I型γ-分泌酶抑制剂对恶性胶质瘤细胞株增殖及凋亡的影响

Objective To investigate the role of gamma secretase inhibitor-I (GSI-I) in cell proliferation and apoptosis of human glioma cell lines U87 and U251.Methods RT-PCR and fluorescent quantitative RT-PCR (qRT-PCR) were employed to evaluate the expressions of Notch receptors and their target gene Hes-I in both U87 and U251 cells treated by GSI-I,respectively.Then,MTT assay was used to examine the effects of GSI-I on cell proliferation of the 2 glioma cells.Meanwhile,flow cytometry technique was also employed to detect the cell cycle changes and apoptosis induced by GSI-I treatment.Results The activity of Notch pathway was inhibited by GSI-I treatment through down-regulating the expression of Notch receptors target gene Hes-I in both U87 and U251 cells.Treatment with 2.5μmol/L GSI-I or above concentrations could significantly induce the cell cycle arrest of U87 and U251 cells and these effects were positively concentration-dependent.Flow cytometry technique showed that GSI-I inhibited the cell proliferation by inducing the cell cycle arrest of U87 cells at GI phase and inducing the apoptosis of U251 cells.Conclusion GSI-I can dramatically inhibit the cell proliferation and induce the apoptosis of U87 and U251 cells,providing a reliable evidence for clinical glioma treatment.     

I型γ-分泌酶抑制剂对恶性胶质瘤细胞株增殖及凋亡的影响

目的 探讨I型γ-分泌酶抑制剂(GSI-I)对U87、U251胶质瘤细胞的增殖抑制及诱导凋亡作用.方法 应用GSI-I作用于U87、U251胶质瘤细胞,通过MTT法观察GSI-I对上述两种细胞的增殖抑制作用,通过流式细胞仪检测GSI-I对该两种细胞细胞周期的影响及诱导凋亡的作用.结果RT-PCR及实时定量荧光RT-PCR结果显示,GSI-T可明显抑制胶质瘤细胞中Notch通路的活性,主要体现为Notch通路的靶基因Hes-1表达明显下调.MTT检测结果显示2.5μmol/L及以上浓度的GSI-I对U87及U251胶质瘤细胞有明显的增殖抑制作用.与对照组比较,差异有统计学意义(P<0.05),且该抑制作用呈剂量依赖型增加.流式细胞检测结果显示GSI-I主要使U87胶质瘤细胞的细胞周期阻滞在G1期而抑制细胞增殖,对于U251胶质瘤细胞则主要通过诱导凋亡来抑制增殖.结论 GSI-I可明显抑制U87及U251胶质瘤细胞的增殖并诱导凋亡,为恶性胶质瘤的临床治疗提供了理论参考依据.     

斑马鱼红系造血缺陷突变体的正向遗传学筛选

目的 化学遗传学方法 大规模筛选并初步鉴定所获得具有不同红系造血缺陷表型的斑马鱼突变体.方法 乙基亚硝基脲(ENU)诱导雄性斑马鱼突变(founder),将其与野生型AB雌性斑马鱼交配产生F1代,源自不同来源的founder的F1代杂交产生F2家族.在F2代同家族内自交所产生的F3代胚胎中,用以βe1为探针,实施整体原位杂交实验,进行红系造血缺陷突变体筛选,并针对所筛选到的突变体在不同造血过程缺陷表型进行分类研究.结果 和结论 筛选得到4个βe1基因表达缺失突变体,其中2个为红系特异性造血缺陷突变体,另外2个突变体同时存在红系和淋系造血缺陷.     

散发性乳腺癌组织中BRCA1选择性调节黄体酮受体A的表达

目的 观察散发性乳腺癌组织中BRCA1、黄体酮受体A(progesterone receptor A,PRA)和黄体酮受体B(progesterone receptor B,PRB)的表达,并分析散发性乳腺癌BRCA1对PRA和PRB表达的影响.方法 选择南方医科大学附属南方医院病理科乳腺腺病和散发性浸润性乳腺导管癌病例蜡块包埋组织各68例,应用免疫组织化学法检测散发性浸润性乳腺导管癌组织BRCA1、PRA和PRB蛋白及乳腺腺病组织BRCA1蛋白的表达,用Wilcoxon两样本检验比较浸润性导管癌和乳腺腺病BRCA1表达的高低,用x2检验分析浸润性导管癌BRCA1对PRA和PRB表达的影响.结果 (1)浸润性导管癌组织BRCA1蛋白阳性表达率为60.3%(41/68),低于乳腺腺病BRCA1蛋白阳性表达率85.3%(58/68),两者之间差异有统计学意义(P<0.01);(2)浸润性导管癌组织中,BRCA1表达阴性标本PRA的阳性率为81.5%(22/27),高于BRCA1表达阳性标本PRA的阳性率53.7%(22/47),差异有统计学意义(P<0.05),提示BRCA1可影响PRA的表达.浸润性导管癌组织中PRB的表达率在BRCA1表达阳性标本与表达阴性标本之间差异无统计学意义(P>0.05),提示BRCA1不影响PRB的表达.结论 在散发性乳腺癌组织中,BRCA1表达阴性选择性地与PRA的高表达相关联,但与PRB表达率不相关联,提示BRCA1在散发性乳腺癌组织中选择性地调节PRA的表达.     

