Circulating natriuretic peptide concentrations in patients with end-stage renal disease: role of brain natriuretic peptide as a biomarker for ventricular remodeling.

OBJECTIVES: To determine levels of natriuretic peptides (NPs) in patients with end-stage renal disease (ESRD) and to examine the relationship of these cardiovascular peptides to left ventricular hypertrophy (LVH) and to cardiac mortality. PATIENTS AND METHODS: One hundred twelve dialysis patients without clinical evidence of congestive heart failure underwent plasma measurement of NP concentrations and echocardiographic investigation for left ventricular mass index (LVMI). RESULTS: Plasma atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) concentrations correlated positively with LVMI and inversely with left ventricular ejection fraction, whereas C-type NP and Dendroaspis NP levels did not correlate with LVMI. In dialysis patients with LVH (LVMI >125 g/m2), plasma ANP and BNP concentrations were increased compared with those in dialysis patients without LVH (both P<001). In a subset of 15 dialysis patients without LVH or other concomitant diseases, plasma BNP concentrations were not significantly increased compared with those in 35 controls (mean +/- SD, 20.1+/-13.4 vs 13.5+/-9.6 pg/mL; P=.06), demonstrating that the BNP concentration was not increased by renal dysfunction alone. Furthermore, the BNP level was significantly higher in the 16 patients who died from cardiovascular causes compared with survivors (mean +/- SD, 129+/-13 vs 57+/-7 pg/mL; P<.003) and was significantly associated with greater risk of cardiovascular death in Cox regression analysis (P<.001), as was the ANP level (P=.002). CONCLUSIONS: Elevation of the plasma BNP concentration is more specifically related to LVH compared with the other NP levels in patients with ESRD independent of congestive heart failure. Thus, BNP serves as an important plasma biomarker for ventricular hypertrophy in dialysis patients with ESRD.

Role of plasma renin activity and the renal nerves in the natriuresis of L-NMMA infusion in rats.

The objective of this study was to determine the effect of N(G)-monomethyl-L-arginine (L-NMMA) infusion on plasma renin activity (PRA) in the presence or absence of the renal nerves in normotensive Wistar-Kyoto (WKY) rats and Okamoto spontaneously hypertensive rats (SHR). All rats were unilaterally nephrectomized two weeks before the acute experiment. On the day of the experiment, acute renal denervation (Dnx) of the remaining kidney was performed in one group of WKY rats (Dnx-WKY; n= 10) and one group of SHRs (Dnx-SHR: n=7). The renal nerves were left intact in a group of WKY rats (Inn-WKY; n=8) and SHRs (Inn-SHR; n=9). After a control clearance period, L-NMMA was administered i.v. (15 mg/kg bolus followed by 500 microg/kg/min infusion) and another clearance period of 20 min was taken. In all experimental groups L-NMMA infusion resulted in a significant natriuresis. L-NMMA infusion increased fractional excretion of sodium (FE(Na)) to a greater extent in the Inn-SHR than in the Inn-WKY (delta FE(Na) = 5.23+/-0.87% vs delta FE(Na) = 2.87+/-0.73% respectively; P=0.05), PRA did not change in the SHR with the infusion of L-NMMA. However, in the Inn-WKY group, the natriuresis of L-NMMA infusion was associated with a tendency for lower PRA levels as compared to a group of time control Inn-WKY rats. In Dnx-WKY, the natriuresis of L-NMMA infusion (delta FE(Na) = 4.60+/-0.52%) was associated with a significantly lower level of PRA (4.26+/-1.18 ng AI/ml/hr) as compared to a group of time control Dnx-WKY rats (9.83+/-1.32 ng AI/ml/hr; P<0.05). In the Dnx-SHR, the natriuretic response to L-NMMA infusion was significantly attenuated by renal denervation (delta FE(Na) = 2.36+/-0.34%) and PRA was unchanged. In conclusion, the natriuretic effect of systemic inhibition of nitric oxide (NO) synthesis was associated with decreased PRA in the Dnx-WKY suggesting that a potential interaction exists between NO and the renal nerves in the modulation of PRA in the normotensive WKY rat. Whereas, the natriuretic effect of L-NMMA infusion in the SHR in the presence and absence of the renal nerves, were independent of changes in PRA.

