Epidemiological studies suggest that hyponutrition during the fetal period increases the risk of mental disorders such as attention deficit hyperactivity disorder and autism-spectrum disorder, which has been experimentally supported using animal models. However, previous experimental hyponutrition or protein-restricted (PR) diets affected stages other than the fetal stage, such as formation of the egg before insemination, milk composition during lactation, and maternal nursing behavior.

This 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.

As a novel approach for disease control and prevention, nutritional modulation of the intestinal health has been proved. However, It is still unknown whether branched-chain amino acid (BCAA) is needed to maintain intestinal immune-related function. The objective of this study was to determine whether BCAA supplementation in protein restricted diet affects growth performance, intestinal barrier function and modulates post-weaning gut disorders. One hundred and eight weaned piglets (7.96±0.26 kg) were randomly fed one of the three diets including a control diet (21% crude protein [CP], CON), a protein restricted diet (17% CP, PR) and a BCAA diet (BCAA supplementation in the PR diet) for 14 d. The growth performance, plasma amino acid concentrations, small intestinal morphology and intestinal immunoglobulins were tested. First, average daily gain (ADG) (p<0.05) and average daily feed intake (ADFI) (p<0.05) of weaned pigs in PR group were lower, while gain:feed ratio was lower than the CON group (p<0.05). Compared with PR group, BCAA group improved ADG (p<0.05), ADFI (p<0.05) and feed:gain ratio (p<0.05) of piglets. The growth performance data between CON and BCAA groups was not different (p>0.05). The PR and BCAA treatments had a higher (p<0.05) plasma concentration of methionine and threonine than the CON treatment. The level of some essential and functional amino acids (such as arginine, phenylalanine, histidine, glutamine etc.) in plasma of the PR group was lower (p<0.05) than that of the CON group. Compared with CON group, BCAA supplementation significantly increased BCAA concentrations (p<0.01) and decreased urea concentration (p<0.01) in pig plasma indicating that the efficiency of dietary nitrogen utilization was increased. Compared with CON group, the small intestine of piglets fed PR diet showed villous atrophy, increasing of intra-epithelial lymphocytes (IELs) number (p<0.05) and declining of the immunoglobulin concentration, including jejunal immunoglobulin A (IgA) (p = 0.04), secreted IgA (sIgA) (p = 0.03) and immunoglobulin M (p = 0.08), and ileal IgA (p = 0.01) and immunoglobulin G (p = 0.08). The BCAA supplementation increased villous height in the duodenum (p<0.01), reversed the trend of an increasing IELs number. Notably, BCAA supplementation increased levels of jejunal and ileal immunoglobulin mentioned above. In conclusion, BCAA supplementation to protein restricted diet improved intestinal immune defense function by protecting villous morphology and by increasing levels of intestinal immunoglobulins in weaned piglets. Our finding has the important implication that BCAA may be used to reduce the negative effects of a protein restricted diet on growth performance and intestinal immunity in weaned piglets.

Between 6% and 11% of the world's population suffers from malnutrition or undernutrition associated with poverty, aging or long-term hospitalization. The present work examined the effect of different types of proteins on the mechanistic target of rapamycin (mTORC1)-signaling pathway in: (1) healthy; and (2) protein restricted rats. (1) In total, 200 rats were divided into eight groups and fed one of the following diets: 20% casein (C), soy (S), black bean (B), B + Corn (BCr), Pea (P), spirulina (Sp), sesame (Se) or Corn (Cr). Rats fed C or BCr had the highest body weight gain; rats fed BCr had the highest pS6K1/S6K1 ratio; rats fed B, BCr or P had the highest eIF4G expression; (2) In total, 84 rats were fed 0.5% C for 21 day and protein rehabilitated with different proteins. The S, soy + Corn (SCr) and BCr groups had the highest body weight gain. Rats fed SCr and BCr had the highest eIF4G expression and liver polysome formation. These findings suggest that the quality of the dietary proteins modulate the mTORC1-signaling pathway. In conclusion, the combination of BCr or SCr are the best proteins for dietary protein rehabilitation due to the significant increase in body weight, activation of the mTORC1-signaling pathway in liver and muscle, and liver polysome formation.

