Urinary (1)H-NMR-based metabolic profiling of children with NAFLD undergoing VSL#3 treatment.

BACKGROUND: Nowadays, non-alcoholic fatty liver disease (NAFLD) is one of the most common chronic liver diseases in children. Our recent clinical trial demonstrated that dietary and VSL#3-based interventions may improve fatty liver by ultrasound and body mass index (BMI) after 4 months. OBJECTIVES: As in this short-term trial, as in others, it is impracticable to monitor response to therapy or treatment by liver biopsy, we aimed to identify a panel of potential non-invasive metabolic biomarkers by a urinary metabolic profiling. METHODS: Urine samples from a group of 31 pediatric NAFLD patients, enrolled in a VSL#3 clinical trial, were analyzed by high-resolution proton nuclear magnetic resonance spectroscopy in combination with analysis of variance-Simultaneous Component Analysis model and multivariate data analyses. Urinary metabolic profiles were interpreted in terms of clinical patient feature, treatment and chronology pattern correlations. RESULTS: VSL#3 treatment induced changes in NAFLD urinary metabolic phenotype mainly at level of host amino-acid metabolism (that is, valine, tyrosine, 3-amino-isobutyrate or ß-aminoisobutyric acid (BAIBA)), nucleic acid degradation (pseudouridine), creatinine metabolism (methylguanidine) and secondarily at the level of gut microbial amino-acid metabolism (that is, 2-hydroxyisobutyrate from valine degradation). Furthermore, some of these metabolites correlated with clinical primary and secondary trial end points after VSL#3 treatment: tyrosine and the organic acid U4 positively with alanine aminotransferase (R=0.399, P=0.026) and BMI (R=0.36, P=0.045); BAIBA and tyrosine negatively with active glucagon-like-peptide 1 (R=-0.51, P=0.003; R=-0.41, P=0.021, respectively). CONCLUSIONS: VSL#3 treatment-dependent urinary metabotypes of NAFLD children may be considered as non-invasive effective biomarkers to evaluate the response to treatment.

The effects of bicarbonate and its combination with chelating agents used for the removal of depleted uranium in rats.

The effects of bicarbonate and its combination with the chelating agents, deferiprone (L1), 4,6-dimethyl-1-hydroxypyrimidin-2(1H)-one (AK-4), catechol-3,6-bis(methyleneimino-diacetic-acid) (CBMIDA), and ethane-1-hydroxy-1,1-bisphoshonate (EHBP) in removing depleted uranium (DU) for radiation emergency medicine were examined. After the intramuscular injection of DU in rats, various time schedules of bicarbonate and chelating agent administration were tested. The results indicate that the bicarbonate helps increase significantly the effects of LI and AK-4, while there were no effects of using bicarbonate alone. The effects of bicarbonate on CBMIDA were unclear, and the effects of EHBP were negative. Further studies are necessary to obtain distinctly synergic effects by the combination of chelating agents with bicarbonate.

Evaluation of renal enzymuria and cellular excretion as an marker of acute nephrotoxicity due to an overdose of paracetamol in Wistar rats.

INTRODUCTION: The present study was conducted to determine whether the urinary levels of excreted enzymes, gamma-glutamyltransferase (GGT), alkaline phosphatase (ALP), lactate dehydrogenase (LDH), aspartate (AST) and alanine aminotransferases (ALT), can efficiently indicate, within 24 h, an acute nephrotoxicity due to an overdose of paracetamol (PAR). METHODS: A baseline urine was collected from the experimental group. Thereafter, blood collected from the orbital sinus (1.0 ml) and paracetamol (650 mg/kg of body weight) was administered by gavage. After the drug administration, animals were returned to the metabolic cages and then urine was collected in the next 22 h. Blood and urine collection was performed at time 0+24 h (T(24)), as well as at times 48 and 72 h (T(48) and T(72)). After the last urine and blood collection, the rats were killed and the kidneys removed and prepared for histological examination. Plasma creatinine and urinary levels of creatinine (to determinate glomerular filtration rate-GFR), GGT, ALP, LDH, ALT and AST were measured. Kidney tissues were stained with hematoxylin and eosin stain for histological assessment. RESULTS: Urinary levels of GGT, ALP and LDH enzymes were significantly higher (P<0.05) at T(24) when compared to the levels at T(0) and returned to basal levels at T(48) and T(72). The number of urinary epithelial cells at T(24) was significantly higher when compared to the control time (T(0)) (P<0.001). The GFR was significantly reduced 24, 48 and 72 h after the drug administration. CONCLUSION: The number of urinary epithelial cells and urinary enzymes levels are a simple and low cost procedure that is available and can help in the detection of renal acute lesions.

