Dietary Protein Restriction Research Articles

Effects of dietary protein restriction followed by realimentation on growth performance and liver transcriptome alterations of lamb

The present study aimed to investigate the compensatory effect of early protein restriction followed by a realimentation on growth performance of lamb and to explore the transcriptomic changes in liver. Thirty-two lambs with an initial birth weight of 2.3 ± 0.20 kg that were weaned on day 15 were randomly divided into two groups. The lambs were fed a basal diet with normal protein level (NPL, protein level in the milk replacer and starter, 25 and 21%, respectively) or low protein level (LPL, protein level in the milk replacer and starter, 19 and 15%, respectively) from 15 to 60 d, after which all lambs consumed the same diet with a normal protein level from 61 to 90 d. Protein restriction led to a significant decrease in average daily gain (ADG), body weight and liver weight (P < 0.05). Transcriptome analysis showed that 302 or 12 differentially expressed genes (DEGs) were identified during the restriction or recovery periods, respectively (P < 0.05). The Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis indicated that DEGs enriched in nutrient metabolism and antioxidant capacity were down-regulated, while vessel development and immunity response-related genes up-regulated. The genes involved in metabolism of tyrosine were still down-regulated in the realimentation phase. Studies in this area indicated the accelerated growth effect of early protein restriction followed by a realimentation on growth performance of lambs and explored the transcriptomics change of liver which can help to develop feeding strategies to optimize the use of feedstuffs and in providing a new perspective for the study of early nutrition and epigenetics in later life.

Protein restricted diet during gestation and/or lactation in mice affects 15N natural isotopic abundance of organs in the offspring: Effect of diet 15N content and growth.

Objectives and studyThis study aimed at measuring the effect in normal to restricted protein diets with specific 15N natural isotopic abundance (NIA) given during gestation and/or lactation on the 15N NIA of fur, liver and muscle in dams and their offspring from birth to adulthood. The secondary aim was to study the effect of growth on the same parameters.MethodsFemale Balb/c mice were fed normal protein diet containing 22% protein or isocaloric low protein diet containing 10% protein throughout gestation. Dam’s diets were either maintained or switched to the other diet until weaning at 30 days. All animals were fed standard chow thereafter. Offspring were sacrificed at 1, 11, 30, 60, 480 days and a group of dams at d1. Growth was modeled as an exponential function on the group followed up until 480 days. Fur, liver and muscle were sampled at sacrifice and analyzed for bulk 15N NIA. Fixed effects and interactions between fixed effects and random elements were tested by three-way ANOVA.ResultsHigher 15N NIA in the diet resulted in higher organ 15N NIA. Switching from one diet to another changed 15N NIA in each organ. Although dam and offspring shared the same isotopic environment during gestation, 15N NIA at day 1 was higher in dams. Growth rate did not differ between groups after 10 days and decreased between 1 and 5 months. 15N NIA differed between organs and was affected by growth and gestation/lactation.ConclusionDietary 15N NIA is a major determinant of the 15N NIA of organs. 15N NIA depended on organ and age (i.e. growth) suggesting an effect of metabolism and/or dilution space. Post-natal normal-protein diet of lactating dams could reverse the effect of a protein-restricted diet during gestation on the offspring growth. Measuring 15N NIA in various matrices may open a field of application particularly useful in studying the pre- and post-natal origins of health and disease.

Low protein diets for non-diabetic adults with chronic kidney disease.

Low protein diets for non-diabetic adults with chronic kidney disease.

Protein-restricted maternal diet during lactation decreases type I and type III tropocollagen synthesis in the skin of mice offspring.

We investigated the effects of a low protein (LP) maternal diet during lactation on type I and III tropocollagen synthesis in infant mouse skin. The LP diet decreased the levels of type I and III tropocollagen proteins and COL1A1 and COL3A1 mRNA. Thus, the protein composition of the maternal perinatal diet may influence the skin health of offspring.

Development of a synthetic live bacterial therapeutic for the human metabolic disease phenylketonuria.