Compensatory growth is a physiological phenomenon found in both humans and animals. However, the underlying mechanisms are unclear. In this study, for the first time, we investigated the role of microbiota in compensatory growth induced by protein restriction using a rat model. Weaned Sprague-Dawley rats were fed a low protein diet (L group), a normal protein diet (N group) and a low protein diet for 2 weeks followed by a normal protein diet (LN group). The results showed that in contrast with the inhibited growth of rats in the L group, compensatory growth was observed in the LN group. Meanwhile, rats in the LN group had increased concentrations of total short chain fatty acids, particularly butyrate, and an altered bacterial composition with modified abundances of Peptostreptococcaceae, Bifidobacteriaceae, Porphyromonadaceae and Prevotellaceae in the colonic content. Furthermore, gene expression analysis indicated that the rats that experienced compensatory growth had improved barrier function and innate immune function in the colon. Our data revealed the importance of colonic microbiota in achieving compensatory growth.

Detrimental exposures during pregnancy have been implicated in programming offspring to develop permanent changes in physiology and metabolism, increasing the risk for developing diseases in adulthood such as hypertension, diabetes, heart disease and obesity. This study investigated the effects of protein restriction on the metabolism of amino acids within the oocyte, liver, and whole organism in a rat model as well as effects on mitochondrial ultrastructure and function in the cumulus oocyte complex.

Nicotine, the well known addictive chemical of tobacco and active medication for several diseases, has proven to be a potential genotoxic compound. Although it is absorbed through lungs with smoking and mainly metabolized in liver, its effect on liver injuries is not clear. This study was designed to evaluate the genotoxicity of nicotine and corresponding the protective role of curcumin against nicotine on liver of female populations particularly who used tobacco but deprived of healthy diet. The effects were investigated by measurement of total DNA concentration of liver tissues and Comet assay of liver tissue DNA damage of female rats maintained under normal and restricted protein diets. Total DNA contents in the liver tissues were observed to decrease more significantly (P<0.001) by nicotine in both dietary conditions. Significant (P<0.01) increase of total DNA content in normal dietary condition and more significant (P<0.001) increase of total DNA content in protein restricted condition of the liver tissues were observed due to curcumin supplementations. Highly significant (P<0.001) DNA damages (37% in normal diet and 56% in protein restricted diet) of the liver tissues were observed due to nicotine treatment. Curcumin reduced the nicotine-induced DNA damage percentage of the liver tissues more significantly (P<0.001) in protein restricted condition. Curcumin proved its potential to function against genotoxic effect by reducing the DNA damage activity of nicotine and minimized the percentage of DNA damage (50-60%) in protein restricted dietary condition. The degree of nicotine-induced genotoxicity therefore can be effectively compensated by the protective effect of curcumin in protein stress condition.

Gestational protein restriction was associated with low birth weight, hypertension and higher prevalence of cardiac disorders in adults. Several mechanisms, including epigenetics, could be related with the cardiovascular phenotype on protein-restricted offspring. Thus, we investigated the morphological cardiac effects of gestational protein restriction and left ventricle miRNAs and target genes expression pattern in both 12-day and 16-week old gestational protein-restricted male offspring. Pregnant Wistar rats were allocated into two groups, according to protein supply during pregnancy: NP (normal protein diet- 17%) or LP (low protein diet-6%). Dams on the gestational protein-restricted diet had lower body weight gain and higher food intake. Gestational protein-restricted offspring had low birth weight, followed by rapidly body weight recovery, hypertension, and increased myocytes cross-sectional area and collagen fraction at 16-week old age. At 12-days old, miR-184, miR-192, miR-376c, miR-380-3p, miR-380-5p, miR-451, and miR-582-3p had increased expression, and miR-547 and miR-743a had decreased expression in the gestational protein-restricted left ventricle. At 16-week old, let-7b, miR-125a-3p, miR-142-3p, miR-182 and miR-188-5p had increased expression and let-7g, miR-107, miR-127, miR-181a, miR-181c, miR-184, miR-324-5p, miR-383, miR-423-5p and miR-484 had decreased expression in gestational protein-restricted left ventricle. Target predicted gene expression analysis showed higher expression of Dnmt3a, Oxct1, Rictor and Trps1 and lower expression of Bbs1 and Calml3 in 12-day old protein-restricted offspring. 16-week old protein-restricted offspring had higher expression of Adrbk1, Bbs1, Dnmt3a, Gpr22, Inppl1, and Oxct1 genes. In conclusion, gestational protein restriction was related to offspring low birth weight, increased systolic blood pressure and morphological heart alterations that could be related to early heart miRNA expression changes that perpetuate into adulthood and which are associated with the regulation of essential genes involved in cardiovascular development, heart morphology, function, and metabolism.

