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Biomedical subjects

R G Price

Publications and source records attributed to R G Price.

At least 73 records · Page 4Linked to original sources
PubMed1982

Colorimetric assays for N-acetyl-beta-D-glucosaminidase and beta-D-galactosidase in human urine using newly-developed omega-nitrostyryl substrates.

(1) The synthesis of 2-methoxy-4-(2'-nitrovinyl)-phenyl 2-acetamido-2-deoxy-beta-D-glucopyranoside (MNP-GlcNAc) and 2-methoxy-4-(2'nitrovinyl)-phenyl beta-D-galactopyranoside (MNP-Gal) as substrates for the assay of NAG and beta-d-galactosidase are described. (2) beta-Glycosidase activities were determined in random urine samples from normal males and females aged between 12 and 87 years and patients with renal disease. (3) Both the MNP N-acetylglucosaminide and MNP galactoside were stable indefinitely, if stored in the solid state at 4 degree C in the dark. (4) The effect of urinary inhibitors was minimized by diluting the urine in the assay procedure. A simple assay procedure has been developed using MNP substrates. A good correlation was found with established assays using 4-methylumbelliferyl and p-nitrophenyl glycosides. (5) The assay was readily automated and a good correlation was found between the automated and manual methods. (6) The assay of urinary glycosidase activity with MNP substrates is simple to perform and has been used successfully in the clinical chemistry laboratory.

Acetylglucosaminidase↗
PubMed1982

Age-related changes in rat glomerular basement membrane.

1. The yield of GBM from rat kidney increased with rat age and the membrane became more collagen-like, as judged by the glycine, hydroxyproline, hydroxylysine content and the degree of glycosylation. 2. GBM became progressively less soluble in sodium dodecyl sulphate (SDS) and 2-mercaptoethanol with age. An increase in the lower molecular weight subunits of membranes prepared from older rats was demonstrated by polyacrylamide gel electrophoresis.

Aging↗
PubMed1981

The assay of arlesterase in serum using two new colorimetric substrates, omega-nitrostyrylacetate and propionate.

(1) The synthesis of 4-acetoxy-3-methoxy-omega-nitrostyrene and 4-propionyloxy-3-methoxy-omega-nitrostyrene as substrates for the assay of arylesterase activity is described. These substrates are stable when stored in the dark for at least one year. (2) Serum isolated from normal males and females aged between 25 and 35 years was used as a source of esterase activity. (3) The chromophore (4-hydroxy-3-methoxy-omega-nitrostyrene) absorbs at 505 nm and has a molar extinction coefficient of 26 800 in sodium carbonate/bicarbonate buffer, pH 9.5. (4) Both acetate and propionate substrates are more soluble in aqueous alcohol than in water or buffer alone and the substrates were therefore prepared in 10% methanol. Since some non-enzymic hydrolysis occurs at 37 degrees C, assays were carried out at 30 degrees C. Details of both single point and reaction rate assay procedures are described. (5) The properties of the arylesterases were determined in human serum using the single point and reaction rate assays. The data are similar to those previously described for these esterases using well-established procedures. (6) The assays described using the omega-nitrostyryl acetate and propionate substrates are simple to perform and can be readily adapted for use in the clinical chemistry laboratory. Large numbers of samples can be rapidly assayed with the minimum number of manipulative steps.

Adult↗
PubMed1980

Modification of the glomerular basement membrane in sucrose-fed and streptozotocin-diabetic rats.

Rats were fed on diets containing either sucrose or starch as the carbohydrate component (55%) for eight months. Diabetes was induced in animals of both groups by injecting streptozotocin (50 mg/kg body weight). Diabetic rats failed to gain weight, had enlarged kidneys, polyuria and elevated blood glucose levels. Starch and sucrose fed rats gained weight normally and had normal blood glucose levels. Sucrose fed rats had enlarged kidneys. Regional thickening of the glomerular basement membrane was present in sucrose-fed and diabetic rats but not in starch-fed controls. Glomerular basement membrane isolated from pooled kidney cortices from rats in the different experimental groups were analysed for amino acid, disaccharide and hexosamine content. Hydroxylysine (9 to 20%), hydroxyproline (21 to 24%), disaccharide (27%) and hexosamine (26%) were increased in membranes insolated from the three experimental groups, compared with starch-fed non-diabetic controls. An increase in low molecular weight components of the glomerular basement membrane of sucrose-fed and diabetic rats was observed using electrophoresis in sodium dodecyl sulphate. Significantly higher (p < 0.001) glucosyltransferase activity was present in kidney supernatants prepared from sucrose-fed (1050 +/- 60 nmol/2h/kidney) compared to starch-fed rats (510 +/- 40 nmol/2h/kidney). Sucrose feeding induces changes similar to those found in diabetes and the induction of diabetes made little difference over the feeding of sucrose alone.

Amino Acids↗
PubMed1980

Renal function in lithium and non-lithium treated patients with affective disorders.

Renal function was examined in 101 patients maintained on sustained-release lithium carbonate for periods ranging from 1 to 12.5 years. A control group of patients with affective disorders who had never been given lithium was also investigated as well as a control group of normal subjects of comparable age and sex. Creatinine clearance, serum creatinine, maximum urine osmolality after DDAVP administration, urine and plasma beta 2-microglobulin and the urinary excretion of N-acetyl-beta-glucosaminidase were not significantly different between patients on lithium and depressed patients never on lithium. Side effects that have been related to lithium therapy were not correlated with duration of lithium therapy. It is concluded that there was little evidence of a serious renal functional impairment attributable to lithium therapy.

Acetylglucosaminidase↗
PubMed1980

Differential solubility and subunit composition of rat glomerular basement membrane.

