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Recently, we have found that two urinary glycoproteins, prostatic acid phosphatase (ACPP) and clusterin (CLU), combined with serum prostate-specific antigen (PSA) can serve as a three-signature panel for detecting aggressive prostate cancer (PCa) based on a quantitative glycoproteomic study. To facilitate the translation of candidates into clinically applicable tests, robust and accurate targeted parallel reaction monitoring (PRM) assays that can be widely adopted in multiple labs were developed in this study. The developed PRM assays for the urinary glycopeptides, FLN*ESYK from ACPP and EDALN*ETR from CLU, demonstrated good repeatability and a sufficient working range covering three to four orders of magnitude, and their performance in differentiating aggressive PCa was assessed by the quantitative analysis of urine specimens collected from 69 nonaggressive (Gleason score = 6) and 73 aggressive (Gleason ≥ 8) PCa patients. When ACPP combined with CLU, the discrimination power was improved from an area under a curve (AUC) of 0.66 to 0.78. By combining ACPP, CLU, and serum PSA to form a three-signature panel, the AUC was further improved to 0.83 (sensitivity: 84.9%, specificity: 66.7%). Since the serum PSA test alone had an AUC of 0.68, our results demonstrated that the new urinary glycopeptide PRM assays can serve as an adjunct to the serum PSA test to achieve better predictive power toward aggressive PCa. In summary, our developed PRM assays for urinary glycopeptides were successfully applied to clinical PCa urine samples with a promising performance in aggressive PCa detection.

Renal dysfunction, a major complication of type 2 diabetes, can be predicted from estimated glomerular filtration rate (eGFR) and protein markers such as albumin concentration. Urinary protein biomarkers may be used to monitor or predict patient status. Urine samples were selected from patients enrolled in the retrospective diabetic kidney disease (DKD) study, including 35 with good and 19 with poor prognosis. After removal of albumin and immunoglobulin, the remaining proteins were reduced, alkylated, digested, and analyzed qualitatively and quantitatively with a nano LC-MS platform. Each protein was identified, and its concentration normalized to that of creatinine. A prognostic model of DKD was formulated based on the adjusted quantities of each protein in the two groups. Of 1296 proteins identified in the 54 urine samples, 66 were differentially abundant in the two groups (area under the curve (AUC): p -value < 0.05), but none showed significantly better performance than albumin. To improve the predictive power by multivariate analysis, five proteins (ACP2, CTSA, GM2A, MUC1, and SPARCL1) were selected as significant by an AUC-based random forest method. The application of two classifiers-support vector machine and random forest-showed that the multivariate model performed better than univariate analysis of mucin-1 (AUC: 0.935 vs. 0.791) and albumin (AUC: 1.0 vs. 0.722). The urinary proteome can reflect kidney function directly and can predict the prognosis of patients with chronic kidney dysfunction. Classification based on five urinary proteins may better predict the prognosis of DKD patients than urinary albumin concentration or eGFR.

Background: To protect the kidney effectively with medication in type 2 diabetics, it is crucial to identify such at-risk patients early for treatment. We investigated whether peptiduria precedes proteinuria (the earliest urinary marker in our model), and thereby serve as an early predictor of diabetic nephropathy., Methods: A longitudinal study was performed in a rat model of diabetic nephropathy. Peptides, defined as degradation products of proteins of < 13 kD size, were quantified by a previously validated method using a combination of Lowry and Biorad protein assays. Peptides in urine were also confirmed by chromatographically separating low molecular weight fractions from urine and quantifying albumin fragments in these fractions by enzyme immunoassay. Also, the mechanism of peptiduria was addressed by measuring acid phosphatase, a marker of lysosomal activity, in urine and on kidney sections (histochemically)., Results: In rats with diabetic nephropathy, proteinuria occurred after 12 weeks of diabetes, while peptiduria occurred as early as 2 weeks after diabetes. Peptiduria was confirmed by showing that the chromatographically separated low molecular weight fractions of urine containing albumin fragments is in proportion to the level of peptiduria. The time course of peptiduria paralleled the increase in urinary acid phosphatase suggesting that the mechanism of early peptiduria could be due to upregulation of lysosomal enzyme activity in the tubules., Conclusions: Our results showing that peptiduria precedes proteinuria in diabetic nephropathy provide a compelling rationale to perform a prospective human clinical trial to investigate whether peptiduria can serve as an early predictor of diabetic nephropathy.