A comparison of methods for measuring serum and urinary markers of bone metabolism in cats.

Biochemical markers of bone cell activity have recently been shown to be useful for monitoring skeletal health in domestic animals, including dogs and horses. The aim of this study was to evaluate a number of biochemical assays, originally developed for use in humans, for their ability to measure indicators of bone cell activity in serum and urine of normal cats over a range of ages. Bone alkaline phosphatase (BAP), a marker of bone formation, was measured in serum using wheatgerm lectin precipitation (WGL) and by ELISA. The curve derived from serial dilution of feline serum was parallel with the ELISA standard curve, indicating species cross-reactivity, and there was a significant relationship between assays (rs = 0.97, P < 0.001). Deoxypyridinoline (DPD), a marker of bone resorption, was measured in its total form in urine by HPLC and ELISA, and in its free form in serum and urine by ELISA. The dilution curve for free DPD in urine showed parallelism with the assay standard curve; however, the curves for total DPD in urine and serum did not. A significant relationship was established between total urinary DPD (HPLC) with total serum DPD (rs = 0.69, P < 0.001), and with free urinary DPD (rs = 0.95, P < 0.001) concentrations. Carboxy-terminal telopeptide of type I collagen (CTX) concentration, another marker of bone resorption, was measured in serum and urine by ELISA, and there was a significant relationship between assays (rs = 0.82, P < 0.001). CTX could not be measured reliably using an auto-analysis method. A significant relationship was established between total urinary DPD (HPLC) with serum CTX (rs = 0.59, P < 0.05), and urinary CTX (rs = 0.65, P < 0.001) concentrations. BAP (ELISA and WGL), total urinary DPD (HPLC), urinary CTX (ELISA), and serum CTX (ELISA) concentrations were significantly inversely correlated with age (rs = -0.66, -0.88, -0.61, -0.70, and -0.51, P < 0.05 respectively). Cats under two years of age had significantly higher BAP, total urinary DPD (HPLC), and urinary CTX concentrations compared to older cats. In conclusion, this study has shown that a number of commercially available assays provide reliable methods for non-invasively monitoring bone cell activity in cats and has shown that bone turnover decreases within the first two years of life, until complete skeletal maturity is attained. Future studies can now be directed at evaluating the potential clinical application of these methods.

Effects of coffee cherry, the residue left after removal of the beans from the coffee fruit, on mammary glands, automatic behavior and related parameters in mice.

To clarify the mechanisms of the anti-mammary tumor activity of coffee cherry (CC), the residue left after the removal of beans from the fruit, the effects in SHN mice of CC on plasma and urine component levels, mammary gland growth, spontaneous motor activity and several related parameters were examined. Hot water extract of CC was given to 2-month-old mice in drinking water (0.5%) for 60 days. The treatment prevented the elevation of plasma and urine levels of alanin amino-transferase and asparate aminotransferase, indicating that CC can protect against metabolic abnormality, which is a cause of the high mammary tumor susceptibility of SHN mice. It also resulted in an inhibition of the formation of precancerous mammary hyperplastic alveolar nodules. Neither food and water intake nor spontaneous motor activity was affected by CC. The findings provide novel information on the mechanism of the protective effect of CC on mammary tumorigenesis and confirm the usefulness of CC as a safe chemopreventive agent of mammary and other types of tumors.

The assessment of nephrotoxic effect of organophosphorous pesticides based on the determination of the activity of some selected enzymes in urine.

