Glutamine supplementation does not improve protein synthesis rate by the jejunal mucosa of the malnourished rat

It has been demonstrated that glutamine, a conditionally essential amino acid, improves nitrogen balance, acts as a stimulant of protein synthesis, and decreases proteolysis in myopathic children. In contrast, other studies have shown no beneficial effect of glutamine supplementation on burn victims or critically ill patients. Nonetheless, we hypothesized that glutamine supplementation would increase the fractional protein synthesis rate (FSR) in the jejunal mucosa of malnourished male Wistar rats. Thus, the objective of the present study was to test the effect of daily oral glutamine supplementation (0.42 g kg(-1) d(-1) for 14 days) on the FSR of the jejunal mucosa of healthy and malnourished rats. A 4-hour kinetic study with L-[1-(13)C]leucine was subsequently performed, and jejunal biopsies were obtained 1.5 cm from the Treitz angle and analyzed. Malnourished rats showed a 25% weight loss and increased urinary nitrogen excretion. Plasma amino acid concentration did not differ between groups. (13)C enrichment in plasma and jejunal cells was higher in the malnourished groups than in the healthy group. The FSR (percent per hour) was similar for the control and experimental groups (P > .05), with a mean range of 220%/h to 27%/h. Oral glutamine supplementation alone did not induce higher protein incorporation by the jejunal mucosa in malnourished rats, regardless of total food intake or the presence or absence of glutamine supplementation. (C) 2009 Elsevier Inc. All rights reserved.

Neonatal Handling Reduces Renal Function in Adult Rats

Background/Aims: To evaluate the effects of neonatal handling on hydroelectrolytic balance in adult rats. Methods: The litters were divided into two groups: nonhandled and handled. The procedure consisted of handling the pups for 1 min/day in the first 10 days postnatally. When adults, animals had their body weight verified and were housed in individual metabolic cages. After a 24-hour period, urine samples were collected and the urinary and water intake volumes measured. Blood samples to determine osmolality, aldosterone, corticosterone, angiotensin II, creatinine, urea, sodium and potassium levels were collected. The kidneys were removed for histological assessment. Urinary osmolality, sodium, urea and creatinine were also measured and the creatinine clearance (CC) calculated. Results: No difference between groups was found in the body weight. Handled animals showed a reduction in the total kidney wet weight, water intake, urinary volume, CC, plasma angiotensin II, corticosterone and aldosterone when compared to the nonhandled and an increase in the urinary osmolality and sodium excretion fraction. No differences in serum potassium and no evidence of structural changes were demonstrated by histological analysis. Conclusion: Neonatal handling induced long-lasting effects decreasing renal function without evidence of kidney structural changes. Copyright (c) 2009 S. Karger AG, Basel

Mast cells, TGF-beta 1 and alpha-SMA expression in IgA nephropathy

IgA nephropathy (IgAN) is a kidney disease with a varying renal prognosis. Recently, many studies have demonstrated that renal alpha-smooth muscle actin (alpha-SMA) and transforming growth factor (TGF-beta 1) expression, as well interstitial mast cell infiltrates could represent a prognostic marker in several renal diseases. The aim of our study was to analyze the prognostic value of mast cell, TGF-beta 1 and alpha-SMA expression in IgAN. A survey of the medical records and renal biopsy reports of 62 patients with a diagnosis of IgAN followed-up from 1987 to 2003 was performed. The mean follow-up time was 74.7 +/- 50.0 months. The immunohistochemical studies were performed using a monoclonal antibody anti-human mast cell tryptase, a polyclonal antibody anti-human TGF-beta 1, and a monoclonal antibody anti-human alpha-SMA. An unfavorable clinical course of IgAN was related to interstitial mast cell infiltrates and alpha-SMA expression in the tubulointerstitial area. Expression of glomerular TGF-beta 1 and alpha-SMA, and interstitial TGF-beta 1 is not correlated with clinical course in IgAN. In conclusion, the increased number of mast cells and higher alpha-SMA expression in the tubulointerstitial area may be predictive factors for the poor prognosis of patients with IgAN.

