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S Sabatini

Publications and source records attributed to S Sabatini.

At least 73 records · Page 4Linked to original sources
PubMed1990

NEM-sensitive ATPase activity in rat nephron: effect of metabolic acidosis and alkalosis.

The present study was designed to quantitate the amount and to map the localization of N-ethylmaleimide (NEM)-sensitive adenosinetriphosphatase (ATPase) activity in microdissected segments of the rat nephron. After complete nephron mapping the effect of chronic metabolic acidosis and alkalosis on enzyme activity was determined. In control animals the highest enzyme activity was found in the early proximal convoluted tubule of juxtamedullary nephrons; superficial early proximal tubule as well as medullary and cortical thick ascending limbs and collecting ducts also contained substantial activity. Enzyme activity in the papillary collecting duct before entry into the ducts of Bellini was 329 +/- 93 pmol.mm-1.h-1 (n = 8); after entry, however, enzyme activity was approximately one-fourth that value (60 +/- 9 pmol.mm-1.h-1, n = 8, P less than 0.01). No NEM-sensitive ATPase activity was found in the thin limbs of the loop of Henle. Enzyme activity increased in both the medullary and cortical thick ascending limbs as well as in the cortical collecting tubule in response to NH4Cl-induced chronic metabolic acidosis; in the cortical collecting duct, metabolic acidosis increased maximum activity (Vmax) but did not change Michaelis-Menten constant (Km). In the proximal convoluted tubule, enzyme activity decreased with metabolic acidosis. Bicarbonate loading had no effect on enzyme activity except in the most distal portion of the collecting duct where it was stimulated. These results show that NEM-sensitive ATPase activity exists throughout much of the rat nephron. These data suggest that both the cortical collecting tubule and thick ascending limb are regulatory sites of distal urinary acidification during acid loading.

Acidosis↗
PubMed1989

Peculiar effects of temperature and polyvinylalcohol on the activity of bovine serum amine oxidase.

An inflexion point of enzyme activity at 38 - 42 degrees C of the bovine serum amineoxidase was found. This result, associated with non-strict Arrhenius curves and slightly different activation energies in various temperature intervals, suggests some conformational transitions at the mentioned temperatures. The high molecular weight polyvinylalcohol (100,000 Da) generated an activatory effect and a sigmoidal (non-Michaelis) curve of the dependence of the activity on the substrate concentrations, while the low molecular weight polyvinylalcohol (20,000 Da) does not produce this effect. The different ratio of the two types of polyvinylalcohol/enzyme monomer sizes is considered to be responsible for these different effects on the enzyme kinetics.

Amine Oxidase (Copper-Containing)↗
PubMed1989

Production of antibodies against the coenzyme pyrrolequinoline quinone.

Polyclonal antibodies against pyrrolequinoline quinone have been elicited in rabbits. These antibodies react with free and protein-bound pyrrolequinoline quinone. In particular they react with native and denatured lentil seedling amine oxidase as detected by dot-blot and ELISA assays. The presence of 1 mol pyrrolequinoline quinone per mol of enzyme was determined by the last method.

Amine Oxidase (Copper-Containing)↗
PubMed1989

Dimethyl sulfoxide affects water flow through a nonosmolar action.

