【作者】 肖建忠；

【导师】 贺修胜；

【作者基本信息】 南华大学 ， 病理学与病理生理学， 2007， 硕士

【摘要】 目的:鼻咽癌相关新基因STGC3是一候选抑癌基因,在Burkitt氏淋巴瘤表达下调,推测STGC3基因表达的改变,可能参与了淋巴瘤的发生发展。本实验在前期研究工作的基础上,采用电穿孔基因转染技术,将STGC3基因导入Daudi细胞系,探讨STGC3高表达,对Burkitt氏淋巴瘤Daudi细胞系生长增殖的影响,为进一步阐明Burkitt氏淋巴瘤发病机理提供实验依据。方法:应用电穿孔基因转染技术,将pCDNA3.1(+)/STGC3真核表达载体导入Daudi细胞系,经G418筛选,挑选阳性克隆,Western blotting方法,检测转染细胞中STGC3的蛋白表达,建立高表达STGC3基因的Daudi细胞系;通过转基因细胞系生长曲线绘制、软琼脂集落形成、流式细胞仪检测、免疫组织化学及HE染色等方法检查,分析STGC3基因高表达,对转基因Daudi细胞倍增时间、增殖速度、集落形成率、细胞周期及凋亡的影响。结果:在转染STGC3基因的Daudi细胞系中,Western blotting检测证实,STGC3的蛋白表达水平显著增高,免疫细胞化学结果显示STGC3定位于Daudi细胞的胞浆与胞核。与未转染的Daudi细胞及转染空载体质粒的Daudi细胞相比,转染STGC3的Daudi细胞增殖速度明显减慢(P<0.05),克隆形成率显著减低(P<0.01);流式细胞仪检测结果显示,瘤细胞群体中处于G0/G1期细胞数增加,S期比例下降,细胞阻滞于G0/G1期。结论1.成功建立高表达STGC3基因的Daudi细胞系2. STGC3基因高表达能使Daudi细胞在体外生长速度减慢,细胞群体倍增时间延长,且软琼脂集落形成率明显降低,证明STGC3基因表达增强对Daudi细胞增殖能力具有一定的抑制作用。3. STGC3基因高表达通过将Daudi细胞阻滞于G0/G1期,进而在调控细胞周期方面发挥重要作用。更多还原

【Abstract】 Objective: STGC3 gene is an anti-oncogene correlation with nasopharyngeal carcinoma. Protein of STGC3 down-expression in Burkitt lymphoma, which is probale. To explore the growth inhibition effect of STGC3 on Burkitt’s lymphoma; to provide experimental proof in elucidating the mechanism of the pathogenisis Burkitt’s lymphoma.Methods: Reconbinant expression vector pcDNA(+)3.1/STGC3 was constructed and STGC3 gene was transfected into Daudi cell line by gene transfection with G418 election.Immunohisochemistry and western blotting were used to identify the positive cell clone and a Daudi cell line highly expressed STGC3 was established. double generation time,proliferation celocity,colony formation rate,cell cycle distribution and cell apoptosis of Daudi cell line highly expressed of STGC 3 were analysed by drawing the growth curves and employing colony formation test,flow cytometry ,immunohistochemistry and HE stainning.Results : The protein level of STGC3 was high in Daudi cell line transfected with STGC3 . Compared with the Daudi cell line and Daudi transfected with the vector,the proliferationn velocity and colony formation rate of Daudi cell line transfected with STGC 3 were decreased (P<0.05), flow cytometry indicated that G0/G1 phase cell rate increased and S phase cell rate decreased and the cells were arrested in G0/G1 phase.Conclusions :1 The Daudi cell line highly expressed STGC3 was established.2 STGC3 re-expression in Daudi cell line could decreased the proliferation velocity and colony formation rate,prolong the double generation time .STGC3 expression could inhibit the proliferation of the Daudi cell line .3 STGC3 re-expression might play a key role in regulating cell cycle through arresting the cells in G0/G1 phase.更多还原