SODIUM PUMP INHIBITORS IN BLOOD PRESSURE REGULATION

我们以前的研究表明，两种内源性钠泵配体（SPL），哇巴因样化合物和海蟾毒配基，在哺乳动物组织中共存。与先前描述的SPL不同，MBG作为Na/K-ATP酶（NKA）的哇巴因抗性α-1同种型的选择性抑制剂，所述Na/K-ATP酶（NKA）是肾脏、血管平滑肌和成人心肌细胞中的主要钠泵同种型。在过去的一年中，我们继续研究SPL在Dahl盐敏感大鼠（DS），其中α-1 NKA的突变和SPL的代偿反应的基础上发展的NaCl敏感性高血压。我们首次证明，HPLC纯化的高血压DS内源性MBG免疫反应性物质有效抑制肾钠泵（α-1亚型），而不是α-3 NKA亚型（胎鼠脑）。此外，在体内施用针对DS的MBG抗体阻断了与NaCl负荷相关的血压升高、利钠反应和肾钠泵抑制。因此，MBG满足Koch假定的钠尿激素参与盐敏感性高血压的发病机制。以前，我们已经表明，蛋白激酶C（PKC）激活导致NKA对SPL的敏感性增强。我们的研究结果表明，上述增效作用的发生是由于诱导调节磷酸化的NKA α-1亚基。这种PKC诱导的NKA磷酸化对α-1（相对于α-3）NKA同种型具有特异性。由于SPL和PKC对NKA活性相互作用，NKA和PKC均参与肥大信号传导，因此我们研究了PKC和MBG对α-1 NKA的调节相互作用是否可能成为Dahl高血压治疗干预的靶点。高NaCl摄入DS患者代偿性左心室肥厚（LVH）的发生与LV心肌中α-1 NKA、PKC β-II和δ的上调相关。上调的α-1 NKA表现出对MBG的敏感性增强。用直接抑制PKC的抗高血压化合物西来他宁预处理DS，可降低血压和LV重量，改善LV性能。LV α-1 NKA仍上调，但对MBG的敏感性较低。与此同时，西来他宁治疗阻止了左室肌膜中PKC β-II和δ的上调，因此，α-1 NKA的内源性配体MBG是Dahl高血压发病机制中的重要因素。PKC对α-1 NKA的磷酸化增强了MBG的作用。因此，SPL和PKC β-II和δ对心血管组织中NKA的调节相互作用是NaCl敏感性高血压的新的治疗靶点。

SUMMARY OF WORK Our previous studies demontrated that two endogenous sodium pump ligands (SPL), ouabain-like compound and marinobufagenin, coexist in mammalian tissues. Unlike previously described SPL, MBG acts as a selective inhibitor of ouabain resistant alpha-1 isoform of Na/K-ATPase (NKA), the main sodium pump isoform in the kidney, vascular smooth muscle and adult myocardiocytes. During the last year, we continued studies of SPL in Dahl salt-sensitive rats (DS), in which the mutation of alpha-1 NKA and compensatory responses of SPL underlie development of NaCl sensitive hypertension. For the first time, we have demonstrated that HPLC purified endogenous MBG immunoreactive material from hypertensive DS potently inhibits the renal sodium pump (alpha-1 isoform) rather than alpha-3 NKA isoform (fetal rat brain). Further, in vivo administration of MBG antibody to DS blocked the elevations of blood pressure, natriuretic response and inhibition of renal sodium pump associated with NaCl loading. Thus, MBG fulfils the Koch's postulate for a putative natriuretic hormone involved in the pathogenesis of salt-sensitive hypertension. Previously, we have shown that protein kinase C (PKC) activation leads to enhanced sensitivity of the NKA to SPL. Our results demonstrate that the above potentiation occurs due to induction of regulatory phosphorylation of the NKA alpha-1 subunit. This PKC induced NKA phosphorylation is specific towards alpha-1 (vs. alpha-3) NKA isoform. Since both SPL and PKC interact on NKA activity, and both NKA and PKC are involved in hypertrophic signaling, we studied whether or not the modulatory interaction of PKC and MBG on alpha-1 NKA may be a target for therapeutic intervention in Dahl hypertension. The development of compensatory left ventricular hypertrophy (LVH) in DS on a high NaCl intake was associated with an upregulation of alpha-1 NKA, and PKC beta-II and delta in LV myocardium. Upregulated alpha-1 NKA exhibited enhanced sensitivity to MBG. Pretreatment of DS with cicletanine, an antihypertensive compound which directly inhibits the PKC, resulted in a decrease in blood pressure and LV weight and improvement of LV performance. LV alpha-1 NKA remained upregulated but exhibited less sensitivity to MBG. In parallel, cicletanine treatment prevented the upregulation of PKC beta-II and delta in LV sarcolemma.Thus, an endogenous ligand of alpha-1 NKA, MBG, is an important factor in pathogenesis of Dahl hypertension. Phosphorylation of alpha-1 NKA by PKC potentiates the effects of MBG. Therefore, a modulatory interaction of SPL and PKC beta-II and delta on NKA in cardiovascular tissues is a novel therapeutic target in NaCl sensitive hypertension.