脂质体介导Clusterin反义寡核苷酸抑制胰腺癌细胞侵袭力及增强5-FU化疗敏感性的实验研究  

Liposome-mediated clusterin antisense oligodeoxynucleotides inhibit invasion and chemosensitize 5-FU.in human pancreatic cancer cells

作　　者：迟绍云[1] 焦学龙[2] 王宝泉[1] 刘春生[2] 王蕾[2] 贾旭亮[2] 牛珉[2] 

机构地区：[1]山东省海阳市人民医院普外科,海阳265101 [2]青岛大学医学院附属医院普外科,青岛266003

出　　处：《临床与实验病理学杂志》2012年第2期173-176,180,共5页Chinese Journal of Clinical and Experimental Pathology

摘　　要：目的研究Clusterin反义寡聚脱氧核苷酸(Clusterin ASO)对胰腺癌细胞侵袭力及5-FU化疗敏感性的影响。方法采用脂质体法将Clusterin ASO转染PANC-1细胞24 h,然后将该细胞暴露于不同浓度的5-FU中,TUNEL法检测72 h后的细胞凋亡指数,MTT检测细胞增殖并计算IC50。观察Clusterin ASO转染对PANC-1细胞体外侵袭力的抑制作用。采用免疫组化,Western blot和RT-PCR技术检测不同处理组细胞中Clusterin和其mRNA和蛋白表达。结果 Clusterin ASO转染可有效沉默PANC-1细胞内Clusterin mRNA及蛋白表达;PANC-1细胞的5-FU IC50为9±0.7,PANC-1/Clusterin ASO的5-FU IC50为1.06±0.3,敏感性增加了9倍。5-FU以剂量依赖方式诱导PANC-1细胞凋亡,但对PANC-1/Clusterin ASO细胞所诱导的凋亡比PANC-1细胞更明显。重组细胞基膜侵袭实验显示,PANC-1细胞与PANC-1/Clusterin ASO细胞的穿透基膜细胞数分别为213±18个和43±8个/高倍镜(×200),差异有统计学意义(P<0.05)。结论靶向Clusterin抑制胰腺癌细胞侵袭力和增殖,并增强胰腺癌细胞对5-FU的化疗敏感性。Purpose To investigate whether clusterin gene silencing by antisense clusterin oligodeoxynucleotides（ASO） can inhibit invasion and metastasis and chemosensitize 5-FU in human pancreatic cancer cell line PNAC-1.Methods Clusterin ASO was transfected into PANC-1 cells for 24 h by Lipofectamine 2000 methods,and then the cells in different groups were treated with serial concentrations（0.01~100 μmol/L） of 5-FU for 72 h.The 50% inhibitory drug concentration（IC50） was obtained by MTT assay.The apoptosis was assessed by TUNEL staining.The inhibitory effects of clusterin ASO on invasion and metastasis were detected by Transwell motility assay.RT-PCR and Western blot were used to detect the clusterin protein and mRNA in PANC-1 cells in vitro.Results Clusterin expression was inhibited in PANC-1 cells in clusterin ASO groups.The inhibitory effect of cell growth was seen in PANC-1 and PANC-1/clusterin ASO cells treated with 5-FU.It could also promote the occurrence of cell apoptosis.5-FU inhibited the proliferation of PANC-1 cells in a concentration-dependent manner.In PANC-1/clusterin ASO cells treated with 5-FU,the apoptosis rate increased greatly and proliferation rate decreased as compared with the control cells treated with 5-FU.PANC-1/clusterin ASO cells presented lower invasion activity compared with those of control cells.Conclusion Clusterin gene silencing by clusterin ASO greatly inhibits the invasion and enhances the sensitivity of PANC-1B cells to 5-FU-induced apoptosis.

关 键 词：胰腺肿瘤 转移 化疗 基因治疗 CLUSTERIN 

分 类 号：R73-3[医药卫生—肿瘤] R735.9[医药卫生—临床医学]