A step toward precision gerontology: Lifespan effects of calorie and protein restriction are consistent with predicted impacts on entropy generation.

Understanding aging is a key biological goal. Precision gerontology aims to predict how long individuals will live under different treatment scenarios. Calorie and protein restriction (CR and PR) extend lifespan in many species. Using data from C57BL/6 male mice under graded CR or PR, we introduce a computational thermodynamic model for entropy generation, which predicted the impact of the manipulations on lifespan. Daily entropy generation decreased significantly with increasing CR level, but not PR. Our predictions indicated the lifespan of CR mice should increase by 13 to 56% with 10 to 40% CR, relative to ad libitum-fed animals. This prediction was broadly consistent with the empirical observation of the lifespan impacts of CR in rodents. Modeling entropy fluxes may be a future strategy to identify antiaging interventions.

Intestinal Amino Acid Transport and Metabolic Health.

Amino acids derived from protein digestion are important nutrients for the growth and maintenance of organisms. Approximately half of the 20 proteinogenic amino acids can be synthesized by mammalian organisms, while the other half are essential and must be acquired from the nutrition. Absorption of amino acids is mediated by a set of amino acid transporters together with transport of di- and tripeptides. They provide amino acids for systemic needs and for enterocyte metabolism. Absorption is largely complete at the end of the small intestine. The large intestine mediates the uptake of amino acids derived from bacterial metabolism and endogenous sources. Lack of amino acid transporters and peptide transporter delays the absorption of amino acids and changes sensing and usage of amino acids by the intestine. This can affect metabolic health through amino acid restriction, sensing of amino acids, and production of antimicrobial peptides.

Dietary protein restriction diminishes sucrose reward and reduces sucrose-evoked mesolimbic dopamine signaling in mice.

A growing literature suggests manipulating dietary protein status decreases sweet consumption in rodents and in humans. Underlying neurocircuit mechanisms have not yet been determined, but previous work points towards hedonic rather than homeostatic pathways. Here we hypothesized that a history of protein restriction reduces sucrose seeking by altering mesolimbic dopamine signaling in mice. We tested this hypothesis using established behavioral tests of palatability and conditioned reward, including the palatability contrast and conditioned place preference (CPP) tests. We used modern optical sensors for measuring real-time nucleus accumbens (NAc) dopamine dynamics during voluntary sucrose consumption, via fiber photometry, in male C57/Bl6J mice maintained on low-protein high-carbohydrate (LPHC) or control (CON) diet for ∼5 weeks. Our results showed that a history of protein restriction decreased the consumption of a sucrose 'dessert' in sated mice by ∼50% compared to controls [T-test, p < 0.05]. The dopamine release in NAc during sucrose consumption was reduced, also by ∼50%, in LPHC-fed mice compared to CON [T-test, p < 0.01]. Furthermore, LPHC-feeding blocked the sucrose-conditioned place preference we observed in CON-fed mice [paired T-test, p < 0.05], indicating reduced sucrose reward. This was accompanied by a 33% decrease in neuronal activation of the NAc core, as measured by c-Fos immunolabeling from brains collected directly after the CPP test [T-test, p < 0.05]. Together, these findings advance our mechanistic understanding of how dietary protein restriction decreases the consumption of sweets-by inhibiting the incentive salience of a sucrose reward, together with reduced sucrose-evoked dopamine release in NAc.

Intermittent protein restriction elevates food intake and plasma ghrelin in male mice.

Low-protein diets affect body weight, body composition, food intake, and food preferences in mice. Furthermore, single periods of protein restriction can have lasting effects on these parameters. We sought to examine the effect of multiple, short, bouts of protein restriction, relative to long-term maintenance on either a control (NR) or protein-restricted (PR) diet. We found that male mice experiencing intermittent protein restriction (IPR) were indistinguishable from NR mice in terms of body weight and composition, but had food intake and plasma ghrelin as high as mice on PR diet, even when they were returned to control diet. This was not found in female mice. The results of this experiment highlight the importance of diet history on food intake and ghrelin levels in male mice, and the difference in how PR diet might affect male and female mice.

Regulation of metabolic health by dietary histidine in mice.

