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(Hypertension. 1995;26:649-655.)
© 1995 American Heart Association, Inc.

Articles

Sodium-Proton Exchange and Primary Hypertension

An Update

Winfried Siffert; Rainer Düsing

From the Institut für Pharmakologie, Universitätsklinikum, Essen (W.S.), and Medizinische Universitäts-Poliklinik, Bonn (R.D.), Germany.

Correspondence to Dr Winfried Siffert, Institut für Pharmakologie, Universitätsklinikum Essen, Hufelandstr 55, D-45122 Essen, FRG.

	Abstract

Top Abstract Introduction Epidemiology and Potential... Intracellular and Systemic... The NHE in Primary... Potential Reasons Underlying... Conclusions and Perspectives References

 
Abstract An enhancement of sodium-proton exchange in blood cells of patients with primary hypertension has been described by various investigators. The present review summarizes some of the most recent findings regarding the enhanced sodium-proton exchanger activity in primary hypertension and discusses the potential mechanisms that may contribute to or explain these findings. Novel evidence has been accumulated on the in vivo regulation of the sodium-proton exchanger in humans, and recent findings suggest that metabolic acidosis, high NaCl intake, and circulating hormones (eg, insulin) can enhance sodium-proton exchanger activity in blood cells. However, the relative roles of such exogenous factors in the stimulation of sodium-proton exchanger activity in primary hypertension remain questionable because enhanced sodium-proton exchanger activity persists in immortalized lymphoblasts from patients with primary hypertension after prolonged cell culture. Therefore, at least in a certain group of hypertensive subjects this abnormality cannot be due to metabolic or hormonal alterations of the "hypertensive" in vivo milieu but appears to be under genetic control. Available evidence strongly argues against intrinsic changes of the sodium-proton exchanger protein itself in primary hypertension, for example, a mutation in the encoding gene. Interestingly, immortalized cells from hypertensive subjects with enhanced sodium-proton exchanger activity display a distinctly enhanced proliferation pattern that appears to be independent of this ion transport. At present we speculate that enhanced sodium-proton exchanger activity and proliferation may represent indicators of a genetically fixed enhanced intracellular signal transduction in primary hypertension that may be caused by an increased activation of pertussis toxin–sensitive G proteins.

Key Words: diabetes mellitus • calcium • G proteins • phosphorylation

	Introduction

Top Abstract Introduction Epidemiology and Potential... Intracellular and Systemic... The NHE in Primary... Potential Reasons Underlying... Conclusions and Perspectives References

 
The Na⁺-H⁺ exchanger (NHE) or Na⁺-H⁺ antiporter is activated in a substantial number of patients with primary (essential) hypertension and therefore has been seen as an intermediate phenotype in this disorder. This transport protein, more precisely the ubiquitously expressed NHE-1 isoform, plays a dominant role in cellular pH and volume homeostasis and may also contribute to the initiation of cell growth and/or proliferation, although this latter issue is controversial.¹ The expression of the other cloned isoforms, that is, NHE-2 through NHE-4, occurs predominantly in epithelial cells in which these transporters seem to be involved in the transepithelial transport of Na⁺ ions.^{2 3} Since it appears likely that most of the studies performed thus far in primary hypertension have measured NHE-1 activity, the present review will focus on the regulation of this particular isoform. The observation that an enhancement of NHE-1 activity does not exist in all subjects with primary hypertension^{4 5} makes it unlikely that enhanced NHE activity develops secondary to increased blood pressure itself.

Several recent reviews are available on the molecular biology, regulation, and potential physiological functions of the NHE family of ion transporters.^{2 3 6 7 8 9 10 11} The present review will concentrate on new insights regarding the intracellular regulation of NHE-1 and the potential contribution of these novel findings to the "hypertensive" NHE phenotype. We will also discuss some of the more recent studies on the in vivo regulation of the NHE in humans that examined the effect of circulating hormones, especially insulin,¹² metabolic acidosis,^{13 14} and NaCl intake,¹⁵ on the activity of the antiporter. The relative importance of these exogenous factors in mediating enhanced NHE-1 activity in primary hypertension appears questionable in the face of the discovery that the hypertensive NHE phenotype persists in immortalized cells from patients with primary hypertension.¹⁶ We will therefore discuss the potential utility of this cell culture model in examining causes and effects of the abnormal NHE regulation in primary hypertension.

