CAJ | 학술논문

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Cyclin A、Cyclin D1、Cyclin E、P21Waf/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0.75-4.0 mmol/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CyclinA、Cyclin D1、Cyclin E、P21Waf/cipl蛋白表达、RT-PCR检测分析Cyclin A、Cyclin DI、Cyclin E、P21Waf/cipl mRNA表达.结果经0.75-4.0 mmol/L VPA作用24、48、72、96 h,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P21Walf/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CyclinA、CyclinE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P21Walf/cipl mBNA、蛋白表达,下调Cyclin D1 mRNA和蛋白表达分别和/或协同实现.
목적 관찰응용조단백탈을선매억제제병무산납(VPA)조절염색체조단백저을선화수평대유선암세포증식적작용,병진일보검측Cyclin A、Cyclin D1、Cyclin E、P21Waf/cipl단백급mRNA표체변화래탐토기분자작용궤제.방법 유선암세포계MCF-7세포경0.75-4.0 mmol/L VPA작용후,MTT법검측세포생장억제、PI표기류식세포술검측세포주기、간접면역형광법분석CyclinA、Cyclin D1、Cyclin E、P21Waf/cipl단백표체、RT-PCR검측분석Cyclin A、Cyclin DI、Cyclin E、P21Waf/cipl mRNA표체.결과 경0.75-4.0 mmol/L VPA작용24、48、72、96 h,시험조세포생장억제솔축점승고,차정시간、제량의뢰추세;대조조세포증식주기G1、S、M기소점비례미견명현개변,이실험조칙수약물농도、작용시간적불동이출현불동정도적세포증식주기G1기조체,G1기비례유56.7%-78.9%불등,여대조조비교기승고차이유통계학의의(P<0.01).시험조P21Walf/cipl단백、mRNA표체피명현상조、Cyclin D1단백급mRNA표체피명현하조,여대조조비교차이유통계학의의(P<0.01);이CyclinA、CyclinE단백화mRNA표체칙미견명현변화(P>0.05).결론 통과응용조단백특이성탈을선매억제제조절조단백을선화수식가명현억제유선암세포생장、유도세포증식주기조체;병무산납작위조단백탈을선매억제제가명현억제유선암세포증식,기작용궤제시통과상조P21Walf/cipl mBNA、단백표체,하조Cyclin D1 mRNA화단백표체분별화/혹협동실현.
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 eytome- try with PI assay, and the protein and mRNA expressions of Cyelin A, Cyclin DI, Cyclin E, P21Waf/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 mmo]/L VPA induced a significant arrest in G1 phrase ( P<0.01), and a total of 55.4% -82.8% G1 phrase ratio were found. P21Waf/cipl was up-regulated both at the mRNA and protein level while Cyclin D1 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 G1 phrase. VPA, as a I class of histone deaeetylase inhibitor, can be used as an option in the treatment of breast cancer. The mechanism may include up-regulating P21Waf/cipl mRNA and protein expression and down-regulating Cyclin D1 mRNA and protein expression.