hTERT基因反义核酸对顺铂诱导Raji细胞凋亡的影响

Effect of hTERT gene antisense oligodeoxynucleotide on cis-diamminedichicloroplatinum-induced apoptosis in Raji cells

目的 利用人类端粒酶逆转录酶 (hTERT)基因的反义寡核苷酸 (ASODN)抑制Raji细胞端粒酶活性后 ,探讨顺铂对Raji细胞凋亡的影响。方法 采用苔盼蓝拒染法观察hTERTASODN与顺铂联合作用对Raji细胞系生长的影响 ;姬姆萨染色法观察凋亡细胞的形态变化 ;琼脂糖凝胶电泳及流式细胞仪分析细胞凋亡。结果 hTERTASODN作用于Raji细胞 2 4h再加入顺铂 ,对细胞抑制明显增强 (P <0 .0 5 )。加入顺铂作用后 4 8h ,细胞出现典型的凋亡形态学改变 ,经琼脂糖凝胶电泳即可见到DNA梯形条带。凋亡细胞百分率 (2 5 .2 4± 1.5 8) % ,与正义寡核苷酸与顺铂联合作用组、单用顺铂作用组比较有显著性差异 (P <0 .0 1)。结论 hTERT基因反义寡核苷酸能促进顺铂诱导Raji细胞凋亡。

Objective To explore the effect of human telomerase reverse transcriptase (hTERT) gene antisense oligodeoxynucleotide (ASODN) on cis-diamminedichicloroplatinum(DDP)-induced apoptosis in Raji cells. Methods Cell viability was determined using the trypan blue dye exclusion assay. Apoptosis was detected by morphological observation,DNA gel electrophoresis and flow cytomertric cell cycle analysis. Results The survival rates of Raji cells cultured with DDP added 24 h later were higher than that cultured with hTERT ASODN and DDP added 24 h later. The survival rates of Raji cells cultured with DDP were similar with that cultured with hTERT sense oligodex ynuleotide (SODN) and DDP. In morphological observation of apoptotic cells using Giemsa staining,cells displayed classic apoptotic changes treated with DDP or DDP combined with hTERT ASODN or SODN at 48 h. Agarose gel electrophoresis of genomic DNA from Raji cells treated with ASODN and DDP combination for 48 h showed typical DNA “ladder”;neither did DNA from Raji cells treated with SODN plus DDP or DDP alone. Apoptosis rates of Raji cells treated with DDP for 48 h after 24 h of exposure to ASODN significantly increased. There was significant difference in the percentage of apoptotic cells of Raji cells between hTERT ASODN plus DDP and SODN plus DDP or DDP alone. Conclusion The hTERT ASODN could enhance DDP-induced apoptosis of Raji cells.