Attenuated natriuretic response to adrenomedullin in experimental heart failure.

BACKGROUND: The recently discovered vasodilating and positive inotropic peptide, adrenomedullin (ADM), has strong natriuretic actions. ADM-induced natriuresis is caused by an increase in glomerular filtration rate and a decrease in distal tubular sodium reabsorption. Although ADM is activated in human and experimental heart failure, the role of ADM in the kidney in heart failure remains undefined. METHODS AND RESULTS: The present study was performed to determine the renal hemodynamic and urinary excretory actions of exogenously administered ADM in a canine model of acute heart failure produced by rapid ventricular pacing. Experimental acute heart failure was characterized by a decrease in cardiac output and an increase in pulmonary capillary wedge pressure with an increase in plasma ADM concentration. Intrarenal infusion of ADM (1 and 25 ng/kg/min) induced an increase in urinary sodium excretion in the normal control dogs (change in urinary sodium excretion [Delta UNaV], +94.5 microEq/min during 1 ng/kg/min ADM infusion and +128.1 microEq/min during 25 ng/kg/min ADM infusion). In the acute heart failure dogs, intrarenal ADM infusion resulted in an attenuated increase in urinary sodium excretion (Delta UNaV, +44.8 microEq/min during 1 ng/kg/min ADM infusion and +51.8 microEq/min during 25 ng/kg/min ADM infusion). Both glomerular and tubular actions of ADM were attenuated in the acute heart failure group compared with responses in the normal control group. CONCLUSION: The present study shows that the renal natriuretic responses to ADM are markedly attenuated in experimental acute heart failure. This study provides insight into humoral mechanisms that may promote sodium retention in heart failure via a renal hyporesponsiveness to natriuretic actions of ADM.

Increase and uncoupling of adrenomedullin from the natriuretic peptide system in aneurysmal subarachnoid hemorrhage.

OBJECT: Natriuresis is a common systemic manifestation of aneurysmal subarachnoid hemorrhage (SAH). Natriuresis and its accompanying hypovolemia may be a major contributing factor in the pathophysiology of symptomatic cerebral vasospasm. METHODS: The authors studied 14 consecutive patients with aneurysmal SAH and compared levels of adrenomedullin (ADM), a novel endogenous natriuretic peptide that possesses additional profound vasodilatory properties, with the natriuretic peptide system by using radioimmunoassay. The mean ADM values on admission were 24.8 pg/ml, a twofold increase over control values, but no correlation was found with atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP), and C-natriuretic peptide (CNP) from the natriuretic peptide system. At Day 5 post-SAH, ADM levels were significantly elevated in patients with vasospasm documented angiographically or on transcranial Doppler studies as compared with those who suffered no vasospasm (mean 61.9 pg/ml compared with 15.3 pg/ml, p < 0.01). CONCLUSIONS: The authors conclude that an elevation of ADM in plasma may indicate a physiological regulatory attempt to induce cerebral vasodilation. The regulation of ADM is uncoupled from ANP, BNP, and CNP.

Modulation of functionally active endothelin-converting enzyme by chronic neutral endopeptidase inhibition in experimental atherosclerosis.

