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Among the lysosomal hydrolytic enzymes, and especially the exoglycosidases which carry out the degradation of the glycan moieties of glycoconjugates, the beta-D-mannosidase (beta-MAN) was the least investigated, up to the discovery of the inherited deficiency, in 1979-1980 for the caprine disease, and in 1986 for the human one. The beta-mannosidosis is characterized by mental retardation in children and by severe osteoarticular damage in kids. In both cases occurs the storage of oligosaccharides which are later excreted in urine: beta-mannosyl (1-4)-N-acetyl-beta-glucosaminyl (1-4)-N-acetylglucosamine, and beta-mannosyl (1-4)-N-acetylglucosamine in caprine disease, but only the last disaccharide in human patients. Having previously studied the serum enzymes, we report here the results obtained in the study of human urinary and renal beta-MAN. In each case we observed the existence of two isoforms of enzyme, B and a more acidic one, A. The main properties of these forms were determined, showing that A form of either origin seems to be identical, as well as the B form. But the ratios of activity B/A are inverted: 0.2-0.3 in urine, versus 15-20 in kidney. This observation led to the examination of the urinary enzyme of patients after a kidney transplantation. The B form is the major one, with a B/A ratio of around 3. The two isoforms are respectively identical to the normal renal and urinary ones. This shows that the B isoform determination way be used as reflecting a renal tubular damage.

The effects of 24R,25-dihydroxyvitamin D3 (24,25-(OH)2D3 on alkaline phosphatase (PAL), gamma-glutamyltransferase (GGT) and acid phosphatase (PAC) activities were investigated on renal cortex slices of hypophysectomized rats. Indeed after hypophysectomy renal 24,25-(OH)2D3 production was increased and renal PAL and GGT activities were decreased. After 5h incubation with physiological concentrations (0.1-10 nM) of 24,25-(OH)2D3 significant increases of PAL and GGT activities were produced. The maximum stimulation obtained with 1 nM was +23% for PAL and +26% for GGT as compared to controls. PAC was not modified. The time course of these effects was studied from 45 min to 8 h. In the presence of 24,25-(OH)2D3 (1 nM), delayed (3h) stimulation of PAL and GGT appeared. It reached the maximal value after 6h, +37% for PAL and +30% for GGT and persisted again at 8h. Cycloheximide added to incubation medium with steroid inhibited the stimulating effect on PAL only. Actinomycin D suppressed the induction of both enzymes, indicating that the observed actions of 24,25-(OH)2D3 depend on protein synthesis whose responsible mechanisms were different. These protein synthesis inhibitors did not modified enzymatic activities. Physiological significance of these renal effects is to be clarified.

Effects in vitro of 1,25-dihydroxycholecalciferol (1,25-(OH)2D3) on alkaline phosphatase (PAL), gamma-glutamyltransferase (gamma-GT) and acid phosphatase (PAC) activities were investigated on renal cortex from hypophysectomized rats. In these animals the biosynthesis of 1,25-(OH)2D3 and the specific activities of kidney PAL and gamma-GT were decreased. The course of these effects was determined from 45 min to 8 h. In the presence of 1,25-(OH)2D3 (2 x 10(-6) M) a delayed (5h) but simultaneous stimulation of the three enzymes was observed. It reached a maximum at 6h and disappeared at 8h. The dose-response relation was studied at 6h. In the presence of 1,25-(OH)2D3 (5 x 10(-7) M), the three enzymes were activated. The effect was maximal at 10(-6) M; it was +22% for PAL, +17% and +15% respectively for gamma-GT and PAC compared with controls. Cycloheximide suppressed the induction of PAL but not of gamma-GT activity. The effects of the secosteroid on renal enzymes seems to be a pharmacological more than a physiological one.

Concentrated and dialyzed 24-h urines of healthy persons were separated by 105000 X g ultracentrifugation into a pellet (P105) and a supernatant (S105) fraction. Chromatography of the P105 fraction on Sepharose 4B and 2B revealed that gamma-glutamyltranspeptidase and aminopeptidase activities had a molecular mass of 20 X 10(5) to 40 X 10(6), whereas in the S105 fraction soluble gamma-glutamyltranspeptidase and aminopeptidase had 86000 and 160000, respectively. Triton X-100 solubilization was performed on the P105 fraction and on a human renal cortex 40000 X g pellet, used as a reference. All the activity was recovered in both cases in a single peak of detergent gamma-glutamyltranspeptidase and detergent aminopeptidase eluted by filtration on Ultrogel Ac A22. Apparent molecular mass of Triton X-100 solubilized urinary and renal enzymes were 250000 and 243000 for gamma-glutamyltranspeptidase, and 298000 for both aminopeptidases. Protease solubilized forms were obtained by trypsic digestion of detergent urinary and renal forms. Both gamma-glutamyltranspeptidases were found to have an apparent molecular mass of 86000 on Sephadex G 150, which is identical to the value found for the S105 urinary gamma-glutamyltranspeptidase. The aminopeptidases had 238000 and 232000, which is a higher value than the molecular mass of the S105 urinary aminopeptidase. This letter could be a degraded form of the renal aminopeptidase. These findings suggest that gamma-glutamyltranspeptidase and aminopeptidase in the P105 fraction are similar to native renal enzymes. Evaluation of the P105 fraction enzymatic activities may thus be useful in the diagnosis of tubular damage.

