长期缺氧对肺腺癌SPCA1细胞谷胱甘肽转移酶表达及耐药性的影响

目的 探讨长期缺氧对SPCA1细胞谷胱甘肽转移酶（GST-π）表达的影响及耐药性的改变。方法 SPCA1细胞常氧（20％O2）和缺氧（0．5％O2）条件下作用16h后，提取RNA和蛋白，用定量RT-PCR的方法检测缺氧诱导因子1α（HIF-1α）和GST-π mRNA的表达情况，用Western blotting法检测HIF-1α蛋白；制备细胞悬液，用流式细胞仪测定GST-π表达；通过克隆形成实验分析SPCA1细胞对阿霉素和丝裂霉素的药物敏感性。通过 RNA干扰技术下调HIF-1α表达后，分为对照组和干扰组，观察GST-π的表达。结果 设常氧组GST-π mRNA表达率为1，缺氧组为12．2。常氧组GST-π蛋白阳性表达率为（35．78±1．25）％，缺氧组为（72．13±2．11）％。与常氧组比较缺氧组HIF-1α表达明显增加。计算1％克隆存活的药物浓度，阿霉素处理细胞，常氧组浓度为（0．29±0．05）μg／ml；缺氧组浓度为（0．48±0．07）μg／ml；丝裂霉素处理细胞，常氧组浓度为（0．71±0．10）μg／ml；缺氧组浓度为（0．79±0．12）μg／ml。干扰HIF-1α后，HIF-π mRNA与对照组比较下降了70％，而GST-π mRNA稍有下降。GST-π蛋白表达常氧条件下对照组为（32．14±1．23）％，干扰组为（30．88±1．65）％；缺氧条件下对照组为（64．78±2．45）％，干扰组为（67．42±2．21）％。结论 长期缺氧可诱导SPCA1细胞HIF-1α和GST-π表达增加，对阿霉素耐药性增加，对丝裂霉素没有影响，HIF-1α和GST-π并没有直接的相关性。

Prolonged hypoxia induces GST-π expression and drug resistance in lung adenocarcinoma cell line SPCA1

OBJECTIVE: To investigate the association of hypoxia with GST-pi expression in lung adenocarcinoma cells SPCA1 and the drug resistance in hypoxic condition. METHODS: RNA and protein in SPCA1 cells treated with hypoxia (0.5% O(2)) for 16 h were isolated. HIF-1alpha and GST-pi mRNA expression was detected by real time RT-PCR, and HIF-1alpha protein was detected by Western blotting analysis. GST-pi protein was finally analyzed on a FACScan flow cytometer using Cellquest software. Sensitivity of Doxorubicin and Mitomycin in hypoxic SPCA1 cells was detected using cell clonogenic assay. RESULTS: HIF-1alpha expression was up-regulated on mRNA and protein levels in hypoxic SPCA1 cells, so as was GST-pi expression. Doxorubicin resistance was increased, but hypoxia had no significant influence on Mitomycin resistance. GST-pi expression had no significant change after HIF-1alpha was down-regulated by RNA interfering. CONCLUSION: Though prolonged hypoxia can induce GST-pi expression and increase drug resistance such as to Doxorubicin, HIF-1alpha expression has no correlation with GST-pi expression.