A mixture of valine (Val) and isoleucine (Ile) not only decreases the negative impact of very low protein (VLP) diets on the growth of pigs, but also influences the nitrogen (N) balance and lipid metabolism; however, the underlying pathways are not well understood. This study aimed to investigate the effect of dietary Val and Ile on lipogenesis, lipolysis, and ureagenesis under protein restriction. After one week of acclimation, forty three-week-old pigs were randomly assigned to following dietary treatments (n = 8/group) for 5 weeks: positive control (PC): normal protein diet; negative control (NC): VLP diet; HV: NC supplemented with Val; HI: NC supplemented with Ile; and HVI: NC supplemented with both Val and Ile. HVI partially improved the body weight and completely recovered the feed intake (FI) of pigs fed with NC. HVI increased thermal radiation and improved the glucose clearance. HVI had a lower blood triglyceride than PC and blood urea N than NC. NC and HV promoted lipogenesis by increasing the transcript of fatty acid synthase (FAS) in the liver and lipoprotein lipase (LPL) in adipose tissue but reducing hormone-sensitive lipase (HSL) in the liver. HVI reduced the increased rate of lipogenesis induced by the NC group through normalizing the mRNA abundance of hepatic FAS, sterol regulatory element binding transcription factor 1, and HSL and LPL in adipose tissue. NC, HV, HI, and HVI reduced the ureagenesis by decreasing the protein abundance of carbamoyl phosphate synthetase I, ornithine transcarboxylase, and arginosuccinate lyase in the liver. Overall, HVI improved the growth, FI, and glucose clearance, and decreased the rate of lipogenesis induced by VLP diets.

A low-protein diet (LPD) is believed to be beneficial in slowing the progression of kidney disease. It is reported that low protein diet can improve protein, sugar and lipid metabolism, and reduce the symptoms and complications of renal insufficiency. However, there has been controversial regarding the effects of protein restriction on diabetic nephropathy (DN).

The liver is an essential regulator of energy metabolism, and its function can be disrupted by nutritional alterations. Since liver development continues during breastfeeding nutritional challenges during this period predispose patients to diseases throughout life. A maternal protein-restricted (PR) diet during lactation promotes reductions in the body weight, adiposity, and plasma glucose and insulin, leptin resistance and an increase in corticosterone and catecholamines in adult male rat offspring. Here, we investigated hepatic metabolism in the offspring (both sexes) of PR (8% protein diet during lactation) and control (23% protein diet) dams. Both male and female offspring were evaluated at 6 months of age. PR males had no liver steatosis and manifested a reduction in lipids in hepatocytes adjacent to the vasculature. These animals had lower levels of esterified cholesterol in hepatocytes, suggesting higher biliary excretion, unchanged glycolysis and gluconeogenesis, and lower contents of the markers of mitochondrial redox balance and endoplasmic reticulum (ER) stress response and estrogen receptor alpha. PR females showed normal hepatic morphology associated with higher uptake of cholesterol esters, normal glycolysis and gluconeogenesis, and lower ER stress parameters without changes in the key markers of the redox balance. Additionally, these animals had lower content of estrogen receptor alpha and higher content of androgen receptor. The maternal PR diet during lactation did not program hepatic lipid accumulation in the adult progeny. However, several repair homeostasis pathways were altered in males and females, possibly compromising maintenance of normal liver function.

