大鼠胰腺癌和非癌胰腺组织Skp2和p27表达研究

目的建立大鼠胰腺癌模型,研究大鼠胰腺癌和非癌胰腺组织中Skp2和p27表达及其意义。方法应用二甲基苯并蒽(DMBA)置入大鼠胰实质内(A组)及设立曲古霉素(TSA)干预组(B组),3~5个月内处死观察胰腺癌发生情况,应用SP免疫组化法研究胰腺癌和非癌胰腺组织中Skp2和p27表达。结果3~5个月内A组发癌率为48.7%(18/37),B组为33.3%(12/36),A组和B组各1例纤维肉瘤外,余均为胰腺导管癌;A组肿块最大径大于B组(P<0.05);非癌胰腺组织中A组导管上皮中至重度不典型增生者(52.6%)高于B组(33.3%),但差异无显著性(P>0.05);C组胰腺和A或B组鼠胰腺外主要脏器均无明显病变。A组和/或B组胰腺导管癌Skp2表达阳性率均明显高于A组和/或B组非癌胰腺组织(P<0.05);但胰腺导管癌p27表达阳性率明显低于非癌胰腺组织(P<0.01);Skp2阳性和p27阴性的非胰腺组织导管上皮均呈中至重度不典型增生;C组胰腺导管上皮Skp2均阴性表达和p27均阳性表达。结论DM-BA置入胰实质内可在短期获得较高的SD鼠胰腺癌发生率,TSA能抑制胰腺癌发生和生长。Skp2和p27表达水平与SD鼠胰腺癌发生发展有密切关系。     

慢加急性肝衰竭小鼠肝组织组蛋白去乙酰化酶mRNA水平变化

目的 探讨组蛋白去乙酰化酶(HDAC)抑制剂干预慢加急性肝衰竭(ACLF)小鼠肝组织组蛋白去乙酰化酶mRNA水平变化.方法 随机将小鼠分为7组,每组4只.A组为对照组,B组为模型组,C组为曲古抑菌素A(TSA)处理组,D组为大剂量正丁酸钠处理组,E组为小剂量正丁酸钠处理组,F组为大剂量吡咯烷二硫代氨基甲酸(PDTC)处...     

曲古抑菌素A对甲状腺鳞癌细胞中Cyclin D1、CDK4、pRB表达的影响

目的:观察曲古抑菌素A(TSA)对甲状腺鳞癌(SW579)细胞中细胞周期蛋白D1(Cyclin D1)、细胞周期蛋白依赖性激酶4(CDK4)、视网膜母细胞瘤基因(RB)的蛋白产物(pRB)表达的影响。方法:体外培养SW579细胞,以二甲基亚砜(DMSO)为溶剂对照组,另设空白对照(未作任何处理)组,观察50、100、200、400nmol·L-1TSA对SW579细胞生长抑制率的影响,及细胞中Cy-clin D1、CDK4、RB基因和蛋白的表达情况。结果:与溶剂对照组和空白对照组比较,各剂量TSA作用后SW579细胞的细胞生长抑制率明显升高(P<0.01),且呈剂量依赖性。与空白对照组比较,各剂量TSA作用后SW579细胞中Cyclin D1 mRNA表达明显降低(P<0.01),且随剂量增加表达降低,但CDK4、RB mRNA表达和溶剂对照组中这2个基因的表达均无明显变化(P>0.05);与空白对照组比较,各剂量TSA作用后细胞中Cyclin D1和pRB蛋白表达明显降低(P<0.01),且随剂量增加表达降低,但CDK4蛋白表达无明显变化(P>0.05)。结论:TSA能明显抑制SW579细胞的生长,且呈剂量依赖性;其机制可能与Cyclin D1基因和蛋白、pRB蛋白的表达有关。     