The analysis of indicators of mineral metabolism in patients with degenerative dystrophic affections of joints demonstrated that under development of osteoarthrosis process the alteration of indicators of concentration of electrolytes in blood serum, urine and synovial fluid occurs. The stage II of process is characterized by maximal alterations of indicators. The indicator of relationship between concentration of phosphate-ion and index of phosphatases of blood serum turned out the significant coefficient of correlation.

Background: Paediatric reference values for novel markers of phosphate homeostasis, bone formation and resorption and their putative relationship to growth are lacking., Methods: A total of 424 healthy children, adolescents and young adults (221 males) aged 0.1-21 y, were enrolled in this cross-sectional study. Height, weight and height velocity were assessed. Plasma/serum samples for determination of C-terminal fragment of fibroblast growth factor-23 (cFGF-23), sclerostin, bone alkaline phosphatase (BAP) and tartrate-resistant acid phosphatase 5b (TRAP5b) were available from 222, 264, 352 and 338 individuals, respectively. Calculation of cross-sectional centiles and z-scores was based on median (M), standard coefficient of variation (S) and the Box-Cox power (L) of transformation (LMS method) per age cohort. Correlations between variables as well as with growth were assessed., Results: cFGF-23, BAP and TRAP5b were significantly correlated with age (each P < 0.01), with highest values during infancy and adolescence. Serum levels of BAP and TRAP5b were significantly higher in adolescent boys compared with girls (each P < 0.01). In contrast, sclerostin levels were independent of age and gender. BAP and TRAP5b were strongly correlated and both were significantly associated with cFGF-23 and sclerostin as well (each P < 0.01). cFGF-23 was positively correlated with serum phosphate and renal phosphate threshold concentration (each P < 0.01). Height, weight, body mass index and height velocity were weakly correlated with BAP and TRAP5b (each P < 0.05)., Conclusions: This study provides age- and gender-related centile charts and z-scores for cFGF-23, BAP, TRAP5b and sclerostin and highlights the link between phosphate homeostasis and markers of bone metabolism during growth.

Objectives: To study the pattern of excretion of enzymes in urine during normal pregnancy., Design and Methods: Primigravidae, with uncomplicated pregnancies, were followed up throughout gestation. Urine samples were collected from them and activities of alanine aminopeptidase (AAP), gamma-glutamyl transferase (GGT), acid phosphatase (ACP) and N-acetyl-beta-d-glucosaminidase (NAG) activities in urine were estimated., Results: Small but significant increases were found in the activities of AAP and NAG excreted through the course of pregnancy. The changes seen in excretion of ACP and GGT were not statistically significant., Conclusions: Changes in excretion of ACP and GGT may be useful indicators of renal dysfunction in pregnancy, as their activities did not vary significantly through the course of normal pregnancy.

Oxalate induced renal calculi formation and the associated renal injury is thought to be caused by free radical mediated mechanisms. An in vivo model was used to investigate the effect of phycocyanin (from Spirulina platensis), a known antioxidant, against calcium oxalate urolithiasis. Male Wistar rats were divided into four groups. Hyperoxaluria was induced in two of these groups by intraperitoneal infusion of sodium oxalate (70 mg/kg) and a pretreatment of phycocyanin (100 mg/kg) as a single oral dosage was given, 1h prior to sodium oxalate infusion. An untreated control and drug control (phycocyanin alone) were also included in the study. We observed that phycocyanin significantly controlled the early biochemical changes in calcium oxalate stone formation. The antiurolithic nature of the drug was evaluated by the assessment of urinary risk factors and light microscopic observation of urinary crystals. Renal tubular damage as divulged by urinary marker enzymes (alkaline phosphatase, acid phosphatase and gamma-glutamyl transferase) and histopathological observations such as decreased tubulointerstitial, tubular dilatation and mononuclear inflammatory cells, indicated that renal damage was minimised in drug-pretreated group. Oxalate levels (P < 0.001) and lipid peroxidation (P < 0.001) in kidney tissue were significantly controlled by drug pretreatment, suggesting the ability of phycocyanin to quench the free radicals, thereby preventing the lipid peroxidation mediated tissue damage and oxalate entry. This accounts for the prevention of CaOx stones. Thus, the present analysis revealed the antioxidant and antiurolithic potential of phycocyanin thereby projecting it as a promising therapeutic agent against renal cell injury associated kidney stone formation.