黄体酮促进培养乳腺癌细胞的增殖和迁移

目的研究黄体酮对乳腺癌细胞增殖和迁移的影响。方法100nM黄体酮刺激黄体酮受体阳性的乳腺癌细胞株T-47D和MCF-748小时后,用MTT方法检测细胞增殖,用细胞划痕愈合实验检测细胞迁移,用Western blotting检测细胞内E-Cadherin的表达。结果黄体酮刺激MCF-7和T-47D细胞的增殖和迁移;并且可以下调MCF-7和T-47DE-Cadherin的表达。结论黄体酮能够促进培养乳腺癌细胞的增殖和迁移,提示抗黄体酮治疗可能有临床意义。     

I型γ-分泌酶抑制剂对恶性胶质瘤细胞株增殖及凋亡的影响

Objective To investigate the role of gamma secretase inhibitor-I (GSI-I) in cell proliferation and apoptosis of human glioma cell lines U87 and U251.Methods RT-PCR and fluorescent quantitative RT-PCR (qRT-PCR) were employed to evaluate the expressions of Notch receptors and their target gene Hes-I in both U87 and U251 cells treated by GSI-I,respectively.Then,MTT assay was used to examine the effects of GSI-I on cell proliferation of the 2 glioma cells.Meanwhile,flow cytometry technique was also employed to detect the cell cycle changes and apoptosis induced by GSI-I treatment.Results The activity of Notch pathway was inhibited by GSI-I treatment through down-regulating the expression of Notch receptors target gene Hes-I in both U87 and U251 cells.Treatment with 2.5μmol/L GSI-I or above concentrations could significantly induce the cell cycle arrest of U87 and U251 cells and these effects were positively concentration-dependent.Flow cytometry technique showed that GSI-I inhibited the cell proliferation by inducing the cell cycle arrest of U87 cells at GI phase and inducing the apoptosis of U251 cells.Conclusion GSI-I can dramatically inhibit the cell proliferation and induce the apoptosis of U87 and U251 cells,providing a reliable evidence for clinical glioma treatment.     

5-氮杂-2’-脱氧胞苷诱导乳腺癌细胞自噬与DNA损伤相关联

目的:探讨5-氮杂-2’-脱氧胞苷对乳腺癌细胞自噬的影响及其可能的机制.方法:用依托泊苷、顺铂和5-氮杂-2’-脱氧胞苷处理乳腺癌细胞后,采用彗星实验测定细胞DNA损伤,蛋白质印迹法检测p53和p21蛋白表达;细胞自噬测定用3种方法:(1)蛋白质印迹法检测微管相关蛋白1轻链3-Ⅱ (microtubule-associated protein1 light chain 3-Ⅱ,LC3-Ⅱ)蛋白的表达;(2)用单丹磺酰戊二胺对乳腺癌细胞染色后,在荧光显微镜下计数自噬泡阳性细胞,并计算自噬泡阳性细胞百分比;(3)将pEGFP-LC3质粒转染至乳腺癌细胞后,在荧光显微镜下计数含绿色荧光蛋白的阳性细胞,并计算绿色荧光蛋白阳性细胞的百分比.结果:依托泊苷和顺铂均可诱导乳腺癌细胞DNA损伤和自噬,与对照组比较,药物处理组细胞的彗星长度增长(P＜0.01),p53、p21和LC3-Ⅱ蛋白表达上调.5-氮杂-2’-脱氧胞苷也可诱导乳腺癌细胞DNA损伤和自噬,与对照组比较,处理组细胞的彗星长度增长(P＜0.01),p53、p21和LC3- Ⅱ蛋白表达上调,自噬泡阳性细胞百分比和绿色荧光蛋白阳性细胞百分比上升,差异均有统计学意义(P＜0.05和P＜0.01).结论:5-氮杂-2’-脱氧胞苷可诱导乳腺癌细胞自噬,其机制可能与其所诱导的DNA损伤有关.