	Epidemiology and Potential Associations of Enhanced NHE Activity in Primary Hypertension

Top Abstract Introduction Epidemiology and Potential... Intracellular and Systemic... The NHE in Primary... Potential Reasons Underlying... Conclusions and Perspectives References

 
Little information exists regarding the question of what proportion of patients with primary hypertension display enhanced NHE activity. Canessa et al⁴ reported a bimodal distribution of NHE activity in red blood cells of 42 hypertensive subjects, 45% displaying enhanced NHE activity. Platelet NHE activity was reported to be enhanced in 14 of 26 patients with primary hypertension.⁵ Recently, Díez et al¹⁷ reported that erythrocyte NHE activity was enhanced in 12 of 27 hypertensive patients. Although the total number of patients categorized in terms of "low" (or "normal") versus "increased" NHE activity is as yet rather limited (total, n=95), on the basis of these data one may speculate that up to 50% of all patients with primary hypertension belong to the "enhanced" NHE phenotype. Thus, studies regarding the epidemiology of NHE activity in normotension and hypertension are highly desirable. Similarly, only a few studies have so far addressed the question of which phenotypes coincide with elevated NHE activity; the proposed associations are summarized in the Table. Since the numbers of patients and normotensive control subjects investigated were rather small, these findings, though interesting, require confirmation in larger studies before being commonly accepted.

View this table: [in this window] [in a new window]   Table 1. Proposed Associations With Enhanced Sodium-Proton Exchanger Activity

The development of transgenic animal models may help to clarify the role of the NHE in essential hypertension. Transgenic mice that overexpress NHE-1 develop salt-sensitive hypertension.²⁶ Furthermore, these animals display reduced fractional Na⁺ excretion, plasma renin activity, and aldosterone levels compared with controls,²⁶ these peculiarities resembling those also proposed for hypertensive patients with elevated NHE activity (Table).

	Intracellular and Systemic Regulation of NHE-1

Top Abstract Introduction Epidemiology and Potential... Intracellular and Systemic... The NHE in Primary... Potential Reasons Underlying... Conclusions and Perspectives References

 
Intracellular Regulation
Role of Phosphorylation
A variety of agents, including thrombin, epidermal growth factor, and activators of protein kinase C (phorbol esters), increase the phosphorylation of NHE-1 exclusively on serine residues.^{27 28 29} Thus, regardless of by which pathway cells are stimulated, via G protein–coupled receptors (eg, thrombin) or receptors with intrinsic tyrosine kinase activity (eg, epidermal growth factor), the same phosphorylation pattern was determined. This has led to the attractive hypothesis that these different signal-transducing pathways converge on a specific kinase, referred to as NHE-1 kinase, that integrates these signals and finally mediates the phosphorylation of NHE-1 (for reviews see References 2, 6, 7, 11, and 29^{2 6 7 11 29} ). Thus, although activators of protein kinase C, such as phorbol esters, have unequivocally been shown to activate the antiporter, it remains questionable whether this activation was actually mediated by protein kinase C itself or by kinases located "downstream" of this enzyme. It should also be stressed that not all stimuli that activate the antiporter do so by increasing its phosphorylation. Thus, intracellular acidification,³⁰ osmotic shrinkage,³¹ and most notably a rise in the concentration of intracellular free Ca²⁺³² activate NHE-1 without an apparent change in its phosphorylation pattern.

Role of Ca²⁺-Calmodulin
Cell stimulation by many physiological agonists, for example, epidermal growth factor or thrombin, not only activates protein kinase C but also induces a rise in cytosolic free Ca²⁺ concentration ([Ca²⁺]_i). It has long been controversial whether Ca²⁺_i can activate the NHE. Thus, Huang et al³³ demonstrated that thrombin induced a rise in pH in rat aortic smooth muscle cells despite the downregulation of protein kinase C. Pretreatment of these cells with pertussis toxin not only blunted the thrombin-induced rise in Ca²⁺_i but also the thrombin-evoked alkalinization, suggesting a role for Ca²⁺ ions in this activation.³³ However, since an artificial increase in Ca²⁺_i evoked by the Ca²⁺ ionophore ionomycin failed to raise pH_i, they concluded that Ca²⁺ ions per se do not activate the antiporter.³³ However, a rise in Ca²⁺_i was reported to activate an amiloride-sensitive Na⁺ influx in human fibroblasts, which is indicative of NHE activation.³⁴

