Further biochemical characterization of an Na<sup>+</sup> pump inhibitor purified from human urine

Crabos, Maryse; Grichois, Marie‐Laure; Guicheney, Pascale; Wainer, Irving W.; Cloix, Jean‐François

doi:10.1111/j.1432-1033.1987.tb10552.x

Cited by 22 publications

(5 citation statements)

References 42 publications

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“…From these studies, digoxin (9) and "iso-ouabain" (10-13) have been isolated; however, digoxin and ouabain lead to kaliuresis (4,(14)(15)(16)(17). Therefore, we hypothesized that this search tool is an inadequate marker for natriuresis.…”

Section: Introductionmentioning

confidence: 99%

A new endogenous natriuretic factor: LLU-alpha.

Wechter

Kantoci

Murray

et al. 1996

Proc. Natl. Acad. Sci. U.S.A.

180

126

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For over three decades, renal physiology has sought a putative natriuretic hormone (third factor) that might control the body's pool of extracellular fluid, an important determinant in hypertension, congestive heart failure, and cirrhosis. In our search for this hormone, we have isolated several pure natriuretic factors from human uremic urine that would appear, alone or in combination, to mark a cluster of phenomena previously presumed to be that of a single "natriuretic hormone." This paper reports the purification, chemical structure, and total synthesis of the first of these compounds, LLU-a, which proved to be 2,7,8-trimethyl-2-(pcarboxyethyl)-6-hydroxychroman, presumably a metabolite of y-tocopherol. Both natural LLU-a and synthetic material are identical (except for optical activity) with respect to structure and biological activity. It appears that the natriuretic activity of LLU-a is mediated by inhibition of the 70 pS K+ channel in the apical membrane of the thick ascending limb of the kidney.As a result of salt-induced plasma-volume expansion in mammals, three concurrent events have been observed: sustained natriuresis, rising plasma concentration of a Na+ transport inhibitor, and pressor activity. It has been presumed that these effects are due to elaboration of a low-molecular-weight "natriuretic hormone," the putative controller of extracellular fluid (1). Over 30 years of effort have failed to characterize the putative hormone (2, 3). Atrial natriuretic factor, when infused, produces a natriuresis and a decrease in blood pressure that are short-lived (4, 5) and whose physiological role is still undefined (6, 7).The approach of essentially all other workers has been to investigate mammalian-derived isolates that inhibit the Na+ pump (2,8). From these studies, digoxin (9) and "iso-ouabain" (10-13) have been isolated; however, digoxin and ouabain lead to kaliuresis (4,(14)(15)(16)(17). Therefore, we hypothesized that this search tool is an inadequate marker for natriuresis. Consequently, a natriuretic assay has been developed in which the in vivo physiological events, urine volume, K+ and Na+ concentrations, and mean arterial pressure are measured (18).In this paper, we report the isolation of pure LLU-a, the determination of its structure by spectroscopy, its synthesis in racemic form, and its biological characterization. Given its structure, we infer that it is the product of in vivo oxidative metabolism of y-tocopherol, a member of the vitamin E complex. MATERIALS AND METHODSPurification of LLU-a. Human uremic urine (800 liters) was collected, processed by ultrafiltration (3 kDa) and lyophilization, and then chromatographed on Sephadex G-25 to obtain the post-salt fraction as described (18 Step 1. Reversed-phase HPLC was performed by gradient elution with 0.2 M pyridinium acetate, pH 5.5, and methanol as described (4). Based on bioassay data, fractions 50-80 were combined for further purification.Step 2. The next column (5 pm; 10 x 250 mm) was eluted at 2 ml/min with a gradient formed from...

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Section: Introductionmentioning

confidence: 99%

A new endogenous natriuretic factor: LLU-alpha.

Wechter

Kantoci

Murray

et al. 1996

Proc. Natl. Acad. Sci. U.S.A.

180

126

View full text Add to dashboard Cite

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“…Kramer et al (30), studying a semi-purified urinary inhibitor, described the NH inhibition of Na-K-ATPase as noncompetitive with respect to potassium. Crabos et al (31) reported that NH, also isolated from urine, inhibited Na-K-ATPase noncompetitively with respect to ATP. Inhibition of the enzyme system was facilitated by sodium.…”

Section: Discussionmentioning

confidence: 99%

“…Inhibition of the enzyme system was facilitated by sodium. Ouabain, a digitalis glycoside and a specific inhibitor of the Na-K-ATPase system, has been shown to be competitive with respect to potassium (32) and noncompetitive with respect to ATP (31). In the present study, we employed a semi-purified NH preparation derived from plasma of a patient with essential hypertension to perform kinetic studies.…”

Section: Discussionmentioning

confidence: 99%

Effects of lead and natriuretic hormone on kinetics of sodium-potassium-activated adenosine triphosphatase: possible relevance to hypertension.

Weiler

Khalil-Manesh

Gonick

1988

Environ Health Perspect

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Inhibition of vascular smooth muscle sodium-potassium-activated adenosine triphosphatase (Na-K-ATPase) has been postulated as a central mechanism in enhancing vascular contractility. In the present study, kinetics of inhibition of Na-K-ATPase by lead, ouabain, and natriuretic hormone (NH) was studied in a purified hog cerebral cortex enzyme preparation. Determination of I50 values for lead, ouabain, and NH revealed that NH is the most potent inhibitor of the enzyme system (0.8 x 10(-6) M ouabain equivalents). Kinetic analyses indicated that lead and NH exhibited different inhibitory mechanisms. The inhibition by lead was noncompetitive with respect to potassium and competitive with respect to sodium and MgATP. Natriuretic hormone was noncompetitive with respect to potassium, uncompetitive with respect to MgATP, and exhibited no inhibitory effect with respect to sodium. Synergism between lead and NH in the inhibition of Na-K-ATPase raises the possibility that lead may be a contributory factor in hypertension via this mechanism.

show abstract

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Effects of Lead and Natriuretic Hormone on Kinetics of Sodium-Potassium-Activated Adenosine Triphosphatase: Possible Relevance to Hypertension

Weiler¹,

Khalil-Manesh²,

Gonick³

1988

Environmental Health Perspectives

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Further biochemical characterization of an Na⁺ pump inhibitor purified from human urine

Cited by 22 publications

References 42 publications

A new endogenous natriuretic factor: LLU-alpha.

A new endogenous natriuretic factor: LLU-alpha.

Effects of lead and natriuretic hormone on kinetics of sodium-potassium-activated adenosine triphosphatase: possible relevance to hypertension.

Effects of Lead and Natriuretic Hormone on Kinetics of Sodium-Potassium-Activated Adenosine Triphosphatase: Possible Relevance to Hypertension

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Further biochemical characterization of an Na+ pump inhibitor purified from human urine

Cited by 22 publications

References 42 publications

A new endogenous natriuretic factor: LLU-alpha.

A new endogenous natriuretic factor: LLU-alpha.

Effects of lead and natriuretic hormone on kinetics of sodium-potassium-activated adenosine triphosphatase: possible relevance to hypertension.

Effects of Lead and Natriuretic Hormone on Kinetics of Sodium-Potassium-Activated Adenosine Triphosphatase: Possible Relevance to Hypertension

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Further biochemical characterization of an Na⁺ pump inhibitor purified from human urine