摘要
目的通过检测川芎嗪(TMP)对NB4细胞增殖、凋亡及端粒酶活性的影响,以及对人端粒酶逆转录酶(hTERT)、c-myc mRNA表达水平的影响,探讨TMP抗肿瘤的作用机制。方法采用MTT法检测细胞增殖;流式细胞术Annexin V-FITC/PI双染法检测细胞凋亡率;PCR-TRAP银染法定性检测端粒酶活性;PCR-TRAP-ELISA法定量检测端粒酶活性;RT-PCR法检测细胞的hTERT、cmycmRNA表达。结果MTF结果显示TMP浓度100～500 ug/ml能明显抑制NB4细胞的增殖(P<0.01),且抑制作用呈剂量时间依赖性;流式细胞术结果显示TMP浓度300ug/ml、500 ug/ml,作用NB4细胞48 h、72 h后,细胞凋亡率明显升高(P<0.05);PCRTRAP银染法定性检测端粒酶活性结果显示TMP各浓度组,作用NB4细胞72h后,条带明显变浅;PCR-TRAP-ELISA法定量检测端粒酶活性结果显示TMP各浓度组,作用NB4细胞72 h后,端粒酶活性(A_(450nm))明显降低(P<0.05);RT-PCR结果显示NB4细胞的hTERT、c-myc mRNA表达水平随TMP浓度增加、作用时间延长逐渐降低,且二者降低程度呈正相关。结论本实验提示TMP可能通过下调NB4细胞的c-myc表达来抑制细胞增殖和促进细胞凋亡,同时通过下调hTERT来降低细胞的端粒酶活性以发挥其抗肿瘤的作用。
Objective To detect the effect of tetranethylpyrazine （TMP） on the proliferation, apoptosis, telomerase activity of NB4 cells and the expression of human telomerase reverse transcriptase （hTERT）, c-myc mRNA, and explore anti-tumor effect of TMP. Methods The cell proliferation was determined by MTT;The cell apoptosis was detected by Annexin V-FITC/PI double staining;The telomerase activity was determined by PCR-TRAP silver staining and PCR-TRAP-ELISA;The expression of hTERT, c-myc mRNA was detected by RT-PCR. Results MTT results showed that the proliferation of NB4 cells was significantly inhibited with TMP at concentrations of 100 - 500 ug/ml （ P 〈 0.01 ） ,and the inhibitory effect was dose and time-dependent. The flow eytometry showed that the rate of apoptosis of NB4 cells was signifi- cantly increased with TMP at the concentration of 300 ug/ml and 500 ug/ml treated 48 h and 72 h （ P 〈 0.05 ） ; PCR-TRAP silver staining showed that the band significantly shallowed with TMP at all concentrations treated 72 h ; PCR-TRAP-ELISA method showed that the telomerase activity （ A450nm） significantly decreased with TMP at all concentrations treated 72 h（ P 〈 0.05 ） ; RT-PCR results showed that the expression of hTERT, c-myc was gradually reduced as the increasing concentration and hours of TMP, and the decrease in the former was positively correlated to that in the latter. Conclusion The study demonstrated that TMP could significantly inhibit the proliferation and telomerase activity of NB4 cells and induce the leukemia cells apoptosis, thereby playing its anti-tumor effect.
出处
《世界科技研究与发展》
CSCD
2012年第6期1004-1007,共4页
World Sci-Tech R&D
关键词
川芎嗪
白血病
端粒酶
tetramethylpyrazine
leukemia
telomerase