Low-protein (LP) diets are associated with a decreased risk of diabetes in humans, and promote leanness and glycaemic control in both rodents and humans. While the effects of an LP diet on glycaemic control are mediated by reduced levels of the branched-chain amino acids, we have observed that reducing dietary levels of the other six essential amino acids leads to changes in body composition. Here, we find that dietary histidine plays a key role in the response to an LP diet in male C57BL/6J mice. Specifically reducing dietary levels of histidine by 67% reduces the weight gain of young, lean male mice, reducing both adipose and lean mass without altering glucose metabolism, and rapidly reverses diet-induced obesity and hepatic steatosis in diet-induced obese male mice, increasing insulin sensitivity. This normalization of metabolic health was associated not with caloric restriction or increased activity, but with increased energy expenditure. Surprisingly, the effects of histidine restriction do not require the energy balance hormone Fgf21. Histidine restriction that was started in midlife promoted leanness and glucose tolerance in aged males but not females, but did not affect frailty or lifespan in either sex. Finally, we demonstrate that variation in dietary histidine levels helps to explain body mass index differences in humans. Overall, our findings demonstrate that dietary histidine is a key regulator of weight and body composition in male mice and in humans, and suggest that reducing dietary histidine may be a translatable option for the treatment of obesity. KEY POINTS: Protein restriction (PR) promotes metabolic health in rodents and humans and extends rodent lifespan. Restriction of specific individual essential amino acids can recapitulate the benefits of PR. Reduced histidine promotes leanness and increased energy expenditure in male mice. Reduced histidine does not extend the lifespan of mice when begun in midlife. Dietary levels of histidine are positively associated with body mass index in humans.

Epigenetic regulation of functional candidate genes for milk production traits in dairy sheep subjected to protein restriction in the prepubertal stage.

BACKGROUND: As the prepubertal stage is a crucial point for the proper development of the mammary gland and milk production, this study aims to evaluate how protein restriction at this stage can affect methylation marks in milk somatic cells. Here, 28 Assaf ewes were subjected to 42.3% nutritional protein restriction (14 animals, NPR) or fed standard diets (14 animals, C) during the prepubertal stage. During the second lactation, the milk somatic cells of these ewes were sampled, and the extracted DNA was subjected to whole-genome bisulfite sequencing. RESULTS: A total of 1154 differentially methylated regions (DMRs) were identified between the NPR and C groups. Indeed, the results of functional enrichment analyses of the genes harboring these DMRs suggested their relevant effects on the development of the mammary gland and lipid metabolism in sheep. The additional analysis of the correlations of the mean methylation levels within these DMRs with fat, protein, and dry extract percentages in the milk and milk somatic cell counts suggested associations between several DMRs and milk production traits. However, there were no phenotypic differences in these traits between the NPR and C groups. CONCLUSION: In light of the above, the results obtained in the current study might suggest potential candidate genes for the regulation of milk production traits in the sheep mammary gland. Further studies focusing on elucidating the genetic mechanisms affected by the identified DMRs may help to better understand the biological mechanisms modified in the mammary gland of dairy sheep as a response to nutritional challenges and their potential effects on milk production.

Dietary Protein Modulates the Efficacy of Taurine Supplementation on Adaptive Islet Function and Morphology in Obesity.

Adaptation of islet ß-cell mass and function under limiting or excess nutrient availability is critical for maintenance of glucose homeostasis. Taurine regulates islet function of obese mice in normal and low dietary protein conditions, but whether this involves remodeling of the endocrine pancreas architecture is not well understood. Here, we carried functional and morphometric evaluation of the endocrine pancreas of normal and protein-restricted mice fed a high-fat diet (HFD) and investigated the role of taurine supplementation. Weaned mice were placed in a normal (C) or a low-protein diet (R) for 6 weeks, followed by HFD for 8 weeks (CH and RH). Half of HFD groups received 5% taurine supplementation since weaning (CHT and RHT) until the end of the experiment. Isolated islets from both CH and RH groups showed increased insulin release in association with increased pancreas weight and independently of changes in islet or ß-cell area. In normal protein CHT mice, taurine supplementation prevented obesity-induced insulin hypersecretion and promoted increased islet and ß-cell areas in association with increased protein expression of the proliferation marker, PCNA. On a low-protein background, taurine effects on islet function and morphology were blunted, but it prevented obesity-induced DNA fragmentation. In summary, taurine regulates islet function and morphology to improve the adaptive response to diet-induced obesity, but these effects are dependent on adequate dietary protein levels.

Transcriptome and Methylome Profiling in Rat Skeletal Muscle: Impact of Post-Weaning Protein Restriction.