Utilizing a commercially available helium-purging device and PEEK tubes for all tubing, especially for connection between the mobile phase and pump, high performance liquid chromatography with an electrochemical detector (ECD/HPLC) at the cathodic mode is a simple and precise method for the determination of p-nitrophenol (NP). Studies with cyclic and hydrodynamic voltammetry indicated that 25% aqueous MeOH containing 0.1% (v/v) CF(3)CO(2)H and -0.8 V vs. Ag/AgCl are the best mobile phase and detection potential for cathodic ECD/HPLC. With the present system, the limits of detection and determination were 0.2 and 0.25 microM, respectively, and up to 50 microM, a linear calibration curve was afforded. Within-day precisions for the analysis of 5 and 50 microM NP were 0.8 and 0.7% (n=6), respectively, and between-day precisions (n=6) for these samples were 3.5 and 2.2%, respectively. Compared with the commonly used Bessey-Lowry-Brock method, cathodic ECD/HPLC was useful for the assay of acid phosphatase in urine samples with p-nitrophenyl phosphate disodium salt as a substrate.

1,1-Dichloroethane (DCE) is a solvent that is often found as a contaminant of drinking water and a pollutant at hazardous waste sites. Information on its short- and long-term toxicity is so limited that the U.S. EPA and ATSDR have not established oral reference doses or minimal risk levels for the volatile organic chemical (VOC). The acute oral LD(50) in male Sprague-Dawley (S-D) rats was estimated in the present study to be 8.2 g/kg of body weight (bw). Deaths appeared to be due to CNS depression and respiratory failure. In an acute/subacute experiment, male S-D rats were given 0, 1, 2, 4, or 8 g DCE/kg in corn oil by gavage for 1, 5, or 10 consecutive days. The animals were housed in metabolism cages for collection of urine and sacrificed for blood and tissue sampling 24 h after their last dose. There were decreases in body weight gain and relative liver weight at all dosage levels, as well as increased renal nonprotein sulfhydryl levels at 2 and 4 g/kg after 5 and 10 days. Elevated serum enzyme levels, histopathological changes, and abnormal urinalyses were not manifest. For the subchronic study, adult male S-D rats were gavaged with 0.5, 1, 2, or 4 g DCE/kg 5 times weekly for up to 13 weeks. Animals receiving 4 g/kg exhibited pronounced CNS depression, with more than one-half dying by week 11. The 2-g/kg rats exhibited moderate CNS depression. One 2-g/kg rat died during week 6. There were very few manifestations of organ damage in animals that succumbed or in survivors at any dosage level. Decreases in bw gain and transient increases in enzymuria were noted at 2 and 4 g/kg. Serum enzyme levels and blood urea nitrogen were not elevated, nor were glycosuria or proteinuria present. Chemically induced histological changes were not seen in the liver, kidney, lung, brain, adrenal, spleen, stomach, epididymis, or testis. Hepatic microsomal cytochrome P450 experiments revealed that single, high oral doses of DCE did not alter total P450 levels, but did induce CYP2E1 levels and activity and inhibit CYP1A1 activity. These effects were reversible and regressed with repeated DCE exposure. There was no apparent progression of organ damage during the 13-week subchronic study, nor appearance of adverse effects not seen in the short-term exposures. One g/kg orally (po) was found to be the acute, subacute, and subchronic LOAEL for DCE, under the conditions of this investigation. In each instance, 0.5 g/kg was the NOAEL.

It has been reported that the urinary excretion of N-acetyl-beta-D-glucosaminidase (NAG), a lysosomal enzyme, transiently increases in human after treatment with human parathyroid hormone (hPTH)(1-34). We report here that hPTH(1-34) caused transient changes in the size and density of rat renal lysosomes following urinary excretion of NAG and other lysosomal enzymes tested. Percoll density gradient centrifugation revealed that hPTH(1-34) slightly but significantly increased the fraction of high density lysosomes (around 1.12 g/ml) 5-10 min after the treatment with hPTH(1-34), with a concomitant decrease in the fraction of intermediate density lysosomes (1.07-1.08 g/ml). On electron micrographs, some lysosomes in proximal tubules but not in distal tubules showed a change in morphology from circular to oval, and became enlarged and electron-dense 5-10 min after the treatment with hPTH(1-34). These responses to hPTH(1-34) were also reversible and transient. NAG excreted in urine after treatment with hPTH(1-34) had the molecular mass of a mature form in lysosomes and/or endosomes and was not a prepro-and/or pro-form of the enzyme. Thus, the changes in the density and size of renal lysosomes appear to be associated with the exocytosis of lysosomal enzymes by hPTH(1-34).