BACKGROUND: Endothelin-converting enzyme-1 (ECE-1) processes big endothelin-1 (ET-1) to ET-1, a peptide implicated in atheroma formation. ECE-1 exists in 2 isoforms (ECE-1alpha and ECE-1beta), the result of alternative splicing of a common gene. Neutral endopeptidase (NEP) is a genetically distinct metallopeptidase that degrades the natriuretic peptides. These peptides mediate antiproliferative and vasodilating actions. We sought to demonstrate the distribution of the 2 ECE-1 isoforms in experimental atherosclerosis, to determine the effects of chronic NEP-I on plasma cGMP concentrations, vascular wall ECE-1 activity, and ET-1 concentration, and to correlate these actions with atheroma formation. Our hypothesis was that chronic NEP-I, in association with augmented cGMP, would inhibit ECE-1 conversion of big ET-1 to active ET-1, thus reducing tissue ET-1 concentrations and associated atheroma formation. METHODS AND RESULTS: Cholesterol-fed New Zealand White rabbits (n=8, 1% cholesterol diet) and NEP-I-treated cholesterol-fed New Zealand White rabbits (n=8; candoxatril, 30 mg/kg per day, Pfizer) were euthanized after 8 weeks of feeding. ECE-1alpha and ECE-1beta immunoreactivity was present in the aortas of both groups. Compared with control values, plasma cGMP concentrations were increased (2.8+/-0.6 versus 8.4+/-1.2 pmol/mL, P<0.05), ECE-1 activity was attenuated (68+/-3% versus 32+/-8%, P<0. 05), aortic tissue ET-1 concentrations were reduced (4.6+/-0.5 versus 3.2+/-0.3 pg/mg protein, P<0.05), and atheroma formation was attenuated (62+/-6% versus 34+/-5%, P<0.01) by NEP-I. CONCLUSIONS: These studies suggest that ECE-1 is present and functionally active in the vascular wall in atherosclerosis. Inhibition of ECE-1 by NEP-I represents a novel approach to interruption of the endothelin system in this cardiovascular disease state.

Renal actions of synthetic dendroaspis natriuretic peptide.

BACKGROUND: Dendroaspis natriuretic peptide (DNP), recently isolated from the venom of the green Mamba snake Dendroaspis angusticeps, is a 38 amino acid peptide containing a 17 amino acid disulfide ring structure similar to that of atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP), and C-type natriuretic peptide (CNP). DNP-like immunoreactivity (DNP-LI) was reported to be present in human plasma and atrial myocardium and to be elevated in human congestive heart failure. Although previously named DNP, it remains unknown if DNP is natriuretic or if is it present in canine plasma, urine, and atrial myocardium. METHOD: Studies were performed in vivo in anesthetized dogs (N = 6) using intravenous infusion of synthetic DNP at 10 and 50 ng/kg/min. Employing a sensitive and specific radioimmunoassay for DNP, the presence of DNP-like peptide was assessed in the canine plasma and urine before, during, and following the administration of exogenous synthetic DNP. Additionally, we performed immunohistochemical studies using the indirect immunoperoxidase method with polyclonal DNP antiserum in normal atrial myocardium (N = 10). Atrial concentrations of DNP-LI were also assessed. RESULTS: We report that DNP is markedly natriuretic and diuretic, which, like ANP and BNP, is associated with the increase in urinary and plasma cGMP. DNP-like peptide is also detected in canine plasma, urine, and atrial myocardium. CONCLUSION: These studies establish that DNP is a potent natriuretic and diuretic peptide with tubular actions linked to cGMP and that DNP may play a physiological role in the regulation of sodium excretion.

Presence of Dendroaspis natriuretic peptide-like immunoreactivity in human plasma and its increase during human heart failure.

OBJECTIVE: To determine whether Dendroaspis natriuretic peptide (DNP), a novel peptide isolated from the venom of the Dendroaspis angusticeps snake that contains a 17-amino acid disulfide ring structure similar to that in atrial, brain, and C-type natriuretic peptides, is present in normal human plasma and myocardium and whether, like the other natriuretic peptides, DNP-like immunoreactivity (DNP-LI) is activated in human congestive heart failure (CHF). MATERIAL AND METHODS: Circulating DNP-LI was assessed in 19 normal human subjects and 19 patients with CHF (New York Heart Association class III or IV) with a specific and sensitive radioimmunoassay for DNP with no cross-reactivity with the other natriuretic peptides. Immunohistochemical studies that used polyclonal rabbit anti-DNP antiserum were performed on human atrial myocardial tissue obtained from four patients with end-stage CHF who were undergoing cardiac transplantation and from three donor hearts at the time of transplantation. RESULTS: We report that DNP-LI circulates in normal human plasma and is present in the normal atrial myocardium. In addition, DNP-LI is increased in the plasma of patients with CHF. CONCLUSION: This study demonstrates, for the first time, the presence of a DNP-like peptide in normal human plasma and in the atrial myocardium. Additionally, these studies demonstrate increased plasma DNP-LI in human CHF. These results support the possible existence of an additional new natriuretic peptide in humans, which may have a role in the neurohumoral activation that characterizes human CHF.

