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目的 检测SAcc83细胞系端粒酶活性并观察人端粒酶模板hTR封闭后细胞生长行为的改变及与凋亡的关系。方法TRAP-PCR-ELISA法检测SAcc83细胞系端粒酶活性,脂质体介导真核表达载体PBBS212-ahTR转染SAcc83,PCNA免疫组化染色以证实其增殖活性改变,TUNEL法及琼脂糖凝胶电泳检测细胞凋亡。结果转染反义hTR后SAcc83细胞增殖能力明显下降,群体倍增时间延长,转染细胞约30天后死亡。各项凋亡检测均证明细胞凋亡的存在。结论hTR反义基因通过封闭端粒酶模板RNA抑制了端粒酶活性,并可能通过某种途径激活细胞的凋亡程序,这为以端粒酶为靶子治疗恶性肿瘤提供了方向。
Objective To detect the telomerase activity of SAcc83 cell line and to observe the change of cell growth behavior and its relationship with apoptosis after human telomerase hTR closure. Methods The telomerase activity of SAcc83 cell line was detected by TRAP-PCR-ELISA. The expression of telomerase activity was detected by liposome-mediated eukaryotic expression vector PBBS212-ahTR. SAcc83 was detected by immunohistochemical staining with PCNA. TUNEL assay and agarose gel electrophoresis Detection of apoptosis by gel electrophoresis. Results After antisense hTR transfection, the proliferation of SAcc83 cells was significantly decreased and the population doubling time was prolonged. The transfected cells died after about 30 days. Each apoptosis test proved the existence of apoptosis. Conclusion The hTR antisense gene inhibits the telomerase activity by blocking the telomerase template RNA and may activate the apoptosis program in some way, which provides a direction for the treatment of malignant tumors with telomerase.