Indicators of lean body mass catabolism: emphasis on the creatinine excretion rate

Background: The major stress response to critical illness leads to a catabolic state and loss of lean body mass. Aims: To test whether an increased rate of creatinine excretion might provide unique and timely information to monitor cell catabolism; to relate this information to balances of cell constituents (nitrogen, potassium, phosphate and magnesium); to evaluate the effectiveness of nutritional therapy to reverse this catabolic process. Design: Prospective observational study. Methods: Children with severe traumatic brain injury admitted to the paediatric critical care units of The Hospital for Sick Children, Toronto, Canada and Hospital das Clnicas, Faculty of Medicine of Ribeiro Preto, University of So Paulo, Brazil were studied. Complete 24 h urine collections were obtained for measurement of creatinine excretion rate and daily balances of nitrogen, potassium, phosphate and magnesium. Results: Seventeen patients were studied for 310 days. On Day 1, all had negative balances for protein and phosphate. Balances for these intracellular constituents became positive when protein intake was >= 1 g/kg/day and energy intake was >= 50% of estimated energy expenditure (P < 0.0001). Creatinine excretion rate was positively correlated with the urea appearance rate (r = 0.60; P < 0.0001), and negatively with protein balance (r = -0.45; P < 0.0001). Sepsis developed in four patients; before its clinical detection, there were negative balances for all intracellular markers and an abrupt rise in the excretion of creatinine. Conclusions: Negative balances of intracellular components and an increase in rate of creatinine excretion heralded the onset of catabolism.

Expression of genes related to apoptosis, cell cycle and signaling pathways are independent of TP53 status in urinary bladder cancer cells

Urinary bladder cancer is the fourth most common malignancy in the Western world. Transitional cell carcinoma (TCC) is the most common subtype, accounting for about 90% of all bladder cancers. The TP53 gene plays an essential role in the regulation of the cell cycle and apoptosis and therefore contributes to cellular transformation and malignancy; however, little is known about the differential gene expression patterns in human tumors that present with the wild-type or mutated TP53 gene. Therefore, because gene profiling can provide new insights into the molecular biology of bladder cancer, the present study aimed to compare the molecular profiles of bladder cancer cell lines with different TP53 alleles, including the wild type (RT4) and two mutants (5637, with mutations in codons 280 and 72; and T24, a TP53 allele encoding an in-frame deletion of tyrosine 126). Unsupervised hierarchical clustering and gene networks were constructed based on data generated by cDNA microarrays using mRNA from the three cell lines. Differentially expressed genes related to the cell cycle, cell division, cell death, and cell proliferation were observed in the three cell lines. However, the cDNA microarray data did not cluster cell lines based on their TP53 allele. The gene profiles of the RT4 cells were more similar to those of T24 than to those of the 5637 cells. While the deregulation of both the cell cycle and the apoptotic pathways was particularly related to TCC, these alterations were not associated with the TP53 status.

Evaluation of hepatobiliary excretion and enterobiliary reflux in rats with biliary obstruction submitted to bilioduodenal or biliojejunal anastomosis

Bilioduodenal and biliojejunal anastomoses are effective for the treatment of biliary obstruction. The objective of this study was to compare the effects of these anastomoses on hepatobiliary excretion and enterobiliary reflux. Enterobiliary reflux and biliary excretion were evaluated respectively after oral administration of technetium (Tc-99m) in combination with sodium phytate and intravenous infusion of Tc-99m with diisopropyl-iminodiacetic acid. Enterobiliary reflux occurred to an equal degree in the bilioduodenal and biliojejunal groups. Maximum hepatic activity time (T-max) and radiotracer clearance half-time (T-1/2) were similar in both groups. However, when compared with that found for the sham-operated group, T-max, and T-1/2 were higher in the biliojejunal group (P = 0.02 and P = 0.01, respectively). Histopathological analysis showed marked reduction in ductal proliferation in both groups. These data undermine the theoretical advantages attributed to biliojejunal anastomosis and further the understanding of the pathophysiology of cholangitis that occurs even with patent anastomosis.

Developing human minor salivary glands: morphological parallel relation between the expression of TGF-beta isoforms and cytoskeletal markers of glandular maturation

Morphogenesis of salivary glands involves complex coordinated events. Synchronisation between cell proliferation, polarisation and differentiation, which are dependent on epithelial-mesenchymal interactions and on the microenvironment, is a requirement. Growth factors mediate many of these orchestrated biological processes and transforming growth factor-beta (TGF-beta) appear to be relevant. Using immunohistochemistry and immunofluorescence, we have mapped the distribution of TGF-beta 1, 2 and 3 and compared it with the expression of maturation markers in human salivary glands obtained from foetuses ranging from weeks 4 to 24 of gestation. TGF-beta 1 first appeared during canalisation stage in the surrounding mesenchyme and, in the more differentiated stages, was expressed in the cytoplasm of acinar cells throughout the adult gland. TGF-beta 2 was detected since the bud stage of the salivary gland. Its expression was observed in ductal cells and increased along gland differentiation, TGF-beta 3 was detected from the canalisation stage of the salivary gland, being weakly expressed on ductal cells, and it was the only factor detected on myoepithelial cells. The data suggest that TGF-beta have a role to play in salivary gland development and differentiation.