Skeletal muscle is programmable, and early-life nutritional stimuli may form epigenetic memory in the skeletal muscle, thus impacting adult muscle function, aging, and longevity. In the present study, we designed a one-month protein restriction model using post-weaning rats, followed by a two-month rebound feeding, to investigate how early-life protein restriction affects overall body growth and muscle development and whether these influences could be corrected by rebound feeding. We observed comprehensive alterations immediately after protein restriction, including retarded growth, altered biochemical indices, and disturbed hormone secretion. Transcriptome profiling of the gastrocnemius muscle followed by gene ontology analyses revealed that "myogenic differentiation functions" were upregulated, while "protein catabolism" was downregulated as a compensatory mechanism, with enhanced endoplasmic reticulum stress and undesired apoptosis. Furthermore, methylome profiling of the gastrocnemius muscle showed that protein restriction altered the methylation of apoptotic and hormone secretion-related genes. Although most of the alterations were reversed after rebound feeding, 17 genes, most of which play roles during muscle development, remained altered at the transcriptional level. In summary, early-life protein restriction may undermine muscle function in the long term and affect skeletal muscle development at the both transcriptional and methylation levels, which may hazard future muscle health.

Postnatal Protein Intake as a Determinant of Skeletal Muscle Structure and Function in Mice-A Pilot Study.

Sarcopenia is characterised by an age-related decrease in the number of muscle fibres and additional weakening of the remaining fibres, resulting in a reduction in muscle mass and function. Many studies associate poor maternal nutrition during gestation and/or lactation with altered skeletal muscle homeostasis in the offspring and the development of sarcopenia. The aim of this study was to determine whether the musculoskeletal physiology in offspring born to mouse dams fed a low-protein diet during pregnancy was altered and whether any physiological changes could be modulated by the nutritional protein content in early postnatal stages. Thy1-YFP female mice were fed ad libitum on either a normal (20%) or a low-protein (5%) diet. Newborn pups were cross-fostered to different lactating dams (maintained on a 20% or 5% diet) to generate three groups analysed at weaning (21 days): Normal-to-Normal (NN), Normal-to-Low (NL) and Low-to-Normal (LN). Further offspring were maintained ad libitum on the same diet as during lactation until 12 weeks of age, creating another three groups (NNN, NLL, LNN). Mice on a low protein diet postnatally (NL, NLL) exhibited a significant reduction in body and muscle weight persisting up to 12 weeks, unlike mice on a low protein diet only prenatally (LN, LNN). Muscle fibre size was reduced in mice from the NL but not LN group, showing recovery at 12 weeks of age. Muscle force was reduced in NLL mice, concomitant with changes in the NMJ site and changes in atrophy-related and myosin genes. In addition, µCT scans of mouse tibiae at 12 weeks of age revealed changes in bone mass and morphology, resulting in a higher bone mass in the NLL group than the control NNN group. Finally, changes in the expression of miR-133 in the muscle of NLL mice suggest a regulatory role for this microRNA in muscle development in response to postnatal diet changes. Overall, this data shows that a low maternal protein diet and early postnatal life low-protein intake in mice can impact skeletal muscle physiology and function in early life while postnatal low protein diet favours bone integrity in adulthood.

A review of treatment modalities in gyrate atrophy of the choroid and retina (GACR).

Gyrate atrophy of the choroid and retina (GACR) is a rare inborn error of amino acid metabolism caused by bi-allelic variations in OAT. GACR is characterised by vision decline in early life eventually leading to complete blindness, and high plasma ornithine levels. There is no curative treatment for GACR, although several therapeutic modalities aim to slow progression of the disease by targeting different steps within the ornithine pathway. No international treatment protocol is available. We systematically collected all international literature on therapeutic interventions in GACR to provide an overview of published treatment effects. METHODS: Following the PRISMA guidelines, we conducted a systematic review of the English literature until December 22nd 2020. PubMed and Embase databases were searched for studies related to therapeutic interventions in patients with GACR. RESULTS: A total of 33 studies (n = 107 patients) met the inclusion criteria. Most studies were designed as case reports (n = 27) or case series (n = 4). No randomised controlled trials or large cohort studies were found. Treatments applied were protein-restricted diets, pyridoxine supplementation, creatine or creatine precursor supplementation, l-lysine supplementation, and proline supplementation. Protein-restricted diets lowered ornithine levels ranging from 16.0-91.2%. Pyridoxine responsiveness was reported in 30% of included mutations. Lysine supplementation decreased ornithine levels with 21-34%. Quality assessment showed low to moderate quality of the articles. CONCLUSIONS: Based primarily on case reports ornithine levels can be reduced by using a protein restricted diet, pyridoxine supplementation (variation-dependent) and/or lysine supplementation. The lack of pre-defined clinical outcome measures and structural follow-up in all included studies impeded conclusions on clinical effectiveness. Future research should be aimed at 1) Unravelling the OAT biochemical pathway to identify other possible pathologic metabolites besides ornithine, 2) Pre-defining GACR specific clinical outcome measures, and 3) Establishing an international historical cohort.

