CAJ | 학술논문

目的研究组蛋白脱乙酰化酶(HDAC)抑制剂丙戊酸钠(VPA)体内给药对白血病细胞系Kasumi-1细胞裸鼠移植瘤生长的影响,探讨可能的作用机制.方法建立Kasumi-1细胞裸鼠移植瘤模型,分为对照组和VPA治疗组进行实验,观察并记录各组裸鼠移植瘤的生长情况,计算肿瘤体积和肿瘤生长抑制率.HE染色观测肿瘤细胞形态.采用末端脱氧核糖核酸转移酶介导的dUTP原位缺口末端标记(TUNEL)法检测肿瘤细胞凋亡.RT-PCR和Western blot方法分析粒-单核细胞集落刺激因子(GM-CSF)、组蛋白脱乙酰化酶1(HDAC1)、乙酰化组蛋白H3(Ac-H3)、Survivin mRNA或蛋白的表达.染色质免疫共沉淀(ChIP)法检测VPA对GM-CSF基因启动子区域染色质内组蛋白H3乙酰化程度的影响.结果 ①VPA体内用药明显抑制裸鼠Kasumi-1细胞移植瘤的生长,VPA治疗组肿瘤体积及重量与对照组相比明显缩小或降低,瘤体抑制率(IR)为57.25%.②HE染色可见肿瘤细胞分化及凋亡程度增加,凋亡指数VPA组[(3.66 ±0.77)%]与对照组[(0.27±0.11)%]相比明显升高.③与对照组比较,VPA治疗组胞核内HDAC1蛋白表达水平明显降低,Ac-H3蛋白表达水平明显升高;GM-CSF mRNA和蛋白表达水平明显升高;GM-CSF启动子区域染色质内组蛋白H3的乙酰化程度明显升高;Survivin的mRNA表达水平明显降低.结论 VPA对Kasumi-1细胞裸鼠移植瘤有明显的生长抑制作用,诱导肿瘤细胞分化及凋亡,其机制可能与通过抑制Survivin基因表达诱导细胞凋亡,和通过抑制细胞核内HDAC表达,增强组蛋白乙酰化程度,影响GM-CSF基因启动子区域染色质内的组蛋白乙酰化修饰,增加AML1靶基因GM-CSF的转录诱导细胞分化有关.
목적 연구조단백탈을선화매(HDAC)억제제병무산납(VPA)체내급약대백혈병세포계Kasumi-1세포라서이식류생장적영향,탐토가능적작용궤제.방법 건립Kasumi-1세포라서이식류모형,분위대조조화VPA치료조진행실험,관찰병기록각조라서이식류적생장정황,계산종류체적화종류생장억제솔.HE염색관측종류세포형태.채용말단탈양핵당핵산전이매개도적dUTP원위결구말단표기(TUNEL)법검측종류세포조망.RT-PCR화Western blot방법분석립-단핵세포집락자격인자(GM-CSF)、조단백탈을선화매1(HDAC1)、을선화조단백H3(Ac-H3)、Survivin mRNA혹단백적표체.염색질면역공침정(ChIP)법검측VPA대GM-CSF기인계동자구역염색질내조단백H3을선화정도적영향.결과 ①VPA체내용약명현억제라서Kasumi-1세포이식류적생장,VPA치료조종류체적급중량여대조조상비명현축소혹강저,류체억제솔(IR)위57.25%.②HE염색가견종류세포분화급조망정도증가,조망지수VPA조[(3.66 ±0.77)%]여대조조[(0.27±0.11)%]상비명현승고.③여대조조비교,VPA치료조포핵내HDAC1단백표체수평명현강저,Ac-H3단백표체수평명현승고;GM-CSF mRNA화단백표체수평명현승고;GM-CSF계동자구역염색질내조단백H3적을선화정도명현승고;Survivin적mRNA표체수평명현강저.결론 VPA대Kasumi-1세포라서이식류유명현적생장억제작용,유도종류세포분화급조망,기궤제가능여통과억제Survivin기인표체유도세포조망,화통과억제세포핵내HDAC표체,증강조단백을선화정도,영향GM-CSF기인계동자구역염색질내적조단백을선화수식,증가AML1파기인GM-CSF적전록유도세포분화유관.
Objective To investigate in vivo inhibitory effect of histone deacetylase ( HDAC) inhibitor valproic acid( VPA) on xenografted Kasumi-1 tumor in nude mice and its mechanism.Methods Xenografted Kasumi-1 tumor mouse model was established by subcutaneous inoculation of Kasumi-1 cells.Xenotransplanted nude mice were assigned into control or VPA treatment groups.Volume of the xenografted tumors was measured and compared between the two groups.Terminal deoxynucleotidyl transferase biotin-dUTP nick end labeling (TUNEL) was applied to detection of tumor cell apoptosis.The gene expression of GM-CSF、 HDAC1 、Ac-H3 and survivin was studied with semi-quantitative RT-PCR and Western blotting.ChIP method was used to assay the effects of VPA on acetylation of histone H3 within GM-CSF promoter region.Results ①VAP significantly inhibited xenografted Kasumi-1 tumor growth.The calculated inhibition rate was 57.25%.②Morphologic study showed that VPA induced differentiation and apoptosis of Kasumi-1 tumor cells.The apoptosis index of VAP treatment group[(3.661 ±0.768) %]was significantly higher than that of control group[(0.267 ±0.110)%].③Comparing to those in control group, the level of nuclear HDAC1 protein was significantly decreased, the Ac-H3 protein expression level was increased, the mRNA and protein expression levels of GM-CSF and acetylation of histoneH3were remarkablyincreased,and the geneexpression level of survivin significantly decreased in VPA treatment group.Conclusion VAP significantly inhibites xenografted Kasumi-1 tumor growth and induces tumor cell differentiation and apoptosis.The mechanism may be decrease of survivin gene expression,inhibition of nuclear expression of HDAC,promotion of histone protein acytelation level and acetylation of histone H3 within GM-CSF promoter region,and increase of GM-CSF transcription.