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.

Collagen type 1 represents more than 90% of bone matrix. Therefore, quantitation of collagen crosslinks, such as deoxypyridinoline, can provide information on bone resorption degree. An evaluation was made of deoxypyridinoline as well as other bone markets, such as alkaline phosphatase, tartrate resistant acid phosphatase, and hydroxyproline in patients with the diagnosis of osteoporosis. Paget's disease, hyperthyroidism, and chronic renal failure on haemodialysis or not. Deoxypyridinoline levels were significantly increased in patients with osteoporosis, Paget's disease, and hyperthyroidism. Hydroxyproline levels were increased in patients with Paget's disease, and tartrate resistant acid phosphatase was increased in all the entities studied. Deoxypyridinoline can be a more sensitive marker than hydroxyproline, with some advantages, such as its quantitation in a urine specimen and its high bone specificity. In patients with renal failure, tartrate resistant acid phosphatase was the only biochemical marker of bone resorption with increased levels.

Most newly synthesized lysosomal enzymes contain a transient carbohydrate modification, mannose 6-phosphate (Man-6-P), which signals their vesicular transport from the Golgi to the lysosome via Man-6-P receptors (MPRs). We have examined Man-6-P glycoproteins in human urine by using a purified soluble fragment of the soluble cation-independent MPR (sCI-MPR) as a preparative and analytical affinity reagent. In a survey of urine samples from seven healthy subjects, the pattern of Man-6-P glycoproteins detected with iodinated sCI-MPR as a probe in a blotting assay was essentially identical in each, regardless of sex or age. Two bands of approx. 100 and 110 kDa were particularly prominent. Man-6-P glycoproteins in human urine were purified by affinity chromatography on immobilized sCI-MPR. Seven distinct bands revealed by SDS/PAGE and Coomassie Blue staining were subjected to N-terminal sequence analysis. The prominent 100 and 110 kDa Man-6-P glycoproteins were identified as N-acetylglucosamine-6-sulphatase and alpha-glucosidase respectively. This identification was confirmed by molecular mass determinations on the two major bands after deglycosylation. Sequence analysis revealed arylsulphatase A and several previously unidentified proteins as minor species. Man-6-P glycoproteins were also purified on an analytical scale to determine the proportion of a number of lysosomal enzyme activities represented by the mannose-6-phosphorylated forms. The lysosomal enzymes in urine containing the highest proportion of mannose-6-phosphorylated form were beta-mannosidase (82%), hexosaminidase (27%) and alpha-glucosidase (24%). The profiles of Man-6-P glycoproteins detected by blotting in urine and plasma were not similar, suggesting that the urinary species are not derived from the bloodstream.

This study was undertaken to observe osteoclast differentiation related to inflammatory progression in aggressive periodontitis induced in beagle dogs by ligature of the gingival sulcus. To monitor osteoclastic activity, we used histochemical methods (staining for tartrate-resistant acid phosphatase [TRAP]) to visualize osteoclasts and their TRAP-positive precursors and biochemical methods (ELISA assay of pyridinium crosslinks) to detect bone matrix degradation products in gingival crevicular fluid (GCF), serum, and urine. For histochemical study, tissue specimens were prepared from 3 adult female beagle dogs induced with experimental periodontitis by silk ligature placement below the gingival margin of mandibular molars ligated for 3, 7, and 21 days. For biochemical study for pyridinoline measurement, the 24 mandibular molars of 4 male beagle dogs were ligated. GCF, urine, and serum were collected at day 0 and at 3, 7, 14, and 21 days after ligation. In the early inflammatory phase of ligature-induced periodontitis (day 3), TRAP+ mononuclear and TRAP+ multinucleated cells were present in the gingival connective tissue, and active bone-resorbing cells were found in excavated lacunae at the alveolar crest, but osteoclasts were not infiltrating the periodontal ligament during this early phase. During later stages of the inflammatory process (7 and 21 days), osteoclasts appeared at both the gingival and ligament side of the alveolar bone. Osteoclastic bone resorption appeared to be more severe on the bone surface at the gingival side than on the bone surface of the periodontal ligament side. Measurement of pyridinoline significantly increased in GCF and urine 3 days after ligation. The results suggested that bone at the crest of the alveolar bone is rapidly resorbed within 3 days of inducing experimental periodontitis.

