维甲酸对人子宫颈癌细胞系HeLa细胞间隙连接蛋白

目的 探讨维甲酸对人子宫颈癌细胞系HeLa细胞间隙连接蛋白（Cx)43信号转导途径的调控作用。方法 应用特异性钙指示剂（Fluo-3 AM)在激光扫描共焦显微镜下动态观察维甲酸作用后的细胞质内信号转导分子游离钙的分布及强度变化。采用充式细胞仪、结合蛋白印迹技术分析，检测外源信号分子对Cx43的表达以及蛋白酪氨酸磷化状态的影响。结果 HeLa细胞内游离钙经维甲酸作用后明显超载，细胞内游离钙浓度（[Ca2 ]i）由静息状态下的35.73μmol/L。流式细胞仪分析，Cx43阳性细胞表达率由1.9%上升至26.3%。蛋白印迹技术分析，HeLa细胞出现Cx43酪氨酸磷化。结论 维甲酸对HeLa细胞Cx43信号转导途径的调控是在细胞质内游离钙的参与下，使Cx43阳性细胞表达率上升，并在酪氨酸位点出现明显的磷酸化用用下实现的。

[关键词] 连接蛋白43；钙；磷酰化；宫颈肿瘤；维甲酸；HeLa细胞

Pho horylation of gap junction co exin 43 protein and di ociated calcium in HeLa cell line by alltra -retinoic acid CHEN BiLiang,MA Xiangdong,WANG Detang,et al. Department of O tetric and Gynecology ,Xijing Ho ital,Fourth Military Medical Universiry,Xi\'an 710032,China

[A tract]Objective To investage the regulation effect on signal tra duction pathway of gap junction gene co exin (Cx)43 in human cervical carcinoma cell line Hela by all -tra -retinoic acid(ATRA).Methods Cell culture,Fluo-3 AM loading and laser sca ing ceonfocal microscope,flowcytometer(FCM)and western blotwere employed to detect expre ion and regulation effect on signal tra duction pathway of gap junction gene co exin Cx43 in HeLa cells by retiontic acid(RA),Results After treated with ATRA,the intercellular second me enger di ociated calcium ([Ca2 ]i)waw much higher in HeLa cells(58.16 μmol/L)than untreated cell (35.73% in RA-treated cells than untreated cell examined by FCM and westem blot .Immunoblot analysis showed that only the treated cells had pho horylated form of Cx43 protein.Conclusion The anti-tumor effect of ATRA in HeLa cells might be due to up-regulation of Cx43 protein .Conchusion The anti-tumor effect of ATRA in HeLa cells might be due to up-regulation of Cx43 gene and its signal tra duction tra duction pathway which mediats gap junction intercellular cimmunication.

[Key words]Co exin43; Calcium ho horylatio Cervix neoplasm Tretinoi HeLa cells

间隙连接蛋白（co exin,Cx)是相邻细胞质膜上的整合膜蛋白，为细胞间通讯提供信号的传递的特殊跨膜亲水性通道。近年业研究发现，Cx基因不达缺失与多种肿瘤的发生密切相关，被认为是非突变型肿瘤抑制基因。其中，Cx43在人子宫颈癌组织和人子宫颈癌细胞系HeLa中表达显著降低或缺失，可能与Cx基因信号转导途径异常明显相关，本工作在此方面进行了研究。