【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(q RT-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m RNA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Hu CCT-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m RNA和蛋白表达水平明显高于对应癌旁组织。经q RT-PCR和Western blot方法证实稳定转染PKM2 sh RNA的胆管癌细胞中PKM2的m RNA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-sh RNA)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 sh RNA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Hu CCT-1、HCCC-9810细胞的迁移、侵袭及增殖。更多还原

【Abstract】 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 q RT-PCR and immunohistochemistry. PKM2 was down-regulated by a lentiviral vector expression system in cholangiocarcinoma cell lines Hu CCT-1 and HCCC-9810. q RT-PCR and Western blot were performed to analyze the m RNA 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 m RNA and protein expressions of PKM2 in the experimental group(PKM2-sh RNA)were significantly lower than those in the two control groups,confirmed by q RT-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-sh RNA)(P < 0.05). Conclusion:Down-regulation of PKM2 by PKM2 sh RNA can inhibit migration,invasion and proliferation of Hu CCT-1 and HCCC-9810 cells.更多还原