The influence of lipoic acid on adriamycin induced nephrotoxicity in rats

Adriamycin, which is widely used in the treatment of various neoplastic conditions, exerts toxic effects in several organs. Adriamycin nephrotoxicity has been recently documented in a variety of animal species. The present study was designed to investigate the effect of lipoic acid on the nephrotoxic potential of adriamycin. The study was carried out with adult male albino rats of Wistar strain. Test animals were divided into four groups of six rats each as follows: Group I (control) received only normal saline throughout the course of the experiment. Group II (ADR) received intravenous injections of adriamycin through the tail vein (1 mg kg^–1 body wt day^–1) once a week for a period of 12 weeks. Group III (LA) received lipoic acid (35 mg kg^–1 body wt day^–1) intraperitoneally once a week for a period of 12 weeks. Group IV (ADR + LA) received a single injection of lipoic acid intraperitoneally 24 h prior to the administration of adriamycin through the tail vein once a week for a period of 12 weeks. Intravenous injections of adriamycin resulted in decreased activities of the glycolytic enzymes; hexokinase, phosphoglucoisomerase, aldolase and lactate dehydrogenase in the rat renal tissue. The gluconeogenic enzymes; glucose-6-phosphatase and fructose-1,6-diphosphatase, showed a decline in their activities on adriamycin administration. The transmembrane enzymes namely the Na⁺,K⁺-ATPase, Ca²⁺-ATPase, Mg²⁺-ATPase and the brush-border enzyme alkaline phosphatase also showed a decrease in their activities. This decrease in the activities of ATPases and alkaline phosphatase suggests basolateral and brush-border membrane damage. Decreased activities of the TCA cycle enzymes isocitrate dehydrogenase, succinate dehydrogenase and malate dehydrogenase, suggest a loss in mitochondrial function and integrity. Nephrotoxicity was evident from the increased excretions of N-acetyl--D-glucosaminidase and -glutamyl transferase in the urine of adriamycin administered rats. These biochemical disturbances were effectively counteracted on pretreatment with lipoic acid, which brought about an increase in the activities of glycolytic enzymes, ATPases and the TCA cycle enzymes. On the other hand, the gluconeogenic enzymes showed a further decrease in their activities on lipoic acid pretreatment. LA pretreatment also restored the activities of the urinary enzymes to normal. These observations shed light on the nephroprotective action of lipoic acid rendered against experimental aminoglycoside toxicity.     

Renal peroxidative changes mediated by oxalate: the protective role of fucoidan

Oxalate, one of the major constituents of renal stones is known to induce free radicals which damage the renal membrane. Damaged epithelia might act as nidi for stone formation aggravating calcium oxalate precipitation during hyperoxaluria. In the present study, the beneficial effects of fucoidan on oxalate-induced free radical injury were investigated. Male Wistar rats were divided into four groups. Hyperoxaluria was induced in two groups by administration of 0.75% ethylene glycol in drinking water for 28 days and one of them was treated with fucoidan from Fucus vesiculosus at a dose of 5 mg/kg b.wt subcutaneously commencing from the 8th day of induction. A control and drug control (fucoidan alone) was also included in the study. The extent of renal injury in hyperoxaluria was evident from the increased activities of alkaline phosphatase, gamma-glutamyl transferase, beta-glucuronidase, N-acetyl-beta-D-glucosaminidase in urine. There was a positive correlation between plasma malondialdehyde levels and renal membrane damage indicating a striking relation between free radical formation and cellular injury. Increased protein carbonyl and decreased thiols further exemplified the oxidative milieu prevailing during hyperoxaluria. Decreased renal membrane ATPases accentuated the renal membrane damage induced by oxalate. Renal microscopic analysis showed abnormal findings in histology as an evidence of oxalate damage. The above biochemical and histopathological discrepancies were abrogated with fucoidan administration, indicating its protective role in oxalate mediated peroxidative injury.     