Radiolabelling of 1-34 N terminal synthetic biological active fragment of bovine parathormone (1-34 bPTH) with high specific activity and without loss of biological activity was studied. Chloramine T and lactoperoxydase methods were used. To assess biological activity of 125I 1-34 bPTH labelled fragment, we use their ability to activate renal cortical adenylate cyclase. Our results show that enzymatic labelling method preserved the biological activity. With the chloramine T method it is possible to keep the biological activity by using DMSO.

Habekacin is a new aminoglycoside antibiotic. In this study we want to know the effect of increasing dose of habekacin on renal function and on renal morphology. We decide to compare the renal alterations induced by habekacin to these provoked by gentamicin, netilmicin and amikacin. Female Wistar rats received intraperitonally a single injection daily of 10, 30, 50, 150 mg/kg of habekacin for seven days. Wistar rats received also 50 mg/kg gentamicin, 50 mg/kg netilmicin and 150 mg/kg amikacin. No mortality was observed in groups treated with 10, 30, 50 mg/kg habekacin but 50 per cent of rats died with 150 mg/kg habekacin. Habekacin--30 mg/kg seven days--induced a decrease of cortical enzymatic activities, an increase of the number of lysosomes, a great accumulation of myeloid bodies, an alteration of lysosomal membranes Habekacin--50 mg/kg seven days and 150 mg/kg--induced a decrease of creatinine clearance and ultrastructural alterations of renal tubular cells. Comparative studies with other aminoglycosides showed that amikacin--150 mg/kg was the lesser nephrotoxic drug. With a same dose of 50 mg/kg, gentamicin appeared lesser nephrotoxic than habekacin and habekacin seemed to induce a same degree of renal modifications than netilmicin. With the dose of 150 mg/kg habekacin this drug was higher nephrotoxic than 50 mg/kg gentamicin. In conclusion, if it could be necessary to use habekacin and to prefer this aminoglycoside to gentamicin from an antibacterial activity point of view it is necessary to keep in mind that this drug is potentially nephrotoxic and that the dosage had to be strictly respected.(ABSTRACT TRUNCATED AT 250 WORDS)

Renal tissue kallikrein and proteins were measured in two kidney-one clip Goldblatt hypertensive rats both in the stenotic and the controlateral kidney and in sham operated rats at either 1 or 2 weeks after clipping. Activity was assessed by the amidolytic activity and by the kininogenase activity. Kallikrein in normotensive controls was 97.4 +/- 13 ng of bradykinin min-1 mg-1 of protein at week 1 and increased up to 116 +/- 18. Kallikrein in the GH rats was 83 +/- 12 in the stenotic kidney and 85,6 +/- 14 in the controlateral one at week 1, these values remained unchanged at week 2. As a consequence renal tissue kallikrein became significantly lower in the GH rats only at week 2 when compared to controls both the clipped and unclipped kidney showed the same magnitude decrease. Protein concentration remained at a steady level through out the 2 weeks of study. The results suggest that the lower renal kallikrein activity secondary to hypertension found in GH rats result from a decreased activation of prekallikrein in both kidney.

The catalytic properties of a neutral alpha-glucosidase purified to homogeneity from human renal cortex are described. The pH optimum was 6 (maltose and starch). It has a broad range of substrate specificity, hydrolysing di- and oligosaccharides with alpha (1 leads to 2), alpha (1 leads to 3), alpha (1 leads to 4) and alpha (1 leads to 6) linkages. Glucosidase action on maltosaccharides was associated with pronounced substrate inhibition at concentrations exceeding 0,5 mM. It also hydrolyses polysaccharides as starch and glycogen. The Km and Vmax values for the various substrates were determined. The enzymes exhibited intrinsic transglucosylase activity. It catalysed glucosyl-transfer reaction from maltose to itself (disproportionation). Mixed substrate inhibition studies, inhibition studies and heat inactivation are interpreted in terms of the existence of at least two interacting sites on the enzyme.