应用VPA调节组蛋白乙酰化修饰抑制乳腺癌细胞增殖的实验研究  

Study of regulating histone acetylizad level with VPA on the proliferation of breast cancer cells

作　　者：时昌文[1] 赵霞[2] 孙京杰[1] 曹莉莉[1] 于振海[3] 顾禾[3] 

机构地区：[1]山东省千佛山医院普外中心实验室,山东济南250014 [2]山东省千佛山医院检验科,山东济南250014 [3]山东省千佛山医院普外中心,山东济南250014

出　　处：《中国医师杂志》2008年第7期907-910,共4页Journal of Chinese Physician

基　　金：基金项目：山东省卫生科技发展计划青年基金资助项目（2007QZ019）

摘　　要：目的观察应用组蛋白脱乙酰酶抑制剂丙戊酸钠（VPA）调节染色体组蛋白低乙酰化水平对乳腺癌细胞增殖的作用，并进一步检测CyclinA、Cyclin D1、Cyclin E、P21^Walf/cipl蛋白及mRNA表达变化来探讨其分子作用机制。方法乳腺癌细胞系MCF-7细胞经0.75-4.0mmol／L VPA作用后，MTT法检测细胞生长抑制、PI标记流式细胞术检测细胞周期、间接免疫荧光法分析Cyclin A、Cyclin D1、Cyclin E、P21^Walf/cipl蛋白表达、RT-PCR检测分析Cyclin A、Cyclin D1、Cyclin E、P21^Walf/cipl mRNA表达。结果经0.75-4.0mmol／L VPA作用24、48、72、96h，试验组细胞生长抑制率逐渐升高，且呈时间、剂量依赖趋势；对照组细胞增殖周期G1、S、M期所占比例未见明显改变，而实验组则随药物浓度、作用时间的不同而出现不同程度的细胞增殖周期G1期阻滞，G1期比例由56．7％．78．9％不等，与对照组比较其升高差异有统计学意义（P〈0．01）。试验组P21^Walf/cipl蛋白、mRNA表达被明显上调、Cyclin D1蛋白及mRNA表达被明显下调，与对照组比较差异有统计学意义（P〈0．01）；而Cyclin A、Cyclin E蛋白和mRNA表达则未见明显变化（P〉0．05）。结论通过应用组蛋白特异性脱乙酰酶抑制剂调节组蛋白乙酰化修饰可明显抑制乳腺癌细胞生长、诱导细胞增殖周期阻滞；丙戊酸钠作为组蛋白脱乙酰酶抑制剂可明显抑制乳腺癌细胞增殖，其作用机制是通过上调P21^Walf/cipl mRNA、蛋白表达，下调Cyclin D1 mRNA和蛋白表达分剐和／或协同实现。Objective To investigate the effect and the mechanism of up-regulating histone acetylizad level with a selective inhibitor of HDACs-Valproate acid sodium （VPA） on breast cancer cell proliferation. Methods MCF-7 cells were cultured with 0.75-4.0 mmol/L valproic acid （VPA） for 24, 48, 72, 96 hours in vitro, the inhibiting rate was tested by MTT assay. Cell cycle was analyzed by flow cytometry with PI assay, and the protein and mRNA expressions of Cyelin A, Cyelin DI, Cyelin E, P21^Walf/cipl of MCF-7 cells after 1.5, 3.0 mmol/ L VPA treated were analyzed by indirect immunofluorescence technique and RT-PCR respectively. Results After cultured with 0.75-4.0 mmol/L valproic acid （VPA） for 24, 48, 72, 96 hours, the inhibiting rate of experimental groups increased significantly（ P 〈 0. 01 ） and a dose and acting time dependent manner was found. As to cell cycle, the percentages of GI, S, M phrase in control groups remained the same. Contrary to control groups, 0.75 -4.0 mmol/L VPA induced a significant arrest in GI phrase （ P 〈0. 01 ）, and a total of 55.4% - 82.8% GI phrase ratio were found. P21^Walf/cipl was up-regulated both at the mRNA and protein level while Cyclin DI was down-regulated （ P 〈 0. 001 ）. Conversely, neither mRNA nor protein expression of Cyclin A, Cyclin E showed difference （ P 〉 0. 05）. Conclusions Up-regulating histone acetylizad level can inhibit breast cancer cell proliferation, induce cell cycle arrest in GI phrase. VPA, as a I class of histone deacetylase inhibitor, can be used as an option in the treatment of breast cancer. The mechanism may include up-regulating P21^Walf/cipl mRNA and protein expression and down-regulating Cyclin DI mRNA and protein expression.

关 键 词：丙戊酸 组蛋白脱乙酰基酶类 乳腺肿瘤 细胞增殖 

分 类 号：R737.9[医药卫生—肿瘤]