Attenuation of oxalate-induced nephrotoxicity by eicosapentaenoate-lipoate (EPA-LA) derivative in experimental rat model

Hyperoxaluria is one of the major risk factors for the formation of urinary calcium oxalate stones. Calcium oxalate crystals and their deposition have been implicated in inducing renal tubular damage. Lipoic acid (LA) and eicosapentaenoic acid (EPA) have been shown to ameliorate the changes associated with hyperoxaluria. This prompted us to investigate the nephroprotectant role of EPA-LA, a new derivative, in vivo in hyperoxaluric rats. Elevation in the levels of calcium, oxalate and phosphorus, the stone-forming constituents, were observed in calculogenic rats as a manifestation of crystal deposition.Tubular damage to the renal tissue was assessed byassaying the excretion of marker enzymes in the urine. Damage to the tubules was indicated by increased excretion of alkaline phosphatase (ALP), lactate dehydrogenase (LDH), gamma-glutamyl transferase (gamma-GT), beta-Glucuronidase (beta-GLU) and N-Acetyl beta-D glucosaminidase (NAG). Fibrinolytic activity was found to be reduced. Administration of EPA, LA and EPA-LA reduced the tubular damage and decreased the markers of crystal deposition markedly, which was substantiated by the reduction in weight of bladder stone formed. Our results highlight that EPA-LA is the most effective drug in inhibiting stone formation and mitigating renal damage caused by oxalate toxicity, thus confirming it as a nephroprotectant. Further work in this direction is warranted to establish the therapeutic effectiveness of this new derivative.     

Protection by selenium against gentamicin-induced acute renal damage in the rat

Gentamicin has been shown to induce renal tubular damage in man and laboratory animals and to result in elevated urinary excretion of some enzymes associated with specific cell regions in the kidney. In the present investigation, the possible protective effect of selenium against gentamicin-induced renal damage was tested by measuring the urinary excretion of some enzymes in the presence and absence of selenium. Our results show that a prior subcutaneous injection of selenium to rats for two days followed by a simultaneous S.C. injection of gentamicin and selenium resulted in a marked reduction in the excretion of such biochemical systems as the urine volume, urinary proteins, alkaline and acid phosphatases, beta-glucuronidase, muramidase, and glutamate dehydrogenase. Renal functional studies revealed that selenium-treated rats suffered less adverse effects compared to rats treated with gentamicin alone. Urinary acid phosphatase, beta-glucuronidase and muramidase, the three lysosomal enzymes tested, appeared to respond most readily to protection by selenium.     

Inhibition of polyamine synthesis blocks urinary secretion of beta-glucuronidase from mouse kidney

The effect of inhibition of polyamine synthesis on castrated male mouse kidney beta-glucuronidase induction and secretion by testosterone was studied. Inhibition of the activities of polyamine synthesis key-enzymes, L-ornithine and S-adenosyl-L-methionine decarboxylases, was performed with the combined treatment of 2-difluoromethylornithine and methylglyoxal' bis(guanylhydrazone). Blockage of polyamine synthesis did not affect testosterone-induced increase in renal beta-glucuronidase but blocked its secretion into the urine. After withdrawal of inhibitor-treatment beta-glucuronidase secretion normalized, and repeated testosterone administration produced undisturbed beta-glucuronidase secretion peak in urine suggesting that blockage of beta-glucuronidase secretion was not due to the tissue damage produced by inhibitors. These results indicate that the stimulation of renal polyamine synthesis by testosterone is not necessary for the induction of beta-glucuronidase but is required for the urinary secretion of this protein.     

Human parathyroid hormone (1-34) increases urinary excretion of lysosomal enzymes in rats

