摘要
目的探讨DNA甲基化转移酶抑制剂5-氮-2′-脱氧胞苷(5-aza-dC)联合组蛋白去乙酰化抑制剂曲古抑菌素A(TSA)对人肺腺癌细胞株A549增殖和凋亡的影响。方法人肺腺癌细胞株A549细胞用含10%胎牛血清的DMEM培养液培养。实验分4组,5-aza-dC组:细胞接种20h后,加入5μmol/L的5-aza-dC;TSA组:细胞接种20h后,加入300nmol/LTSA;5-aza-dC+TSA组:细胞接种20h后,加入5μmol/L5-aza-dC,24h后再加入300nmol/L TSA;对照组:DMEM培养液中加入等量的二甲基亚砜(DMSO)。MTT比色法测定各处理因素对A549细胞生长的影响;Annexin V/PI双染法测定各组细胞培养72h后的凋亡情况;应用RT-PCR检测各组细胞培养72h后细胞CCAAT增强子启动蛋白α(C/EBPα)mRNA的表达。结果 (1)5-aza-dC及TSA呈时间依赖性地抑制A549细胞的生长,而在同一时间不同组别比较联合用药组细胞生长抑制率较单用药组明显增加[12h:35.51%vs21.63%,19.59%;24h:43.19%vs32.39%,30.60%;36h:54.85%vs38.83%,37.35%;48h:60.69%vs45.79%,43.84%;60h:68.92%vs50.60%,50.20%;72h:74.54%vs59.23%,57.82,P<0.05];(2)5-aza-dC和TSA能够诱导A549细胞凋亡,且联合用药组A549细胞凋亡率(75.35±1.46)%明显高于5-aza-dC(49.76±1.32)%(P<0.05)和TSA组(50.88±1.06)%(P<0.05);(3)5-aza-dC和TSA能够上调C/EBPαmRNA的表达,联合用药组中C/EBPαmRNA表达量(0.581±0.024)明显高于5-aza-dC组(0.402±0.016)(P<0.05)和TSA组(0.394±0.031)(P<0.05),有统计学意义;(4)5-aza-dC组与TSA组在抑制A549细胞生长、诱导细胞凋亡、增强C/EBPαmRNA的表达方面比较,差别均无统计学意义(P>0.05)。结论与单用5-aza-dC或TSA相比,联合应用两种药物能够更好的抑制人肺腺癌A549细胞的增殖、诱导细胞凋亡和增强C/EBPα基因mRNA的表达。
Objective To study the effect of methyltransferase inhibitor 5-aza-2-deoxycytidine （5-aza-dC） and histone deacetylase inhibitor trichostatin A （TSA） on proliferation and apoptosis of lung epithelial carcinoma A549 cells. Methods Human epithelial carcinoma A549 cells cultured in DMEM containing 10% fetal bovine serum were divided into control group,5-aza-dC group,TSA group,and 5-aza-dC ＋TSA group. Dimethyl sulfoxide （DMSO） was added into DMEM for control group. A549 cells were treated with 5 μmol/L 5-aza-dC and 300 nmol/L TSA 20 h after inoculation in 5-aza-dC group and TSA group,while A549 cells were treated with 5 μmol/L 5-Aza-CdR 20 h after inoculation and 300 nmol/L TSA after 24 h after inoculation in 5-aza-dC ＋TSA group. Effects of various processing factors on growth and apoptosis of A549 cells were detected by MTT assay with Annexin V/PI double staining 72 h after inoculation. Expression of CCAAT/enhancer-binding protein-a （C/EBPα） mRNA in different groups was detected by RT-PCR 72 h after inoculation. Results 5-aza-dC and TSA inhibited the growth of A549 cells in a time-dependent manner. The cell growth inhibitory rate was significantly higher in combination group than in single drug group （P0.05）. 5-aza-dC and TSA induced the apoptosis of A549 cells. The apoptotic rate of A549 cells was significantly higher in the combined treatment group than in single drug treatment group （P0.05）. The expression level of C/EBPα mRNA was significantly higher in combination group （0.581±0.024） than in 5-Aza-CdR group （0.402±0.016） and TSA group （0.394±0.031） after treatment with 5-aza-dC and TSA （P0.05）. No significant difference was found in growth and apoptosis of A549 cells,and in expression of C/EBPα mRNA between 5-aza-dC group and TSA group. Conclusion Combined 5-aza-CdR and TSA can more significantly inhibit the proliferation and induce the apoptosis of A549 cells,and increase the expression of C/EBPα mRNA than 5-aza-CdR or TSA alone,thus laying a theoretical basi
出处
《第三军医大学学报》
CAS
CSCD
北大核心
2010年第17期1857-1860,共4页
Journal of Third Military Medical University
基金
江西省科技支撑计划项目(837)~~