Activation of myocardial and renal natriuretic peptides during acute intravascular volume overload in dogs: functional cardiorenal responses to receptor antagonism.

1. A family of structurally related but genetically distinct natriuretic peptides exist which include atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) of myocardial cell origin and C-type natriuretic peptide (CNP) of endothelial and renal epithelial cell origin. All three exert actions via cGMP, with ANP and BNP functioning via the natriuretic peptide A receptor and CNP via the natriuretic peptide B receptor. 2. Circulating and urinary natriuretic peptides were determined in response to acute intravascular volume overload (AVO). Additionally, their functional role in cardiorenal regulation during AVO was investigated by utilizing the natriuretic peptide receptor antagonist HS-142-1. Control (n=5) and study dogs (HS-142-1, n=9) underwent AVO with normal saline equal to 10% of body weight over 1 h. Both groups demonstrated similar significant increases in right atrial pressure, pulmonary capillary wedge pressure, pulmonary artery pressure and cardiac output. Circulating ANP paralleled increases in right atrial pressure and pulmonary capillary wedge pressure, with no changes in plasma BNP or CNP. At peak AVO, urinary CNP excretion was increased compared with baseline (7.0+/-4.2 versus 62+/-8.0 pg/min, P<0.05). 3. In the HS-142-1-treated group, plasma cGMP was decreased compared with the control group (9.6+/-1.1 to 5.0+/-1.2 pmol/ml, P<0.05). A significant attenuation of natriuresis (566+/-91 versus 1241+/-198 microEq/min, P<0.05) and diuresis (4.8+/-0.7 versus 10.1+/-2.0 ml/min, P<0.05) was also observed at peak AVO in the HS-142-1 treated group. 4. These findings support differential and selective responses of the three natriuretic peptides to AVO, in which plasma ANP and urinary CNP are markers for AVO. Secondly, these studies confirm the role of ANP and CNP but not BNP in the natriuretic and diuretic response to acute volume overload.

Enhanced endothelin-converting enzyme immunoreactivity in early atherosclerosis.

Endothelin-1 (ET-1) is a 21-amino-acid local and circulating factor whose plasma concentrations are increased in advanced atherosclerosis. ET-1 is cleaved from a prohormone (big ET-1) by endothelin-converting enzymes (ECEs) into the biologically active mature form which mediates vasoconstriction and cell proliferation. This study was designed to test by immunohistochemistry the hypothesis that ECE is present locally in the neointima of atherosclerotic vessels. Two groups of rabbits, control (n = 6) and cholesterol-fed (1% cholesterol diet for 8 weeks; n = 6) were sacrificed. Aortas were excised and divided for determination of tissue ET-1 concentration by RIA and immunohistochemical analysis of ECE. Vascular wall ET-1 was increased in the atherosclerotic aorta (6.1 +/- 0.8 vs. 9.8 +/- 0.9 pg/mg protein; p < 0.05), whereas circulating ET-1 concentrations were similar in the two groups (3.8 +/- 0.4 vs. 2.4 +/- 1.4 pg/ml). Immunostaining revealed the presence of ECE in endothelial and vascular smooth-muscle cells of the control group. Enhanced ECE immunoreactivity was present in atherosclerotic aortas, particularly in the neointimal macrophages and smooth-muscle cells. We conclude that local vascular wall, but not circulating ET-1, is increased in early atherosclerosis. In addition, ECE immunoreactivity is increased in early atherosclerosis and may therefore contribute to the generation of local ET-1 in early experimental atherosclerosis. These studies provide important insights into the regulation of ET-1 in early atherosclerosis, which may contribute to the elucidation of factors involved in the progression of atherosclerosis.

Role of prostaglandins and renal nerves in the renal actions of adrenomedullin.

