Protein restriction reprograms the multi-organ proteomic landscape of mouse aging.

FGF21, not GCN2, influences bone morphology due to dietary protein restrictions

The highly conserved effect of dietary protein restriction on lifespan and ageing is observed in both sexes and across a vast range of taxa. This extension of lifespan is frequently accompanied by a reduction in female fecundity, and it has been hypothesized that individuals may reallocate resources away from reproduction and into somatic maintenance. However, effects of dietary protein restriction on male reproduction are less consistent, suggesting that these effects may depend on other environmental parameters. Using the neriid fly, Telostylinus angusticollis, we examined age-specific effects of adult dietary protein restriction on male post-copulatory reproductive performance (fecundity and offspring viability). To explore the context dependence of these effects, we simultaneously manipulated male larval diet and adult mating history. We found that protein-restricted males sired less viable offspring at young ages, but offspring viability increased with paternal age and eventually exceeded that of fully fed males. The number of eggs laid by females was not affected by male dietary protein, whereas egg hatching success was subject to a complex interaction of male adult diet, age, larval diet and mating history. These findings suggest that effects of protein restriction on male reproduction are highly context dependent and cannot be explained by a simple reallocation of resources from reproduction to somatic maintenance. Rather, these effects appear to involve changes in the scheduling of male reproductive investment with age.

Effects of aging and dietary protein restriction on nitrogen balance and cardiovascular functions were examined. Male Fischer 344 rats 6-7 months old were fed ad libitum until 24 or 25 months old with either a 12% or a 23% protein diet, respectively. The nitrogen balance measured at the age of 24 months old demonstrated a significant difference between the two groups: the group with the 23% protein diet had a negative balance, while the group with the 12% protein diet had a positive balance. Endothelium-dependent relaxation with acetylcholine of the thoracic aortas was impaired by age but to a lesser extent to the protein-restricted animals. In addition, increased ability of platelet aggregation according to aging was also significantly suppressed by protein restriction. These observations demonstrate that protein restriction delays the aging effects of nitrogen loss and reduced cardiovascular function.

Dietary protein restriction has been demonstrated to improve metabolic health under various conditions. However, the relevance of ageing and age-related decline in metabolic flexibility on the effects of dietary protein restriction has not been addressed. Therefore, we investigated the effect of short-term dietary protein restriction on metabolic health in young and aged mice. Young adult (3months old) and aged (18months old) C57Bl/6J mice were subjected to a 3-month dietary protein restriction. Outcome parameters included fibroblast growth factor 21 (FGF21) levels, muscle strength, glucose tolerance, energy expenditure (EE) and transcriptomics of brown and white adipose tissue (WAT). Here, we report that a low-protein diet had beneficial effects in aged mice by reducing some aspects of age-related metabolic decline. These effects were characterized by increased plasma levels of FGF21, browning of subcutaneous WAT, increased body temperature and EE, while no changes were observed in glucose homeostasis and insulin sensitivity. Moreover, the low-protein diet used in this study was well-tolerated in aged mice indicated by the absence of adverse effects on body weight, locomotor activity and muscle performance. In conclusion, our study demonstrates that a short-term reduction in dietary protein intake can impact age-related metabolic health alongside increased FGF21 signalling, without negatively affecting muscle function. These findings highlight the potential of protein restriction as a strategy to induce EE and browning of WAT in aged individuals.

Effect of Dietary Protein Restriction or Food Restriction on Oxygen Consumption and Mitochondrial Distribution in Cardiac and Red and White Skeletal Muscle of Rats

