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目的:探讨人胃腺癌细胞系SGC7901及其多药耐药亚系SGC7901/VCR中端粒酶活性变化及其机制,为胃癌多药耐药机制研究提供新靶点。方法:采用TRAP银染方法检测SGC7901和SGC7901/VCR中端粒酶的活性;应用反转录聚合酶链反应(RT-PCR)技术检测SGC7901和SGC7901/VCR中端粒酶hTERT、Mad1及c-mycmRNA的表达;将hTERT启动子构建的报告基因质粒(pGL3B-TRTP),转染入SGC7901和SGC7901/VCR中,应用双荧光素酶报告基因检测系统(Dual-Luciferase reporter assay system)检测端粒酶hTERT启动子的活性。结果:SGC7901/VCR细胞中的端粒酶活性及端粒酶hTERTmRNA表达均显著高于SGC7901细胞,且SGC7901/VCR中hTERT启动子的活性显著高于SGC7901,在SGC7901/VCR中检测到c-mycmRNA的表达,但未检测到Mad1mRNA的表达,而在SGC7901细胞中分别检测到Mad1mRNA和c-mycmRNA的表达,且SGC7901细胞中c-mycmRNA的表达也高于SGC7901/VCR。结论:端粒酶活性及端粒酶hTERT转录水平升高与胃癌细胞的多药耐药性密切相关,转录因子Mad1/c-myc的表达高低是其作用机制之一。
OBJECTIVE: To investigate the changes of telomerase activity and its mechanism in human gastric adenocarcinoma cell line SGC7901 and its multidrug-resistant subline SGC7901 / VCR and provide a new target for the study of multidrug resistance in gastric cancer. Methods: The telomerase activity in SGC7901 and SGC7901 / VCR was detected by TRAP silver staining. Telomerase hTERT, Mad1 and c-Myc were detected in SGC7901 and SGC7901 / VCR by reverse transcription-polymerase chain reaction (RT-PCR) (pGL3B-TRTP) constructed by hTERT promoter was transfected into SGC7901 and SGC7901 / VCR cells and the expression of telomerase was detected by Dual-Luciferase reporter assay system hTERT promoter activity. Results: The telomerase activity and telomerase hTERT mRNA expression in SGC7901 / VCR cells were significantly higher than those in SGC7901 cells, and the hTERT promoter activity in SGC7901 / VCR was significantly higher than that in SGC7901 and c-myCMRNA was detected in SGC7901 / VCR But no expression of Mad1 mRNA was detected. However, the expression of Mad1 mRNA and c-mycRNA was detected in SGC7901 cells, and the expression of c-myc mRNA in SGC7901 cells was also higher than that of SGC7901 / VCR. CONCLUSION: The increase of telomerase activity and telomerase hTERT transcription is closely related to the multidrug resistance of gastric cancer cells. The expression of Mad1 / c-myc is one of the mechanisms.