Elucdidation of physiological and pathophysiological roles of Na^+/Ca^<2+> exchanger using drug discovery and genetic engineering

Na ^+/Ca ^2+交换器（NCX）是一种膜蛋白，它能用3~4个Na ^+交换1个Ca ^2+。哺乳动物NCX形成同源蛋白质的多基因家族，其包括三种同种型，NCX1、NCX2和NCX3。NCX1在心脏、脑和肾中高度表达，在许多其他组织中以低得多的水平表达，而NCX2和NCX3的表达主要限于脑。在这项研究中，我们试图阐明NCX的生理和病理生理作用的NCX抑制剂和NCX基因工程小鼠。通过表征NCX抑制剂，我们发现KB-R7943和SN-6分别优先阻断NCX3和NCX1，并且都特异性地抑制Na ^+/Ca ^<2 +>交换的反向模式。对NCX1/NCX3嵌合体的分析表明，这两种抑制剂可能与NCX分子上的不同位点相互作用。最近的数据表明，NCX抑制剂的效力与高Na ^+ i_1浓度引起的NCX失活程度有关。我们以前报道过NCX抑制剂可以改善缺血/再灌注引起的肾损伤。在这项研究中，我们分析了缺血/再灌注诱导的肾损伤的NCX1基因敲除小鼠（杂合子）。与野生型小鼠相比，杂合子小鼠中缺血/再灌注诱导的肾功能障碍明显减弱。这些结果表明，通过NCX1的反向模式的Ca ^<2 +>超载可能在缺血/再灌注诱导的肾功能衰竭的发病机制中起重要作用。

The Na^+/Ca^<2+> exchanger (NCX) is a membrane protein that exchanges 3 to 4 Na^+ ions for each Ca^<2+> ion. The mammalian NCX forms a multigene family of homologous proteins comprising three isoforms, NCX1, NCX2, and NCX3. NCX1 is highly expressed in the heart, brain, and kidney and expressed at much lower levels in many other tissues, whereas the expression of NCX2 and NCX3 is limited mainly to the brain. In this study, we tried to elucidate physiological and pathophysiological roles of NCX using NCX inhibitors and NCX-engineered mice. From characterizing NCX inhibitors, we found that KB-R7943 and SN-6 preferentially block NCX3 and NCX1, respectively, and both specifically inhibit the reverse mode of Na^+/Ca^<2+> exchange. The following analyses of NCX1/NCX3 chimeras revealed that these two inhibitors might interact with different sites on NCX molecule. More recent data have shown that the potency of NCX inhibitors is related to the extent of NCX inactivation caused by high Na^+i_1 concentration. We have reported before that NCX inhibitors can improve the ischemia/reperfusion-induced renal injury. In this study, we analyzed the ischemia/reperfusion-induced renal injury in NCX1-knockout mice (heterozygous). Expectedly, the ischemia/reperfusion-induced renal dysfunction in the heterozygous mice was markedly attenuated compared with cases in the wild-type mice. These results suggest that Ca^<2+> overload via the reverse mode of NCX1 seems to play an important role in the pathogenesis of the ischemia/reperfusion-induced renal failure.