Proteomic Analysis of Urine in Rats Chronically Exposed to Fluoride

Urine is an ideal source of materials to search for potential disease-related biomarkers as it is produced by the affected tissues and can be easily obtained by noninvasive methods. 2-DE-based proteomic approach was used to better understand the molecular mechanisms of injury induced by fluoride (F(-)) and define potential biomarkers of dental fluorosis. Three groups of weanling male Wistar rats were treated with drinking water containing 0 (control), 5, or 50 ppm F(-) for 60 days (n = 15/group). During the experimental period, the animals were kept individually in metabolic cages, to analyze the water and food consumption, as well as fecal and urinary F excretion. Urinary proteome profiles were examined using 2-DE and Colloidal Coomassie Brilliant Blue staining. A dose-response regarding F(-) intake and excretion was detected. Quantitative intensity analysis revealed 8, 11, and 8 significantly altered proteins between control vs. 5 ppm F(-), control vs. 50 ppm F(-) and 5 ppm F(-) vs. 50 ppm F(-) groups, respectively. Two proteins regulated by androgens (androgen-regulated 20-KDa protein and 0c-2,1-globulin) and one related to detoxification (aflatoxin-Bl-aldehyde-reductase) were identified by MALDI-TOF-TOF MS/MS. Thus, proteomic analysis can help to better understand the mechanisms underlying F(-) toxicity, even in low doses. 2010 Wiley Periodicals, Inc. J Biochem Mol Toxicol 25:8-14, 2011; View this article online at wileyonlinelibrary.com. DOI 10:1002/jbt.20353

Influence of Genetic Background on Fluoride Metabolism in Mice

A/J and 129P3/J mouse strains have different susceptibilities to dental fluorosis, due to their genetic backgrounds. This study tested whether these differences are due to variations in water intake and/or F metabolism. A/J (susceptible to dental fluorosis) and 129P3/J mice (resistant) received drinking water containing 0, 10, or 50 ppm F. Weekly F intake, excretion and retention, and terminal plasma and femur F levels were determined. Dental fluorosis was evaluated clinically and by quantitative fluorescence (QF). Data were tested by two-way ANOVA. Although F intakes by the strains were similar, excretion by A/J mice was significantly higher due to greater urinary F excretion, which resulted in lower plasma and femur F levels. Compared with 129P3/J mice given 50 ppm F, significantly higher QF scores were recorded for A/J mice. In conclusion, these strains differ with respect to several features of F metabolism, and amelogenesis in the 129P3/J strain seems to be unaffected by high F exposure.

Daily variations in human plasma fluoride concentrations

This study investigated the variations in human plasma fluoride concentrations ([F]) and sought to determine the causes. Five subjects (27-33 years old) received a low-F diet during the 5 days of the study. Plasma samples and urine were collected every 3 h from 8 a.m. to 8 p.m. F, PTH, Ca and P were analyzed with the electrode, by chemiluminescence, AAS and colorimetry, respectively. A trend for the plasma [F] was found. The peak [F], 0.55 +/- 0.11 mu mol L(-1), occurred at 11 a.m. and the lowest [F], 0.50 +/- 0.06 mu mol L(-1) occurred between 5 and 8 p.m. Plasma [F] were positively correlated with urinary F excretion rates and with serum PTH levels, but not with the Ca or P levels. Serum PTH levels were positively correlated with urinary F excretion rates and negatively correlated with plasma Ca. The results suggest that the renal system seems to control the daily fluctuations in plasma [F]. (c) 2008 Elsevier B.V. All rights reserved.

