Eid A H, Chotani M A, Mitra S, Miller T J, Flavahan N A
Lebanese International University, Beirut, Lebanon.
Am J Physiol Heart Circ Physiol. 2008 Jul;295(1):H266-72. doi: 10.1152/ajpheart.00084.2008. Epub 2008 May 16.
Cold increases cutaneous vasoconstriction by unmasking the contractile activity of alpha(2C)-adrenoceptors (alpha(2C)-ARs) in vascular smooth muscle cells (VSMCs), which is mediated by the cold-induced mobilization of alpha(2C)-ARs from the transGolgi to the cell surface. The expression of alpha(2C)-ARs in human cutaneous VSMCs is under dual regulation by cyclic AMP: gene transcription is inhibited by cyclic AMP acting through protein kinase A but is increased by cyclic AMP acting through the exchange protein directly activated by cyclic AMP (EPAC) and the GTP-binding protein Rap1. Experiments were performed to further characterize the Rap1 signaling pathway. Forskolin (10 muM), the selective EPAC activator, 8-pCPT-2'-O-Me-cyclic AMP (CMC; 100 microM), or a constitutively active mutant of Rap1 (Rap1CA) increased the activity of c-Jun NH(2)-terminal kinase (JNK) in human cutaneous VSMCs. This was associated with the increased phosphorylation of c-Jun and activation of an activator protein (AP)-1 reporter construct, which were inhibited by the JNK inhibitor SP600125 (3 microM). Rap1CA increased the activity of an alpha(2C)-AR promoter-reporter construct, which was inhibited by SP600125 (3 microM) or by the mutation of an AP-1 binding site in the alpha(2C)-AR promoter. Furthermore, forskolin (10 microM) or CMC (100 microM) increased the expression of the alpha(2C)-AR protein, and these effects were inhibited by SP600125 (3 microM). Therefore, cyclic AMP increases the expression of alpha(2C)-ARs in cutaneous VSMCs by activating a novel Rap1 signaling pathway, mediated by the activation of JNK, AP-1, and the subsequent transcriptional activation of the alpha(2C)-AR gene. By increasing the expression of cold-responsive alpha(2C)-ARs, this pathway may contribute to enhanced cold-induced vasoconstriction in the cutaneous circulation, including Raynaud's phenomenon.
寒冷通过揭示血管平滑肌细胞（VSMC）中α(2C)-肾上腺素能受体（α(2C)-ARs）的收缩活性来增加皮肤血管收缩，这是由寒冷诱导的α(2C)-ARs从反式高尔基体向细胞表面的转运介导的。人皮肤VSMC中α(2C)-ARs的表达受环磷酸腺苷（cAMP）的双重调节：通过蛋白激酶A起作用的cAMP抑制基因转录，但通过环磷酸腺苷直接激活的交换蛋白（EPAC）和GTP结合蛋白Rap1起作用的cAMP则增加基因转录。进行了实验以进一步表征Rap1信号通路。福斯可林（10 μM）、选择性EPAC激活剂8-pCPT-2'-O-Me-环磷酸腺苷（CMC；100 μM）或Rap1的组成型活性突变体（Rap1CA）增加了人皮肤VSMC中c-Jun氨基末端激酶（JNK）的活性。这与c-Jun磷酸化增加和激活蛋白（AP）-1报告基因构建体的激活有关，而这两者均被JNK抑制剂SP600125（3 μM）抑制。Rap1CA增加了α(2C)-AR启动子-报告基因构建体的活性，该活性被SP600125（3 μM）或α(2C)-AR启动子中AP-1结合位点的突变抑制。此外，福斯可林（10 μM）或CMC（100 μM）增加了α(2C)-AR蛋白的表达，且这些作用被SP600125（3 μM）抑制。因此，cAMP通过激活由JNK、AP-1激活介导的新的Rap1信号通路以及随后α(2C)-AR基因的转录激活来增加皮肤VSMC中α(2C)-ARs的表达。通过增加冷反应性α(2C)-ARs的表达，该通路可能有助于增强皮肤循环中包括雷诺现象在内的寒冷诱导的血管收缩。