The aim of this study was to investigate the reproductive, biochemical, and hematological outcomes of pregnant rats exposed to protein restriction. Wistar rat dams were fed a control normal-protein (NP, 17% protein, n=8) or a low-protein (LP, 8% protein, n=14) diet from the 1st to the 20th day of pregnancy. On the 20th day, the clinical signs of toxicity were evaluated. The pregnant rats were then anesthetized and blood samples were collected for biochemical-hematological analyses, and laparotomy was performed to evaluate reproductive parameters. No sign of toxicity, or differences (P>0.05) in body weight gain and biochemical parameters (urea, creatinine, albumin, globulin, and total protein) between NP and LP pregnant dams were observed. Similarly, hematological data, including red blood cell count, white blood cell count, hemoglobin, hematocrit, red blood cell distribution width (coefficient of variation), mean corpuscular volume, mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, % lymphocytes, absolute lymphocyte count, platelet count, and mean platelet volume were similar (P>0.05) at the end of pregnancy. Reproductive parameters (the dam-offspring relationship, ovary mass, placenta mass, number of corpora lutea, implantation index, resorption index, and the pre- and post-implantation loss rates) were also not different (P>0.05) between NP and LP pregnant dams. The present data showed that a protein-restricted diet during pregnancy did not alter reproductive, biochemical, and hematological parameters and seems not to have any toxic effect on pregnant Wistar rats.

We have examined maternal mechanisms for adult-onset glucose intolerance, increased adiposity, and atherosclerosis using two mouse models for intrauterine growth restriction (IUGR): maternal protein restriction and hypercholesterolemia.

Elevated blood pressure is an important global health problem, and in-utero under-nutrition may be an important factor in the pathogenesis of hypertension. In the present study, we tested the hypothesis that antenatal maternal low protein diet (MLPD) leads to sexually dimorphic developmental programming of the components of the pulmonary renin-angiotensin system. This may be important in the antenatal MLPD-associated development of hypertension. In pregnant mice, we administered normal (control) and isocaloric 50% protein restricted diet, commencing one week before mating and continuing until delivery of the pups. From the 18th to 24th week postnatal, we measured blood pressure in the offspring by use of a non-invasive tail-cuff method. In the same mice, we examined the mRNA and protein expression of the key components of the pulmonary renin-angiotensin system. Also, we examined microRNA complementary to angiotensin converting enzymes (ACE) 2 in the offspring lungs. Our results demonstrate that as a consequence of antenatal MLPD: 1) pup birthweight was significantly reduced in both sexes. 2) female offspring developed hypertension, but males did not. 3) In female offspring, ACE-2 protein expression was significantly reduced without any change in the mRNA levels. 4) miRNA 429, which has a binding site on ACE-2 - 3' UTR was significantly upregulated in the female antenatal MLPD offspring. 5) In males, ACE-2 mRNA and protein expression were unaltered. We conclude that in the mouse, antenatal MLPD-induced reduction of ACE-2 in the female offspring lung may be an important mechanisms in sexually dimorphic programming of hypertension.

Considering the prevalence of obesity and global aging, the consumption of a high-protein diet (HPD) may be advantageous. However, an HPD aggravates kidney dysfunction in patients with chronic kidney disease (CKD). Moreover, the effects of an HPD on kidney function in healthy individuals are controversial. In this study, we employed a remnant kidney mouse model as a CKD model and aimed to evaluate the effects of an HPD on kidney injury under conditions of non-CKD and CKD. Mice were divided into four groups: a sham surgery (sham) + normal diet (ND) group, a sham + HPD group, a 5/6 nephrectomy (Nx) + ND group and a 5/6 Nx + HPD group. Blood pressure, kidney function and kidney tissue injury were compared after 12 weeks of diet loading among the four groups. The 5/6 Nx groups displayed blood pressure elevation, kidney function decline, glomerular injury and tubular injury compared with the sham groups. Furthermore, an HPD exacerbated glomerular injury only in the 5/6 Nx group; however, an HPD did not cause kidney injury in the sham group. Clinical application of these results suggests that patients with CKD should follow a protein-restricted diet to prevent the exacerbation of kidney injury, while healthy individuals can maintain an HPD without worrying about the adverse effects.