BackgroundTo investigate the effects of dietary crude protein (CP) restriction on muscle fiber characteristics and key regulators related to protein deposition in skeletal muscle, a total of 18 growing-finishing pigs (62.30 ± 0.88 kg) were allotted to 3 groups and fed with the recommended adequate protein (AP, 16 % CP) diet, moderately restricted protein (MP, 13 % CP) diet and low protein (LP, 10 % CP) diet, respectively. The skeletal muscle of different locations in pigs, including longissimus dorsi muscle (LDM), psoas major muscle (PMM) and biceps femoris muscle (BFM) were collected and analyzed.ResultsResults showed that growing-finishing pigs fed the MP or AP diet improved (P < 0.01) the average daily gain and feed: gain ratio compared with those fed the LP diet, and the MP diet tended to increase (P = 0.09) the weight of LDM. Moreover, the ATP content and energy charge value were varied among muscle samples from different locations of pigs fed the reduced protein diets. We also observed that pigs fed the MP diet up-regulated (P < 0.05) muscular mRNA expression of all the selected key genes, except that myosin heavy chain (MyHC) IIb, MyHC IIx, while mRNA expression of ubiquitin ligases genes was not affected by dietary CP level. Additionally, the activation of mammalian target of rapamycin complex 1 (mTORC1) pathway was stimulated (P < 0.05) in skeletal muscle of the pigs fed the MP or AP diet compared with those fed the LP diet.ConclusionThe results suggest that the pigs fed the MP diet could catch up to the growth performance and the LDM weight of the pigs fed the AP diet, and the underlying mechanism may be partly due to the alteration in energy status, modulation of muscle fiber characteristics and mTORC1 activation as well as its downstream effectors in skeletal muscle of different locations in growing-finishing pigs.Electronic supplementary materialThe online version of this article (doi:10.1186/s40104-016-0106-8) contains supplementary material, which is available to authorized users.

Studies in fruit flies, Drosophila melanogaster, have observed considerable variation in the effect of dietary protein restriction (PR) on various fitness traits. In addition, not only are there inconsistent results relating lifespan to stress resistance, but also the long-term effects of PR are unexplored. We study PR implementation across generations (long term) hypothesizing that it will be beneficial for fitness traits, stress resistance and storage reserves due to nutritional plasticity transferred by parents to offspring in earlier Drosophila studies. By imposing two concentrations of PR diets (50% and 70% of control protein) from the pre-adult and adult (age 1 day) stages of the flies, we assessed the stage-specific and long-term effect of the imposed PR. All long-term PR flies showed increased resistance against the tested stressors (starvation, desiccation, H2O2-induced oxidative stress). In addition, we also found long-term PR-induced increased stress resistance across generations. The PR flies also possessed higher protein and triglyceride (TG) content, reduced glucose and unaffected glycogen levels. We also assayed the effect of returning the PR flies to control (AL) food for a single generation and assessed their biochemical parameters to witness the transient PR effect. It was seen that TG content upon reversal was similar to AL flies except for PRI70 males; however, the glucose levels of PR males increased, while they were consistently lower in females. Taken altogether, our study suggests that long-term PR implementation contributes to increased stress resistance and was found to influence storage reserves in D. melanogaster.

It has been reported that loss of ovarian oestrogen after menopause or by ovariectomy causes osteoporosis. In order to elucidate the effect of dietary protein restriction on bone metabolism after ovariectomy, we fed ovariectomized young female rats on a casein-based diet (50 g/kg diet (protein restriction) or 200 g/kg diet (control)) for 3 weeks and measured mRNA contents of bone-matrix proteins such as osteocalcin, osteopontin and alpha 1 type I collagen, insulin-like growth factors (IGF) and IGF-binding proteins (IGFBP) in femur. Ovariectomy decreased the weight of fat-free dry bone and increased urinary excretion of pyridinium cross-links significantly, although dietary protein restriction did not affect them. Neither ovariectomy nor protein restriction affected the content of mRNA of osteopontin and osteocalcin; however, ovariectomy increased and protein restriction extensively decreased the alpha 1 type I collagen mRNA content in bone tissues. Ovariectomy increased IGF-I mRNA only in the rats fed on the control diet. Conversely, protein restriction increased and ovariectomy decreased the IGF-II mRNA content in femur. Furthermore, the contents of IGFBP-2, IGFBP-4 and IGFBP-5 mRNA increased, but the content of IGFBP-3 mRNA decreased in femur of the rats fed on the protein-restricted diet. In particular, ovariectomy decreased the IGFBP-2 mRNA content in the protein-restricted rats and the IGFBP-6 mRNA content in the rats fed on the control diet. These results clearly show that the mRNA for some of the proteins which have been shown to be involved in bone formation are regulated by both quantity of dietary proteins and ovarian hormones.