Phenylketonuria (PKU) is a genetic disease that is characterized by an inability to metabolize phenylalanine (Phe), which can result in neurotoxicity. To provide a potential alternative to a protein-restricted diet, we engineered Escherichia coli Nissle to express genes encoding Phe-metabolizing enzymes in response to anoxic conditions in the mammalian gut. Administration of our synthetic strain, SYNB1618, to the Pahenu2/enu2 PKU mouse model reduced blood Phe concentration by 38% compared with the control, independent of dietary protein intake. In healthy Cynomolgus monkeys, we found that SYNB1618 inhibited increases in serum Phe after an oral Phe dietary challenge. In mice and primates, Phe was converted to trans-cinnamate by SYNB1618, quantitatively metabolized by the host to hippurate and excreted in the urine, acting as a predictive biomarker for strain activity. SYNB1618 was detectable in murine or primate feces after a single oral dose, permitting the evaluation of pharmacodynamic properties. Our results define a strategy for translation of live bacterial therapeutics to treat metabolic disorders.

Pharmacokinetics of glycerol phenylbutyrate in pediatric patients 2 months to 2 years of age with urea cycle disorders

IntroductionGlycerol phenylbutyrate (GPB) is approved in the US and EU for the chronic management of patients ≥2 months of age with urea cycle disorders (UCDs) who cannot be managed by dietary protein restriction and/or amino acid supplementation alone. GPB is a pre-prodrug, hydrolyzed by lipases to phenylbutyric acid (PBA) that upon absorption is beta-oxidized to the active nitrogen scavenger phenylacetic acid (PAA), which is conjugated to glutamine (PAGN) and excreted as urinary PAGN (UPAGN). Pharmacokinetics (PK) of GPB were examined to see if hydrolysis is impaired in very young patients who may lack lipase activity. MethodsPatients 2 months to <2 years of age with UCDs from two open label studies (n = 17, median age 10 months) predominantly on stable doses of nitrogen scavengers (n = 14) were switched to GPB. Primary assessments included traditional plasma PK analyses of PBA, PAA, and PAGN, using noncompartmental methods with WinNonlin™. UPAGN was collected periodically throughout the study up to 12 months. ResultsPBA, PAA and PAGN rapidly appeared in plasma after GPB dosing, demonstrating evidence of GPB cleavage with subsequent PBA absorption. Median concentrations of PBA, PAA and PAGN did not increase over time and were similar to or lower than the values observed in older UCD patients. The median PAA/PAGN ratio was well below one over time, demonstrating that conjugation of PAA with glutamine to form PAGN did not reach saturation. Covariate analyses indicated that age did not influence the PK parameters, with body surface area (BSA) being the most significant covariate, reinforcing current BSA based dosing recommendations as seen in older patients. ConclusionThese observations demonstrate that UCD patients aged 2 months to <2 years have sufficient lipase activity to adequately convert the pre-prodrug GPB to PBA. PBA is then converted to its active moiety (PAA) providing successful nitrogen scavenging even in very young children.

Effect of dietary protein level and conjugated linoleic acid supply on milk secretion and fecal excretion of fatty acids