The steroids corticosterone and dehydroepiandrosterone (DHEA) perform multiple life course functions. Rodent life-course circulating corticosterone and DHEA trajectories are unknown. We studied life course basal corticosterone and DHEA in offspring of rats fed protein-restricted (10% protein, R) or control (20% protein, C), pregnancy diet first letter, and/or lactation second letter, producing four offspring groups-CC, RR, CR, and RC. We hypothesize that 1. maternal diet programs are sexually dimorphic, offspring life course steroid concentrations, and 2. an aging-related steroid will fall. Both changes differ with the plastic developmental period offspring experienced R, fetal life or postnatally, pre-weaning. Corticosterone was measured by radioimmunoassay and DHEA by ELISA. Steroid trajectories were evaluated by quadratic analysis. Female corticosterone was higher than male in all groups. Male and female corticosterone were highest in RR, peaked at 450 days, and fell thereafter. DHEA declined with aging in all-male groups. DHEA: corticosterone fell in three male groups but increased in all-female groups with age. In conclusion, life course and sexually dimorphic steroid developmental programming-aging interactions may explain differences in steroid studies at different life stages and between colonies experiencing different early-life programming. These data support our hypotheses of sex and programming influences and aging-related fall in rat life course serum steroids. Life course studies should address developmental programming-aging interactions.

This experiment was conducted to investigate the effects of branched-chain amino acids (BCAA) supplemented in protein-restricted diets on the growth performance and the expression profile of amino acid transporters and energy metabolism related regulators in the white adipose tissue (WAT) of different regional depots including dorsal subcutaneous adipose (DSA) and abdominal subcutaneous adipose (ASA). A total of 24 crossbred barrows (7.40 ± 0.70 kg) were randomly divided into 4 groups and were fed the following isocaloric diets for 33 days: 1) a recommended adequate protein diet (AP, 20% CP, as a positive control); 2) a low protein diet (LP, 17% CP); 3) the LP diet supplemented with BCAA (LP + B, 17% CP) to reach the same level of the AP diet group; 4) the LP diet supplemented with 2 times the amount of BCAA (LP + 2B, 17% CP). The daily gain and daily feed intake of the LP diet group were the lowest among all the treatments (P < 0.01). The feed conversion was improved markedly in the group of LP + B compared with the LP diet group (P < 0.05). No significant difference was noted for the serum biochemical parameter concentrations of glucose, triglyceride, nonesterified fatty acid and insulin among the groups (P > 0.05). Moreover, BCAA supplementation down-regulated the expression levels of amino acid transporters including L-type amino acid transporter 1 and sodium-coupled neutral amino acid transporter 2 in DSA, but up-regulated the expression level of L-type amino acid transporter 4 in ASA (P < 0.05). Meanwhile, the energy sensor AMP-activated protein kinase α was activated in the DSA of pigs fed LP diet and in the ASA of the pigs fed AP or LP + 2B diets (P < 0.05). The mRNA expression profile of the selected mitochondrial component and mitochondrial biogenesis associated regulators in DSA and ASA also responded differently to dietary BCAA supplementation. These results suggested that the growth performance of growing pigs fed protein restricted diets supplemented with BCAA could catch up to that of the pigs fed AP diets. The results also partly demonstrated that the regulation mechanisms of BCAA are different in the adipose tissues of different depots.

Prior study shows that maternal protein-restricted (LP) 16-wk-old offspring have pronounced reduction of nephron number and arterial hypertension associated with unchanged glomerular filtration rate, besides enhanced glomerular area, which may be related to glomerular hyperfiltration/overflow and which accounts for the glomerular filtration barrier breakdown and early glomerulosclerosis. In the current study, LP rats showed heavy proteinuria associated with podocyte simplification and foot process effacement. TGF-β1 glomerular expression was significantly enhanced in LP. Isolated LP glomeruli show a reduced level of miR-200a, miR-141, miR-429 and ZEB2 mRNA and upregulated collagen 1α1/2 mRNA expression. By western blot analyzes of whole kidney tissue, we found significant reduction of both podocin and nephrin and enhanced expression of mesenchymal protein markers such as desmin, collagen type I and fibronectin. From our present knowledge, these are the first data showing renal miRNA modulation in the protein restriction model of fetal programming. The fetal-programmed adult offspring showed pronounced structural glomerular disorders with an accentuated and advanced stage of fibrosis, which led us to state that the glomerular miR-200 family would be downregulated by TGF-β1 action inducing ZEB 2 expression that may subsequently cause glomeruli epithelial-to-mesenchymal transition.