Dimethyl sulfoxide (DMSO) is a dipolar organic compound commonly used as a solvent in studies of membrane transport. DMSO also has many effects on cell function and, although the precise mechanism of action is not known completely, it has been stated to exert its effect on transport solely through osmolality. The present study was designed to examine the effects of serosal DMSO at three osmolar concentrations on Basal Water Flow and vasopressin (AVP)- and cyclic AMP-stimulated water flow (Maximal Water Flow) in the toad bladder. The results obtained were compared to equi-osmolar concentrations of mannitol and NaCl. All three agents significantly enhanced Basal Water Flow after 60 min. The results obtained on Maximal Water Flow were different depending on the final osmolality. At 300 mOsm final concentration, all three agents increased AVP-stimulated water flow. When the serosal osmolality was 600 or 900 mOsm DMSO increased Maximal Water Flow, whereas mannitol and NaCl decreased it. When 300 mOsm DMSO plus 300 mOsm mannitol (600 mOsm total)-treated hemibladders were challenged with AVP, the water flow response was similar to that of 600 mOsm DMSO alone. In the presence of verapamil, AVP-stimulated water flow was decreased markedly; when DMSO was added to verapamil-pretreated hemibladders, and they were then challenged with AVP, water flow increased significantly. In similar experiments with mannitol, water flow remained inhibited. Dimethylsulfone did not enhance AVP-stimulated water flow as compared to the same concentration of DMSO. These results demonstrate that the effects of DMSO on water transport are not mediated solely by its osmolar action.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1989

Vanadate stimulates the N-ethylmaleimide-sensitive adenosine triphosphatase in rat nephron.

Vanadate has been used in many cellular systems to elucidate mechanisms of enzyme action. Vanadate inhibits Na-K adenosine triphosphatase (ATPase) activity in many tissues. In isolated collecting tubule it inhibits sodium transport and vasopressin-stimulated water flux, the latter presumably distal to cyclic AMP formation. Depending upon the tissue studied, vanadate also stimulates a variety of cellular reactions including adenylate cyclase, glucose oxidation and glycogen synthesis. We studied the effect of varying concentrations of vanadate on N-ethylmaleimide (NEM)-sensitive ATPase activity in microdissected segments of rat nephron. In proximal convoluted tubule and in cortical, medullary and papillary collecting ducts vanadate had no effect on enzyme activity. In medullary and cortical thick ascending limbs, however, vanadate significantly stimulated NEM-sensitive ATPase activity (medullary thick ascending limb, 241 +/- 14 pmol/mm/hr vs. 531 +/- 74 pmol/mm/hr; control vs. (1 mM) vanadate, respectively; n = 14, P less than 0.01). The stimulatory effect of vanadate on NEM-sensitive ATPase activity was present at 5 microM vanadate, a concentration that inhibited Na-K ATPase activity approximately 80%. Metabolic acidosis also stimulated enzyme activity in the thick ascending limb, and the effect of vanadate was not additive. Metabolic alkalosis had no effect on NEM-sensitive ATPase in the thick ascending limb, but the stimulatory effect of vanadate was still seen. These data document that the NEM-sensitive ATPase in thick ascending limb is different from that found in other nonmammalian proton secretory epithelia which are vanadate inhibitable. The results with vanadate plus metabolic acidosis suggest that both are acting via the same mechanism.

Acidosis↗
PubMed1989

Effect of phthalate acid esters on transport in toad bladder membrane.

Sclerosing peritonitis is a serious complication in patients on long-term peritoneal dialysis; it markedly decreases transport of water and solute across the peritoneal membrane. Although the precise mechanism is unknown, organic compounds (i.e., plasticizers) from plastic tubing and dialysis bags have been suggested to be a cause of the syndrome. The effects of three such compounds on water and sodium transport in vitro were studied in the toad bladder. The compounds studied were didodecylphthalate, dioctylphthalate, and benzylbutylphthalate. After 4 hr incubation in vitro, dioctylphthalate and benzylbutylphthalate significantly inhibited vasopressin-stimulated water flow in toad bladder. Basal water flow was not affected by any of the three compounds. Sodium transport, as measured using short-circuit current, was decreased to an equivalent degree by all compounds; inhibition of short-circuit current was dose dependent and was approximately 30% at 10(-3) M. The onset of action was between 3.5 and 4 hr, and the effect on short-circuit current was not reversible. These results demonstrate that the plasticizers (to which patients of all sorts are commonly exposed) inhibit transport across living membranes. In the toad bladder these compounds decrease sodium transport and maximal water flow. Although other evidence suggests that the cumulative toxic effects of these compounds may play a causal role in sclerosing peritonitis in patients on peritoneal dialysis, our study suggests that chronic exposure to the phthalate acid esters in patients with normal renal function may result in sodium wastage, polyuria, and a concentrating defect resistant to AVP.

