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目的研究重组TFAR19(TF-1cellapoptsis-relatedgene19)蛋白对鼻咽癌细胞株CNE-1端粒酶活性及端粒酶逆转录酶(hTERT)mRNA表达的影响,探讨重组TFAR19蛋白促CNE-1细胞凋亡的作用机制。方法将不同浓度的高纯度重组TFAR19蛋白分别和人类鼻咽癌细胞株CNE-1共同培养后,通过7-AAD及AnnexinV-PE标记进行流式细胞术分析,检测TFAR19蛋白对细胞的促凋亡效应;RT-PCR法比较试验组和对照组CNE-1细胞株端粒酶hTERTmRNA的表达水平;TRAP-银染法检测重组TFAR19蛋白作用下CNE-1细胞株端粒酶活性的改变。结果(1)一定浓度的TFAR19蛋白在未加载体的情况下,可促进细胞凋亡,加入5、10、15、20、30mg/L的TFAR19蛋白后,CNE-1细胞的凋亡率分别是15.26%,29.48%,37·11%,54·20%,72.36%。阳性对照组(凋亡诱导剂stuanrosporin处理)与阴性对照组(PBS处理)的细胞凋亡率分别是98·37%、13·40%。(2)试验组与对照组CNE-1细胞株端粒酶hTERTmRNA表达无显著差异。(3)与20mg/L以上浓度的重组TFAR19蛋白共同培养后,CNE-1细胞株端粒酶活性明显降低。结论重组TFAR19蛋白在较高浓度下可降低鼻咽癌细胞株CNE-1的端粒酶活性,明显促进肿瘤细胞凋亡。
Objective To investigate the effect of recombinant TFAR19 (TF-1cellapoptsis-related gene19) on telomerase activity and telomerase reverse transcriptase (hTERT) mRNA expression in nasopharyngeal carcinoma cell line CNE-1 and to investigate the effect of recombinant TFAR19 protein on CNE-1 cell apoptosis The mechanism of death. Methods Different concentrations of highly purified recombinant TFAR19 protein were co-cultured with human nasopharyngeal carcinoma cell line CNE-1, and then analyzed by flow cytometry with 7-AAD and Annexin V-PE to detect the pro-apoptotic effect of TFAR19 on cells The expression of telomerase hTERT mRNA in CNE-1 cell line was detected by RT-PCR. The telomerase activity of CNE-1 cell line was detected by TRAP-silver staining. Results (1) A certain concentration of TFAR19 protein could promote apoptosis in the absence of vector, and the apoptotic rate of CNE-1 cells after adding 5,10,15,20,30 mg / L TFAR19 protein were 15.26%, 29.48%, 37.11%, 54.20%, 72.36%. The apoptotic rate of positive control group (stuanosporin-induced apoptosis) and negative control group (PBS-treated) were 98.37% and 13.4% respectively. (2) The telomerase hTERT mRNA expression in CNE-1 cells in the experimental group and the control group showed no significant difference. (3) The telomerase activity of CNE-1 cell line was significantly decreased after co-cultured with recombinant TFAR19 protein at the concentration above 20mg / L. Conclusion The recombinant TFAR19 protein can reduce telomerase activity of nasopharyngeal carcinoma cell line CNE-1 at higher concentration and significantly promote the apoptosis of tumor cells.