Biomarkers of Fluoride in Children Exposed to Different Sources of Systemic Fluoride

There has been no comparison between fluoride concentrations in urine and nails of children exposed to different sources of systemic fluoride. The aim of this study was to compare the relationship between fluoride intake with urinary fluoride excretion and fluoride concentrations in fingernails and toenails of children receiving fluoride from artificially fluoridated water (0.6-0.8 mg F/L, n = 25), naturally fluoridated water (0.6-0.9 mg F/L, n = 21), fluoridated salt (180-200 mg F/Kg, n = 26), and fluoridated milk (0.25 mg F, n = 25). A control population was included (no systemic fluoride, n = 24). Fluoride intake from diet and dentifrice, urinary fluoride excretion, and fluoride concentrations in fingernails/toenails were evaluated. Fluoride was analyzed with an ion-selective electrode. Urinary fluoride excretion in the control community was significantly lower when compared with that in the fluoridated cities, except for the naturally fluoridated community. However, the same pattern was not as evident for nails. Both urinary fluoride output and fluoride concentrations in fingernails/toenails were significantly correlated to total fluoride intake. However, the correlation coefficients for fluoride intake and urinary fluoride output were lower (r = 0.28, p < 0.01) than those observed for fingernails/toenails (r = 0.36, p < 0.001), suggesting that nails might be slightly better indicators of fluoride intake at the individual level.

Improved endothelial function with simvastatin but unchanged insulin sensitivity with simvastatin or ezetimibe

In addition to their expected effects on lipid profile, lipid-lowering agents may reduce cardiovascular events because of effects on nonclassic risk factors such as insulin resistance and inflammation. Ezetimibe specifically blocks the absorption of dietary and biliary cholesterol as well as plant sterols. Although it is known that an additional reduction of low-density lipoprotein cholesterol (LDL-C) levels can be induced by the combination of ezetimibe with statins, it is not known if this can enhance some pleiotropic effects, which may be useful in slowing the atherosclerotic process. This study assessed the effects of simvastatin and ezetimibe, in monotherapy or in combination, on markers of endothelial function and insulin sensitivity. Fifty prediabetic subjects with normo- or mild-to-moderate hypercholesterolemia were randomly allocated to 2 groups receiving either ezetimibe (10 mg/d) or simvastatin (20 mg/d) for 12 weeks, after which the drugs were combined for both groups for an additional 12-week period. Clinical and laboratory parameters were measured at baseline and after 12 and 24 weeks of therapy. Homeostasis model assessment of insulin resistance index and the area under the curve of insulin were calculated. As expected, both groups receiving drugs in isolation significantly reduced total cholesterol, LDL-C, apolipoprotein B, and triglyceride levels; and additional reductions were found after the combination period (P <.05). After 12 weeks of monotherapy, plasminogen activator inhibitor-1 levels and urinary albumin excretion were lower in the simvastatin than in the ezetimibe group. No change in homeostasis model assessment of insulin resistance index, area under the curve of insulin, and adiponectin levels was observed tiller either the monotherapies or the combined therapy. However, simvastatin combined with ezetimibe provoked significant reductions in E-selectin and intravascular cellular adhesion molecule-1 levels that were independent of LDL-C changes. Our findings support claims that simvastatin may be beneficial in preserving endothelial function in prediabetic subjects with normo- or mild-to-moderate hypercholesterolemia. Alternatively, a deleterious effect of ezetimibe on the endothelial function is suggested, considering the increase in intravascular cellular adhesion molecule I and E-selectin levels. Simvastatin and ezetimibe, in isolation or in combination, do not interfere with insulin sensitivity. (C) 2010 Elsevier Inc. All rights reserved.

N-acetylcysteine protects against renal injury following bilateral ureteral obstruction