Urinary alkaline phosphatase (ALP), acid phosphatase (ACP), aryl sulphatase (Ar. sulph.), beta-glucuronidase (beta-gluc.) and galactosidase were assayed in a group of Bilharzia haematobium patients and another group of healthy subjects (control group). The results for most of the determined enzymes revealed high activities as compared to the controls. The activity of acid phosphatase in male urine samples increased also, though not significantly. These elevated enzyme activities could be used to establish the diagnosis of schistosomiasis in patients whose urine contains no ova or when it is difficult to detect them. The results are discussed in the light of localization of each enzyme in the urinary tract as well as in other organs like the liver.

The loop diuretic, furosemide, which has been known to elicit renin release in vivo as well as in vitro, has also been shown to induce lysosomal enzyme release. After administration of furosemide (10 mg/kg and 300 mg/kg), a significantly increased acid phosphatase activity (1.33 x 10(-4) and 1.73 x 10(-4) vs. 0.23 x 10(-4) U, p < 0.001) was detected in the urine of the drug-treated mice, accompanied by a reduction of the residual enzyme activity in the kidney (5.0% for 10 mg/kg and 18.4%--p < 0.05--for 300 mg/kg dosage). Two marker enzymes, acid phosphatase and beta-D-glucuronidase, were assayed to demonstrate that furosemide also exerts its effect in in vitro systems like renal cortex suspension (corr. coeff.: 0.899 for acid phosphatase and 0.908 for beta-D-glucuronidase, p < 0.001) and isolated lysosomes (corr. coeff.: 0.981 for acid phosphatase and 0.989 for beta-D-glucuronidase, p < 0.001 for both). This action of the drug seems to be different from the well-known furosemide-sensitive inhibition of ion transport systems and may become of clinical relevance for patients receiving high doses of furosemide.

To study the mechanism of bone loss in physical unloading, we examined indices of bone formation and bone resorption in the serum and urine of eight healthy men during a 7 day -6 degrees head-down tilt bed rest. Prompt increases in markers of resorption--pyridinoline (PD), deoxypyridinoline (DPD), and hydroxyproline (Hyp)/g creatinine--during the first few days of inactivity were paralleled by tartrate-resistant acid phosphatase (TRAP) with significant increases in all these markers by day 4 of bed rest. An index of formation, skeletal alkaline phosphatase (SALP), did not change during bed rest and showed a moderate 15% increase 1 week after reambulation. In contrast to SALP, serum osteocalcin (OC) began increasing the day preceding the increase in Hyp, remained elevated for the duration of the bed rest, and returned to pre-bed rest values within 5 days of reambulation. Similarly, DPD increased significantly at the onset of bed rest, remained elevated for the duration of bed rest, and returned to pre-bed rest levels upon reambulation. On the other hand, the other three indices of resorption, Hyp, PD, and TRAP, remained elevated for 2 weeks after reambulation. The most sensitive indices of the levels of physical activity proved to be the noncollagenous protein, OC, and the collagen crosslinker, DPD. The bed rest values of both these markers were significantly elevated compared to both the pre-bed rest values and the post-bed rest values. The sequence of changes in the circulating markers of bone metabolism indicated that increases in serum OC are the earliest responses of bone to head-down tilt bed rest.

Two premenopausal women with rheumatoid arthritis of 16 years' duration were enrolled in a one year conditioning exercise program consisting of walking and low impact aerobics. The effect of exercise on the markers of bone formation (serum bone specific alkaline phosphatase and osteocalcin), bone resorption (urinary calcium/creatine and serum tartrate resistant acid phosphatase), and vertebral bone mineral density as determined by dual photon absorptiometry are discussed.