The kidney is the major target of parathyroid hormone (PTH), and PTH influences the urinary excretion of calcium, phosphate and hydrogen ions. It was previously reported that the urinary, excretion of N-acetyl-beta-D-glucosaminidase (NAG), a lysosomal enzyme, transiently increases after human PTH (hPTH) (1-34) infusion in normal subjects and idiopathic hypoparathyroidism patients, but not in pseudohypoparathyroidism type I patients. Here we report that intravenous infusion of hPTH(1-34) to rats transiently increased the urinary excretion of various lysosomal enzymes, such as beta-glucuronidase and acid phosphatase as well as NAG. However, it did not affect the urinary excretion of tubular brush border membrane enzymes, i.e. alkaline phosphatase, leucine aminopeptidase and gamma-glutamyl transpeptidase. Human PTH(1-34) dose-dependently increased the urinary excretion of NAG in rats with a peak at 30 min, which returned to a baseline within 60 min. The increase in the urinary NAG excretion caused by hPTH(1-34) positively correlated with the increase in the urinary cAMP excretion (r = 0.844, p < 0.01), and infusion of dibutyryl cAMP at a dose of 20 mg/kg similarly increased the urinary excretion of NAG. These results suggested that the increase in the urinary excretion of lysosomal enzymes caused by hPTH(1-34) may be a functional response to hPTH(1-34) occurring in the renal tubules via PTH signaling pathway.     

Evaluation of the effect of lipoic acid administered along with gentamicin in rats rendered bacteremic

Gentamicin is an aminoglycosidic antibiotic widely used in the treatment of many gram-negative bacterial infections. The present study was designed to investigate the extent of nephrotoxicity and the degree of protection afforded by lipoic acid under E. coli infected conditions and to note its effect on the antimicrobial activity of gentamicin. The study was carried out with adult male albino rats of Wistar strain. Group I animals served as controls. Group II animals were injected intraperitoneally for 2 successive days with 0.2 ml inoculum containing 10¹⁰ colony forming units of E. coli. Group III animals were injected E. coli as those in group II, in addition gentamicin 100 mg kg^–1 was administered intraperitoneally for 10 successive days. Group IV animals received intraperitoneal injections of E. coli as above plus gentamicin and also received lipoic acid (25 mg kg^–1) for 10 days by oral gavage. Rats subjected to E. coli administration showed a decline in the thiol content of the cell accompanied by high malondialdehyde levels along with lowered activities of catalase, superoxide dismutase and glutathione peroxidase with an added effect observed when gentamicin was administered along with it. The extent of nephrotoxicity induced by gentamicin was clearly evident with the decline in the activities of lactate dehydrogenase, alkaline phosphatase and N-acetyl--D-glucosaminidase in the rat renal tissues. A significant decrease was also observed in the activities of the transmembrane enzymes upon gentamicin administration. Treatment with lipoic acid decreased lipid peroxidation thereby maintaining the antioxidant status of the cell. The activities of the renal and transmembrane enzymes were also restored on lipoic acid treatment. The study has highlighted the beneficial effects of lipoic acid against experimental aminoglycoside toxicity in rats rendered bacteremic.     

Chelation therapy and vanadium: effect on reproductive organs in rats

Present investigation was planned to evaluate the therapeutic effectiveness of chelating agents against vanadium intoxication on blood and reproductive organs of rats. Male and female albino rats were injected vanadyl sulphate (7.5 mg/kg, po, for 21 days, 5 days in a week). Chelating agents tiron (T) alone and in combination with lipoic acid (LA), vitamin E (vit E) and selenium (Se) were given for 2 days/week. With the administration of vanadyl sulphate to rats fructose level in seminal vesicles was significantly (P< or =0.05) declined. The activities of alkaline phosphatase and adenosine triphosphatase were also decreased, whereas glycogen content and acid phosphatase activity increased in testis, seminal vesicles, ovaries and uterus after toxicant exposure. Significant changes in serum transaminases, serum alkaline phosphatase and lactate dehydrogenase were recouped by chelation therapy. Lipid peroxidation, reduced glutathione level and triglycerides levels altered significantly after exposure to vanadium in rats. The ultrastructural damage in spermatogenic stages in treated animals showed recovery pattern after therapy. Co-treatment with antioxidants restored these activities. The most effective combination was tiron + selenium followed by tiron + vitamin E, and tiron + lipoic acid.     

