重组TFAR19蛋白对鼻咽癌CNE-1细胞端粒酶活性和细胞凋亡的影响

Effect of recombinant TFAR19 protein on telomerase activity and apoptosis of human nasopharyngeal carcinoma CNE-1 cells

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 rTFAR19 protein on the expression of hTERT mRNA and telomerase activity in human nasopharyngeal carcinoma CNE-1 cells, so as to understand the mechanism by which rTFAR19 protein promotes the apoptosis of CNE-1 cells. Methods： Different concentrations of rTFAR19 protein were co-cultured with human CNE-1 cells. The apoptosis of CNE-1 cells was analyzed by 7-AAD and Annexin V-PE-labeled flow cytometry （FACS） assay. The expression of hTERT mRNA was detected by RT-PCR and was compared between the 2 groups. The changes of telomerase activity after TFAR19 treatment were examined by TRAP-sliver staining. Results： rTFAR19 protein incorporated into human nasopharyngeal carcinoma CNE-1 cells without a carrier. When the concentrations of rTFAR19 protein were 5mg/L, 10 mg/L, 15 mg/L,20 mg/L, and 30 mg/L, the apoptosis rates of CNE-1 cells were 15.26%, 29.48% ,37.11% ,54.20% ,and 72.36%, respecitively. The apoptosis rate of the positive control group （treated with stuanrosporin） and the negative control group （treated with PBS） were 98.37% and 13.40% , respectively. The expression of hTERT mRNA in CNE-1 cells was similar between the experimental group and the control group. The telomerase activity of CNE-1 cells was obviously decreased after cultured with rTFAR19 protein （ 〉20 mg/L）. Conclusion： High concentration of rTFAR19 protein can depress the telomerase activity of human nasopharyngeal carcinoma CNE-1 cells and subsequently accelerate the apoptosis of CNE-1 cells .