Little is known about the influence of dietary crude protein (CP) restriction and rumen-protected conjugated linoleic acid (rpCLA) supply on digestibility and metabolism of nutrients in dairy cows. This study investigated the effect of low dietary CP level combined with rpCLA supply on FA secretion into milk and excretion via feces in dairy cows. Twenty Friesian cows were involved in a 4 × 4 Latin square design over 3-week periods. Cows were fed 4 different rations based on 2 dietary CP levels (150 and 123 g/kg dry matter of CP), each one with or without the addition of a 80 g/d supplement of rpCLA. Feed, feces and milk samples were analyzed for their fatty acid content (FA) and data were used to calculate apparent FA transfer as the ratios between the daily FA secretion in milk, or excretion in feces, and intake. Dietary CP restriction had little influence on apparent FA transfer from the feed to the feces and the milk, but slightly increased the apparent transfer to the milk of 14:0iso (P = 0.031) and 15:0iso (P = 0.006), and tended to increase transfer of other branched FAs of microbial origin, i.e. 15:0anteiso (P = 0.06), 16:0iso (P = 0.06) and 17:0iso (P = 0.07). The rpCLA supply lowered the secretion/intake ratio of total FAs in the milk (P = 0.02), particularly those with the shortest chains and with even numbers of carbons, but had no effect on apparent transfer of branched and odd FAs of microbial origin. The rpCLA supply greatly reduced (P < 0.001) the secretion/intake ratio of 18:2c9,t11 and 18:2t10,c12 in the milk but also increased (P < 0.001) the milk secretion of 18:2c11,t13 (not supplied with the rpCLA) leaving no trace of it in the feces. In conclusion, the CP × rpCLA interaction had negligible effects of FA profile of both milk and feces. Our results suggested that rpCLA supply alters milk FA profile, mainly reducing the de novo synthesis of short-chain FAs and the Δ9 desaturase activity in the bovine mammary gland. Little influence of the dietary CP on animal digestive and metabolic processing of nutrients at the levels used in this experiment were suggested by small variations in the transfer of FA from feed to milk and feed to feces.

Medical Nutrition Therapy for Patients with Non–Dialysis-Dependent Chronic Kidney Disease: Barriers and Solutions

Medical Nutrition Therapy for Patients with Non–Dialysis-Dependent Chronic Kidney Disease: Barriers and Solutions

Moderate Dietary Protein Restriction Optimized Gut Microbiota and Mucosal Barrier in Growing Pig Model.

Appropriate protein concentration is essential for animal at certain stage. This study evaluated the effects of different percentages of dietary protein restriction on intestinal health of growing pigs. Eighteen barrows were randomly assigned to a normal (18%), low (15%), and extremely low (12%) dietary protein concentration group for 30 days. Intestinal morphology and permeability, bacterial communities, expressions, and distributions of intestinal tight junction proteins, expressions of biomarkers of intestinal stem cells (ISCs) and chymous bacterial metabolites in ileum and colon were detected. The richness and diversity of bacterial community analysis with Chao and Shannon index were highest in the ileum of the 15% crude protein (CP) group. Ileal abundances of Streptococcaceae and Enterobacteriaceae decreased respectively, while beneficial Lactobacillaceae, Clostridiaceae_1, Actinomycetaceae, and Micrococcaceae increased their proportions with a protein reduction of 3 percentage points. Colonic abundances of Ruminococcaceae, Christensenellaceae, Clostridiaceae_1, Spirochaetaceae, and Bacterodales_S24-7_group declined respectively, while proportions of Lachnospiraceae, Prevotellaceae, and Veillonellaceae increased with dietary protein reduction. Concentrations of most bacterial metabolites decreased with decreasing dietary protein concentration. Ileal barrier function reflected by expressions of tight junction proteins (occludin, zo-3, claudin-3, and claudin-7) did not show significant decrease in the 15% CP group while sharply reduced in the 12% CP group compared to that in the 18% CP group. And in the 15% CP group, ileal distribution of claudin-3 mainly located in the cell membrane with complete morphological structure. In low-protein treatments, developments of intestinal villi and crypts were insufficient. The intestinal permeability reflected by serous lipopolysaccharide (LPS) kept stable in the 15% CP group while increased significantly in the 12% CP group. The expression of ISCs marked by Lgr5 slightly increased in ileum of the 15% CP group. Colonic expressions of tight junction proteins declined in extremely low protein levels. In conclusion, moderate protein restriction (15% CP) can optimize the ileal microbiota structure via strengthening beneficial microbial populations and suppressing harmful bacterial growth and altering the function of ileal tight junction proteins as well as epithelial cell proliferation.

Vegetarian diets and chronic kidney disease.

