Department of Respiratory Medicine, Nagoya University Graduate School of Medicine, Nagoya 466-8550, Japan.
Am J Respir Cell Mol Biol. 2011 Oct;45(4):684-91. doi: 10.1165/rcmb.2010-0332OC. Epub 2011 Jan 28.
To investigate the effects of capsaicinoids on airway anion transporters, we recorded and analyzed transepithelial currents in human airway epithelial Calu-3 cells. Application of capsaicin (100 μM) attenuated vectorial anion transport, estimated as short-circuit currents (I(SC)), before and after stimulation by forskolin (10 μM) with concomitant reduction of cytosolic cyclic AMP (cAMP) levels. The capsaicin-induced inhibition of I(SC) was also observed in the response to 8-bromo-cAMP (1 mM, a cell-permeable cAMP analog) and 3-isobutyl-1-methylxanthine (1 mM, an inhibitor of phosphodiesterases). The capsaicin-induced inhibition of I(SC) was attributed to suppression of bumetanide (an inhibitor of the basolateral Na(+)-K(+)-2 Cl(-) cotransporter 1)- and 4,4'-dinitrostilbene-2,2'-disulfonic acid (an inhibitor of basolateral HCO(3)(-)-dependent anion transporters)-sensitive components, which reflect anion uptake via basolateral cAMP-dependent anion transporters. In contrast, capsaicin potentiated apical Cl(-) conductance, which reflects conductivity through the cystic fibrosis transmembrane conductance regulator, a cAMP-regulated Cl(-) channel. All these paradoxical effects of capsaicin were mimicked by capsazepine. Forskolin application also increased phosphorylated myosin phosphatase target subunit 1, and the phosphorylation was prevented by capsaicin and capsazepine, suggesting that these capsaicinoids assume aspects of Rho kinase inhibitors. We also found that the increments in apical Cl(-) conductance were caused by conventional Rho kinase inhibitors, Y-27632 (20 μM) and HA-1077 (20 μM), with selective inhibition of basolateral Na(+)-K(+)-2 Cl(-) cotransporter 1. Collectively, capsaicinoids inhibit cAMP-mediated anion transport through down-regulation of basolateral anion uptake, paradoxically accompanied by up-regulation of apical cystic fibrosis transmembrane conductance regulator-mediated anion conductance. The latter is mediated by inhibition of Rho-kinase, which is believed to interact with actin cytoskeleton.
为了研究辣椒素对气道阴离子转运体的影响，我们记录并分析了人气道上皮细胞 Calu-3 的跨上皮电流。应用辣椒素（100 μM）可减弱短电路电流（I（SC）），这是在 forskolin（10 μM）刺激前后的变化，同时降低细胞浆环磷酸腺苷（cAMP）水平。在对 8-溴-cAMP（1 mM，一种细胞可渗透的 cAMP 类似物）和 3-异丁基-1-甲基黄嘌呤（1 mM，一种磷酸二酯酶抑制剂）的反应中，也观察到辣椒素诱导的 I（SC）抑制。辣椒素诱导的 I（SC）抑制归因于对布美他尼（基底外侧 Na（+）-K（+）-2 Cl（-）共转运蛋白 1 的抑制剂）和 4,4'-二硝基苯-2,2'-二磺酸（基底外侧 HCO（3）（-）依赖性阴离子转运蛋白的抑制剂）敏感成分的抑制，这反映了通过基底外侧 cAMP 依赖性阴离子转运体的阴离子摄取。相比之下，辣椒素增强了顶端 Cl（-）电导，这反映了通过囊性纤维化跨膜电导调节剂（一种 cAMP 调节的 Cl（-）通道）的电导。辣椒素的所有这些矛盾作用都被辣椒素模拟。应用 forskolin 还增加了磷酸化肌球蛋白磷酸酶靶亚基 1，并且辣椒素和辣椒素模拟物可阻止磷酸化，表明这些辣椒素类具有 Rho 激酶抑制剂的某些特性。我们还发现，顶端 Cl（-）电导的增加是由传统的 Rho 激酶抑制剂 Y-27632（20 μM）和 HA-1077（20 μM）引起的，同时对基底外侧 Na（+）-K（+）-2 Cl（-）共转运蛋白 1 有选择性抑制。总之，辣椒素通过下调基底外侧阴离子摄取来抑制 cAMP 介导的阴离子转运，同时令人惊讶地伴随着顶端囊性纤维化跨膜电导调节剂介导的阴离子电导的上调。后者是由 Rho-激酶抑制介导的，据信 Rho-激酶与肌动蛋白细胞骨架相互作用。