Endocrine Active Compounds: From Biology to Dose Response Assessment

Histone deacetylase (HDAC) inhibitors target key steps of tumor development: They inhibit proliferation, induce differentiation and/or apoptosis, and exhibit potent antimetastatic and antiangiogenic properties in transformed cells in vitro and in vivo. Preliminary studies in animal models have revealed a relatively high tumor selectivity of HDAC inhibitors, strenghtening their promising potential in cancer chemotherapy. Until now, preclinical in vitro research has almost exclusively been performed in cancer cell lines and oncogene-transformed cells. However, as cell proliferation and apoptosis are essential for normal tissue and organ homeostasis, it is important to investigate how HDAC inhibitors influence the regulation of and interplay between proliferation, differentiation, and apoptosis in primary cells as well. This review highlights the discrepancies in molecular events triggered by trichostatin A, the reference compound of hydroxamic acid-containing HDAC inhibitors, in hepatoma cells and primary hepatocytes (which are key targets for drug-induced toxicity). The implications of these differential outcomes in both cell types are discussed with respect to both toxicology and drug development. In view of the future use of HDAC inhibitors as cytostatic drugs, it is highly recommended to include both tumor cells and their healthy counterparts in preclinical developmental studies. Screening the toxicological properties of compounds early in their development process, using a battery of different cell types, will enable researchers to discard those compounds bearing undesirable adverse activity before entering into expensive clinical trials. This will not only reduce the risk for harmful exposure of patients but also save time and money.     

曲古抑菌素A对乙型肝炎病毒HBx蛋白与组蛋白去乙酰化酶1相互作用的影响

目的 探讨曲古抑菌素A对乙型肝炎病毒HBx蛋白与组蛋白去乙酰化酶1(HDAC1)相互作用的影响. 方法 常规分子克隆技术构建HBx蛋白及HDAC1表达载体,Western blot分别验证蛋白质表达;谷胱甘肽转移酶沉降试验及免疫共沉淀技术分别于体内外验证HBx蛋白与HDAC1存在的相互作用. 结果成功构建了HBx蛋白及HDAC1表达载体,体内外实验均证实HBx蛋白与HDACI间存在相互作用,但HDACI抑制剂曲古抑菌素A不影响两者间相互作用.结论 HBx蛋白以曲古抑菌素A非依赖方式与HDAC1存在相互作用.     

Effect of histone deacetylase inhibitor on proliferation of biliary tract cancer cell lines

AIM： To explore the effect of histone deacetylase inhibitor, trichostatin A （TSA） on the growth of biliary tract cancer cell lines （gallbladder carcinoma cell line and cholangiocarcinoma cell line） in vivo and in vitro, and to investigate the perspective of histone deacetylase inhibitor in its clinical application. METHODS： The survival rates of gallbladder carcinoma cell line （Mz-ChA-l cell line） and cholangiocarcinoma cell lines （QBC939, KMBC and OZ cell lines） treated with various doses of TSA were detected by methylthiazoy tetrazolium （MTT） assay. A nude mouse model of transplanted gallbladder carcinoma （Mz-ChA-l cell line） was successfully established, and changes in the growth of transplanted tumor after treated with TSA were measured. RESULTS： TSA could inhibit the proliferation of gallbladder carcinoma cell line （Mz-ChA-l cell line） and cholangiocarcinoma cell lines （QBC939, KMBC and OZ cell lines） in a dose-dependent manner. After the nude mouse model of transplanted gallbladder carcinoma （Mz- ChA-l cell line） was successfully established, the growth of cancer was inhibited in the model after treated with TSA. CONCLUSION： TSA can inhibit the growth of cholangiocarcinoma and gallbladder carcinoma cell lines in vitro and in vivo.     