While dietary restriction of protein intake has long been proposed as a possible kidney-protective treatment, the effects of changes in the quality of ingested proteins on the prevalence and risk of progression of chronic kidney disease (CKD) have been scarcely studied; these two aspects are reviewed in the present article. The prevalence of hypertension, type 2 diabetes and metabolic syndrome, which are the main causes of CKD in Western countries, is lower in vegetarian populations. Moreover, there is a negative relationship between several components of plant-based diets and numerous factors related to CKD progression such as uraemic toxins, inflammation, oxidative stress, metabolic acidosis, phosphate load and insulin resistance. In fact, results from different studies seem to confirm a kidney-protective effect of plant-based diets in the primary prevention of CKD and the secondary prevention of CKD progression. Various studies have determined the nutritional safety of plant-based diets in CKD patients, despite the combination of a more or less severe dietary protein restriction. As observed in the healthy population, this dietary pattern is associated with a reduced risk of all-cause mortality in CKD patients. We propose that plant-based diets should be included as part of the clinical recommendations for both the prevention and management of CKD.

The impact of phenylketonuria on PKU patients’ quality of life: Using of the phenylketonuria-quality of life (PKU-QOL) questionnaires

Phenylketonuria (PKU) has a very high prevalence throughout the world. Nowadays, number of studies about impact of this metabolic disease on patients increasing. The aim of our study is to examine PKU patients' quality of life according to PKU-QOL questionnaires. Patients (n=63) diagnosed with PKU were included this study; child (9-11 years (n=20)), adolescent (12-15 years (n=22)) and adult (18-35 years (n=21)). PKU-QOL questionnaires (include 4 modules) developed for PKU patients were used. In accordance with purpose, data were analysed by nonparametric tests (Kruskal Wallis One-Way Analysis of Variance Test and Mann-Whitney U Test), according to results of normality tests. Most of the individuals were female (65,1%) and mean age was 15,7±6,4 years. Symptoms; there were statistically significant differences in all domains excluding tiredness. Especially, median score of slow thinking was very frequent symptom in children as 100,0. PKU in general; there were found that median scores were higher in children. Phe-free amino acid supplement administration: as the age increased, scores were lower. Dietary protein restriction: Overall difficulty following dietary protein restriction and Food enjoyment were found similar in groups (p>0,05). It was concluded that PKU affects younger people more negatively.

Protein-Restricted Diet Is Effective in Decreasing Glycemia, HbA1c, and Cholesterol in Type 2 Diabetic Subjects by the Activation of the GCN2 Pathway

Dietary restriction increases lifespan across the evolutionary spectrum and decreases metabolic disease risk in mammals. It has been shown that amino acid restriction is sufficient to promote some of the beneficial effects of dietary restriction in rodents. This phenomenon is dependent on GCN2 (general control nonderepressible 2), a kinase activated by amino acid restriction that controls protein synthesis and stress response. Our results show that treatment of type 2 diabetic subjects on a protein restriction diet for 4 weeks exhibited a 61% reduction in fasting glucose levels and a 14% decrease in Hb1Ac. These patients had a 32% reduction in Total Cholesterol with a 33% decrease in LDL. We noticed a reduction of 4.5% of body weight, with a decrease of 11% of fat mass and maintenance of lean mass. In addition, all subjects had systemic blood pressure normalization, with a decrease of 23% in systolic pressure and 44% in diastolic pressure. In order to understand in detail the action of protein restriction we evaluated the gene expression in the subcutaneous adipose tissue of these individuals. We observed the activation of GCN2 expression (p=0.0394) and consequently the activation of ATF4 (Activating transcription factor 4) and CHOP (C/EBP homologous protein) (p=0.0360 and p=0.0011 respectively). This findings suggesting that in fact, protein restricted diet affects the GCN2 pathway in humans, modulating the energy metabolism and enabling glycemic and lipidemic control. Disclosure R.C. Ferraz: None. R.A. Beraldo: None. P. Gomes: None. M.C. Foss: None. M. Foss-Freitas: None.

