Nihei Oscar K, Fonseca Paula C, Rubim Nara M, Bonavita Andre G, Lyra Jurandy S P O, Neves-dos-Santos Sandra, de Carvalho Antonio C Campos, Spray David C, Savino Wilson, Alves Luiz A
Laboratory of Cellular Communication, Oswaldo Cruz Institute, The Oswaldo Cruz Foundation, Rio de Janeiro, Brazil.
BMC Cell Biol. 2010 Jan 15;11:3. doi: 10.1186/1471-2121-11-3.
We investigated the effects of the signaling molecules, cyclic AMP (cAMP) and protein-kinase C (PKC), on gap junctional intercellular communication (GJIC) between thymic epithelial cells (TEC).
Treatment with 8-Br-cAMP, a cAMP analog; or forskolin, which stimulates cAMP production, resulted in an increase in dye transfer between adjacent TEC, inducing a three-fold enhancement in the mean fluorescence of coupled cells, ascertained by flow cytometry after calcein transfer. These treatments also increased Cx43 mRNA expression, and stimulated Cx43 protein accumulation in regions of intercellular contacts. VIP, adenosine, and epinephrine which may also signal through cyclic nucleotides were tested. The first two molecules did not mimic the effects of 8-Br-cAMP, however epinephrine was able to increase GJIC suggesting that this molecule functions as an endogenous inter-TEC GJIC modulators. Stimulation of PKC by phorbol-myristate-acetate inhibited inter-TEC GJIC. Importantly, both the enhancing and the decreasing effects, respectively induced by cAMP and PKC, were observed in both mouse and human TEC preparations. Lastly, experiments using mouse thymocyte/TEC heterocellular co-cultures suggested that the presence of thymocytes does not affect the degree of inter-TEC GJIC.
Overall, our data indicate that cAMP and PKC intracellular pathways are involved in the homeostatic control of the gap junction-mediated communication in the thymic epithelium, exerting respectively a positive and negative role upon cell coupling. This control is phylogenetically conserved in the thymus, since it was seen in both mouse and human TEC preparations. Lastly, our work provides new clues for a better understanding of how the thymic epithelial network can work as a physiological syncytium.
我们研究了信号分子环磷酸腺苷（cAMP）和蛋白激酶C（PKC）对胸腺上皮细胞（TEC）间缝隙连接细胞间通讯（GJIC）的影响。
用cAMP类似物8-溴-cAMP或刺激cAMP产生的福斯可林处理后，相邻TEC之间的染料转移增加，通过钙黄绿素转移后流式细胞术确定，偶联细胞的平均荧光增强了三倍。这些处理还增加了Cx43 mRNA表达，并刺激了细胞间接触区域的Cx43蛋白积累。对可能也通过环核苷酸信号传导的血管活性肠肽（VIP）、腺苷和肾上腺素进行了测试。前两种分子没有模拟8-溴-cAMP的作用，然而肾上腺素能够增加GJIC，表明该分子作为一种内源性TEC间GJIC调节剂发挥作用。佛波醇-肉豆蔻酸酯-乙酸酯刺激PKC抑制了TEC间GJIC。重要的是，在小鼠和人TEC制剂中均观察到分别由cAMP和PKC诱导的增强和减弱作用。最后，使用小鼠胸腺细胞/TEC异细胞共培养的实验表明，胸腺细胞的存在不影响TEC间GJIC的程度。
总体而言，我们的数据表明，cAMP和PKC细胞内途径参与胸腺上皮中缝隙连接介导的通讯的稳态控制，分别对细胞偶联发挥正向和负向作用。这种控制在胸腺中具有系统发育保守性，因为在小鼠和人TEC制剂中均可见到。最后，我们的工作为更好地理解胸腺上皮网络如何作为一个生理合体发挥作用提供了新线索。