REVIEW: Practical strategies to maintain anabolism by intravenous nutritional management in children with inborn metabolic diseases.

One of the most vital elements of management for patients with inborn errors of intermediary metabolism is the promotion of anabolism, the state in which the body builds new components, and avoidance of catabolism, the state in which the body breaks down its own stores for energy. Anabolism is maintained through the provision of a sufficient supply of substrates for energy, as well as critical building blocks of essential amino acids, essential fatty acids, and vitamins for synthetic function and growth. Patients with metabolic diseases are at risk for decompensation during prolonged fasting, which often occurs during illnesses in which enteral intake is compromised. During these times, intravenous nutrition must be supplied to fully meet the specific nutritional needs of the patient. We detail our approach to intravenous management for metabolic patients and its underlying rationale. This generally entails a combination of intravenous glucose and lipid as well as early introduction of protein and essential vitamins. We exemplify the utility of our approach in case studies, as well as scenarios and specific disorders which require a more careful administration of nutritional substrates or a modification of macronutrient ratios.

Protein restriction during puberty alters nutritional parameters and affects ovarian and uterine histomorphometry in adulthood in rats.

In a large part of the population inefficient ingestion of proteins, whether for cultural, aesthetic or economic reasons, is a global concern. Low-protein diets can cause severe functional complications, mainly during the development and maturation of organs and systems, including the female reproductive system. The present study investigated the effect of nutritional protein restriction during puberty on the oestrous cycle and expression of sex steroid receptors (AR, ERα e ERß) in ovarian and uterine tissues of adult rats. Protein restriction promoted lower body weight gain, feed efficiency and higher caloric intake. There was an increase in the oestrus phase arrest without changing the total length of the oestrous cycle. The consumption of low-protein diet also reduced the thickness of the uterine endometrium (uterine epithelium and endometrial stroma) in addition to increasing the number of primary and atretic follicles in the ovaries. Furthermore, the low-protein diet reduced the levels of androgen receptor (AR) and increased the oestrogen receptor ß (ERß) in the ovary, while no significant changes were observed in the uterus. Our study reinforces the importance of adequate protein intake during puberty, since physiological changes in this developmental period interfere with the histomorphometry of the ovaries and uteri, possibly resulting in impaired folliculogenesis and fertility in the reproductive period.

Low protein intake during reproduction compromises the recovery of lactation-induced bone loss in female mouse dams without affecting skeletal muscles.

Lactation-induced bone loss occurs due to high calcium requirements for fetal growth but skeletal recovery is normally achieved promptly postweaning. Dietary protein is vital for fetus and mother but the effects of protein undernutrition on the maternal skeleton and skeletal muscles are largely unknown. We used mouse dams fed with normal (N, 20%) or low (L, 8%) protein diet during gestation and lactation and maintained on the same diets (NN, LL) or switched from low to normal (LN) during a 28 d skeletal restoration period post lactation. Skeletal muscle morphology and neuromuscular junction integrity was not different between any of the groups. However, dams fed the low protein diet showed extensive bone loss by the end of lactation, followed by full skeletal recovery in NN dams, partial recovery in LN and poor bone recovery in LL dams. Primary osteoblasts from low protein diet fed mice showed decreased in vitro bone formation and decreased osteogenic marker gene expression; promoter methylation analysis by pyrosequencing showed no differences in Bmpr1a, Ptch1, Sirt1, Osx, and Igf1r osteoregulators, while miR-26a, -34a, and -125b expression was found altered in low protein fed mice. Therefore, normal protein diet is indispensable for maternal musculoskeletal health during the reproductive period.

Protein restriction and cancer.

Protein restriction without malnutrition is currently an effective nutritional intervention known to prevent diseases and promote health span from yeast to human. Recently, low protein diets are reported to be associated with lowered cancer incidence and mortality risk of cancers in human. In murine models, protein restriction inhibits tumor growth via mTOR signaling pathway. IGF-1, amino acid metabolic programing, FGF21, and autophagy may also serve as potential mechanisms of protein restriction mediated cancer prevention. Together, dietary intervention aimed at reducing protein intake can be beneficial and has the potential to be widely adopted and effective in preventing and treating cancers.