Neuronal FGF-21 Signaling–A Sensor of Dietary Protein Restriction

The central nervous system is acknowledged as a major regulator of both energy and glucose homeostasis. Our data demonstrate that dietary protein restriction increases energy expenditure and improves glucose homeostasis, and that the metabolic hormone FGF-21 coordinates these adaptive responses. However, the tissue site mediating these FGF-21-dependent effects is unclear. In this study, mice with dysfunctional FGF-21 signaling in either the central nervous system (CNS) or adipose tissue were fed a control or low protein (LP) diet to assess changes in body weight and associated metabolic endpoints. We first deleted the required FGF co-receptor beta-Klotho (Klb) from neurons to test whether CNS FGF-21 signaling is required for increased energy expenditure or improved glucose homeostasis in mice consuming a low protein diet. Our data show that LP diet increased energy expenditure (p≤0.01) and reduced body weight (p≤0.001) in control littermates, but these effects were lost in mice bearing CNS-specific deletion of Klb (p=0.65). Protein restriction in the context of a high fat diet also improved glucose tolerance (P≤0.001) and insulin sensitivity (p≤0.001) in control littermates, but these effects were also lost in mice lacking Klb in the CNS. We next tested whether FGF-21 signaling within adipose tissue contributed to the metabolic effects of protein restriction. However, in this case LP diet reduced body weight (p=0.90) and increased food intake (p= 0.98) similarly in control littermates and mice bearing adipose-specific deletion of Klb. Collectively, these data highlight CNS FGF-21 signaling as the first known neuroendocrine mechanism to explain the coordinated metabolic changes induced by dietary protein restriction. Disclosure C.M. Hill: None. M.V. Dehner: None. D. McDougal: None. H. Berthoud: None. H. Muenzberg: None. C. Morrison: None.

A Low-Protein Calorie-Restricted Diet Attenuates Renal Injury and Facilitates Podocyte Autophagy in Type 2 Diabetic Rats

Purpose: Caloric restrictionhas been shown to exert a renoprotective effect in diabetic rats. Dietary protein restriction slows the progression of nephropathy in diabetic patients.We explored whether a low-protein calorie-restricted (LPCR) diet reduces renal injury and whether the renoprotective effect of LPCR diet isrelated to the activation of podocyte autophagy. Methods: The type 2 diabeticrat model was induced by a combination of high fat diet and low dose streptozotocin injection. The diabetic rats were randomly divided into 3 groups, normal protein (NP, 20% protein) group, low protein(LP, 8% protein) group and LPCR (8% protein, 30% restriction)group. Nondiabetic rats were used as normal control (NC). After 4 weeks of dietary intervention, urine albumin-to-creatinine ratio,creatinine clearance, serum creatinine, and kidney-to-body weight ratio, renal histologies, podocyte density, autophagy activity in glomeruli as well as glomerular basement membrane thickness and autophagic vacuoles in podocytesunder electron microscopy were estimated. Results: TheLPCR-diettreated diabetic ratsexhibited reduced body weight, serum triglycerides, and fasting blood glucose levels, as well as improved insulin resistance comparedwith NP-diettreated diabetic rats. LPCR diet attenuated renal injury, includingpodocyte injury, in type 2 diabetic rats. The reduced number of autophagic vacuolesin podocytesand insufficient podocyte autophagyin type 2 diabetic rats were reversed by LPCR diet. Conclusions: The findings suggest that a LPCR diet can ameliorate renal injury in type 2 diabetic rats, which maybe linked to the activation of podocyte autophagy. Disclosure G. Chen: None. Y. Geng: None. C. Liu: None.

Homeostatic sensing of dietary protein restriction: A case for FGF21.

Restriction of dietary protein intake increases food intake and energy expenditure, reduces growth, and alters amino acid, lipid, and glucose metabolism. While these responses suggest that animals 'sense' variations in amino acid consumption, the basic physiological mechanism mediating the adaptive response to protein restriction has been largely undescribed. In this review we make the case that the liver-derived metabolic hormone FGF21 is the key signal which communicates and coordinates the homeostatic response to dietary protein restriction. Support for this model centers on the evidence that FGF21 is induced by the restriction of dietary protein or amino acid intake and is required for adaptive changes in metabolism and behavior. FGF21 occupies a unique endocrine niche, being induced when energy intake is adequate but protein and carbohydrate are imbalanced. Collectively, the evidence thus suggests that FGF21 is the first known endocrine signal of dietary protein restriction.

