MCPH1在肺组织的表达及其对人肺癌细胞株H1299凋亡的影响.docVIP

MCPH1在肺组织的表达及其对人肺癌细胞株H1299凋亡的影响 张冀1，麦力 1，吴晓彬1，肖明1，袁成福1，卜友泉1,2，宋方洲1,2 （400016 重庆 重庆医科大学分子医学与肿瘤研究中心1；400016 重庆 重庆医科大学生物化学与分子生物学教研室2） [摘要] 目的 探讨MCPH1基因在肺癌组织和正常组织的蛋白表达分布及其过表达后对人肺腺癌细胞系H1299凋亡的影响。方法 临床收集20例肺癌组织和8例正常肺组织标本，免疫组化法检测MCPH1蛋白在肺组织的表达分布，真核表达质粒pcDNA3.1(－)/MCPH1和空白质粒pcDNA3. 1瞬时转染肺癌细胞株H1299，并设立未处理组。转染质粒48h后，采用Real-time PCR法检测细胞中MCPH1基因mRNA的转录表达情况；转染72h后提取细胞总蛋白，Western blot检测细胞中MCPH1蛋白表达水平，并进行流式细胞术检测细胞凋亡的情况。结果 MCPH1蛋白在肺癌组织中表达低于正常组织，而且MCPH1蛋白在癌组织中核质低表达，在正常组织中为核质高表达；H1299转染pcDNA3.1(－)/MCPH1重组质粒后，可有效上调H1299细胞中MCPH1基因的 mRNA和蛋白表达，而处理组的细胞凋亡率(18.10±1.87)％较阴性对照组(5.76±1.85)％和未处理组(5.85±0.57)％显著增加(P＜0. 05)。结论 本实验观察到MCPH1在人肺癌组织表达降低，过表达之后可诱导肺腺癌细胞发生凋亡这一现象值得进行机制探讨。ung tissues and its effect of over-expression of MCPH1 on apoptosis of Human lung cancer cells ZHANG Ji1, MAI Li1, WU Xiao-Bin1, XIAO Ming1, YUAN Chen-Fu1, BU You-Quan1,2, SONG Fang-Zhou1,2 (1Molecular Medicine Cancer Research Center，Chongqing Medical University, Chongqing, China; 2Department of Biochemistry Molecular Biology, Chongqing Medical University, Chongqing, China) [Abstract] Objective To evaluated the expression of protein MCPH1 in lung tissues and investigate the effect of over-expression of MCPH1 gene on apoptosis of Human lung cancer cells. Methods We evaluated the expression of protein MCPH1 in 20 lung?cancer?and 8 normal lung tissues by immunohistochemistry; Eukaryotic expression plasmid pcDNA3.1(－)/MCPH1 and blank plasmid pcDNA3.1 were transfected to lung cells H1299, and using the cells untransfected as control. 48 hours after transfection，the cells were detected for transcription and expression of MCPH1 mRNA by real-time PCR; 72 hours after transfection, the total protein was extracted, the MCPH1 protein were determined by Western blot，and the apoptosis of H1299 cells were determined by flow cytometry. Results Compared with normal tissues, MCPH1 expression showed much lower in lung cancer tissues, and MCPH1 was expressed mainly in nucleus in normal tissues. Both the expressions of MCPH1 mRNA and prote