Estrogen deficiency-induced bone loss has been associated with accelerated bone turnover. Levels of some biochemical markers, such as serum osteocalcin (BGP), tartrate-resistant acid phosphatase (TRAP), and urinary hydroxyproline (OHP), have been shown to be related to the rate of bone turnover. They may therefore be useful in identifying the individual at risk for osteoporosis and monitoring the efficacy of the treatment. Two recently discovered markers, urinary pyridinoline (PYD) and deoxypyridinoline (DPD), are apparently directly related to bone matrix degradation and may be more accurate markers of bone resorption than OHP or TRAP. To evaluate the effects of menopause, osteoporosis, and estrogen replacement on the excretion of these new markers, we measured the levels of PYD and DPD and other biochemical markers of bone turnover in four groups of women, premenopausal healthy (PRE), postmenopausal healthy (POST), postmenopausal osteoporotic (UTO), and postmenopausal osteoporotic with estrogen treatment (ETO). Significant increases in PYD, DPD, BGP, TRAP, and OHP were found in POST and UTO groups compared with PRE. These increases were blunted by estrogen treatment when the levels of each of the markers returned to PRE levels. When comparing POST and UTO groups, significant increases were observed in UTO only for PYD, DPD, and urinary calcium but not for OHP, BGP, or TRAP. With subgroups matched for age and years from menopause, only DPD discriminated between POST and UTO. Indices of bone formation covaried with markers of bone resorption in the total population.(ABSTRACT TRUNCATED AT 250 WORDS)

Both sexes of F344 rats were gavaged with maximal tolerated doses of mercuric chloride for periods from 2 wk to up to 2 yr to investigate chronic nephrotoxicity and potential carcinogenicity. The toxicity of mercuric chloride was excessive after 2 wk of exposure to doses ranging from 1.25 to 20 mg/kg, compromising renal function by selectively destroying cells of the proximal tubules, and eliciting marked elevations in urinary biomarker enzymes diagnostic for acute renal tubule necrosis. In the 2-wk studies, urinary alkaline phosphatase and aspartate amino-transferase were most sensitive to renal mercury toxicity among a panel of six enzymes, exhibiting twofold increases above controls at the 5.0 mg/kg dose, before changes in the other enzymes occurred. Urinary lactate dehydrogenase was the most responsive enzyme, with up to 11-fold increases in activity above controls. In response to mercuric chloride exposure of 5.0 mg/kg for 2-6 mo, the greatest and most persistent increases in elevation of urinary enzyme activities were exhibited by alkaline phosphatase and gamma-glutamyl transferase, which increased two-to threefold above controls. At this interval, the maximal severity of the renal lesions in both sexes of rats was graded as minimal to mild. Beyond 6 mo none of the urinary enzymes measured in this study was adequate as biomarkers of nephrotoxicity, although the severity of the renal lesions had progressed. Mercury accumulated in a dose-related fashion primarily in the kidney, and to a lesser extent in the liver. The severity of the renal lesions was increased by continued exposure to mercuric chloride, as tissue concentrations of mercury rose in proportion to dose. Mercuric chloride treatment for 2 yr clearly exacerbated the severity of the spontaneous nephrotoxicity prevalent in aging F344 rats. The excessive mortality that occurred in the male rats was probably due to a combination of these factors. No renal tumors were detected in rats, possibly because the potential for their development was reduced; however, direct tissue contact with mercury induced squamous-cell papillomas of the forestomach in both sexes.

To study the significance of prostatic acid phosphatase (PAP), gamma-seminoprotein (gamma-Sm) and prostatic specific antigen (PA) in urine, we have determined the urinary levels of these proteins in women and infants, in patients without prostatic disease, in patients with benign prostatic hypertrophy, and in patients with prostatic adenocarcinoma. Women and infants were found to excrete little PAP (27.9 +/- 4.8 ng/mg) and undetectable levels of gamma-Sm except one case, and undetectable levels of PA in the urine. The excretion of PAP in patients with prostatic carcinoma who were either castrated, or treated with endocrine therapy was lower than the levels in women and infants, or the levels in patients without prostatic diseases, or the levels in patients with BPH. Urinary excretion levels of gamma-Sm and PA were undetectable in the patients with well-controlled prostatic carcinoma. The present study suggests that the determination of PAP, gamma-Sm and PA in the urine of patients with prostatic carcinoma may become a useful tool for monitoring of the primary locus of the carcinoma, but additional assays of urinary PAP, gamma-Sm and PA should be measured at regular intervals to be concluded.