Protective effect of lipoic acid on adriamycin induced lipid peroxidation in rat kidney

Adriamycin, which is widely used in the treatment of various neoplastic conditions, exerts toxic effects in many organs. The present study was designed to investigate the effect of lipoic acid upon adriamycin induced peroxidative damages in rat kidney. The increase in peroxidated lipids on adriamycin administration was accompanied by alterations in the antioxidant defense systems. The extent of nephrotoxicity induced by adriamycin was evident from the decreased activities of the enzymes -glutamyl transferase and -glucuronidase in the rat renal tissues. The study was carried out with adult male albino rats of Wistar strain, which comprised of one control and three experimental groups. Group I rats served as controls. GroupII rats received adriamycin (1 mg kg^–1 body wt day^–1) intravenously through the tail vein. Group III rats were given lipoic acid (35 mg kg^–1 body wt day^–1) intraperitoneally. Group IV rats were given lipoic acid 24 h before the administration of adriamycin. Rats subjected to adriamycin administration showed a decline in the thiol capacity of the cell accompanied by high malondialdehyde levels along with lowered activities of catalase, superoxide dismutase, glutathione peroxidase and glutathione metabolizing enzymes (glutathione reductase, glucose-6-phosphate dehydrogenase, glutathione-S-transferase). Lipoic acid pretreatment also restored the activities of -glutamyl transferase and -glucuronidase nearly to control levels thereby suggesting nephroprotection. The study has highlighted the beneficial effects of lipoic acid pretreatment in reversing the damages caused by adriamycin and thereby bringing about an improvement in the oxidative stress parameters.     

Studies on the enzymology of purified preparations of brush border from rabbit kidney

1. A method for the preparation of brush border from rabbit kidneys is described. Contamination by other organelles was checked by electron microscopy and by the assay of marker enzymes and was low. 2. Seven enzymes, all hydrolases, were substantially enriched in the brush-border preparation and are considered to be primarily located in this structure. They are: alkaline phosphatase, maltase, trehalase, aminopeptidase A, aminopeptidase M, gamma-glutamyl transpeptidase and a neutral peptidase assayed by its ability to hydrolyse [(125)I]iodoinsulin B chain. 3. Adenosine triphosphatases were also present in the preparation, but showed lower enrichments. 4. Alkaline phosphatase was the most active phosphatase present in the preparation. The weak hydrolysis of AMP may well have been due to this enzyme rather than a specific 5'-nucleotidase. 5. The two disaccharidases in brush border were distinguished by the relative heat-stability of trehalase compared with that of maltase. 6. The individuality of the four peptidases was established by several means. The neutral peptidase and aminopeptidase M, both of which can attack insulin B chain, differed not only in response to inhibitors and activators but also in the inhibitory effect of a guinea-pig antiserum raised to rabbit aminopeptidase M. This antiserum inhibited both the purified and the brush-border activities of aminopeptidase M. The neutral peptidase and gamma-glutamyl transpeptidase were unaffected but aminopeptidase A was weakly inhibited. The characteristic responses to Ca(2+) and serine with borate served to distinguish aminopeptidase A and gamma-glutamyl transpeptidase from other peptidases. 7. No dipeptidases, tripeptidases or carboxypeptidases were identified as brush-border enzymes. 8. Incubation of brush border with papain released almost all the aminopeptidase M activity but only about half the activities of maltase, gamma-glutamyl transpeptidase and aminopeptidase A. No release of alkaline phosphatase, trehalase or the neutral peptidase was observed.     

Antioxidant DL-alpha lipoic acid as an attenuator of adriamycin induced hepatotoxicity in rat model

Protective efficacy of DL-alpha lipoic acid on adriamycin induced hepatotoxicity was evaluated in rats. Adriamycin toxicity, induced by a single injection (ip; 15 mg/kg body wt), was expressed by an elevation in alanine transaminase, aspartate transaminase, bilirubin levels in serum and alkaline phosphatase, lactate dehydrogenase, alanine transaminase, aspartate transaminase activity in hepatic tissue. Adriamycin produced significant increase in malondialdehyde levels indicating tissue lipid peroxidation and potentially inhibiting the activity of antioxidant, reduced glutathione and antioxidant enzymes, catalase, superoxide dismutase, glutathione peroxidase, glutathione reductase, glutathione-S-transferase, glucose-6-phosphate dehydrogenase. The present results showed that pretreatment with lipoic acid [75 mg/kg body wt/day (ip), 24 h prior to administration of adriamycin] significantly restored various cellular activity suggesting the antioxidant potential of lipoic acid in ameliorating the hepatotoxicity induced by adriamycin.     