Background. Obstructive nephropathy decreases renal blood flow (RBF) and glomerular filtration rate (GFR), causing tubular abnormalities, such as urinary concentrating defect, as well as increasing oxidative stress. This study aimed to evaluate the effects of N-acetylcysteine (NAC) on renal function, as well as on the protein expression of aquaporin 2 (AQP2) and endothelial nitric oxide synthase (eNOS), after the relief of bilateral ureteral obstruction (BUO). Methods. Adult male Wistar rats were divided into four groups: sham (sham operated); sham operated + 440 mg/kg body weight (BW) of NAC daily in drinking water, started 2 days before and maintained until 48 h after the surgery; BUO (24-h BUO only); BUO + NAC-pre (24-h BUO plus 440 mg/kg BW of NAC daily in drinking water started 2 days before BUO); and BUO + NAC-post (24-h BUO plus 440 mg/kg BW of NAC daily in drinking water started on the day of BUO relief). Experiments were conducted 48 h after BUO relief. Results. Serum levels of thiobarbituric reactive substances, which are markers of lipid peroxidation, were significantly lower in NAC-treated rats than in the BUO group rats. The administration of NAC provided significant protection against post-BUO GFR drops and reductions in RBF. Renal cortices and BUO rats presented decreased eNOS protein expression of eNOS in the renal cortex of BUO group rats, whereas it was partially recovered in BUO + NAC-pre group rats. Urine osmolality was significantly lower in BUO rats than in sham group rats or NAC-treated rats, the last also presenting less interstitial fibrosis. Post-BUO downregulation of AQP2 protein expression was averted in the BUO + NAC-pre group rats. Conclusions. This study demonstrates that NAC administration ameliorates the renal function impairment observed 48 h after the relief of 24-h BUO. Oxidative stress is important for the suppression of GFR, RBF, tissue AQP2 and eNOS in the polyuric phase after the release of BUO.

Dietary copper absorption and excretion in three semi-terrestrial grapsoid crabs with different levels of terrestrial adaptation

Three species of phylogenetically related semi-terrestrial crabs (Superfamily Grapsoidea - Sesarma rectum, Goniopsis cruentata and Neohelice granulata (formerly: Chasmagnathus granulatus) with different degrees of terrestriality were studied to quantify the accumulation of copper (Cu) in hemolymph, gills, hepatopancreas and antennal gland, and its excretion through the faeces. These crabs were fed for 15 days practical diets containing 0 (A), 0.5 (B), 1.0 (C), and 1.5% (D) of added CuCl2 (corresponding to 0, 0.2, 0.5 and 0.7% of Cu2+, respectively). The amount of food ingested was directly proportional to the degree of terrestriality: S. rectum, the most terrestrial species, ate around 2-3 times more than the other crabs, whereas G. cruentata ate 1.5-2 times more than N. granulata, the least terrestrial. The amount of Cu excreted in the feces was proportional to Cu ingestion, and was 76.8% and 64.2% higher for Sesarma fed diet D compared to G. cruentata and N. granulata, respectively. Sesarma also displayed higher Cu concentration in the haemolymph, gills and antennal glands, but not in the hepatopancreas. A detoxifying mechanism followed by elimination was probably present at this last organ, preventing Cu accumulation. More terrestrial crabs, such as Sesarma, may accumulate more Cu in hemolymph and tissues, showing a correlation between metal accumulation and increased terrestriality. In this aspect, contaminated feed sources with Cu may have more impact in conservation of terrestrial crabs. (C) 2008 Elsevier Inc. All rights reserved.

Opposite effects of fasting on TGF-beta 3 and T beta RI distribution in the gastric mucosa of suckling and early weanling rats

Objective: Our aim was to evaluate the effects of a dietary regimen (suckling or early weaning) and feeding status (fed or fasted) on the distribution of transforming growth factor-beta 3 (TGF-beta 3) and TGF receptor-I (T beta RI) in the gastric epithelium of pups Methods: Wistar rats were used At 15 d, half of the pups were separated from dams and fed with hydrated powered chow On day 17, suckling and early weanling rats were subjected to fasting (17 h). Four different conditions were established. suckling fed and fasted and early weanling fed and fasted At 18 d stomachs were collected under anesthesia and were fixed in 4% formaldehyde for immunohistochemistry The number of immunostained epithelial cells per microscopic field was determined for TGF-beta 3 and T beta RI in longitudinal sections from the gastric mucosa Results: We found that during suckling, fasting reduced the number of immunolabeled cells per field of both molecules when compared with the fed group (P < 0.05), whereas in early weaning, food restriction increased TGF-beta 3 and T beta RI distributions (P < 0.05) We also observed that TGF-beta 3 and T beta RI were more concentrated in parietal cells in the upper gland in suckling pups, whereas after early weaning these were displaced to parietal and chief cells at the bottom of the gland Conclusion: Suckling and early weaning directly influence TGF-beta 3 and T beta RI distributions in the gastric epithelium in response to fasting, such that early weaning anticipates the effects observed in adult rats. Furthermore, the differential concentrations of TGF-beta 3 and T beta RI indicate that they might be important for cell proliferation events in growth control (C) 2010 Elsevier Inc. All rights reserved