Kimura et al³⁵ characterized the activation of the platelet NHE in more detail. These authors demonstrated that an inhibitor of protein kinase C completely prevented the phorbol ester–induced activation of the NHE, whereas thrombin- and vasopressin-induced activation was only partially inhibited. In contrast, intracellular Ca²⁺ depletion completely blunted the thrombin- and vasopressin-induced NHE activation. Moreover, raising Ca²⁺_i by ionomycin activated the antiporter via a protein kinase C–independent mechanism. Recent findings on the Ca²⁺-calmodulin regulation of NHE-1 underscore the importance of the above-mentioned findings by different investigators. To our knowledge, Fliegel et al³⁶ were the first to demonstrate a Ca²⁺-calmodulin–dependent phosphorylation of the C-terminal domain of NHE-1. Recently, the existence of specific calmodulin binding sites in the cytoplasmic domain of NHE-1³⁷ has been shown. Their importance for NHE-1 activation is illustrated by the finding that deletions or point mutations in this site reduce the growth factor– or hypertonicity-induced cytosolic alkalinization by 50% to 80%.³² The major role of this Ca²⁺-calmodulin binding site is to regulate the activity of NHE-1 in unstimulated and stimulated cells.³² If this site is deleted, the pH_i activation curve of NHE-1 undergoes a strong alkaline shift which resembles that seen after agonist stimulation. In fact, thrombin stimulation of cells expressing an NHE-1 mutant that lacks the Ca²⁺-calmodulin binding sites fails to further activate the antiporter.

Wakabayashi et al³² have proposed an appealing concept regarding the physiological role of this Ca²⁺-calmodulin binding domain. In resting cells, in which Ca²⁺_i is low and calmodulin is not activated, this domain prevents an alkaline shift of the pH_i activation curve, thereby keeping NHE-1 in its resting conformation. Thus, this domain, when unbound by calmodulin, functions as an intrinsic "autoinhibitor" of NHE-1. This inhibitory control is physiologically important, as the continuous Na⁺ overload that would be imposed on cells by a highly active antiporter would lead to a dramatic cellular energy expenditure, since intracellular Na⁺ must be removed by the Na,K-ATPase. The increase in [Ca²⁺]_i after cell stimulation induces a Ca²⁺-calmodulin complex to bind to this autoinhibitory region in the cytoplasmic domain of the antiporter, thereby switching off this intrinsic inhibition. Thereafter, the ion transporter can efficiently counteract cytosolic acidification in stimulated cells.

These novel findings of a Ca²⁺-calmodulin–dependent activation of NHE-1 could contribute to the understanding of the abnormal regulation of this transporter in primary hypertension, since many investigators have described an increased basal or stimulated [Ca²⁺]_i in platelets of patients with this disorder.^{38 39 40 41 42} This issue will be discussed in more detail below.

Direct Regulation by G Proteins
Intracellular signal transduction by many agonists is initiated via receptors that activate one or more G proteins. The observation that some ion channels, for example, Ca²⁺ and K⁺ channels, are directly regulated by activated G proteins^{43 44 45 46} raises the question of whether ion transporters, for example, the NHE, might also be under the direct control of G proteins. In fact, some evidence exists that the NHE can be directly inhibited or activated by G proteins. Using brush border membrane vesicles from rat kidney, Brunskill et al⁴⁷ demonstrated an apparently specific inhibition by nonhydrolyzable GTP analogues in these membranes. It remains unknown which of the known NHE isoforms was affected by this treatment. We have previously reported that treatment of platelets with aluminum fluoride, which unspecifically activates G proteins, failed to activate the NHE and also prevented the alkalinization normally seen in response to thrombin or phorbol ester stimulation.⁴⁸ Subsequent studies demonstrated that nonhydrolyzable GTP analogues inhibited the NHE in platelet membranes⁴⁹ in a manner consistent with that observed in brush border membrane vesicles. Although they are rather circumstantial, these findings are suggestive of a G protein–mediated inhibition of the NHE. To our knowledge, no further attempts have been made to strengthen this hypothesis or identify the G protein isoforms involved.