Glomerular basement membrane (GBM) was prepared from 2- to 3-month-old Sprague-Dawley rats by differential sieving and sonication. 80% of the membrane was soluble in 1% sodium dodecyl sulphate (SDS) and 1% 2-mercaptoethanol. The soluble fraction was resolved into 15 bands in the molecular weight range 30,000 to 300,000 by SDS-polyacrylamide gel electrophoresis. The major bands present had apparent molecular weights of 100,000 and 148,000. Treatment of GBM with SDS alone solubilised mainly low molecular weight components (45,000-150,000) but when the residue was treated with SDS and 2-mercaptoethanol higher molecular weight material was solubilised. Partial solubilisation of GBM was also achieved with pepsin. Digestion for 18 h at 4 degrees C resulted in 20% of the membrane being solubilised but this was increased to 55% at 10 degrees C. The amino-acid composition of pepsin-soluble GBM was more collagen-like than the residue remaining after enzyme digestion. Although the residue was more polar than whole GBM it still contained significant amounts of glycine and hydroxyproline and could be further subfractionated with SDS into a soluble fraction, the amino acid content of which was similar to whole GBM and a collagenous residue containing 317 residues/1,000 of glycine. When pepsin solubilised GBM was subjected to horizontal electrophoresis in SDS-polyacrylamide slab gels the principal bands migrated in the pro-alpha-chain region. This material was heterogeneous and in addition to the principal components, 6 components with apparent molecular weights less than 95,000 were present together with high molecular weight material in the gamma and beta regions of the gel. The band pattern of the pepsin-insoluble material was similar although the intensities of some individual bands varied significantly from that of the pepsin-soluble material. Rat GBM can therefore be fractionated by treatment with SDS alone, SDS together with 2-mercaptoethanol and pepsin digestion. The data reported is compatible with the presence of a mixture of collagen-like and polar regions rather than a major single collagenous component (type IV collagen).

Amino Acids↗
PubMed1980

Composition and biosynthesis of glomerular basement membrane in rats fed diets rich in sucrose.

1-month-old Sprague-Dawley rats were fed synthetic diets containing 55% sucrose (SU) or starch (ST) as the sole source of carbohydrate for 2, 3 or 8 months. Both the ST and SU fed rats gained weight normally, but SU fed rats had enlarged kidneys. A higher yield of glomerular basement membrane (GBM) was recovered from 9-month-old SU rats. An increase in the hydroxylated amino acid content was found in GBM prepared from SU fed rats and the glycine content was also higher. The increase in the hydroxylation of lysine was accompanied by increased glycosylation and there was 30% more Glc-Gal-Hyl present in GBM from 9-month-old SU rats. GBM was solubilised with sodium dodecyl sulphate (SDS) and 2-mercaptoethanol and subjected to electrophoresis on 5% polyacrylamide gels. There was an apparent fall in the intensity of the bands with molecular weights greater than 200,000 and a concomitant rise in low molecular weight components (50,000-100,000) in GBM from 4-month-old SU rats. These differences between ST and SU membrane were accentuated when the membranes from 9-month-old rats were compared. No significant differences were found in the glucosyl transferase activities of renal cortical homogenates prepared from 3-month-old SU and ST rats, but the activities in SU rats were significantly higher at 4 and 9 months. The feeding of SU-rich diets to rats induces a number of biochemical changes in the kidney which are similar to those found in diabetes. The feeding of SU diets provides a useful animal model with which to study the effect of dietary carbohydrate on renal GBM. SU should not be included in diets fed to diabetic rats because of the similarity of some of its effects and those seen in chemically induced diabetes.

Amino Acids↗
PubMed1979

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.

Animals↗
PubMed1979

Urinary enzyme assays in toxicological studies in the rat and marmoset.

The relative merits of the automated, fluorimetric assay of urinary enzymes and cell exfoliation were compared with other commonly used tests for renal damage. Two types of nephrotoxic agent were used, causing crystal nephropathy and acute tubular necrosis respectively. Groups of marmosets were given one of two drugs known to cause crystal nephropathy. One agent caused intermittent increases in urinary enzyme excretion and an early increase in cell excretion which was not sustained. The second agent in contrast caused elevated cell and enzyme excretion, increasing throughout the period of administration. A nephrotoxic anti-tumour agent also caused increases in cell and enzyme excretion when given to marmosets. The early changes produced by this agent were studied using catheterised rats. Hourly samples of urine were collected and urinary beta-glycosidase excretion was found to give an early indication of renal damage, which correlated with albuminuria and glycosuria. The fluorimetric assay of urinary enzymes provides a sensitive, non-invasive test of nephrotoxicity.

Acetylglucosaminidase↗
PubMed1979

Urinary N-acetyl-beta-D-glucosaminidase (NAG) as an indicator of renal disease.

The automated fluorimetric assay of N-acetyl-beta-D-glucosaminidase (NAG) has proved to be of value in the detection of rejection in transplant patients and in monitoring the course of renal disease. The urine can be diluted prior to assay, due to the sensitivity of the fluorimetric method and this reduces the effect of endogenous inhibitors. Time consuming dialysis or gel filtration steps are therefore unnecessary. Results are available to the physician within hours of the collection of the urine sample, since factoring the enzyme activity by the creatinine concentration avoids the necessity of collecting twenty-four hour or timed urine samples. Over one hundred samples can be assayed in a day using the automated procedure. The excretion of NAG in normal individuals varies with age and the activity found in pathological samples should always be compared with age-matched controls. The development of a new series of colorimetric substrates has allowed the production of a simple 'Dipstick' which should be of value in the screening of at-risk populations for renal disease. The diagnostic potential of the estimation of urinary NAG activity is enhanced by the separation of its isoenzymic forms by ion-exchange chromatography on DEAE-cellulose.

Acetylglucosaminidase↗