Animals↗
PubMed1989

Cyanide poisoning: pathophysiology and current approaches to therapy.

Considering the difficulties following the administration of nitrites (or aminophenols) or cobalt-EDTA as well as the ineffectivenes of hydroxycobalamin and pyruvate, we feel that a more sensible treatment for acute cyanide intoxication is hemodialysis combined with the intravenous administration of sodium thiosulfate. The addition of hemodialysis to such a regimen is helpful in three ways. First, it removes the small extracellular reservoir of cyanide, particularly if the poison is still being absorbed from the gastrointestinal tract. Second, it corrects the severe lactic acidosis seen in virtually all cases of cyanide toxicity. As with most poisons death from metabolic acidosis alone is likely. Third, the removal of thiocyanate, the end product of cyanide metabolism, results in a maneuver which should decrease both tissue and plasma cyanide levels. The immediate treatment of acute cyanide intoxication is supportive, as it is with most all drugs and poisons. Gastric lavage using activated charcoal should be initiated immediately to remove any remaining cyanide in the gastrointestinal tract. Simultaneously, high flow oxygen should be administered either by nasal cannula or by endotracheal intubation. Correction of the metabolic acidosis should be instituted with bicarbonate. Immediate hemodialysis should be performed with the concomitant administration of thiosulfate. Animal studies suggest that continuous infusion of thiosulfate (12 mg/kg/hr) is more effective for treating cyanide intoxication than is bolus administration (41, 51). Bolus administration is the currently recommended form of thiosulfate therapy in humans. The Lilly kit contains a 50 ml ampule of thiosulfate having 12.5 gm, which in adults may be repeated once at one-half the dose.(ABSTRACT TRUNCATED AT 250 WORDS)

Cyanides↗
PubMed1988

Spectroscopic studies of the reaction between bovine serum amine oxidase (copper-containing) and some hydrazides and hydrazines.

The carbonyl cofactor of bovine serum amine oxidase, recently identified as pyrroloquinoline quinone [Ameyama, Hayashi, Matsushita, Shinagawa & Adachi (1984) Agric. Biol. Chem. 48, 561-565; Lobenstein-Verbeek, Jongejan, Frank & Duine (1984) FEBS Lett. 170, 305-309], reacts stoichiometrically and irreversibly with hydrazides of phenylacetic acid and of benzoic acid. With the phenylacetic hydrazides a reversible intermediate step was detected by competition with substrate, carbonylic reagents or phenylhydrazine, a typical inhibitor of the enzyme. All hydrazides form an intense broad band with maximum absorbance in a narrow wavelength range (350-360 nm), irrespective of the acyl group, suggesting that the transition is located on the organic cofactor. A different situation is found with some phenylhydrazines, where extended conjugation can occur between the cofactor and the phenyl pi-electron system via the azo group, as shown by the lower energy and higher intensity of the transition. In this case the transition is sensitive to substituents in the phenyl ring. The c.d. spectrum of the adducts is influenced by the type of hydrazide (derived from phenylacetic acid or benzoic acid), by pH and by NN-diethyldithiocarbamate binding to copper, probably as a result of shifts of equilibria between hydrazone-azo tautomers.

Amine Oxidase (Copper-Containing)↗
PubMed1988

Inhibition of copper-dependent amine oxidases by some hydrazides of pyrrol-1-ylbenzoic and pyrrol-1-ylphenylacetic acids.