Effects of pre- and postnatal protein restriction on maternal and offspring metabolism in the nonhuman primate.

Women in low- and middle-income countries frequently consume a protein-deficient diet during pregnancy and breastfeeding. The effects of gestational malnutrition on fetal and early postnatal development can have lasting adverse effects on offspring metabolism. Expanding on previous studies in rodent models, we utilized a nonhuman primate model of gestational and early-life protein restriction (PR) to evaluate effects on the organ development and glucose metabolism of juvenile offspring. Offspring were born to dams that had consumed a control diet containing 26% protein or a PR diet containing 13% protein. Offspring were maintained on the PR diet and studied [body and serum measurements, intravenous glucose tolerance tests (ivGTTs), and dual-energy X-ray absorptiometry scans] up to 7 mo of age, at which time tissues were collected for analysis. PR offspring had age-appropriate body weight and were euglycemic but exhibited elevated fasting insulin and reduced initial, but increased total, insulin secretion during an ivGTT at 6 mo of age. No changes were detected in pancreatic islets of PR juveniles; however, PR did induce changes, including reduced kidney size, and changes in liver, adipose tissue, and muscle gene expression in other peripheral organs. Serum osteocalcin was elevated and bone mineral content and density were reduced in PR juveniles, indicating a significant impact of PR on early postnatal bone development.

Dietary protein insufficiency: an important consideration in fatty liver disease?

Dietary protein insufficiency has been linked to excessive TAG storage and non-alcoholic fatty liver disease (NAFLD) in developing countries. Hepatic TAG accumulation following a low-protein diet may be due to altered peroxisomal, mitochondrial and gut microbiota function. Hepatic peroxisomes and mitochondria normally mediate metabolism of nutrients to provide energy and substrates for lipogenesis. Peroxisome biogenesis and activities can be modulated by odd-chain fatty acids (OCFA) and SCFA that are derived from gut bacteria, for example, propionate and butyrate. Also produced during amino acid metabolism by peroxisomes and mitochondria, propionate and butyrate concentrations correlate inversely with risk of obesity, insulin resistance and NAFLD. In this horizon-scanning review, we have compiled available evidence on the effects of protein malnutrition on OCFA production, arising from loss in mitochondrial, peroxisomal and gut microbiota function, and its association with lipid accumulation in the liver. The methyl donor amino acid composition of dietary protein is an important contributor to liver function and lipid storage; the presence and abundance of dietary branched-chain amino acids can modulate the composition and metabolic activity of the gut microbiome and, on the other hand, can affect protective OCFA and SCFA production in the liver. In preclinical animal models fed with low-protein diets, specific amino acid supplementation can ameliorate fatty liver disease. The association between low dietary protein intake and fatty liver disease is underexplored and merits further investigation, particularly in vulnerable groups with dietary protein restriction in developing countries.

Early protein restriction increases intra-islet GLP-1 production and pancreatic ß-cell proliferation mediated by the ß-catenin pathway.

PURPOSE: In the present study, we investigated whether intra-islet GLP-1 production and its modulation have a role in apoptosis, proliferation or neogenesis that is compromised by protein restriction during the foetal and suckling periods. METHODS: Exendin-4, a GLP-1 receptor agonist (treated groups), or saline (non-treated groups) was intraperitoneally administered for 15 days from 75 to 90 days of age in female adult rats consisting of offspring born to and suckled by mothers fed a control diet (control groups) and who had the same diet until 90 days of age or offspring born to and suckled by mothers fed a low-protein diet and who were fed the control diet after weaning until 90 days of age (protein-restricted group). RESULTS: The ß-cell mass was lower in the protein-restricted groups than in the control groups. Exendin-4 increased ß-cell mass, regardless of the mother's protein intake. The colocalization of GLP-1/glucagon was higher in the protein-restricted rats than in control rats in both the exendin-4-treated and non-treated groups. The frequency of cleaved caspase-3-labelled cells was higher in the non-treated protein-restricted group than in the non-treated control group and was similar in the treated protein-restricted and treated control groups. Regardless of treatment with exendin-4, Ki67-labelled cell frequency and ß-catenin/DAPI colocalization were elevated in the protein-restricted groups. Exendin-4 increased the area of endocrine cell clusters and ß-catenin/DAPI and FoxO1/DAPI colocalization regardless of the mother's protein intake. CONCLUSIONS: Protein restriction in early life increased intra-islet GLP-1 production and ß-cell proliferation, possibly mediated by the ß-catenin pathway.