Liver transplantation from a live donor to a patient with maple syrup urine disease: Two case reports

Liver transplantation were reported in patients with classic maple syrup urine disease in the literature. Branched chain alpha keto acid dehydrogenase activity can be improved in patients after transplant, and a protein-restricted diet is usually not needed. The first patient was a boy aged 2,5 years who presented with frequent ketosis attacks and epileptic seizures, and the second patient was an 11-month-old boy who also presented with frequent ketosis episodes, both despite adherence to diet therapy. Both patients received liver transplantations from live donors. A low protein diet was no longer required and no decline in cognitive functions was observed in either patient in the follow-up. We wanted to present these cases to show that despite a normal diet, plasma levels of branched- chain amino acids remained normal without any decline in cognitive function after liver transplantation in patients with classic maple syrup urine disease patients.

The Protein Status of Rats Affects the Rewarding Value of Meals Due to their Protein Content

Protein status is controlled by the brain, which modulates feeding behavior to prevent protein deficiency. This study tested in rats whether protein status modulates feeding behavior through brain reward pathways. Experiments were conducted in male Wistar rats (mean ± SD weight; 230±16 g). In experiment 1, rats adapted for 2 wk to a low-protein (LP; 6% of energy) or a normal-protein (NP; 14% of energy) diet were offered a choice between 3 cups containing high-protein (HP; 50% of energy), NP, or LP feed; their intake was measured for 24 h. In 2 other experiments, the rats were adapted for 2 wk to NP and either HP or LP diets and received, after overnight feed deprivation, a calibrated HP, NP, or LP meal daily. After the meal, on the last day, rats were killed and body composition and blood protein, triglycerides, gut neuropeptides, and hormones were determined. In the brain, neuropeptide mRNAs in the hypothalamus and c-Fos protein and opioid and dopaminergic receptor mRNAs in the nucleus accumbens (NAcc) were measured. Rats fed an LP compared with an NP diet had 7% lower body weight, significantly higher protein intake in a choice experiment (mean ± SD: 30.5%±0.05% compared with 20.5%±0.05% of energy), higher feed-deprived blood ghrelin, lower postmeal blood leptin, and higher neuropeptide Y (Npy) and corticotropin-releasing hormone (Crh) mRNA expression in the hypothalamus. In contrast to NP, rats fed an LP diet showed postmeal c-Fos protein expression in the NAcc, which was significantly different between meals, with LP < NP < HP. In contrast, in rats adapted to an HP diet compared with an NP diet, energy intake was lower; and in the NAcc, meal-induced c-Fos protein expression was 20% lower, and mRNA expression was 17% higher for dopamine receptor 2 (Drd2) receptors and 38% lower for κ opioid receptor (Oprk1) receptors. A protein-restricted diet induced a reward system-driven appetite for protein, whereas a protein-rich diet reduced the meal-induced activation of reward pathways and lowered energy intake in male rats.

Adherence to special diets and its association with meeting the nutrient recommendations in individuals with type 1 diabetes.

Not much is known about adherence to special diets in type 1 diabetes, characteristics of individuals with special diets, and whether such practices should raise concerns with respect to meeting the dietary recommendations. In this study, we assessed the frequencies of adherence to special diets, in a population of individuals with type 1 diabetes, and investigated the association between special diet adherence and dietary intake, measured as dietary patterns and nutrient intakes. During the Finnish Diabetic Nephropathy Study visit, participants with type 1 diabetes (n = 1429) were instructed to complete a diet questionnaire inquiring about the adherence to special diets. The participants also completed a food record, from which energy and nutrient intakes were calculated. In all, 36.6% participants reported adhering to some special diet. Most commonly reported special diets were lactose-free (17.1%), protein restriction (10.0%), vegetarian (7.0%), and gluten-free (5.6%) diet. Special diet adherents were more frequently women, older, had longer diabetes duration, and more frequently had various diabetes complications. Mean carbohydrate intakes were close to the lower levels of the recommendation in all diet groups, which was reflected in low mean fibre intakes but high frequencies of meeting the sucrose recommendations. The recommendation for saturated fatty acid intake was frequently unmet, with the highest frequencies observed in vegetarians. Of the micronutrients, vitamin D, folate, and iron recommendations were most frequently unmet, with some differences between the diet groups. Special diets are frequently followed by individuals with type 1 diabetes. The adherents are more frequently women, and have longer diabetes duration and more diabetes complications. Achieving the dietary recommendations differed between diets, and depended on the nutrient in question. Overall, intakes of fibre, vitamin D, folate, and iron fell short of the recommendations.