Pesticides and their metabolites are excreted mainly by the kidneys. The effect of these compounds on the kidney parenchyma was evaluated on the basis of determinations of the activity of the following enzymes: alkaline phosphate, N-acetylglucosaminidase, lactate dehydrogenase, alanine aminotransferase, aspartate aminotransferase and arginase in urine of workers employed at the department producing organophosphorous pesticides (32 males and 53 females) as well as those employed at the production of chlorfenvinphos (35 males). The activity of most of the estimated enzymes was significantly higher as compared to control groups. The dynamic of changes of enzyme activity was traced in the workers employed at the department producing chlorfenvinphos over their first 18 months of employment.

Possible role of thioformamide as a proximate toxicant in the nephrotoxicity of thiabendazole and related thiazoles in glutathione-depleted mice: structure-toxicity and metabolic studies.

In mice depleted of GSH by treatment with buthionine sulfoximine (BSO), thiabendazole (TBZ) causes renal injury characterized by an increase in serum urea nitrogen (SUN) concentration and by tubular necrosis. Previous studies have shown that TBZ requires metabolic activation before it produces nephrotoxicity and that the structure contributing to the toxicity of TBZ is the thiazole moiety of the molecule. TBZ and its thiazole analogues were examined for the ability to increase SUN concentration and serum alanine aminotransferase activity in GSH-depleted mice. Unsubstituted thiazole and thiazoles with 4- and/or 5-, and no 2-, substituents caused marked increases in SUN concentration, suggesting nephrotoxicity. Furthermore, the nephrotoxic potency of these thiazoles decreased with the increasing number and bulk of the 4- and/or 5-substituents. On the other hand, the target organ (the kidney or liver) and the toxic potency of 4-methylthiazoles were markedly altered with the type of substituents at the 2-position. These observations and the known toxicity of thiono-sulfur compounds led us to the hypothesis that the nephrotoxic thiazoles, which lack 2-substituents, would undergo microsomal epoxidation of the C-4,5 double bond and, after being hydrolyzed, the resulting epoxide would then be decomposed to form thioformamide, a possibly toxic metabolite. Evidence for this hypothesis was provided by the results that thioformamide and tert-butylglyoxal as the accompanying fragment were identified as urinary metabolites in mice dosed with 4-tert-butylthiazole and that thioformamide caused a marked increase in SUN concentration when administered to mice in combination with BSO.

Nephrotoxicity of uranyl fluoride in uninephrectomized and sham-operated rats.

The aim of the present study was to determine whether the nephrotoxicity of the uranium-containing compound uranyl fluoride (UO2F2) is enhanced after unilateral nephrectomy. Unilaterally nephrectomized (NPX) and sham-operated (SO) rats were given single intravenous injections of UO2F2 at doses delivering 100 or 250 micrograms U/kg 16 days after surgery. Between the second and third day after the administration of either dose of UO2F2, the urinary excretion of the cellular enzymes lactate dehydrogenase (LDH) and aspartate aminotransferase (AST) and the plasma solute albumin began to increase significantly in both the NPX and SO rats. The urinary excretion of the plasma solute glucose did not begin to increase significantly in the NPX and SO rats until 4 days after the administration of either dose of UO2F2. During the fifth day following the administration of either dose of UO2F2 (which was also the last day that urinary data were collected) the urinary excretion of LDH, AST, and glucose in the NPX and SO rats was greater than that during any previous day. The urinary excretion of these three compounds during this fifth day was greater in the SO rats than in the NPX rats. Also during the fifth day following the injection of either dose of UO2F2, the fractional excretion of glucose was higher in the SO rats than in the NPX rats. By the end of the fifth day, the level of histologically demonstrable cellular necrosis in the pars recta of proximal tubules in the renal cortex and outer medulla of the NPX and SO rats was statistically similar. Therefore, the nephropathy in rats induced by UO2F2 is not made more severe as a result of unilateral nephrectomy.

[Relevance of urinary enzyme determination for the diagnosis and follow-up of kidney injuries and secondary kidney damage--results of a prospective study]. / Relevanz von Harnenzymbestimmungen für die Diagnostik und Verlaufskontrolle von Nierenverletzungen und sekundären Nierenschäden--Ergebnisse einer prospektiven Studie.