曲古抑菌素A促进小鼠骨髓间充质干细胞分化为胰岛素分泌细胞

 目的： 观察曲古抑菌素A(TSA)诱导小鼠骨髓间充质干细胞分化为胰岛素分泌细胞的适宜浓度。方法: C57BL/6小鼠骨髓间充质干细胞系体外培养。实验分为5组：对照组（DMSO,A组)和TSA干预组（25 nmol/L,B组;50 nmol/L,C组;100 nmol/L,D组;200 nmol/L,E组）。各组分两阶段用相应的培养基培养10 d后进行检测。双硫腙染色鉴定各组的胰岛素分泌细胞,结果进行半定量分析。免疫荧光染色鉴定各组的胰岛素分泌,比较各组胰岛素平均荧光强度。酶联免疫吸附试验检测各组胰岛素分泌细胞胰岛素的分泌量。结果: TSA作用10 d能够诱导C57BL/6 小鼠骨髓间充质干细胞分化成为胰岛素分泌细胞并分泌胰岛素。 B组胰岛素染色阳性面积、阳性率、胰岛素染色积分吸光度以及胰岛素平均荧光强度显著高于其它干预组,差异有统计学意义（均P<0.05）。 随着各干预组TSA浓度的升高,胰岛素的分泌量减少。B组胰岛素含量显著高于其它干预组,差异有统计学意义（均P<0.05）。结论：TSA处理10 d能诱导C57BL/6 小鼠骨髓间充质干细胞分化成为胰岛素分泌细胞,并且25 nmol/L为TSA的适宜浓度。     

曲古霉素A和5-氮杂胞苷对高糖诱导损伤的胰岛β细胞的保护作用

目的 探讨高糖诱导下曲古霉素A（TSA）和5-氮杂胞苷（5-AzaC）对胰岛β细胞增殖、凋亡及功能的影响. 方法 体外培养大鼠胰岛素瘤（RIN-m5f）细胞至指数增长期,根据干预方案分为空白对照（NC）组、高糖诱导（HG）组、0.05、0.10 μmol/L TSA干预组、0.63、1.25 μmol/L 5-AzaC干预组、0.10 μmol/L TSA＋1.25 μmol/L 5-AzaC联合干预组.检测RIN-m5f细胞增殖活性、细胞凋亡率、葡萄糖刺激胰岛素分泌能力. 结果 0.05、0.10 μmol/L TSA干预组、0.63、1.25 μmol/L 5-AzaC干预组及联合干预组的细胞增殖活性均高于HG组,而细胞凋亡率均低于HG组（P＜0.05）.在3.3 mmol/L及25 mmol/L葡萄糖刺激时,各组葡萄糖刺激胰岛素分泌量均高于HG组（P＜0.05）. 结论 TSA和5-AzaC可抑制高糖诱导的胰岛β细胞凋亡和恢复β细胞的胰岛素分泌功能.     

曲古菌素A协同GX15-070诱导胃癌细胞SGC-7901凋亡机制的研究

目的探讨联合应用曲古菌素(Trichostatin A,TSA)和Bcl-2抑制剂GX15-070诱导人胃癌细胞SGC-7901凋亡作用及其机制。方法以联用或单用TSA、GX15-070作用于SGC-7901细胞,采用MTT法检测细胞存活率;采用Western blot法检测经处理后细胞SGC-7901中cleaved caspase 3和Bcl-2蛋白表达的变化。结果在一定浓度范围内(2.5～10μmol/L)GX15-070以浓度依赖性诱导SGC-7901细胞存活率降低,各浓度组间细胞存活率差异有统计学意义(P<0.01),且随作用时间延长,SGC-7901细胞存活率降低(P<0.05);TSA和GX15-070联用组较单用TSA组、GX15-070组SGC-7901细胞存活率差异有统计学意义(P<0.01);West-ern blot检测显示TSA能上调凋亡蛋白cleaved caspase 3的表达和下调Bcl-2蛋白的表达。结论 TSA具有协同诱导胃癌细胞SGC-7901凋亡的作用,其机制可能是通过上调cleaved caspase 3和下调Bcl-2蛋白的表达有关。