Adrenomedullin (ADM), originally discovered in human pheochromocytoma, is also of renal cell origin and has natriuretic and diuretic actions. The present study was designed to investigate the role of prostaglandins and renal nerves in the renal hemodynamic and natriuretic actions ofADM. ADM was administered intrarenally (1, 5 and 25 ng x kg(-1) x min(-1)) with and without prostaglandin inhibition (meclofenamate, 5 mg/kg intravenous bolus) in anesthetized normal mongrel dogs (n = 5, each). To elucidate the role of renal nerves, ADM was administered intrarenally to the denervated kidney in five dogs. ADM mediated a natriuretic action via increases in glomerular filtration rate and decreases in distal tubular sodium reabsorption, which was attenuated by renal denervation and completely abolished by prostaglandin inhibition. The renal vasodilatation induced by ADM was attenuated by meclofenamate, as well as by renal denervation, although not significantly. Additionally, renal nerves mediated hemodynamic effects of hypertension that were produced by intrarenal infusion of ADM. This study establishes an important mechanistic role for renal prostaglandins as a mediator of ADM-mediated natriuresis at the level of the glomerulus and terminal nephron.

Blunted cGMP response to agonists and enhanced glomerular cyclic 3',5'-nucleotide phosphodiesterase activities in experimental congestive heart failure.

The natriuretic peptide (NP) and nitric oxide (NO) systems are activated in congestive heart failure (CHF), resulting in increased synthesis of cGMP, which serves as a second messenger for both humoral systems. These two regulatory systems play functional roles in the preservation of glomerular filtration rate (GFR) and sodium excretion in both acute and chronic CHF. A progressive decline in glomerular responsiveness to atrial natriuretic peptide (ANP) characterizes the terminal stage of chronic CHF despite elevation of plasma ANP. Phosphodiesterase isozymes (PDEs) are integral factors in determining cellular content and accumulation of cGMP, and up-regulation of PDE activity could participate in the glomerular resistance to ANP in severe CHF. To date, characterization of possible alteration of glomerular PDE isozyme activities in CHF is unknown, as is the in vitro glomerular response to the nitric oxide-soluble guanylyl cyclase pathway. We, therefore, first determined cGMP generation in response to particulate and soluble guanylyl cyclase activation by ANP and sodium nitroprusside (SNP) in isolated glomeruli from normal (N = 6) and CHF dogs (N = 5) in which CHF was induced by rapid ventricular pacing for 18 to 28 days. Secondly, we explored the presence of major PDE isozymes in glomeruli isolated from the control and CHF dogs. When ANP or SNP (10(-10) to 10(-4) M) were incubated with the suspension of isolated glomeruli, cGMP accumulation was lower by -72 to -96% with ANP and -42 to -77% with SNP in all glomerular medias obtained from CHF compared to controls. PDE hydrolyzing activity of both cAMP and cGMP were higher in the glomerular homogenates obtained from the kidneys of the CHF group (N = 5) compared to those of the control group (N = 5). We conclude that in severe chronic experimental CHF, glomerular cGMP accumulation decreases in response to both ANP and SNP, and CHF is characterized by enhanced cGMP- and cGMP-PDE activities that may participate in glomerular maladaptation to this cardiovascular syndrome.

Angiotensin II in the evolution of experimental heart failure.

Although angiotensin II (Ang II) has been implicated in the pathophysiology of congestive heart failure, its temporal and regional changes during the development and progression of the disease are poorly defined. Our objective was to assess circulating, renal, cardiac, and vascular Ang II in a canine model of rapid ventricular pacing-induced heart failure that evolves from early left ventricular dysfunction to overt congestive heart failure. Ang II was measured by radioimmunoassay with low cross-reactivity to other angiotensins. Control, early left ventricular dysfunction, and overt congestive heart failure dogs were studied. Early left ventricular dysfunction was characterized by impaired cardiac function, cardiac enlargement, preserved renal perfusion pressure, maintained urinary sodium excretion, and normal plasma renin activity. Overt congestive heart failure was characterized by further impaired cardiac function and cardiac enlargement, reduced renal perfusion pressure, urinary sodium retention, and increased plasma renin activity and plasma Ang II. In early left ventricular dysfunction dogs, renal cortical, renal medullary, ventricular, and aortic Ang II were unchanged, and atrial Ang II was decreased. In overt congestive heart failure dogs, Ang II was increased in the kidney and heart compared with normal dogs and in all tissues compared with early left ventricular dysfunction dogs. The greatest increase in tissue Ang II occurred in the renal medulla. We conclude that early increases in local renal, myocardial, and vascular Ang II do not occur in this model of early left ventricular dysfunction and may even be suppressed. In contrast, increased myocardial and particularly renal Ang II in association with increased circulating Ang II are hallmarks of overt experimental congestive heart failure. These studies provide new insights into the temporal and regional alterations in Ang II during the progression of experimental congestive heart failure.

