Na+−H+ exchanger as a target for intervention in cardiovascular remodelling

During the last two decades, several laboratories have reported that enhanced permeability for Na+, initially discovered as an enhanced rate of isotope transmembrane exchange in aortic strips and erythrocytes from spontaneously hypertensive rats (SHR), is at least partially caused by augmented amiloride-sensitive Na+/H+ exchange (NHE) [1–3]. However, in spite of more than 200 publications on NHE and hypertension cited by the NIH library, two major questions remain unanswered. First, which mechanisms underline the increased activity of this carrier detected in blood cells from patients with essential hypertension and in blood cells, renal epithelial cells, cultured vascular smooth muscle cells (VSMC) and cardiomyocytes from SHR? Second, does elevated NHE contribute to the maintenance of high blood pressure or/and the development of renal and cardiovascular complications of this disease? Aviv was probably the first to propose that NHE increases vascular tone via heightened [Ca2+]i as a consequence of elevation of [Na+]i and activation of Na+i/Ca2+o exchange [4]. However, it should be stressed that, by contrast to other monovalent ion transporters, such as Na+,K+-ATPase, Na+,K+,Cl− cotransport and K+ channels, to date, there are no data showing modulation of vascular tone by selective NHE inhibitors. Moreover, the presence of the calmodulin-binding domain in the C-terminus of ubiquitous (NHE1) and several other NHE isoforms [5,6] suggests that activation of this carrier might be a consequence of elevated [Ca2+]i triggered by abnormal activity of Ca2+ transport systems or enhanced efficiency of signalling evoked by angiotensin II and other [Ca2+]i-raising vasoconstrictors. Numerous studies performed during the last decade have demonstrated that, irrespective of animal species and experimental design, NHE inhibitors protect the myocardium from ischemic reperfusion injury [2]. In the current issue of the journal, Ji et al. [7] report that amiloride treatment decreased left ventricular hypertrophy (LVH) in SHR. These data suggest the involvement of augmented NHE activity in the development of cardiovascular remodelling observed in hypertension, and also highlight several unresolved issues. First, the relative contributions of amiloride-sensitive isoforms of NHE and amiloride-sensitive epithelial Na+ channels (ENaC), possessing the same or even higher affinity for amiloride, in the antihypertensive action of this compound remain unknown. The potent contribution of ENaC in the maintenance of elevated blood pressure in monogenic or Mendelian forms of volume-expanded hypertension has been documented in studies by Lifton et al. [8], whereas the role of the renal-specific NHE3 isoform in salt-sensitive hypertension has been demonstrated in transgenic mice overexpressing this carrier [9] and in NHE−/− mice possessing reduced blood pressure on a low-salt diet [10]. By contrast to the above-mentioned Na+ transporters, to date, there is no direct evidence for the involvement of enhanced activity of NHE1 (i.e. the only isoform of the carrier expressed in cardiomyocytes) in the maintenance of elevated blood pressure. This latter issue becomes especially important because identification of the amiloride-binding molecule that triggers the antihypertensive effect should lead to the development of more potent and selective ligands of this target. Indeed, the identification of NHE1 as an amiloride target in the therapy of ischemic reperfusion injury led to the synthesis of compound Hoe 694, with sharply reduced side-effects caused by inhibition of ENaC and NHE isoforms distinct from NHE1 [11]. We do not know if protection by amiloride from LVH is exclusively caused by blood pressure attenuation, or whether this effect is at least partially mediated by blood pressure-independent inhibition of NHE1. The more potent antihypertensive effect of the angiotensin- converting enzyme (ACE) inhibitor enalapril and the more complete protection from LVH in SHR treated with this compound [7], as well as the attenuation of NHE activity in lymphocytes from patients with essential hypertension by the other ACE inhibitor quinapril [12], favour the first hypothesis. On the other hand, the decreased pHi-dependent BCECF fluorescence demonstrated in cardiac myocytes from amiloride-treated SHR only, and the attenuation of hydroxyproline content (i.e. a well-documented marker of collagen production) observed in enalapril-treated but not in amiloride-treated SHR, allowed Ji et al. [7] to propose the role of a specific (i.e. NHE-mediated) pathway in the development of LVH. The direct measurement of NHE activity in freshly isolated cardiac myocytes from control and amiloride (enalapril)-treated SHR, as well as model experiments with cultured cells subjected to specific long-term modulation of NHE activity, is required to dissect this alternative hypothesis. The role of NHE in enhanced cell proliferation and tissue hyperplasia was proposed long ago. This hypothesis is based on several observations. First, growth factors lead to transient NHE activation [13,14]. Second, elevated NHE activity in cultured VSMC from SHR [15,16] and fibroblasts from patients with insulin-dependent diabetes mellitus [17,18] was associated with enhanced DNA synthesis and cell proliferation. Third, amiloride and other NHE inhibitors suppress cell growth in culture [19]. However, it should be stressed that LVH in SHR is caused by increased cell size and enlargement of the extracellular matrix rather than by augmented cardiomyocyte proliferation [7]. A priori, NHE1 involvement in the triggering of cardiac hypertrophy caused by enhanced protein content per cell can be mediated by the signals dicussed subsequently. Parallel activation of NHE and HCO3−/Cl− exchanger leads to elevation of intracellular content of NaCl and osmotically obliged water. In accordance with this hypothesis, initial cell swelling caused by accumulation of inorganic osmolites is sufficient to trigger protein synthesis [20]. To test this hypothesis, precise measurement of the activity of these carriers and intracellular content of monovalent ions should be carried out. NHE activation leads to cytoplasm alkalinization. However, it should be stressed that sustained pHi elevation, known as a marker of transformation-associated phenotypes such as anchorage-independent growth, increased cell proliferation in the absence of growth factors and glycolytic metabolism [21] rather than cellular hypertrophy [22]. Because of the abundance of cardiomyocytes with Na+/Ca2+ exchanger, NHE activation in these cells is accompanied by [Ca2+]i elevation which, in turn, leads to expression of genes containing (Ca2+ + cAMP) response element [23,24], including genes whose augmented expression contributes to cardiomyocyte enlargement. The latter statement is consistent with data obtained in studies demonstrating a Ca2+-dependent mechanism of expression of early response genes (c-fos) and skeletal actin under modest elevation of [Na+]i in primary cultured cardiomyocytes with partially inhibited Na+/K+-ATPase [25,26]. By contrast to cardiomyocytes, cultured VSMC exhibit low activity of Na+/Ca2+ exchanger. In these cells, c-Fos expression is triggered by Na+i-mediated Ca2+i-independent signalling [27]. The relative contributions of these ionic mechanisms of excitation–transcription coupling in the development of LVH under sustained activation of NHE deserve further investigation.