Offspring of murine dams chronically fed a protein-restricted diet have an increased risk for metabolic and neurobehavioral disorders. Previously we showed that adult offspring, developmentally exposed to a chronic maternal low-protein (MLP) diet, had lower body and hind-leg muscle weights and decreased liver enzyme serum levels. We conducted energy expenditure, neurobehavioral and circadian rhythm assays in male offspring to examine mechanisms for the body-weight phenotype and assess neurodevelopmental implications of MLP exposure. C57BL/6J dams were fed a protein restricted (8%protein, MLP) or a control protein (20% protein, C) diet from four weeks before mating until weaning of offspring. Male offspring were weaned to standard rodent diet (20% protein) and single-housed until 8-12 weeks of age. We examined body composition, food intake, energy expenditure, spontaneous rearing activity and sleep patterns and performed behavioral assays for anxiety (open field activity, elevated plus maze [EPM], light/dark exploration), depression (tail suspension and forced swim test), sociability (three-chamber), repetitive (marble burying), learning and memory (fear conditioning), and circadian behavior (wheel-running activity during light-dark and constant dark cycles). We also measured circadian gene expression in hypothalamus and liver at different Zeitgeber times (ZT). Male offspring from separate MLP exposed dams had significantly greater body fat (P = 0.03), less energy expenditure (P = 0.004), less rearing activity (P = 0.04) and a greater number of night-time rest/sleep bouts (P = 0.03) compared to control. MLP offspring displayed greater anxiety-like behavior in the EPM (P<0.01) but had no learning and memory deficit in fear-conditioning assay (P = 0.02). There was an effect of time on Per1, Per 2 and Clock circadian gene expression in the hypothalamus but not on circadian behavior. Thus, transplacental and early developmental exposure of dams to chronic MLP reduces food intake and energy expenditure, increases anxiety like behavior and disturbs sleep patterns but not circadian rhythm in adult male offspring.

Obesity is a growing health crisis of pandemic proportions. Numerous animal and human studies have confirmed that obesity and related metabolic abnormalities, such as insulin resistance and cardiovascular disease, may be programmed during development by adverse maternal nutrition. We previously documented that offspring of female mice who were protein-restricted during pregnancy alone had no alterations to their body weights, but did display a considerable reduction in food intake, a finding which was linked to reduced expression levels of appetite regulatory genes in the hypothalamus. Whether such observations were accompanied by changes in metabolic and phenotypic parameters remained to be determined. Female pregnant MF-1 mice were fed, exclusively during the pregnancy period, a normal protein diet containing 18% casein (C) or an isocaloric protein-restricted diet containing 9% casein (PR). From birth, the lactating dams were fed a normal protein diet. At weaning, offspring were fed either the standard chow which contain 7% kcal fat (C) or high-fat diet (HF, 45% kcal fat). This yielded 4 experimental groups denoted by maternal diet/offspring diet: C/C, C/HF, PR/C, PR/HF. Our results showed that offspring adiposity was significantly increased in HF-fed offspring, and was not affected by the 50% reduction in protein content of the maternal diet fed during pregnancy. Similarly, blood glucose levels were higher in HF-fed offspring, regardless of protein content of the maternal diet. Systolic blood pressure, on the other hand, was significantly increased in both male and female offspring of dams fed the PR diet, and this was exacerbated by a postweaning HF diet. Our results show that maternal protein restriction leads to elevations in systolic blood pressure, which is exacerbated by a postweaning HF-diet. Our present findings suggest that, while changes in offspring adiposity brought about by exposure to maternal protein restriction during pregnancy may be restored by adequate maternal protein content during lactation, the same may not be true for systolic blood pressure, which was similarly impaired, regardless of the timing of maternal low-protein exposure.