Gestational low-protein intake enhances whole-kidney miR-192 and miR-200 family expression and epithelial-to-mesenchymal transition in rat adult male offspring.

Studies have shown that adult offspring of mothers fed a protein-restricted diet during pregnancy present a pronounced reduction of nephron number associated with decreased fractional urinary sodium excretion and arterial hypertension. Additionally, recent advances in our understanding of the molecular pathways that govern the association of gestational nutritional restriction, intrauterine growth retardation and inflammation with impaired nephrogenesis, nephron underdosing and kidney fibrosis point to the epithelial to mesenchymal transition (EMT) as a common factor. In the current study, protein and sodium urinary excretion rates were evaluated in rats, and immunohistochemistry and western blot techniques were used to characterize kidney structure changes in 16week old male offspring of mothers fed a low-protein diet during pregnancy (LP group) compared with age-matched (NP) controls. We also verified the expression of miRNA, mRNA and protein markers of fibrosis and the EMT in whole kidney prepared from LP offspring. We found, surprisingly, that arterial hypertension and long-term hyperfiltration, manifest by proteinuria, were associated with increased renal miR-192 and miR-200 family expression in 16week old LP relative to age-matched NP rats. Measurement of kidney fibrosis and EMT-related protein markers, by histochemistry and immunoblot techniques, showed a significant rise of TGF-β1 and type-I collagen content in glomeruli and tubulointerstitial areas, accompanied by enhanced fibronectin and ZEB1 and decreased E-cadherin immunoreactivity in 16week old LP offspring. The results were partially confirmed by increased gene (mRNA) expression of collagen 1α1, collagen 1α2 and ZEB1 in LP whole kidneys compared with those of age-matched NP offspring. In view of the presumed functional overload in the remaining nephrons, we suggest that hypertension and proteinuria development following maternal protein restriction may be a preponderant factor for EMT and structural kidney changes in LP offspring. However, our study was not wholly able to establish the precise role of miRNAs in LP kidney disorders. Thus, further studies will be required to assess the contribution of the miR family to renal injury in a gestational protein-restricted model of fetal programming.

Evaluation of plasma biomarkers of inflammation in patients with maple syrup urine disease.

Maple syrup urine disease (MSUD) is an autosomal recessive inherited disorder that affects branched-chain amino acid (BCAA) catabolism and is associated with acute and chronic brain dysfunction. Recent studies have shown that inflammation may be involved in the neuropathology of MSUD. However, these studies have mainly focused on single or small subsets of proteins or molecules. Here we performed a case-control study, including 12 treated-MSUD patients, in order to investigate the plasmatic biomarkers of inflammation, to help to establish a possible relationship between these biomarkers and the disease. Our results showed that MSUD patients in treatment with restricted protein diets have high levels of pro-inflammatory cytokines [IFN-γ, TNF-α, IL-1β and IL-6] and cell adhesion molecules [sICAM-1 and sVCAM-1] compared to the control group. However, no significant alterations were found in the levels of IL-2, IL-4, IL-5, IL-7, IL-8, and IL-10 between healthy controls and MSUD patients. Moreover, we found a positive correlation between number of metabolic crisis and IL-1β levels and sICAM-1 in MSUD patients. In conclusion, our findings in plasma of patients with MSUD suggest that inflammation may play an important role in the pathogenesis of MSUD, although this process is not directly associated with BCAA blood levels. Overall, data reported here are consistent with the working hypothesis that inflammation may be involved in the pathophysiological mechanism underlying the brain damage observed in MSUD patients.