On the other hand, some evidence points to an activation of the NHE by specific G protein -subunits. Voyno-Yasenetskaya et al⁵⁰ expressed mutationally activated -subunits of the G proteins G₁₃, G₁₂, G_q, and G_s in human embryonic kidney cells and determined the effects of these expressions on NHE activity as deduced from resting pH_i and pH_i recovery from cytosolic acidification. Both activated ₁₃- and _q-subunits activated the NHE, whereas the other activated subunits left the antiporter unaffected. Although signaling via the cAMP and inositol phosphate pathways could be ruled out, the mechanism or mechanisms underlying this NHE activation remain to be defined. Dhanasekaran et al⁵¹ expressed mutationally activated _s, _i2, _q, ₁₂, and ₁₃ G protein subunits in COS-1 cells. Only the _q, ₁₂, and ₁₃ G protein subunits enhanced the NHE activity as determined from an enhanced pH_i recovery and more alkaline pH_i, whereas expression of the other -subunits did not produce such an effect. Whereas the stimulating effect of G₁₂ and G_q was apparently mediated by the protein kinase C pathway, the activation by G₁₃ was preserved in cells in which protein kinase C was downregulated by phorbol ester pretreatment. The molecular basis of this novel G protein–mediated NHE activation remains to be elucidated.

Recent findings suggest that two typical platelet agonists, thrombin and thromboxane A₂, which were shown to activate the NHE,⁵² also activate G₁₂,⁵³ that is, one of the G proteins that may contribute to NHE regulation. Thus, changes in the expression pattern and/or amounts of certain G proteins in primary hypertension could at least theoretically contribute to the hypertensive NHE phenotype.

Systemic Regulation of the NHE
Regulation by Insulin
In view of the fact that insulin resistance and subsequent hyperinsulinemia are frequently reported abnormalities in primary hypertension,^{54 55 56} Pontremoli et al¹² determined the effect of insulin on the human red blood cell NHE. Treatment of erythrocytes with physiological concentrations of insulin increased NHE activity almost twofold. This effect of insulin requires intracellular Ca²⁺ ions and seems to be mediated by phosphorylation. However, it should be noted that Carr et al⁵⁷ denied an effect of insulin on the NHE in human lymphocytes.

Effect of Metabolic Acidosis
Several in vivo and in vitro studies have shown that metabolic acidosis activates the NHE in renal cells.^{58 59} However, until recently no such information was available regarding the effects of metabolic acidosis on the NHE activity in human blood cells. Corry et al⁶⁰ described an enhanced NHE activity in erythrocytes of patients with end-stage renal failure and acidosis that correlated with an enhanced expression of the NHE-1 protein. Reusch et al¹³ determined NHE activity in healthy control subjects and patients with metabolic acidosis caused by chronic renal failure. Patients with renal insufficiency displayed a distinctly enhanced NHE activity in lymphocytes that could not be explained by a rise in [Ca²⁺]_i.¹³ Moreover, induction of metabolic acidosis in otherwise healthy subjects by means of NH₄Cl ingestion was associated with an enhanced NHE activity in lymphocytes but not in platelets, suggesting an involvement of de novo synthesis of antiporter protein.¹³ This hypothesis was strengthened by findings of Quednau et al,¹⁴ who demonstrated that this increase in lymphocyte antiporter activity during experimentally induced metabolic acidosis was paralleled by an increase in the steady-state levels of NHE-1 mRNA. It thus appears that metabolic acidosis increases NHE-1 protein expression in blood cells capable of protein synthesis and that this results in a Ca²⁺-independent increase in antiporter activity. Whether lymphocytes possess a "pH_i sensor" and whether this effect is mediated by an altered hormonal milieu remains to be investigated.