In rats, maternal protein restriction reduces nephron endowment and often leads to adult hypertension. Sex differences in these responses have been identified. The molecular and genetic bases of these phenomena can best be identified in a mouse model, but effects of maternal protein restriction on kidney development have not been examined in mice. Therefore, we determined how combined prenatal and postnatal protein restriction in mice affects organ weight, glomerular number and dimensions, and renal expression of angiotensin receptor mRNA, in both male and female offspring. C57/BL6/129sv mice received either a normal (20% wt/wt; NP) or low (9% wt/wt; LP) protein diet during gestation and postnatal life. Offspring were examined at postnatal day 30. Protein restriction retarded growth of the kidney, liver, spleen, heart, and brain. All organs except the brain weighed less in female than male offspring. Protein restriction increased normalized (to body weight) brain weight, with females having relatively heavier brains than males. The effects of protein restriction were not sex dependent, except that normalized liver weight was reduced in males but increased in females. Glomerular volume, but not number, was greater in female than in male mice. Maternal protein restriction reduced nephron endowment similarly in male and female mice. Renal expression of AT(1A) receptor mRNA was approximately sixfold greater in female than male NP mice, but similar in male LP and female LP mice. We conclude that maternal protein restriction reduces nephron endowment in mice. This effect provides a basis for future studies of developmental programming in the mouse.

The transcription of several genes that are preferentially expressed in the liver, including the serum albumin, transthyretin and carbamyl phosphate synthetase-I genes, is specifically decreased in animals consuming inadequate amounts of dietary protein. The high level of transcription of these genes in the liver is directed in part by a number of liver-enriched transcription factors, including hepatocyte nuclear factors (HNF)-1, -3, and -4, and proteins of the CCAAT/enhancer-binding protein (C/EBP) family. In the present study, we investigated the possibility that the co-ordinate decrease in transcription of the nutritionally sensitive genes in protein-deprived rats results from altered activity of one or more of the liver-enriched transcription factors. For HNF-4, Western blots indicated no change in the level of nuclear HNF-4 protein in liver of protein-deprived animals, whereas we observed a 40% reduction in the DNA binding activity of HNF-4 as measured by electrophoretic mobility shift assay (EMSA). Furthermore, the binding affinity of HNF-4 for DNA was unaltered by dietary protein deprivation, while the number of HNF-4 molecules able to bind to DNA (Bmax) was reduced, as determined by Scatchard analysis. This indicates that in the protein-restricted rats a portion of the pool of HNF-4 protein is inactivated or otherwise prevented from binding to DNA. The overall DNA binding activity of C/EBP alpha and beta was increased in protein-restricted animals. This change occurred in the absence of a change in the amount of the full-length forms of these two proteins, quantified by Western blotting. Interestingly, dietary protein restriction specifically increased the level of a truncated form of C/EBP beta (liver-enriched transcriptional inhibitory protein, LIP), which is a protein dominant negative inhibitor of C/EBP function. Analysis of HNF-3 DNA-binding activity by EMSA revealed that HNF-3 alpha and beta DNA binding was increased and that HNF-3 gamma DNA-binding activity was unchanged in protein-restricted animals. We also detected two apparently novel shift complexes with the HNF-3 probe by EMSA, both of which were decreased in protein-restricted animals. HNF-1 DNA-binding activity was increased by dietary protein restriction. We also examined the effect of protein restriction on the DNA-binding activity of two ubiquitous transcription factors, NF1 and Sp1. The DNA binding activity of the major NF1 isoforms was unchanged whereas the binding activity of Sp1 was increased in the protein-restricted animals. In summary, restriction of dietary protein resulted in a number of specific changes in the DNA-binding activity of various transcription factors. Because transcriptional activation typically involves the synergistic action of more than one transcription factor, small changes in the amount/activity of several factors, could have a strong net effect on the transcription of many genes.

Beverages, diet, and prevention of kidney stones.

