Department of Pharmacology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore.
J Pharmacol Exp Ther. 2011 Nov;339(2):726-35. doi: 10.1124/jpet.111.184754. Epub 2011 Aug 24.
Intracellular pH (pH(i)) is an important endogenous modulator of cardiac function. Inhibition of Na(+)/H(+) exchanger-1 (NHE-1) protects the heart by preventing Ca(2+) overload during ischemia/reperfusion. Hydrogen sulfide (H(2)S) has been reported to produce cardioprotection. The present study was designed to investigate the pH regulatory effect of H(2)S in rat cardiac myocytes and evaluate its contribution to cardioprotection. It was found that sodium hydrosulfide (NaHS), at a concentration range of 10 to 1000 μM, produced sustained decreases in pH(i) in the rat myocytes in a concentration-dependent manner. NaHS also abolished the intracellular alkalinization caused by trans-(±)-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)-cyclohexyl]benzeneacetamide methane-sulfonate hydrate (U50,488H), which activates NHEs. Moreover, when measured with an NHCl(4) prepulse method, NaHS was found to significantly suppress NHE-1 activity. Both NaHS and cariporide or [5-(2-methyl-5-fluorophenyl)furan-2-ylcarbonyl]guanidine (KR-32568), two NHE inhibitors, protected the myocytes against ischemia/reperfusion injury. However, coadministration of NaHS with KR-32568 did not produce any synergistic effect. Functional study showed that perfusion with NaHS significantly improved postischemic contractile function in isolated rat hearts subjected to ischemia/reperfusion. Blockade of phosphoinositide 3-kinase (PI3K) with 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one (LY294002), Akt with Akt VIII, or protein kinase G (PKG) with (9S,10R,12R)-2,3,9,10,11,12-hexahydro-10-methoxy-2,9-dimethyl-1-oxo-9,12-epoxy-1H-diindolo[1,2,3-fg:3',2',1'-kl]pyrrolo[3,4-i][1,6]]enzodiazocine-10-carboxylic acid, methyl ester (KT5823) significantly attenuated NaHS-suppressed NHE-1 activity and/or NaHS-induced cardioprotection. Although KT5823 failed to affect NaHS-induced Akt phosphorylation, Akt inhibitor did attenuate NaHS-stimulated PKG activity. In conclusion, this work demonstrated for the first time that H(2)S produced cardioprotection via the suppression of NHE-1 activity involving a PI3K/Akt/PKG-dependent mechanism.
细胞内 pH 值（pH(i)）是心脏功能的重要内源性调节剂。在缺血/再灌注过程中，抑制钠/氢交换蛋白-1（NHE-1）可防止钙超载，从而起到保护心脏的作用。已有研究表明，硫化氢（H(2)S）可产生心脏保护作用。本研究旨在探讨 H(2)S 在大鼠心肌细胞中的 pH 调节作用，并评估其对心脏保护的贡献。结果发现，浓度范围在 10 到 1000 μM 的硫氢化钠（NaHS）可浓度依赖性地持续降低大鼠心肌细胞内的 pH(i)。NaHS 还消除了 trans-(±)-3,4-二氯-N-甲基-N-[2-(1-吡咯烷基)-环己基]苯乙酰胺甲磺酸盐水合物（U50,488H）引起的细胞内碱化，而 U50,488H 可激活 NHEs。此外，通过 NHCl(4) 脉冲前处理法检测发现，NaHS 可显著抑制 NHE-1 活性。NaHS 和 NHE 抑制剂 cariporide 或 [5-(2-甲基-5-氟苯基)呋喃-2-羰基]胍（KR-32568）均可保护心肌细胞免受缺血/再灌注损伤。然而，NaHS 与 KR-32568 联合给药并未产生协同作用。功能研究表明，NaHS 灌注可显著改善缺血/再灌注后分离的大鼠心脏的缺血后收缩功能。用 2-(4-吗啉基)-8-苯基-4H-1-苯并吡喃-4-酮（LY294002）抑制磷酸肌醇 3-激酶（PI3K）、用 Akt VIII 抑制 Akt 或用 (9S,10R,12R)-2,3,9,10,11,12-六氢-10-甲氧基-2,9-二甲基-1-氧代-9,12-环氧-1H-二吲哚[1,2,3-fg:3',2',1'-kl]吡咯并[3,4-i][1,6]]并恶嗪-10-羧酸甲酯（KT5823）抑制 NHE-1 活性和/或 NaHS 诱导的心脏保护作用，可显著减弱 NaHS 的作用。虽然 KT5823 未能影响 NaHS 诱导的 Akt 磷酸化，但 Akt 抑制剂可减弱 NaHS 刺激的蛋白激酶 G（PKG）活性。综上所述，本研究首次证明 H(2)S 通过抑制 NHE-1 活性发挥心脏保护作用，涉及 PI3K/Akt/PKG 依赖性机制。