Effect of NaCl Intake
Göbel et al¹⁵ determined the effect of dietary NaCl on the activity of the lymphocyte antiporter. Extreme changes in NaCl intake from 20 to 300 mmol/d increased NHE activity in lymphocytes from normotensive volunteers by approximately 35%, whereas basal pH_i values were unaffected. On the other hand, moderate increases in daily NaCl intake to 85 and 200 mmol were without effect. A tendency for an increased activity of the lymphocyte NHE was also observed in hypertensive subjects on switching NaCl intake from 20 to 300 mmol/d. Since the total number of patients enrolled in this latter study was small, it was not possible to establish whether this enhancement of antiporter activity coincided with salt sensitivity or salt resistance. Furthermore, the mechanism or mechanisms mediating this increase in NHE activity remained obscure. Alexiewicz et al⁶¹ have shown that a dietary Na⁺ load increases [Ca²⁺]_i in lymphocytes of salt-sensitive hypertensive patients. Resnick et al⁶² recently described the effect of dietary NaCl loading on intracellular ions in red blood cells from salt-sensitive and salt-resistant subjects with primary hypertension. Salt-sensitive subjects responded with a significant increase in Ca²⁺_i and a decrease in pH_i after a dietary NaCl load. These findings leave room for two alternative hypotheses regarding the mechanisms by which an NaCl load could modify the Na⁺-H⁺ antiport. Given the Ca²⁺_i dependency of NHE-1 (see above), one could argue that an NaCl-induced rise in [Ca²⁺]_i directly activates NHE-1. Alternatively, the NaCl-induced fall in pH_i, possibly secondary to extracellular "dilution" acidosis,⁶³ could regulate the NHE in a manner similar to that observed in metabolic acidosis (see above). On the other hand, enhanced NHE activity could be a genetic factor that contributes to salt sensitivity. Transgenic mice that overexpress NHE-1 display a marked increase in blood pressure on NaCl loading,¹⁸ as noted above.

	The NHE in Primary Hypertension

Top Abstract Introduction Epidemiology and Potential... Intracellular and Systemic... The NHE in Primary... Potential Reasons Underlying... Conclusions and Perspectives References

 
Many reports have demonstrated enhanced NHE activity in platelets, leukocytes, and erythrocytes from patients with primary hypertension (for review see Reference 64⁶⁴ ). However, the reasons underlying these peculiarities have remained obscure.

Fig 1 summarizes some of the potential reasons that could induce the hypertensive NHE phenotype, and these can be grouped into systemic, intrinsic, and intracellular factors. Thus, the hypertensive NHE could reflect systemic changes of the neurohumoral milieu, for example, a tendency toward metabolic acidosis or hyperinsulinemia. Alternatively, the intracellular regulation of the NHE might be different in primary hypertension because of altered activation by, for instance, protein kinases, Ca²⁺ calmodulin, or G proteins. Finally, the possibility exists that intrinsic changes are present in the hypertensive NHE, for example, a mutation in the encoding gene, a posttranslational modification, or overexpression of the NHE protein.

View larger version (17K): [in this window] [in a new window]   Figure 1. Diagram shows theoretical reasons for enhanced Na+-H+ exchanger activity in primary hypertension. The Na+-H+ exchanger could be enhanced in vivo by systemic hormonal or metabolic abnormalities in primary hypertension. Alternatively, intrinsic alterations may be present, including overexpression, mutation, or altered posttranslational modification of the Na+-H+ exchanger in primary hypertension. Finally, the intracellular regulation of the Na+-H+ exchanger could be modified in primary hypertension.

Garciandia et al⁶⁵ have determined NHE activity and NHE-1–specific mRNA (by reverse transcription–polymerase chain reaction) in lymphocytes of normotensive and hypertensive individuals. Although these authors determined slightly increased NHE-1 mRNA levels by approximately 1.3-fold in lymphocytes of hypertensive patients, they also stated that "no correlations were found between intracellular NHE-1 mRNA levels and either intracellular pH or Na⁺-H⁺exchange activity" (see page 361 of Reference 65⁶⁵ ). Thus, in our view the pathophysiological significance of their findings remains obscure. At present, no strong evidence exists in favor of an overexpression of the NHE in primary hypertension.