Significant changes in caecal microbial composition and metabolites of weaned piglets after protein restriction and succedent realimentation.

The present study aimed to investigate the effects of protein restriction and subsequent realimentation on caecal barrier function, caecal microbial composition and metabolites in weaned piglets. Thirty-six 28-day-old weaned piglets were randomly assigned to a control group and a treatment group. The piglets were fed diets containing 18.83% (normal) or 13.05% (low) of crude protein from the 1st to 14th day, after which all piglets were fed diets containing 18.83% of crude protein from the 15th to 28th day. The results showed that protein restriction increased caecal bacterial diversity and richness as well as the abundance of Ruminococcus 2, Faecalibacterium and Lachnospiraceae_uncultured, but reduced the abundance and the gene copies of Lactobacillus in the treatment group compared with the control group on day 14. Protein restriction also decreased the concentrations of isovaleric acid and total branched-chain fatty acids. During the succedent protein realimentation stage, the abundance of Ruminococcaceae UCG-014 and the concentrations of lactic acid, acetic acid, butyric acid and total short-chain fatty acids were increased in the treatment group on day 28. Furthermore, the ammonia concentration was reduced, while the gene mRNA levels of caecal barrier function were increased in the treatment group both on days 14 and 28. In conclusion, dietary protein restriction and realimentation could change caecal microbial composition and metabolites, and eventually influence caecal barrier function. The present study may provide a new insight into protein restriction and realimentation in weaned piglets.

Protein malnutrition mitigates the effects of a high-fat diet on glucose homeostasis in mice.

Nutrient malnutrition, during the early stages of development, may facilitate the onset of metabolic diseases later in life. However, the consequences of nutritional insults, such as a high-fat diet (HFD) after protein restriction, are still controversial. We assessed overall glucose homeostasis and molecular markers of mitochondrial function in the gastrocnemius muscle of protein-restricted mice fed an HFD until early adulthood. Male C57BL/6 mice were fed a control (14% protein-control diet) or a protein-restricted (6% protein-restricted diet) diet for 6 weeks. Afterward, mice received an HFD or not for 8 weeks (mice fed a control diet and HFD [CH] and mice fed a protein-restricted diet and HFD [RH]). RH mice showed lower weight gain and fat accumulation and did not show an increase in fasting plasma glucose and insulin levels compared with CH mice. RH mice showed higher energy expenditure, increased citrate synthase, peroxisome-proliferator-activated receptor gamma coactivator 1-alpha protein content, and higher levels of malate and α-ketoglutarate compared with CH mice. Moreover, RH mice showed increased AMPc-dependent kinase and acetyl coenzyme-A (CoA) carboxylase phosphorylation, lower intramuscular triacylglycerol content, and similar malonyl-CoA levels. In conclusion, protein undernourishment after weaning does not potentiate fat accumulation and insulin resistance in adult young mice fed an HFD. This outcome seems to be associated with increased skeletal muscle mitochondrial oxidative capacity and reduced lipids accumulation.

Low-protein and methionine, high-starch diets increase energy intake and expenditure, increase FGF21, decrease IGF-1, and have little effect on adiposity in mice.

Low-protein diets most often induce increased energy intake in an attempt to increase protein intake to meet protein needs with a risk of accumulation as fat of the excess energy intake. In female adult BALB/c mice, a decrease in dietary casein from 20% to 6% and 3% increased energy intake and slightly increased adiposity, and this response was exacerbated with soy proteins with low methionine content. The effect on fat mass was however limited because total energy expenditure increased to the same extent as energy intake. Lean body mass was preserved in all 6% fed mice and reduced only in 3% casein-fed animals. Insulin response to an oral glucose tolerance test was reduced in soy-fed mice and in low-protein-fed mice. Low-protein diets did not affect uncoupling protein 1 and increased fibroblast growth factor 21 (FGF21) in brown adipose tissue and increased FGF21, fatty acid synthase, and cluster of differentiation 36 in the liver. In the hypothalamus, neuropeptide Y was increased and proopiomelanocortin was decreased only in 3% casein-fed mice. In plasma, when protein was decreased, insulin-like growth factor-1 decreased and FGF21 increased and plasma FGF21 was best described by using a combination of dietary protein level, protein-to-carbohydrate ratio, and protein-to-methionine ratio in the diet. In conclusion, reducing dietary protein and protein quality increases energy intake but also energy expenditure resulting in an only slight increase in adiposity. In this process, FGF21 is probably an important signal that responds to a complex combination of protein restriction, protein quality, and carbohydrate content of the diet.