Angiotensin converting enzyme inhibition modulates endogenous endothelin in chronic canine thoracic inferior vena caval constriction.

Endothelin (ET) is a potent vasoconstrictor peptide which is elevated in plasma in congestive heart failure. Recent studies suggest an important role for angiotensin II (AII) in the activation of ET in cultured cardiomyocytes. Chronic thoracic inferior vena caval constriction (TIVCC) is a model of reduced cardiac output that mimics the neurohumoral activation observed in congestive heart failure. We hypothesized that activation of the renin-angiotensin system in TIVCC plays a role in the activation of ET and that the elevation of endogenous ET contributes to the systemic and renal vasoconstriction that characterizes this model of venous congestion. We studied conscious dogs after 7 d of TIVCC in the presence or absence of chronic angiotensin converting enzyme inhibition with enalapril. TIVCC resulted in marked activation of plasma AII and ET in plasma, right atrium, lung, and renal medulla which was further localized to cardiomyocytes, pulmonary, and renal epithelial cells. Chronic angiotensin converting enzyme inhibition abolished the increases in plasma AII and ET during TIVCC. Acute endothelin A receptor blockade with FR-139317 resulted in significant decreases in mean arterial pressure and systemic vascular resistance in TIVCC. We conclude that activation of the renin-angiotensin system contributes to the activation of circulating and local ET in TIVCC and that this activation plays an important role in the regulation of arterial pressure and systemic vascular resistance in this model of congestive failure.

Role of natriuretic peptide clearance receptor in in vivo control of C-type natriuretic peptide.

C-type natriuretic peptide (CNP) is a newly described 22-amino acid peptide of endothelial cell origin, which has selective cardiovascular actions and is structurally related to atrial natriuretic peptide (ANP). Recent in vitro studies have demonstrated that an important regulatory pathway for the clearance of natriuretic peptides involves binding to a common clearance receptor [natriuretic peptide C receptor (NPR-C)]. Although CNP has also been identified as a ligand for NPR-C in binding assays, no studies have defined the in vivo interaction of CNP with NPR-C. CNP (10 ng.kg-1.min-1) followed by C-ANP-(4-23), a specific ligand for NPR-C blockade, was infused intravenously in two groups (both n = 7) of anesthetized dogs at two different doses (0.1 or 1.0 micrograms.kg-1.min-1) to permit calculation of total metabolic clearance rate (TMCR). C-ANP-(4-23) increased circulating CNP and reduced TMCR in both groups. Pulmonary metabolic clearance rate was negative at baseline, suggesting a net secretion of CNP across the lung, which was increased during CNP infusion and was abolished with NPR-C blockade. Renal and femoral metabolic clearance rates were positive at baseline and increased with CNP infusion. A decrease in cardiac output and cardiac filling pressures in response to CNP administration was potentiated by NPR-C blockade. We conclude that 1) circulating CNP achieved by CNP infusion is regulated by NPR-C in vivo, 2) the pulmonary circulation is a possible site of CNP secretion, 3) the renal and peripheral circulations are sites of CNP clearance, and 4) NPR-C blockade potentiates the selective cardiovascular actions of CNP.

Renal localization and actions of adrenomedullin: a natriuretic peptide.

