Ruamyod Katesirin, Watanapa Wattana B, Shayakul Chairat
Department of Physiology Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, 10700, Thailand.
Department of Medicine, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, 10700, Thailand.
J Steroid Biochem Mol Biol. 2017 Apr;168:118-126. doi: 10.1016/j.jsbmb.2017.02.014. Epub 2017 Feb 20.
Testosterone has endothelium-dependent vasodilatory effects on the coronary artery, with some reports suggesting endothelial ion channel involvement. This study employed the whole-cell patch clamp technique to investigate the effect of testosterone on ion channels in human coronary artery endothelial cells (HCAECs) and the mechanisms involved. We found that 0.03-3μM testosterone significantly induced a rapid, concentration-dependent increase in total HCAEC current (EC, 71.96±1.66nM; maximum increase, 59.13±8.37%; mean±SEM). The testosterone-enhanced currents consisted of small- and large-conductance Ca-activated K currents (SK and BK currents), but not Cl and nonselective cation currents. Either a non-permeant testosterone conjugate or the non-aromatizable androgen dihydrotestosterone (DHT) could increase HCAEC currents as well. The androgen receptor antagonist flutamide prevented this testosterone, testosterone conjugate, and DHT effect, while the estrogen receptor antagonist fulvestrant did not. Incubating HCAECs with pertussis toxin or protein kinase A inhibitor H-89 largely inhibited the testosterone effect, while pre-incubation with phospholipase C inhibitor U-73122, prostacyclin inhibitor indomethacin, nitric oxide synthase inhibitor L-NAME or cytochrome P450 inhibitor MS-PPOH, did not. Finally, testosterone application induced HCAEC hyperpolarization within minutes; this effect was prevented by SK and BK current inhibitors apamin and iberiotoxin. This is the first electrophysiological demonstration of androgen-induced K current increase, leading to hyperpolarization, in any endothelial cell, and the first report of SK as a testosterone target. Our data show that testosterone rapidly increased whole-cell HCAEC SK and BK currents via a surface androgen receptor, G protein, and protein kinase A. This mechanism may explain rapid testosterone-induced coronary vasodilation seen in vivo.
睾酮对冠状动脉具有内皮依赖性血管舒张作用，一些报告表明内皮离子通道参与其中。本研究采用全细胞膜片钳技术，研究睾酮对人冠状动脉内皮细胞（HCAECs）离子通道的影响及其相关机制。我们发现，0.03 - 3μM睾酮显著诱导HCAEC总电流快速、浓度依赖性增加（EC，71.96±1.66nM；最大增加，59.13±8.37%；平均值±标准误）。睾酮增强的电流由小电导和大电导钙激活钾电流（SK和BK电流）组成，但不包括氯电流和非选择性阳离子电流。非渗透性睾酮共轭物或不可芳香化的雄激素二氢睾酮（DHT）也可增加HCAEC电流。雄激素受体拮抗剂氟他胺可阻止这种睾酮、睾酮共轭物和DHT的作用，而雌激素受体拮抗剂氟维司群则不能。用百日咳毒素或蛋白激酶A抑制剂H - 89孵育HCAECs可大大抑制睾酮的作用，而预先用磷脂酶C抑制剂U - 73122、前列环素抑制剂吲哚美辛、一氧化氮合酶抑制剂L - NAME或细胞色素P450抑制剂MS - PPOH孵育则无此作用。最后，应用睾酮在数分钟内诱导HCAEC超极化；SK和BK电流抑制剂蜂毒明肽和埃博毒素可阻止这种作用。这是首次在任何内皮细胞中通过电生理学证明雄激素诱导钾电流增加并导致超极化，也是首次报道SK是睾酮的作用靶点。我们的数据表明，睾酮通过表面雄激素受体、G蛋白和蛋白激酶A快速增加HCAEC全细胞SK和BK电流。这一机制可能解释了体内观察到的睾酮快速诱导的冠状动脉血管舒张。
