曲古菌素A对肾癌细胞增殖及对去乙酰化酶1、金属蛋白酶9表达的影响

Effects of trichostatin A on cell growth and histone deacetylase 1, matrix metalloproteinase 9 gene expression in renal carcinoma cells

目的 观察曲古菌素A（TSA）对肾癌细胞7860增殖抑制作用及对组蛋白去乙酰化酶1（HDAC1）、基质金属蛋白酶9（MMP9）表达的影响.方法 噻唑蓝（MTT）比色法测定不同浓度（100、200、400、800nmol/L）、不同时间（12、24、36、48 h）TSA作用后7860细胞的抑制率.吖啶橙/溴乙锭（AO/EB）双重染色检测细胞凋亡.逆转录-聚合酶链反应（RT-PCR）及Western blot检测HDAC1、MMP9的表达.结果 不同浓度TSA作用24h后,细胞生长抑制率分别为32%、45%、58%、62%.TSA作用后部分细胞发生凋亡形态改变.HDAC1、MMP9基因在TSA作用后mRNA表达降低（P〈0.01）.与空白组、阴性组比较,HDAC1蛋白在TSA作用后表达分别下调18.6%～87.5%（P〈0.01）、21.3%～91.4%（P〈0.01）,MMP9蛋白表达分别下调24.6%～95.7%（P〈0.01）、20.8%～89.2%（P〈0.01）.结论 TSA可抑制肾癌细胞增殖,并可能通过下调HDAC1、MMP9的表达抑制细胞的增殖.

Objective To study the depressant effect on proliferation and the expression of histone deacetylase 1 （HDAC1） and matrix metalloproteinase 9 （MMP9） when the trichostatin A （TSA） was added to human renal carcinoma cell line 7860. Methods Four concentrations of TSA （100, 200, 400, 800 nmol/L） on7860 at different time durations of 12, 24, 36, 48 h were applied. The proliferation of 7860 was determined by methyl thiazolyl tetrazolium （MTT） method. The apoptosis was observed by AO/EB staining. The expression of HDAC1 and MMP9 in 7860 was examined by reverse transcription-polymerase chain reaction （RT-PCR） with TSA （400 nmol/L） at 24 h. Under the same testing condition, the protein expression level of HDAC1 and MMP9 in 7860 was detected by Western blotting, and the data were analyzed. Results The inhibition ratio was 32% , 45% , 58% , 62% in 7860 cells treated with TSA of 100, 200, 400, 800 nmol/L at 24 h. The apoptosis rate was increased after TSA treatment. The expression of HDAC1 and MMP9 mRNA in cancer cells was reduced after TSA treatment. The expression of HDAC1 protein was reduced by 18. 6%-87. 5% （P〈0.01） and 21. 3%-91.4% （P〈0.01）, and that of MMP9 protein was decreased by 24. 6% -95. 7% （P〈0.01） and 20. 8% -89. 2% （P〈0.01） respectively as compared with control group and negative group. Conclusion TSA could inhibit the expression of HDAC1 and MMP9, and proliferation of 7860 simultaneously.