Some hydrazides of pyrrol-1-ylbenzoic and pyrrol-1-ylphenylacetic acids were prepared, and their effect on copper-dependent amine oxidases (Cu-AOs) and FAD monoamine oxidases (MAOs) activities was tested. The compounds were not substrates for Cu-AO enzymes but acted as noncompetitive inhibitors. Hydrazides of pyrrol-1-ylphenylacetic acids were highly specific for plasma amine oxidase (Ki = 0.5-1 microM). In contrast, all the hydrazides were weak inhibitors of MAO activity. Incubation with the hydrazide derivatives led to irreversible inactivation of Cu-AOs. Therefore, the inhibition implied two distinct steps. The first one consisted of the rapid formation of the enzyme-inhibitor complex and was reversed by dialysis. In the second step, the complex was irreversibly transformed, probably by the formation of a Schiff base between the hydrazide and the prosthetic carbonyl group of the enzyme.

Amine Oxidase (Copper-Containing)↗
PubMed1988

Thiazides stimulate calcium absorption in the turtle bladder.

The mechanism of action of the thiazide diuretics on calcium transport is not completely understood. The present study was designed to examine the effect of hydrochlorothiazide (HTZ) on Na transport, proton secretion, and Ca45 flux in the turtle bladder, a high resistance membrane. When added to the mucosal solution, 1 mM HTZ had no effect on Na transport or proton secretion, but significantly increased mucosa-to-serosa Ca45 flux at 60 minutes (control 118.9 +/- 39.7 pmol/mg/60 min versus thiazide 286.0 +/- 64.9 pmol/mg/60 min, N = 16, P less than 0.02). In the presence of 5 X 10(-4) M ouabain, a maneuver which inhibits active Na transport, HTZ again significantly enhanced mucosa-to-serosa Ca45 flux. The increment of calcium transport under these conditions was 83.4 +/- 35.2 pmol/mg/60 min (N = 8, P less than 0.05). Mucosal HTZ had no effect on serosa-to-mucosa (that is, bath-to-lumen) Ca45 flux after a 60 minute incubation. Serosal addition of HTZ had no effect on either of the unidirectional Ca45 fluxes or on Na transport. Mucosal tissue Ca45 content was enhanced by HTZ (mucosal) in the presence or absence of ouabain. These results provide support for the view that thiazides have a direct stimulatory effect on calcium absorption at the luminal membrane, perhaps secondary to increased mucosal calcium permeability.

Animals↗
PubMed1988

On the mechanism of toluene-induced renal tubular acidosis.

This study was aimed to investigate the pathogenesis of toluene-induced renal tubular acidosis (RTA). In 5 individuals addicted to toluene sniffing we documented the occurrence of hypokalemia and hyperchloremic metabolic acidosis associated with inability to lower urine pH below 5.5 (6.06 +/- 0.24). Overall kidney bicarbonate reabsorption was normal or enhanced, a feature characteristic of the distal form of RTA (DRTA). These findings resemble those found during the administration of amphotericin B, a drug felt to cause DRTA by increasing hydrogen ion (H+) back-diffusion in the collecting tubule. In toluene sniffers, the urine pCO2 measured in a highly alkaline urine was reduced (47 +/- 8.8 mm Hg), suggesting a decrease in the rate of collecting tubule H+ secretion rather than H+ back-diffusion. To investigate these two mechanisms of altered distal acidification more directly we studied the effect of toluene on acidification by the urinary turtle bladder, an epithelial analogue of the mammalian collecting tubule. In this preparation, toluene resulted in a decrease in the rate of H+ secretion measured by either the pH stat technique or the reverse short circuit current. When mucosal pH was progressively lowered to examine H+ secretion against an H+ gradient, toluene-treated bladders displayed a significant decrease in proton conductance but the lowest mucosal pH required to nullify H+ secretion, (MpH) JH = O, was not different from that of control bladders (4.05 +/- 0.29 and 3.90 +/- 0.13, respectively).(ABSTRACT TRUNCATED AT 250 WORDS)

Acidosis, Renal Tubular↗