Hypertensive NHE-1 Phenotype Is Conserved in Immortalized Cells
To determine whether the enhanced hypertensive NHE phenotype is somehow under genetic control, we pursued a novel experimental strategy.¹⁶ We immortalized lymphocytes from previously characterized normotensive and hypertensive subjects with "low" and "high" NHE activity, respectively, using Epstein-Barr virus and cultured them for several weeks. Since all cell lines in culture are treated equally, this procedure removes all potential abnormalities of the hypertensive milieu (referred to as systemic factors in Fig 1) that might influence the kinetic properties of the NHE in vivo and that might precipitate in the hypertensive NHE phenotype detectable in blood cells ex vivo. The enhanced NHE activity persisted in the immortalized hypertensive cells under different conditions, for example, in serum-stimulated cells,¹⁶ and even after cell activation with phorbol ester.¹⁶

Several lines of evidence suggest that the abnormalities in primary hypertension involve the NHE-1 isoform of antiporters: (1) Blood cells express NHE-1 protein^{23 60} and NHE-1 mRNA^{16 66} but not NHE-3 and NHE-4 mRNA³⁰ ; (2) the NHE expressed in lymphoblasts is completely inhibited by low concentrations of ethylisopropylamiloride (K_i 20 nmol/L),⁶⁷ whereas the NHE-2 and NHE-3 isoforms exhibit a lower affinity for amiloride analogues^{11 68} ; and (3) stimulation by phorbol ester leaves the V_max of NHE-1 in lymphoblasts unchanged,¹⁶ whereas the V_max values of NHE-2 and NHE-3 are increased and decreased, respectively, by such treatment.⁶⁹ Similar considerations apply for vascular smooth muscle cells (VSMCs) from spontaneously hypertensive rats (SHR) and normotensive Wistar-Kyoto rats (WKY), which display differences in NHE activity⁷⁰ that are very similar to those observed in immortalized lymphoblasts from normotensive and hypertensive subjects. Lucchesi et al⁷¹ could rule out mRNA expression of the NHE-2, NHE-3, and NHE-4 isoforms in VSMCs from both strains and confirmed the sole expression of NHE-1 mRNA.

The NHE Is Neither Overexpressed Nor Mutated in Primary Hypertension
Increased V_max of the NHE in immortalized lymphoblasts from patients with primary hypertension could be due to overexpression of the transport protein under cell culture conditions. We could rule out an increased steady-state level of the NHE-1 mRNA in immortalized lymphoblasts from patients with primary hypertension, which strongly suggests but does not ultimately exclude an overexpression of the antiporter.¹⁶ Similar results were obtained in VSMCs of SHR and WKY. Lucchesi et al⁷¹ and LaPointe et al⁷² found no differential expression of NHE-1 mRNA in VSMCs from WKY and SHR, respectively. Using an antibody raised against NHE-1, Siczkowski et al⁷³ could rule out an overexpression of NHE-1 in VSMCs from SHR on the protein level. Thus, available evidence argues against an overexpression of NHE-1 as the major reason for the enhanced NHE activity in primary hypertension and in SHR.

This raises the question regarding potential structural changes of NHE-1 in primary hypertension. We have sequenced the complete cDNAs encoding for NHE-1 from immortalized hypertensive and normotensive cell lines with low and enhanced NHE activity without finding any sequence changes.¹⁶ These findings are in agreement with linkage analysis that argued against a mutation in the NHE gene in primary hypertension.^{74 75} Taking these findings together, we propose that the enhanced NHE-1 activity in primary hypertension is caused by neither the hypertensive neurohumoral or metabolic environment nor intrinsic changes in the NHE-1 protein. Therefore, the enhanced NHE activity in primary hypertension can best be explained by altered intracellular regulation (Fig 1).

Enhanced Proliferation of Hypertensive Cell Lines With Enhanced NHE Activity
Epstein Barr virus–immortalized lymphocytes with enhanced NHE-1 activity from patients with primary hypertension proliferate distinctly faster,¹⁶ display enhanced DNA synthesis,⁶⁷ and are more frequently found in the G₂/M phase of the cell cycle⁶⁷ than normotensive cell lines. This raises the question of whether enhanced NHE activity constitutes a major growth-permissive factor in primary hypertension. We reported recently that the enhanced proliferation of hypertensive cell lines persists under cell culture conditions in which the contribution of NHE-1 to pH regulation is minimized.⁶⁷ Thus, at least in our particular model it appears that enhanced NHE-1 activity and cell proliferation are separate and not interdependent phenomena in primary hypertension, both most likely reflecting cellular alterations in more proximal components of intracellular signal transduction.