Adrenomedullin (ADM) is a newly described 52-amino acid peptide originally isolated from extracts of human pheochromocytoma and, more recently, detected in human plasma. Based on the report that ADM mRNA and immunoreactivity are present in the kidney, the current study was designed to determine the renal distribution of ADM by immunohistochemistry and the renal biological actions of ADM. In the immunohistochemical studies, the present investigation demonstrated the localization of ADM in glomeruli, cortical distal tubules, and medullary collecting duct cells of the normal canine kidney. In the in vivo studies, ADM was administered (0.25 ng.kg-1.min-1 in group I and 1, 5, and 25 ng.kg-1.min-1 in group II) intrarenally in normal mongrel dogs with the contralateral kidney receiving only saline vehicle. Intrarenal infusion of ADM resulted in a marked diuretic and natriuretic response, whereas the contralateral kidney showed no renal effects. These significant natriuresis and diuresis in the ADM kidney were associated with increases in glomerular filtration rate and fractional sodium excretion and with a decrease in distal tubular sodium reabsorption. Intrarenal infusion of ADM also caused an increase in mean arterial blood pressure and a decrease in heart rate. Plasma concentrations of atrial natriuretic peptide, renin activity, aldosterone, and guanosine 3',5'-cyclic monophosphate were not changed during the infusion of ADM. The current study demonstrates that ADM is present in renal glomerular and tubular cells and is a potent natriuretic peptide that may play an important role in the regulation of sodium excretion.

Presence and secretion of atrial natriuretic peptide from cultured human aortic endothelial cells.

The endothelium is the production site of several potent vasoactive substances that modulate vascular tone and growth. The present study was undertaken to investigate the presence and secretion of atrial natriuretic peptide (ANP) immunoreactivity from vascular endothelial cells. ANP immunoreactivity was present in cultured human aortic endothelial cells by both immunohistochemical staining and radioimmunoassay. ANP immunoreactivity was also detectable in culture medium from human aortic endothelial cells in low picogram concentrations. These findings suggest that vascular endothelium is a site of ANP production and secretion of ANP. There was a differential distribution of ANP and endothelin-1 (ET-1), with a higher ANP concentration in cell extracts and a higher ET-1 concentration in cell culture media. Although ANP has been conceived as a circulating endocrine hormone, these findings are consistent with ANP functioning also as an autocrine and paracrine modulator in the regulation of vascular tone and growth.

Immunohistochemical localization of adrenomedullin in canine heart and aorta.

Adrenomedullin (ADM) is a new endogenous hypotensive and vasorelaxing peptide that may play an important role in the regulation of cardiovascular function. Although ADM was originally isolated from pheochromocytoma, ADM-like immunoreactivity has also been widely detected in various tissues, including the cardiovascular system. Based upon the reports that ADM mRNA and ADM-like immunoreactivity are present in the heart, the present study was designed to investigate the immunohistochemical localization of ADM in the canine heart and aorta. In the canine heart, immunohistochemical examination revealed positive immunostaining within the myocardia in both atria and ventricles. ADM immunoreactivity was observed within the cytoplasm of myocardium, and was widely distributed in the peripheral cytoplasm. ADM immunoreactivity was more intense in the atria than in the ventricles. In the canine aorta, vascular smooth muscle cells of the aorta and vasa vasorum were also immunopositive for ADM. ADM immunoreactivity was mostly localized in the perinuclear position within the smooth muscle cells. There was no immunoreactivity in endothelium, endocardium, epicardium, adventitia, or connective tissues. The current study demonstrates for the first time that immunoreactive ADM by immunohistochemistry is present in the cardiovascular system. As ADM has hypotensive and vasorelaxing actions and circulates in the body, ADM is a cardiovascular peptide hormone that may play an important role in the regulation of cardiovascular system.

CNP is present in canine renal tubular cells and secreted by cultured opossum kidney cells.

