摘要
目的 探讨组蛋白去乙酰化酶抑制剂抑制神经胶质瘤细胞株增殖的机制. 方法 分别用0.1、0.2、0.5、1.0、2.0 μmol/L曲古菌素A（TSA）处理U251细胞48 h,0.5 μmol/L TSA处理细胞8、16、24、36、48 h,1μmol/L M344、0.5 μmol/L LBH589、6 mmol/L NaBu、6 mmol/L VPA处理细胞48 h,对照组加入等量溶剂,MTT法检测细胞存活率;Western blotting检测0.5 μmol/L TSA处理U251细胞8、16、24 h后c-Jun氨基末端激酶（JNK）、磷酸化JNK （p-JNK）蛋白的表达;Westernblotting、MTT法分别检测对照组、10 μmol/L SP600125组和1μmol/L CEP11004组细胞c-Jun、p-c-Jun蛋白的表达和细胞存活率;Western blotting、MTT法分别检测转染pcDNA3.1组、转染pcDNA3.1 ＋TSA组、转染pMKK7-JNK1组、转染pMKK7-JNK1 ＋TSA组细胞p-c-Jun、Flag蛋白的表达和细胞存活率. 结果 0.1、0.2、0.5、1.0、2.0 μmol/L TSA组细胞存活率低于对照组,且TSA浓度越高,细胞存活率越低,半数抑制浓度（IC50）为0.5μmol/L;0.5 μmol/L TSA处理16、24、36、48 h后细胞存活率低于对照组,且处理时间越长,细胞存活率越低;1 μmol/L M344、0.5 μmol/L LBH589、6 mmol/L NaBu、6 mmol/L VPA组细胞存活率低于对照组,差异均有统计学意义（P＜0.05）.0.5 μmol/L TSA处理16h和24 h后细胞p-JNK蛋白水平显著降低;10 μμmol/L SP600125、1μmol/L CEP11004组细胞p-c-Jun蛋白的表达降低;10 μmol/L SP600125、1μmol/L CEP11004组细胞存活率低于对照组,差异有统计学意义（P＜0.05）;与转染pcDNA3.1组比较、转染pMKK7-JNK1组细胞存活率增加,与转染pcDNA3.1 ＋TSA组比较,转染pMKK7-JNK1 ＋TSA组细胞存活率增加,差异有统计学意义（P＜0.05）. 结论 组蛋白去乙酰化酶抑制剂抑制神经胶质瘤增殖的机制包含抑制JNK活性.
Objective To investigate the mechanism of histone deacetylase inhibitors in proliferation ofglioma cells.Methods （1） Glioma cell line U251 was cultured in vitro and treated with Trichostatin A （TSA） at 0.1,0.2,0.5,1.0 or 2.0 μmol/L for 48 h to determine the IC50 for TSA,and then,cells were treated with TSA at the IC50 concentration （0.5 μ mol/L） for 8,16,24,48 h;1 μmol/L M344,0.5 μmol/L LBH589,6 mmol/L NaBu and 6 mmol/L VPA were used to treat the cells for 48 h and same volume of solvent was given to cells for 48 h as control group;MTT assay was performed to determine cell viability.（2） Western blotting was performed to test the Jun N-terminal kinase （JNK）expression and phosphorylated JNK （p-JNK） level in the U251 cells after being treated with 0.5 μmol/L TSA for 8,16 and 24 h.（3） Western blotting and MTT assay were employed to detect the c-Jun expression and phosphorylated c-Jun （p-c-Jun） level,and cell viability in the U251 cells of control group,10 μmol/L SP600125 treatment group （JNK inhibitor） and 1 μmol/L CEP11004 treatment group （MLK3,a direct upstream kinase of JNK）.（4） Western blotting and MTT assay were employed to detect the c-Jun and Flag expressions,and cell viability in the U251 cells of pcDNA3.1 transfected group,pcDNA3.1＋TSA transfected group,pMKK7-JNK1 transfected group and pMKK7-JNK1＋TSA transfected group.Results （1） The viability of cells of 0.1,0.2,0.5,1.0 or 2.0 μmol/L TSA treated group was significantly lower than that in the control group,and the higher the TSA concentration,the lower the viability of cells;the viability of cells of 0.5 μmol/L TSA treated for 16,24,36 and 48 h groups was significantly lower than that in the control group,and the longer the TSA treatment,the lower the viability of cells.（2） The viability of 1 μmol/L M344,0.5 μmol/L LBH589,6 mmol/L NaBu and 6 mmol/L VPA treatment groups was significantly lower than that in the control group （P〈0.05）.（3） As compared with those in the control group,the
出处
《中华神经医学杂志》
CAS
CSCD
北大核心
2015年第6期547-552,共6页
Chinese Journal of Neuromedicine
基金
国家自然科学基金（31171021）
广东省医学科学技术研究基金（WSTJJ20111115440104197005103717）
广东省自然科学基金（S2011010003392）
广东省中医药局基金（20121135）
广州市属高校科研计划（10A23）.