	Potential Reasons Underlying Enhanced NHE-1 Activity and Cell Growth in Primary Hypertension

Top Abstract Introduction Epidemiology and Potential... Intracellular and Systemic... The NHE in Primary... Potential Reasons Underlying... Conclusions and Perspectives References

 
At present, only speculations are possible regarding the reasons for the enhanced NHE activity and cell proliferation in primary hypertension. Recent results strongly support the frequently made proposal of Aviv^{76 77 78 79 80} of a tight link between enhanced Ca²⁺ and Na⁺-H⁺ exchange in primary hypertension. Touyz and Schiffrin⁴¹ found both enhanced angiotensin II–induced [Ca²⁺]_i and pH_i rises in platelets from hypertensive subjects compared with those from normotensive control subjects. Similarly, Poch et al⁴² described a linear relationship between [Ca²⁺]_i and pH_i rises in thrombin-stimulated platelets, both responses being enhanced in platelets from hypertensive patients. In view of these findings we have recently analyzed platelet-activating factor–induced [Ca²⁺]_i rises in immortalized lymphoblasts from normotensive and hypertensive subjects (Fig 2). Platelet activating factor–induced [Ca²⁺]_i rises were enhanced almost twofold in the fast-growing hypertensive cells that display the enhanced NHE phenotype⁸¹ ; similar findings were obtained by other researchers.⁸² Given the Ca²⁺-calmodulin activation of NHE-1 (see above), one could speculate that the enhanced agonist-evoked Ca²⁺ signals in hypertension are the main reasons for the enhanced NHE-1 activity. However, this remains to be shown. Alternatively, both phenomena could be independent indicators of a disturbance in intracellular signal transduction that could be located proximal to both Ca²⁺ mobilization and NHE activity control, for example, activation of G proteins. We actually found a more pronounced reduction of cell proliferation by pertussis toxin in hypertensive cell lines with the enhanced NHE phenotype.⁶⁷ Furthermore, we determined a distinctly enhanced activation of pertussis toxin–sensitive G proteins in these hypertensive cell lines.⁸¹ On the basis of these novel findings, we find it attractive to speculate that an enhanced activation of pertussis toxin–sensitive G proteins could be the common denominator for altered signal transduction in hypertension.

View larger version (21K): [in this window] [in a new window]   Figure 2. Graphs show Ca2+ rises in immortalized lymphoblasts from a normotensive subject (low Na+-H+ exchanger activity, top) and hypertensive subject (enhanced Na+-H+ exchanger activity, bottom). Cells were immortalized as described in detail previously.16 Displayed are responses of fura 2–loaded cells suspended in Ca2+-containing buffer (1 mmol/L) on stimulation with 0.1 µmol/L platelet-activating factor. Similar differences in agonist-evoked Ca2+ signals could be determined in all established normotensive and hypertensive cell lines. For full details see Reference 8181 .

	Conclusions and Perspectives

Top Abstract Introduction Epidemiology and Potential... Intracellular and Systemic... The NHE in Primary... Potential Reasons Underlying... Conclusions and Perspectives References

 
New insights have been gained regarding the in vivo regulation of NHE-1 by metabolic acidosis,^{13 14} NaCl intake,¹⁵ and insulin.¹² Studies on immortalized cells from patients with hypertension have shown that enhanced NHE-1 activity is genetically fixed and associated with an enhanced proliferation tendency.^{16 67} Both phenomena appear to be caused by an enhanced cellular reactivity in primary hypertension. The cell culture model that we have established for primary hypertension now allows one to systematically investigate the signal transduction pathways that control both NHE-1 activity and cell proliferation. The main advantage of this system is that all these determinations can now be conducted without the confounding influences of the hypertensive in vivo environment.

	Acknowledgments

 
Studies in the laboratory of W.S. were financially supported by the Deutsche Forschungsgemeinschaft and the Ministerium für Wissenschaft und Forschung NRW.

Received March 24, 1995; first decision May 19, 1995; accepted July 18, 1995.

	References

Top Abstract Introduction Epidemiology and Potential... Intracellular and Systemic... The NHE in Primary... Potential Reasons Underlying... Conclusions and Perspectives References

 
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