control组和Vector组相比,ARHI组p-AKT、MDM2表达降低,p53、p21^(WAF1)表达增多,而AKT表达没有变化;ARHI组加入LY294002与未加入LY294002相比,p-AKT、MDM2表达降低,p53、p21^(WAF1)表达增加,而ARHI、AKT表达没有变化。结论 ARHI基因通过抑制PI-3K/AKT/MDM2,导致p53、p21^(WAF1)表达上调,在胰腺癌发生中起着重要的作用。
Objective To investigate impact of ARHI gene on the proliferation of pancreatic cancer cells and the mechanism of ARHI gene in inhibiting cell proliferation.Methods From March to September in 2009,the study selected human pancreatic cancer cell lines from Peking Union Medical College Hospital,and divided them into three groups：Control group（PANC-1 cells without plasmid transfection）,Vector group（PANC- 1 cells with stable expression of pIRES2- EGFP plasmid）,ARHI group（PANC- 1 cells with stable expression of pIRES2- EGFP- ARHI plasmid）.Using methyl thiazolyl tetrazolium（MTT） method,the proliferation of these pancreatic cancer cells on day 0,day 1,day 2,day 3,day 4 and day 5were analyzed;flow cytometry was applied to detect cell cycle;using western blotting method,the expression levels of ARHI,p-AKT,MDM2,p53,p21^（WAF1） and AKT of the three groups were detected;PI-3K/AKT inhibitor LY294002 was used to conduct intervention on ARHI group,and the expression levels of ARHI,p- AKT,MDM2,p53,p21^（WAF1） and AKT were detected.Results The three groups were significantly different in cell proliferation on day 1,day 2,day 3,day 4 and day 5（P0.05）.Vector group and Control group were significantly different in cell proliferation on day 4 and day 5（P〈0.05）;ARHI group was significantly different from Control group and Vector group in the cell proliferation on day 1,day 2,day 3,day 4 and day 5（P0.05）.The three groups were significantly different in cell cycle（P〈0.05）.Vector group and Control group were significantly different in the cell proportion in stage G_0- G_1 and stage S（P〈0.05）;ARHI group was significantly different from Control group and Vector group in stage G_0- G_1 and stage S（P〈0.05）;ARHI group and Control group were significantly different in cell proportion in stage G_2- M（P〈0.05）.Compared with Control group and Vector group,the expression levels of p- AKT and MDM2 decreased and the expression levels of p53 and p21^（WAF1） increased,while the expression
出处
《中国全科医学》
CAS
CSCD
北大核心
2015年第36期4455-4458,共4页
Chinese General Practice
基金
国家自然科学基金资助项目(30670963)
河北省科技支撑计划项目(13277744D
14277735D)
