目的探讨葡萄糖神经酰胺合成酶(glucosylceramide synthase,GCS)基因在人胰腺癌细胞SW1990及其耐药亚株中的表达和意义。方法体外培养人胰腺癌细胞SW1990和其耐药亚株SW1990/FU、SW1990/ADM、SW1990/GEM,以细胞活性测定(CCK-8法)检测耐药亚株的耐药指数,相对定量荧光实时PCR(RT—PCR)法检测GCS mRNA的表达,免疫印迹法检测GCS蛋白表达水平。结果与亲本株相比,SW1990/FU、SW1990/ADM、SW1990/GEM的耐药指数分别为339.7、11.9、56.6。RT—PCR和免疫印迹检测结果显示亲本株和耐药亚株均有GCS mRNA和蛋白的表达,GCS mRNA在3株耐药亚株中的表达均比亲本株有增高趋势,且SW1990/ADM、SW1990/GEM的GCS蛋白表达水平较亲本株升高,差异有统计学意义(P〈0.05)。结论GCS基因在人胰腺癌细胞株SW1990和其3株耐药亚株中表达阳性,GCS蛋白在SW1990/ADM、SW1990/GEM中呈高表达,表明GCS可能参与胰腺癌耐药株细胞对ADM、GEM的耐药。
Objective To study the expression and significance of GCS gene in human pancreatic cancer cell line SW1990 and its drug-resistant sublines. Methods SW1990 and its drug-resistant sublines, SW1990/FU, SW1990/ADM and SW1990/GEM were cultured in vitro. CCK-8 (Cell Counting kit-8) was used to detect the drug resistance of the sublines. Relative quantitation of GCS mRNA expression was evaluated by real-time PCR and Western blot was adopted to evaluate the expression of GCS protein. Results The drug resistance indexes of SW1990/FU, SW1990/ADM and SW1990/GEM were 339. 7, 11.9 and 56. 6, respectively. The GCS mRNA and protein were expressed in all SW1990 and its drug-resistant sublines. There was a higher expression of GCS mRNA in all the sublines and a significant difference of GCS protein expression was detected in SW1990/ADM and SW1990/GEM compared with SW1990. Conclusions GCS gene is expressed in SW1990 and its drug-resistance sublines. The high expression of GCS protein in SW1990/ADM and SW1990/GEM might be one reason of resistance to ADM and GEM in the sublines.