Diacetylbenzidine was used to induce a nephrotic syndrome in female rats. Enzymes involved in glycoprotein metabolism were evaluated during an early stage of induced renal disease before extensive histologic changes occurred. The results show that lysosomal acid hydrolases are not activated or released to any measurable degree during the early stages of the disease. Minimal differences in the composition of glomerular basement membrane of nephrotic rats were found despite heavy proteinuria. Glomerular specific activities of certain glycoprotein:glycosyl transferases were depressed in nephrotic animals. A new viewpoint to explain the pathology of glomerular proteinuria is presented based on the phenomenon of sublethal autolysis affecting cell surface structure and function, of which activity levels of glycoprotein:glycosyl transferases are an example. Increased activities of glycosyl transferases and Na-D ATPase were noted in the cortex from nephrotic animals. These studies involving cortex indicate that the pathologic process is not confined to the glomerulus and may contribute information concerning Na+ transport in the nephrotic rat.

Biochemical changes in rat urine and tissues treated with five consecutive daily doses of ochratoxin A (10 mg/kg body weight) were studied. Urine volume and urinary proteins were moderately raised during the first few days of ochratoxin treatment, and were then highly elevated towards the end of the investigation. Urinary muramidase excretion was significantly raised (p less than 0.01) 24 h after the first insult with the toxin. The urinary output of alkaline and acid phosphatases, lactate dehydrogenase (LDH) and glutamate dehydrogenase (GDH) were all elevated but very much later, during the course of injections with ochratoxin A. Kidney alkaline and acid phosphatases, LDH and GDH were correspondingly reduced 7 days from the beginning of ochratoxin A administration. Liver LDH activity was reduced while serum LDH was raised. Liver glycogen level was significantly (p less than 0.0001) increased. Experimental evidence was presented to show that the initial point of interaction of ochratoxin A with the rat renal system may be at the first portion of the proximal convoluted tubular cell region.

The renal effect of the daily administration of gentamicin to male albino rats (20 mg/kg body weight) for 6 consecutive days on some biochemical systems of the kidney was examined. Urine volume and urinary protein levels were found to be progressively raised following gentamicin. Urinary alkaline phosphatase, acid phosphatase, muramidase and glutamate dehydrogenase (GDH) activities were found to be markedly elevated. Acid phosphatase and GDH activities in urine were raised 24 h and 48 h, respectively, from the onset of gentamicin administration. The sequence in which some regions of the renal cells were involved in gentamicin nephrotoxicity was determined and the probable mechanism of interaction of gentamicin antibiotic with the renal tubular cells is proposed.

Ultrafiltration of urine and rapidly separates urinary enzymes from inhibitors, activators, and heat-stable, spurious activities present in urine. Using eight enzymes of clinical interest as examples, we verified the reliability of results obtained after ultrafiltration by comparsion with results obtained after gel filtration.

Using a modified model of Masugi's nephritis of rats, various enzymatic activities in urine, serum and renal tissue (glomeruli or cortex) were determined at appropriate intervals after the administration of anti-kidney serum and compared with the urinary protein content and the kidney weight. In the urine, alkaline phosphatase (Al-Phosase), acid phosphatase (Ac-Phosase) and N-acetyl-beta-glucosaminidase (NA-beta-Gase) activities remarkably increased after the induction of nephritis, reached their peaks on the 10th day and reverted to almost the normal levels on the 30th day. The patterns of time course of these enzymatic activities were similar to patterns seen in the urinary protein content and the kidney weight. In the serum, the Al-Phosase activity decreased slightly, while NA-beta-Gase activity increased slightly. The Ac-Phosase activity in serum remained at normal levels during the experimental periods. In the glomeruli, the bound activities of these three enzymes decreased with nephritis, showing a negative correlation with results in the urine. On the other hand, fibrinolytic activities in the urine (plasmin-like enzyme) and renal cortex (plasminogen activator) also paralleled the urinary protein content and the kidney weight in the course of the disease. These results suggest that the Al-Phosase, Ac-Phosase and NA-beta-Gase excreted into urine in cases of nephritis may be mostly derived from damaged renal cells and one part of Al-Phosase may also come from the plasma. Moreover, the increase of plasmin-like enzyme in urine is considered to be due to the increase of plasminogen activator in the renal cortex. Thus, the determination of these enzymatic activities in the urine should be useful for evaluating effects of drugs for the treatment of nephritis.