C-type natriuretic peptide (CNP) is a vasoactive and antimitogenic peptide that is structurally similar but genetically distinct from atrial natriuretic peptide. While first discovered in the brain, CNP has been shown to be produced by endothelial cells and may function in a paracrine and autocrine fashion in the control of vascular tone. Recently, CNP immunoreactivity and B-type natriuretic peptide receptors (NPR-B), for which CNP is a specific ligand, have been identified in the kidney. The present study was designed to determine whether renal epithelial cells produce and secrete CNP and whether CNP immunoreactivity is present in canine kidney. Opossum kidney (OK) cells that express proximal tubular cell characteristics were incubated for 6 h in fetal calf serum-free Dulbecco's modified Eagle's medium (DMEM). CNP immunoreactivity was measured in the preincubation and 6-h conditioned media by radioimmunoassay (RIA) using a specific antibody to CNP-22. Furthermore the molecular form of this CNP-like protein was determined by reverse-phase high-performance liquid chromatography (HPLC), and intracellular localization of the CNP immunoreactivity was determined by immunohistochemical staining. CNP immunoreactivity was also determined in renal tissue from dogs subjected to saline or endothelin infusion. Six-hour incubation in DMEM resulted in accumulation of CNP immunoreactivity (baseline below detection level vs. 6 h = 117.3 +/- 8.3 pg/ml, P < 0.001). Intracellular CNP concentration determined after sonication was 1.9 +/- 0.2 micrograms/g protein, and immunohistochemical staining for CNP was markedly positive in the cytoplasm.(ABSTRACT TRUNCATED AT 250 WORDS)

Presence of C-type natriuretic peptide in human kidney and urine.

The current study was undertaken to investigate the presence of CNP immunoreactivity in both human kidney and urine. Immunohistochemical staining with an indirect immunoperoxidase method utilizing an antibody which is 100% cross-reactive to both CNP-53 and CNP-22 was performed on five human kidney specimens (three biopsies of normal cadaveric donor kidneys and two of normal autopsy specimens). CNP immunoreactivity was positive in proximal, distal and medullary collecting duct tubular cells in a cytoplasmic and granular staining pattern. CNP immunoreactivity was also determined in the urine of five healthy volunteers utilizing a sensitive and specific double-antibody radioimmunoassay with a mean concentration of 10.8 +/- 1.0 pg/ml. With the utilization of high pressure liquid chromatography, this immunoreactivity proved to be consistent with both the low molecular weight form, CNP-22, as well as the high molecular weight form, CNP-53. Urinary excretion of CNP was also measured in normal subjects (N = 5) and in patients with congestive heart failure (CHF, N = 6). CHF patients excreted over three times more CNP than normals (27.2 +/- 2.8 vs. 8.7 +/- 0.81 pg/min, P < 0.004) despite no difference between the two groups in plasma CNP concentrations (6.97 +/- 0.28 vs. 8.08 +/- 1.52 pg/ml, P = NS). This study demonstrates for the first time the presence of CNP immunoreactivity in human kidney and suggests that renal tubular cells may be an additional non-vascular site of synthesis for this cardiorenal acting peptide. This study also demonstrates an increase in urinary CNP excretion in congestive heart failure.

Acute hypoxia and endogenous renal endothelin.

Endothelin (ET) is a potent vasoconstrictor peptide of endothelial cell origin. Recent studies have suggested a nonvascular paracrine and/or autocrine role for endothelin in the kidney. This study was designed to elucidate the renal ET response to acute moderate hypoxia, as reflected by urinary ET excretory rate and renal tissue ET immunoreactivity, and to correlate these responses to the hemodynamic and excretory changes during hypoxia. Experiments were conducted in two groups of anesthetized dogs: hypoxic group (10% O2 ventilation: PO2, 44 mm Hg; N = 7) and time control group (room air ventilation: PO2, 111 mm Hg; N = 6). After 60 min of hypoxia or room air ventilation, kidneys were harvested and stained immunohistochemically for ET. Acute moderate hypoxia was associated with significant increases in urinary ET excretion, urine flow, urinary sodium excretion, and fractional excretion of sodium (P < 0.05). There was no significant change in GFR, RBF, renal vascular resistance, or mean arterial pressure. Renal immunohistochemistry for ET revealed increased staining in the proximal and distal tubules in the hypoxic group as compared with controls. This study demonstrates that acute moderate hypoxia results in increased urinary ET excretion and renal tubular ET immunoreactivity, in association with diuresis and natriuresis, and suggests a nonvascular role of endogenously produced renal ET in the regulation of sodium homeostasis during hypoxia.