Protein Restriction Specifically Decreases the Abundance of Serum Albumin and Transthyretin Nuclear Transcripts in Rat Liver

Diabetic renal disease (diabetic nephropathy) is a leading cause of end-stage renal failure. Once the process has started, it cannot be reversed by glycaemic control, but progression might be slowed by control of blood pressure and protein restriction. To assess the effects of dietary protein restriction on the progression of diabetic nephropathy in patients with diabetes. We searched The Cochrane Library, MEDLINE, EMBASE, ISI Proceedings, Science Citation Index Expanded and bibliographies of included studies. Randomised controlled trials (RCTs) and before and after studies of the effects of a modified or restricted protein diet on diabetic renal function in people with type 1 or type 2 diabetes following diet for at least four months were considered. Two reviewers performed data extraction and evaluation of quality independently. Pooling of results was done by means of random-effects model. Twelve studies were included, nine RCTs and three before and after studies. Only one study explored all-cause mortality and end-stage renal disease (ESRD) as endpoints. The relative risk (RR) of ESRD or death was 0.23 (95% confidence interval (CI) 0.07 to 0.72) for patients assigned to a low protein diet (LPD). Pooling of the seven RCTs in patients with type 1 diabetes resulted in a non-significant reduction in the decline of glomerular filtration rate (GFR) of 0.1 ml/min/month (95% CI -0.1 to 0.3) in the LPD group. For type 2 diabetes, one trial showed a small insignificant improvement in the rate of decline of GFR in the protein-restricted group and a second found a similar decline in both the intervention and control groups. Actual protein intake in the intervention groups ranged from 0.7 to 1.1 g/kg/day. One study noted malnutrition in the LPD group. We found no data on the effects of LPDs on health-related quality of life and costs. The results show that reducing protein intake appears to slightly slow progression to renal failure but not statistically significantly so. However, questions concerning the level of protein intake and compliance remain. Further longer-term research on large representative groups of patients with both type 1 and type 2 diabetes mellitus is necessary. Because of the variability amongst patients, there might perhaps be a six month therapeutic trial of protein restriction in all individuals, with continuation only in those who responded best. Trials are required of different types of protein.

1. A low protein diet prevents the development of proteinuria and glomerular damage in adriamycin experimental nephrosis without affecting renal haemodynamics. In this study the hypothesis was tested as to whether protein restriction is able to modulate the purine metabolic cycle and related enzymes such as xanthine oxidase, one of the putative effectors of adriamycin nephrotoxicity. 2. Renal activities of xanthine oxidase and purine nucleoside phosphorylase were markedly depressed in adriamycin-treated rats fed a 9% casein (low protein) diet compared with the group fed a 22% casein (normal protein) diet both 1 day after adriamycin administration and at the time of appearance of heavy proteinuria (day 15), whereas the activity of renal adenosine deaminase was unchanged. 3. The concentrations of the metabolic substrates of xanthine oxidase, i.e. hypoxanthine and xanthine, were constantly lower in renal homogenates of rats fed a low protein diet compared with those on a normal protein diet. In urine, uric acid, the product of hypoxanthine-xanthine transformation, was lower 1 day after adriamycin injection in protein-restricted rats compared with the group on a normal protein diet which showed a marked increase in its excretion. At the same time, the urinary efflux of adenosine 5'-monophosphate, which is the precursor nucleotide of the above-mentioned nucleosides and bases, was very high in rats fed a low protein diet, whereas it was absent in the group on a normal protein diet. 4. The progressive increment in proteinuria of glomerular origin (i.e. increased excretion of albumin and transferrin) typical of adriamycin-treated rats fed a normal protein diet was inhibited in the protein-restricted animals, which were normoproteinuric on day 10 and were only slightly proteinuric on day 15. 5. Like protein restriction, the pharmacological suppression of renal xanthine oxidase by dietary tungstate and the scavenging by dimethylthiourea of the putative free radical deriving from the action of xanthine oxidase, were associated with a similar (quantitative and qualitative) inhibition of glomerular proteinuria. 6. These data demonstrate that dietary protein restriction is associated with a block in purine metabolism within the kidney due to a marked reduction in the activities of two main enzymes of the cycle, i.e. purine nucleoside phosphorylase and xanthine oxidase, the latter being a putative effector of adriamycin nephrotoxicity. The partial reduction of proteinuria induced by a low protein diet is quantitatively and qualitatively comparable with the reduction induced by the specific block of renal xanthine oxidase or by the scavenging of OH.deriving from hypoxanthine and xanthine transformation.(ABSTRACT TRUNCATED AT 400 WORDS)

Effects of dietary protein restriction and glucocorticoid administration on urea excretion in rats