转染连接蛋白基因对人脑胶质瘤细胞间隙连接通讯及其生长变化的研究

目的 研究连接蛋白（Cx)基因对人脑胶质瘤细胞的细胞间隙连接通讯（GJIC）及其增殖的抑制作用,探索以Cx43基因治疗胶质瘤的可行性。方法 将含Cx43cDNA的质粒,以脂质体介导转染Cx43表达缺失的TJ905人胶质母细胞瘤细胞,通过Northern印染杂交、原位杂交及免疫组化染色检测Cx43mRNA及蛋白表达,划痕标记荧光染料示踪技术（SLDT）检测GJIC,MTT法测定细胞增殖率,核仁组成区嗜银蛋白（AgNOR)染色检测细胞增殖活性,TUNEL法检测细胞凋亡。结果 转染后TJ905细胞有不同程度的Cx43mRNA和蛋白表达及GJIC恢复。Cx43表达水平高的克隆细胞增殖明显下降,细胞凋亡并未增加。结论 Cx43基因及GJIC在恶性胶质瘤的发生发展过程中起重要作用,可能成为恶性胶质瘤基因治疗的优选靶的之一。

Effect of transfected Cx43 gene on the gap junction intercellular communication and the human glioma cells proliferation

Objective To investigate the role of connexin 43 (Cx43) gene in the gap junction intercellular communication (GJIC) and the development of glioma so as to demonstrate the feasibility of using Cx43cDNA as the target of gene therapy for gliomas.Methods TJ905 human glioblastoma cell line without Cx43 gene expression was transfected with Cx43cDNA mediated by lipofectamine. Northern blot, in situ hybridization and immunocytochemistry were used for examination of Cx43mRNA and its protein examination; MTT assay and silver colloid stain were used to detect cell proliferation. TUNEL method was used to determine the cell apoptosis. Scrape loading dye transfer (SLDT) was used to monitor GJIC.Results The TJ905 transfectants resulted in dramatic upregulation of Cx43mRNA and its protein, with the GJIC of transfected glioma cells restored. Clones with high Cx43 expression also showed reduced proliferation in vitro. However, the cell apoptosis did not increase following the transfection with Cx43 gene.Conclusion These results indicate that Cx gene and gap junotion intercellular communication are of crucial importance in malignant glioma cell growth and development. Cx43 gene may be a target for gene therapy of gliomas.