Carruba Giuseppe, Stefano Rosalba, Cocciadiferro Letizia, Saladino Francesca, Di Cristina Antonietta, Tokar Erik, Quader Salmann T A, Webber Mukta M, Castagnetta Luigi
Department of Experimental Oncology and Clinical Application, University Medical School, Palermo, Italy.
Ann N Y Acad Sci. 2002 Jun;963:156-68. doi: 10.1111/j.1749-6632.2002.tb04107.x.
Gap-junction-mediated intercellular communication (GJIC) is required for completion of embryonic development, tissue homeostasis, and regulation of cell proliferation and death. Although, as emphasized in several reports, defects or disruption of GJIC may be important in carcinogenesis, the potential role of GJIC in the onset and progression of human prostate cancer remains ill-defined. The gap junction channel-forming connexins (Cx) comprise a multigene family of highly conserved proteins that are differentially expressed in a tissue- and development-specific manner; changes in connexin expression are also commonly seen during cellular differentiation. However, when multiple connexins are concurrently expressed, gap junction channels may consist of more than one connexin species. This is important, because only certain pairings give rise to functional channels. In our studies, we investigated GJIC in a panel of both nontumorigenic (RWPE-1) and malignant (RWPE-2, LNCaP, DU-145) human prostate epithelial cells, compared to a normal rat liver epithelial F344 (WB-1) cell line, as it was found to be junctionally proficient. In addition, expression and regulation of Cx43 and Cx32 were also inspected using western blot analysis. The ability of hormones, antihormones, and the antihypertensive drug forskolin to restore GJIC in nontumorigenic and malignant human prostate epithelial cells was examined by the scrape-loading/dye transfer (SL/DT) or fluorescence recovery after photobleaching (FRAP) methods using an Ultima laser cytometer. Results from both assays showed that neither nontumorigenic nor malignant prostate cells have functional GJIC. However, both estrone (E1) and forskolin (FK) induced a significant increase (4.4- and 2.8-fold, respectively) in cell-cell communication only in the RWPE-1 cells. Interestingly, the use of Matrigel, a solubilized basement membrane, as substrate for cell attachment and growth resulted in the rescue of GJIC activity in RWPE-1 cells, as revealed by the SL/DT method. Furthermore, E1 induced a twofold increase in connexin 43 (Cx43), whereas forskolin caused a 50% reduction in Cx32 expression in RWPE-1 cells. These data suggest that agents that increase Cx43:Cx32 ratio may restore GJIC in junctionally deficient cells, providing a basis for the development of new strategies for the prevention and treatment of human prostate cancer.
间隙连接介导的细胞间通讯（GJIC）对于胚胎发育的完成、组织稳态以及细胞增殖和死亡的调节是必需的。尽管如几份报告所强调的，GJIC的缺陷或破坏在致癌过程中可能很重要，但GJIC在人类前列腺癌发生和发展中的潜在作用仍不明确。间隙连接通道形成蛋白（Cx）构成了一个高度保守的蛋白质多基因家族，它们以组织和发育特异性的方式差异表达；在细胞分化过程中也常见连接蛋白表达的变化。然而，当多种连接蛋白同时表达时，间隙连接通道可能由不止一种连接蛋白组成。这很重要，因为只有某些配对才能产生功能性通道。在我们的研究中，我们研究了一组非致瘤性（RWPE - 1）和恶性（RWPE - 2、LNCaP、DU - 145）人类前列腺上皮细胞中的GJIC，并与正常大鼠肝上皮F344（WB - 1）细胞系进行了比较，因为发现该细胞系具有良好的连接功能。此外，还使用蛋白质印迹分析检测了Cx43和Cx32的表达及调控。使用Ultima激光细胞仪通过刮擦加载/染料转移（SL/DT）或光漂白后荧光恢复（FRAP）方法，研究了激素、抗激素和抗高血压药物福斯可林恢复非致瘤性和恶性人类前列腺上皮细胞中GJIC的能力。两种检测方法的结果均显示，非致瘤性和恶性前列腺细胞均无功能性GJIC。然而，仅在RWPE - 1细胞中，雌酮（E1）和福斯可林（FK）均显著诱导了细胞间通讯的增加（分别增加4.4倍和2.8倍）。有趣的是，使用基质胶（一种溶解的基底膜）作为细胞附着和生长的底物，如SL/DT方法所示，可挽救RWPE - 1细胞中的GJIC活性。此外，E1诱导RWPE - 1细胞中连接蛋白43（Cx43）增加两倍，而福斯可林导致Cx32表达降低50%。这些数据表明，增加Cx43:Cx32比值的药物可能恢复连接功能缺陷细胞中的GJIC，为开发预防和治疗人类前列腺癌的新策略提供了依据。
