文章摘要
柴 浩,熊新魁,孙道一,单文刚,浦立勇,俞 悦,成 峰.PKM2基因对胆管细胞癌迁移、侵袭及增殖的影响[J].南京医科大学学报,2015,(5):615~621
PKM2基因对胆管细胞癌迁移、侵袭及增殖的影响
Au experimental research on PKM2 gene on migration, invasion and proliferation of cholangiocarcinoma cell line
投稿时间:2015-01-13  
DOI:10.7655/NYDXBNS20150504
中文关键词: shRNA干扰  PKM2基因  生物学功能
英文关键词: shRNA interference  PKM2 gene  biological function
基金项目:国家自然科学基金面上项目(81070324);江苏省卫生厅重点项目(H201102);卫生厅开放课题(ZX05200906);江苏省六大人才高峰项目(2009)
作者单位
柴 浩 南京医科大学第一附属医院肝脏移植中心,江苏 南京 210029 
熊新魁 南京医科大学第一附属医院肝脏移植中心,江苏 南京 210029 
孙道一 南京医科大学第一附属医院肝脏移植中心,江苏 南京 210029 
单文刚 南京医科大学第一附属医院肝脏移植中心,江苏 南京 210029 
浦立勇 南京医科大学第一附属医院肝脏移植中心,江苏 南京 210029 
俞 悦 南京医科大学第一附属医院肝脏移植中心,江苏 南京 210029 
成 峰 南京医科大学第一附属医院肝脏移植中心,江苏 南京 210029 
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中文摘要:
      目的:检测M2-型丙酮酸激酶(pyruvate kinase M2,PKM2)在胆管癌(cholangiocarcinoma,CCA)组织中的表达情况,探讨PKM2下调对胆管癌细胞迁移?侵袭及增殖的影响。方法:实时定量逆转录聚合酶链反应(qRT-PCR)和免疫组织化学(immunohistochemistry,IHC)染色分别检测胆管癌及对应癌旁组织标本中PKM2的mRNA和蛋白表达水平。利用慢病毒表达载体系统在胆管癌细胞株HuCCT-1?HCCC-9810中下调PKM2,分别用划痕实验?Transwell细胞侵袭实验和CCK-8比色法检测细胞迁移?侵袭及增殖能力。结果:胆管癌组织中PKM2的mRNA和蛋白表达水平明显高于对应癌旁组织。经qRT-PCR和Western blot方法证实稳定转染PKM2 shRNA的胆管癌细胞中PKM2的mRNA和蛋白水平较对照组均明显下降(P < 0.05),与空载对照组(PKM2-NC)和正常对照组(PKM2)相比,实验组细胞(PKM2-shRNA)的迁移?侵袭及增殖能力明显减弱,差异有统计学意义(P < 0.05)。结论:胆管癌组织中PKM2表达明显高于癌旁组织,PKM2 shRNA能有效地降低胆管癌细胞中PKM2基因的表达,PKM2基因沉默可以抑制HuCCT-1?HCCC-9810细胞的迁移?侵袭及增殖。
英文摘要:
      Objective:To investigate the expression level of PKM2 in cholangiocarcinoma (CCA)tissues,then study the effect of PKM2 down-regulation on migration,invasion and proliferation in cholangiocarcinoma cell lines. Methods:RNA and protein expressions of PKM2 in CCA tissues and paired adjacent tissues were detected by qRT-PCR and immunohistochemistry. PKM2 was down-regulated by a lentiviral vector expression system in cholangiocarcinoma cell lines HuCCT-1 and HCCC-9810. qRT-PCR and Western blot were performed to analyze the mRNA and protein expression of PKM2 in both cell lines. Cell migration, invasion and proliferation were assessed by wound-healing experiment, matrigel invasion and Cell Counting Kit-8(CCK-8). Results:The expression of PKM2 in CCA tissues had a higher level than that in paired adjacent tissues. The mRNA and protein expressions of PKM2 in the experimental group (PKM2-shRNA)were significantly lower than those in the two control groups,confirmed by qRT-PCR and Westen blot (P < 0.05). Compared to the empty vector group (PKM2-NC)and the normal control group (PKM2),the cell invasion,migration and proliferation were significantly decreased in the experimental group (PKM2-shRNA)(P < 0.05). Conclusion:Down-regulation of PKM2 by PKM2 shRNA can inhibit migration,invasion and proliferation of HuCCT-1 and HCCC-9810 cells.
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