In a prospective study we examined and confirmed the suitability of urinary enzyme determinations for the diagnosis and control of kidney injuries. Overall, 186 patients were included in the study. 68 patients had blunt lumbar, thoracic and abdominal traumata. Renal operations (n = 14), in particular nephropexy, served as clinical model. Moreover, analyses were performed in 15 polytraumatized patients, 20 patients with fractures of the extremities, 10 patients with thermic injuries, 10 operations on the intervertebral disks, 35 abdominal operations, 1 extirpation of brain tumour, 2 meniscectomies and 11 patients with renal diseases without surgical intervention. We determined the catalytic activities of alanine aminopeptidase (AAP), gamma glutamyltransferase (GGT) and alkaline phosphatase (AP) in the gel-filtered spontaneous urine. The urinary enzyme activities were related to the symptoms, laboratory findings and other parameters. With high sensitivity and without depending on the occurrence and duration of a haematuria the urine enzymes reacted to a traumatic damage to the renal parenchyma by an increased efflux of enzymes. The amount of information from intravenous urography was small. However, there was considerable agreement between nuclear medical findings (isotope nephrography and renal sequence scintigraphy) and enzymuria registered. The methods of investigation complement each other and should be employed in combination. A wide use of urinary enzyme diagnosis to identify mild and moderate renal injuries can be recommended.

Vitamin B complex in treatment of cadmium intoxication.

The effect of vitamin B-complex on cadmium nephrotoxicity and hepatotoxicity was investigated in rats. The administration of Cd (3 mg per kg, s.c., three days) increased the urinary excretions of lactic dehydrogenase (LDH), glutamic oxalacetic transaminase (GOT), and total proteins, decreased renal activities LDH and GOT and increased concentration in kidney tissue of Cd, Cu, and Zn, Cadmium also increased serum BOT and glutamic pyruvic transaminase (GPT), decreased hepatic activities of GOT and GPT, and increased hepatic levels of Cd and Zn. The supplementation of vitamin B-complex (10 mg per kg, orally) simultaneously with Cd caused less marked biological alterations. Cadmium concentration in renal tissue was significantly less on the eighth day whereas the hepatic level of Cd was unaffected by vitamin supplementation. The protective effect of vitamin B-complex in Cd toxicity may be attributed to the interference by the constituents of vitamin B-complex in body absorption of Cd, possibly through forming readily excretable complexes. The results suggest that Cd toxicity can be reduced by vitamin B-complex supplementation.

[Activity of various urinary enzymes in workers engaged in the production of polychlorinated pesticides]. / Aktywnosc niektórych enzymów w moczu pracowników zatrudnionych przy produkcji pestycydów polichlorowych.

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.

[Urinary enzyme excretion after prophylactic antibiotic therapy in patients following colorectal surgery (author's transl)]. / Harnenzymausscheidung nach präventiver antibiotikatherapie bei dickdarmresezierten patienten.

The urinary excretion of N-acetyl-beta-glucosaminade (NAG) and alanine aminopeptidase (AAP) was measured as an indicator for nephrotoxicity to neomycin and gentamycin in patients after colorectal surgery. We found that the continuous determination of urinary enzymes is a good indicator for the degree of kidney damage. The dose dependence of nephrotoxicity was influenced by the severity of colorectal surgery. The simple method of urinary enzyme determination seems to be a very valuable test to detect parenchymal damage of the kidney after prophylactic antibiotic therapy.

Neomycin toxicosis in calves.

Calves (n = 4) were given neomycin (2.25 or 4.5 mg/kg) twice daily IM and were compared with 2 calves given penicillin IM. The 2 hallmarks of aminoglycoside toxicosis, nephrotoxicosis and ototoxicosis, were seen with both dosages of parenterally administered neomycin. Nephrotoxicosis was confirmed by abnormal findings in urinalysis (granular casts, proteinuria, low specific gravity), renal biopsy results (tubular degeneration and necrosis), and increased 24-hour amounts of urinary enzymes (alanine aminopeptidase and gamma-glutamyltranspeptidase). Azotemia, decreased creatinine clearance, polyuria, and polydipsia also were documented in calves given neomycin. Clinically, deafness was suspected in 2 calves and was documented by electrical auditory-evoked response tests. Abnormalities in partial thromboplastin times and renal residues of neomycin were seen in all 4 calves that were given neomycin, but not in calves that were given penicillin.