Role of DL α-lipoic acid in gentamicin induced nephrotoxicity

The effect of DL -lipoic acid on the nephrotoxic potential of gentamicin was examined. Intraperitoneal injection of gentamicin (100 mg/kg/day) to rats resulted in decreased activity of the glycolytic enzymes-hexokinase, phosphoglucoisomerase, aldolase and lactate dehydrogenase. The two gluconeogenic enzymes—glucose-6-phosphatase and fructose-1, 6-diphosphatase, the transmembrane enzymes namely the Na⁺, K⁺-ATPase, Ca²⁺-ATPase, Mg²⁺-ATPase and the brushborder enzyme alkaline phosphatase, also showed decreased activities. This decrease in the activities of ATPases and alkaline phosphatase suggests basolateral and brush border membrane damage. Decreased activity of the TCA cycle enzymes isocitrate dehydrogenase (ICDH), succinate dehydrogenase (SDH) and malate dehydrogenase (MDH), suggests a loss in mitochondrial integrity. These biochemical disturbances were effectively counteracted by lipoic acid administration. Lipoic acid administration by gastric intubation at two different concentrations (10 mg and 25 mg/kg/day) brought about an increase in the activity of the glycolytic enzymes, ATPases and the TCA cycle enzymes. The gluconeogenic enzymes however showed a further decrease in their activities at both the concentrations of lipoic acid administered. These observations shed light on the nephroprotective action of lipoic acid against experimental aminoglycoside toxicity and the protection afforded at 25 mg/kg/day of lipoic acid was noted to be higher than that at 10 mg level.     

Mechanism of biliary secretion of membranous enzymes: bile acids are important factors for biliary occurrence of gamma-glutamyltransferase and other hydrolases

To elucidate the mechanism of biliary occurrence of gamma-glutamyl transferase [EC 2.3.2.2] and alkaline phosphatase [EC 3.1.3.1], the effect of bile acids on the biliary level of these enzymes was studied in vivo and in vitro. Following intravenous administration of taurocholate, the activities of both enzymes in rat bile increased markedly with a concomitant increase in the excretion of the bile acid. The biliary levels of these enzymes increased to reach a maximum at 10-20 min after administration of the bile acid and decreased thereafter. Right-side-out oriented rat liver canalicular membrane vesicles which localize gamma-glutamyltransferase, aminopeptidase M and alkaline phosphatase on their outer surface (Inoue, M., Kinne, R., Tran, T., Biempica, L., & Arias, I.M. (1983) J. Biol. Chem. 258, 5183-5188) were prepared. Upon incubation of the vesicles with either intact or heat-treated bile samples, the membranous enzymes were released from the vesicles in a time-dependent manner. Incubation of these vesicles with physiological concentrations of taurocholate also solubilized these enzymes from the membranes. Affinity chromatographic analysis on concanavalin A-Sepharose revealed that the transferase thus solubilized retained the hydrophobic domain responsible for anchoring the enzyme to membrane/lipid bilayers. These results indicate that bile acid(s) excreted into the bile canalicular lumen solubilized these enzymes from the apical membrane surface of the biliary tract cells by their detergent action.     

A combined assay of three lysosomal marker enzymes: acid phosphatase, beta-D-glucuronidase, and beta-N-acetyl-D-hexosaminidase

A simplified and rapid method for simultaneous activity measurements of three lysosomal marker enzymes, acid phosphatase, beta-glucuronidase, and beta-N-acetyl-D-hexosaminidase is described. The incubation is carried out in a single test tube and stopped by adding an alkaline sodium dodecyl sulfate solution, thus avoiding centrifugations and allowing for higher Triton X-100 concentrations in the incubation media. Two products of the beta-glycosidases (phenolphthalein and 2-nitrophenolate) are measured spectrophotometrically at the respective wavelengths (555 and 420 nm), and one of the acid phosphatase products is quantitatively determined by measuring inorganic phosphate.     