Manipulations of metabolic acid-base status were used in an attempt to modify the renal handling and toxicity of the anticancer drug cisplatinum. Rats were orally pretreated with either tap water (TW), ammonium chloride (AC), or sodium bicarbonate (SB) for 3 days prior to intraperitoneal administration of a high cisplatinum dose (7.5 mg/kg b.w.). Urine was collected daily for 4 days between drug dosing and killing of the animals. AC-pretreated rats did not exhibit the characteristic cisplatinum-induced diuresis and were unable to maintain an acid urinary pH following drug administration. AC rats had a significantly lower, and SB rats a significantly higher, urinary excretion of platinum than did TW rats. Platinum excretion was found to be correlated with urinary pH (r = 0.88) and not urinary volume (r = 0.30). The renal concentration of platinum was greater in AC animals than in SB or TW animals, but no significant difference was observed in liver or plasma concentrations between the groups. Both pretreated groups had equal percent of free vs bound platinum. Proteinuria was more severe in AC-pretreated rats, but histologic evidence of renal tubular damage was present in all the three groups. It is concluded that metabolic acidosis can seriously impair the renal handling of high dose cisplatinum but that metabolic alkalosis offers no evident advantages over nonpretreatment.

Enzymuria observed in PCM, especially in severe cases, does not seem to be a simple reflection of the increased serum values and suggests renal involvements. A number of factors among which are :- lowered clearance threshold for certain enzymes as well as rapid wear and tear or renal cells seem to participate in the causation of such enzymuria. Urine enzyme analysis may reflect the extent and state of renal pathology among different forms of PCM.

In 44 workers of a chemical plant producing chlorinated hydrocarbon pesticides, protein concentration and activity of acid phosphatase, lactic dehydrogenase and aspartate and alanine aminotransferase in urine were estimated. Protein concentration and activity of all test enzymes were significantly higher as compared to controls: the serum creatinine and uric acid concentration was normal in all examined persons.

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.

Generally acid phosphatase (ACP) assay is used for testing cases of alleged rape. Comparison of three different chemical methods for Acid Phosphatase (Andersch's method with p-nitrophenyl-phosphate substrate and tartrate inhibitor (A-Tart), Roy's method with thymolphthalein phosphate substrate (R-TMP), and Babson's method with alphanaphthyl phosphate substrate) indicated that Roy's method (R-TMP) should be the preferred one. This method had both acceptable sensitivity and confirmed specificity. Our data confirmed that the vaginal wash of normal healthy women has a very low level of ACP activity. Because of inconclusive data in the literature regarding this ACP level, a normal and equivocal range of ACP was suggested until more is known about causes and interferences. Possible sources of normal ACP activity in the wash fluids were also indicated.

We have studied the mode of excretion of three prostatic secretory proteins, namely acid phosphatase (PAP), prostate-specific antigen (PSA) and beta-inhibin, in the urine of normal adult men, and we have determined the urinary levels of these proteins in men with benign prostatic hypertrophy (BPH) or adenocarcinoma. The output of the three proteins was highly variable during the day. In order to minimize these variations, 24-hour urine samples were collected thereafter. Our study showed that PAP concentrations in 50% of men with or without symptomatic BPH were similar to those of normal young men. In the remaining 50%, PAP was undetectable. In contrast, average PSA and beta-inhibin concentrations were higher in patients with BPH than in young men (p less than 0.05). The three markers were decreased or nondetectable in about half of the patients with untreated prostatic cancer. This phenomenon was even more pronounced in patients receiving hormonal treatment (castration or diethylstilbestrol). However, some of these patients still excreted normal amounts of PAP, PSA, and beta-inhibin. Urinary and serum PAP levels showed no correlation. These results indicate that urinary prostatic markers provide an easy means to study the behavior of the primary prostatic tumor. This information may be of potential value since it is not obtained with serum markers which originate mostly from metastatic cells.

The effect of administration of suramin on the urinary levels of some lysosomal enzymes in rats was investigated. The urinary levels of acid phosphatase and lysozyme were significantly elevated 24 h after drug administration. The high levels were sustained for 9 days after drug administration was terminated. These actions were indicative of labilization of kidney lysosomal membrane by the drug molecules, the membrane being repaired and integrity restored some days following termination of drug administration.

The acid phosphatase level of irrigant solution used during transurethral prostate resections was measured in 20 patients. The mean acid phosphatase of the irrigant solution was 45.3 IU/L (S.D. 50.7). The degree of elevation of irrigant acid phosphatase was related to the weight of tissue resected, the duration of the resection, and the total volume of irrigant used. A transient elevation of serum acid phosphatase occurred postoperatively which was associated with the level of urinary acid phosphatase elevation and the amount of absorbed irrigant solution based on changes in serum sodium.