The H₁-H₂ domain of the 伪₁ isoform of Na⁺-K⁺-ATPase is involved in ouabain toxicity in rat ventricular myocytes

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• 作者：Chen Xiong ; Jun-xia Li ; Hui-cai Guo ; Li-nan Zhang ; Wei Guo ; Jing Meng ; Yong-li Wang ; ^{wangyongli@gmail.com}
• 关键词：Na+-K+-ATPase ; [Ca2+]i ; Mitochondrial ; Ventricular myocytes ; Ouabain ; Toxicity
• 刊名：Toxicology and Applied Pharmacology
• 出版年：2012
• 期刊代码：58_0041008x
• 类别：pha
• 出版时间：1 July, 2012
• 卷：262
• 期：1
• 页码：32-42
• 文件大小：1121 K

摘要

The composition of different isoforms of Na⁺-K⁺-ATPase (NKA, Na/K pump) in ventricular myocytes is an important factor in determining the therapeutic effect and toxicity of cardiac glycosides (CGs) on heart failure. The mechanism whereby CGs cause these effects is still not completely clear. In the present study, we prepared two site-specific antibodies (SSA78 and WJS) against the H₁-H₂ domain of 伪₁ and 伪₂ isoforms of NKA in rat heart, respectively, and compared their influences on the effect of ouabain (OUA) in isolated rat ventricular myocytes. SSA78 or WJS, which can specifically bind with the 伪₁ or 伪₂ isoform, were assessed with enzyme linked immunosorbent assay (ELISA), Western blot and immunofluorescent staining methods. Preincubation of myocytes with SSA78 inhibited low OUA affinity pump current but not high OUA affinity pump current, reduced the rise in cytosolic calcium concentration ([Ca²⁺]_i), attenuated mitochondrial Ca²⁺ overload, restored mitochondrial membrane potential reduction, and delayed the decrease of the myocardial contractile force as well as the occurrence of arrhythmic contraction induced by high concentrations (1 mM) but not low concentrations (1 渭M) of OUA. Similarly, preincubation of myocytes with WJS inhibited high OUA affinity pump current, reduced the increase of [Ca²⁺]_i and the contractility induced by 1 渭M but not that induced by 1 mM OUA. These results indicate that the H₁-H₂ domain of the NKA 伪₁ isoform mediates OUA-induced cardiac toxicity in rat ventricular myocytes, and inhibitors for this binding site may be used as an adjunct to CGs treatment for cardiovascular disease.