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目的探讨缺氧诱导因子1α(HIF-1α)在微环境诱导的肝癌多药耐药(MDR)表型形成过程中的作用和地位,并为预防和逆转肝癌耐药提供有效的新的分子靶点。方法运用免疫细胞化学技术分别检测在缺氧、低糖环境下生长或稳定整合HBX基因的HepG2细胞内HIF-1α蛋白的表达及定位。应用荧光定量PCR技术和Western-blot技术检测上述不同条件下HepG2细胞内HIF-1α在mRNA和蛋白水平的表达。转染 pcDNA3/HIF-1α质粒于HepG2细胞,检测转染细胞中多药耐药相关基因mdr1、LRP、MRP1的表达。结果在缺氧、低糖环境下生长或稳定整合了HBX基因的HepG2细胞中HIF-1α的mRNA和蛋白均不同程度地高表达,并发生了核转位。pcDNA3/HIF-1α质粒转染HepG2细胞中多药耐药相关基因mdr1、MRP1、LRP的mRNA水平较未转染组分别增加了2．4± 0．2*、2．2±0．3#、2．3±0．4&倍(*t=3．75,P<0．01;#t=3.42,P<0．01;&t=3．26,P<0．01),它们在蛋白水平的改变与之一致。结论肝癌生长的微环境可通过核转录因子HIF-1α在转录水平调控多药耐药相关基因的表达,从而诱导肝癌多药耐药表型的形成;HIF-1α的表达上调是微环境诱导肝癌多药耐药彤成的中心环节;HIF-1α有望成为预防和逆转肝癌耐药的新的分子靶点。
Objective To investigate the role and location of hypoxia-inducible factor-1α (HIF-1α) in the microenvironment-induced multidrug resistance (MDR) phenotype of hepatocellular carcinoma and to provide an effective new molecular target for the prevention and reversal of drug resistance in hepatocellular carcinoma point. Methods Immunocytochemistry was used to detect the expression of HIF-1α protein in HepG2 cells which were stably integrated with HBX gene under hypoxia and hypoglycemic conditions respectively. Fluorescent quantitative PCR and Western-blot were used to detect the expression of HIF-1α in HepG2 cells at different mRNA and protein levels. The pcDNA3 / HIF-1α plasmid was transfected into HepG2 cells to detect the expression of multidrug resistance-related genes mdr1, LRP and MRP1 in transfected cells. Results The mRNA and protein of HIF-1α in HepG2 cells that grew or stably integrated with HBX gene under hypoxic and hypoglycemic conditions were both highly expressed and translocated. The mRNA levels of multidrug resistance-related genes mdr1, MRP1 and LRP in pcDNA3 / HIF-1α plasmid transfected HepG2 cells increased by 2.4 ± 0.2 * and 2.2 ± 0.3 # , 2.3 ± 0.4 times (* t = 3.75, P <0.01; # t = 3.42, P <0.01; The same level of change. Conclusion The microenvironment of HCC can induce the multidrug resistance phenotype of hepatocellular carcinoma by regulating the expression of multidrug resistance-related genes at the transcriptional level by HIF-1α. The up-regulation of HIF-1α expression is induced by microenvironment in HCC Multi-drug resistance Tung into the center of the link; HIF-1α is expected to become a new molecular target of prevention and reversal of drug resistance in liver cancer.