Studies on the nephrotoxicity of p-nitrophenylarsonic acid: changes in rat kidney and urinary enzyme activities following the administration of p-nitrophenylarsonic acid.

Histological studies showed that the administration of p-nitrophenylarsonic acid to rats resulted in renal tubular necrosis. The nephrotoxin was administered intraperitoneally and doses greater than 30 mg/kg were found to be fatal. The severity of the renal lesion depended on the amount of the nephrotoxin used. Elevated serum urea levels, urinary protein and volume were recorded over an 8-day period following the injection of the nephrotoxin. These changes were paralleled by an increase in the activity of lactate dehydrogenase, acid and alkaline phosphatase, N-acetyl-beta-glucosaminidase and beta-glucosidase in the urine. beta-Glycosidase activities increased in kidney homogenates, immediately after the injection of the nephrotoxin, but this eventually fell to well below the normal range. Subcellular fractions were prepared from sucrose homogenates by differential centrifugation and beta-glycosidases and cytochrome oxidase were used as enzyme markers. Only minor changes in the activity of cytochrome oxidase activity resulted from the administration of p-nitrophenylarsonic acid. One of the earliest indications of renal damage was a decrease in lysosomal latency. The activities of the lysosomal and soluble enzymes were elevated above normal during the first two days after the injection of p-nitrophenylarsonic acid, but they fell to values, significantly lower than normal, on the third day. The isoenzymic forms of beta-galactosidase, beta-glucosidase and N-acetyl-beta-glucosaminidase in normal and damaged kidneys were studied, using starch gel electrophoresis. The activities of both the lysosomal and the soluble forms of these enzymes decreased following the injection of the nephrotoxin, confirming the results obtained with whole homogenates. The relationship between the changes in renal enzyme activity and urinary enzyme excretion during the nephrotoxic process is discussed.     

Regulation of the activity of choline acetyl transferase by lipoic acid

The observations reported in this article demonstrate that lipoic acid strongly influences the activity of a purified preparation of choline acetyl transferase. The reduced form, dihydrolipoic acid, is a powerful activator of the enzyme while lipoic acid itself has an inhibitory effect and counteracts the stimulatory effect of dihydrolipoic acid. It is proposed that dihydrolipoic acid serves an essential function in the action of this enzyme and that the ratio of reduced to oxidized lipoic acid in the cell may play an important role in the regulation of the activity of the enzyme. The implications of these findings for cell function and acetyl choline formation are discussed.Affiliation     

20-Hydroxyeicosatetraenoic acid is excreted as a glucuronide conjugate in human urine.

20-hydroxyeicosatetraenoic acid, a major renal P-450 metabolite of arachidonic acid, has been quantified in human urine using capillary gas chromatography/electron capture negative ion chemical ionization mass spectrometry. The urinary excretion of 20-hydroxyeicosatetraenoic acid was in the low pg/ml range. However, treatment of urine with beta-glucuronidase resulted in a 13- to 28-fold increase in its concentration. This suggests 20-hydroxyeicosatetraenoic acid differs from other eicosanoids in that it is excreted primarily as a glucuronide conjugate.     

Isoelectric-focusing behavior of acid hydrolases in rat kidney lysosomes. Effects of the pH gradient, autolysis and neuraminidase.

Isoelectric focusing was used to study the multiple forms of acid phosphatase, arylsulfatase, beta-glucuronidase and beta-N-acetylhexosaminidase in lysosomes isolated from rat kidney. The isoelectric points of the main protein and hydrolase peaks were 1-1.5 units lower when electrofocusing was done in a pH 3-10 gradient than in a pH 10-3 gradient, apparently because the lysosomal constituents aggregated strongly at their isoelectric points and tended to settle somewhat in the gradient due to gravity. In the extended pH gradient the acidic form of each hydrolase occurred as asingle, relatively discrete peak. However, when pooled acidic fractions were refocused in a restricted pH gradient (pH 6-3 or 3-5) multiple acidic enzyme and protein components were resolved with isoelectric points between 2.7 and 5.1. When autolysis was minimized by extracting lysosomal fractions at alkaline pH (0.2% Triton X-100, 0.1%p-nitrophenyloxamic acid, 0.1 M glycine buffer, pH9) and including 0.1%p-NITROPHENYLOXAMIC ACID, AN INHIBITOR OF LYSOSOMAL NEURAMINIDASE AND CATHEPSIN D, in the pH gradient, arylsulfatase, beta-glucuronidase and beta-N-acetylhexosaminidase occurred in two forms, an acidic form with an isoelectric point of about 4.4, and a basic form with an isoelectric point close to 6.2, 6.7 and 8.0, respectively. Acid phosphatase occurred in three forms with isoelectric points of 4.1, 5.6 and 7.4. When some autolytic digestion was permitted by extracting lysosomal fractions in an acidic medium (0.2% Triton X-100, 0.1 M sodium acetate buffer, pH 5.2) AT 0-4DEGREES C and omitting p-nitrophenyloxamic acid from the gradient, the acidic form of beta-glucuronidase and the intermediate form of acid phosphatase were lost, the isoelectric points of the acidic forms of acid phosphatase, arylsulfatase and beta-N-acetylhexosaminidase were increased 0.6-1.2 units, and the isoelectric point of the basic forms of acid phosphatase, arylsulfatase and beta-glucuronidase was increased 0.5 unit. When lysosomal extracts were incubated with bacterial neuraminidase before electrofocusing, the acidic forms of acid phosphatase, arylsulfatase and beta-glucuronidase were largely lost, the isoelectric point of the acidic form of beta-N-acetylhexosaminidase was increased from 4.5 to 6.4, and the isoelectric points of the basic forms of all four hydrolases were increased 0.5-1.5 units. Autoincubation of lysosomal extracts in vitro at pH 5.2 PRODUCED SIMILAR, THOUGH LESS MARKED, effects. cont'd     

Acid hydrolase and glycoprotein: glycosyl transferase activities in experimental renal disease in the rat

Changes in the activity levels of kidney and urinary glycoprotein : glycosyl transferases and acid hydrolases were studied in ethylene glycol-induced hyperoxaluria and glycerol-induced renal failure in the rat. Hyperoxaluria resulted in increased urine activity levels of several glycosidases and acid phosphatase; changes in tissue levels of neuraminidase and CMP-sialic acid synthetase were also found. In the glycerol experimental model, there was stimulation in the activity of collagen : glycosyl transferase and neuraminidase in the rat kidney.     

Mechanism of enhanced bioavailability and diuretic effect of azosemide by ascorbic acid in rats

To increase the extent of comparative oral bioavailability (F) value and the diuretic and natriuretic effects of orally administered azosemide, ascorbic acid was coadministered to rats. The rationales for this study are that ascorbic acid might inhibit intestinal first-pass effect of azosemide and might increase the unionized fraction of azosemide at the receptor sites. After oral administration of azosemide (20 mg/kg) with 100 mg of ascorbic acid, the F value (138% vs. 100%), 8-h urinary excretion of azosemide (5.18% vs. 1.32% of oral dose), 8-h urine output (41.3 vs. 23.0 ml), and 8-h urinary excretion of sodium (24.6 vs. 15.3 mmol/kg) were greater than controls (without ascorbic acid). The amount of spiked azosemide remaining after 30 min incubation of 50 mug of azosemide with the 9000 g supernatant fraction of rat small intestine was significantly greater by 100 microg of ascorbic acid (45.3 vs. 40.9 microg) than controls (without ascorbic acid). After oral administration of azosemide with NH4Cl, the urine pH decreased by 0.5 U, and 8-h urine output (25.8 vs. 11.0 ml) and 8-h urinary excretion of sodium (13.3 vs. 6.89 mmol/kg) were significantly greater than controls (without NH4Cl). The increase in F value and diuretic and natriuretic effects of azosemide with coadministration of ascorbic acid seemed to be due to reduced intestinal first-pass metabolism of azosemide, increased urinary excretion of azosemide, and increased unionized fraction of azosemide at the renal tubular receptor sites.