Leucine alleviates cytokine storm syndrome by regulating macrophage polarization via the mTORC1/LXRα signaling pathway.

Cytokine storms are associated with severe pathological damage and death in some diseases. Excessive activation of M1 macrophages and the subsequent secretion of pro-inflammatory cytokines are a major cause of cytokine storms. Therefore, promoting the polarization of M2 macrophages to restore immune balance is a promising therapeutic strategy for treating cytokine storm syndrome (CSS). This study was aimed at investigating the potential protective effects of leucine on lipopolysaccharide (LPS)-induced CSS in mice and exploring the underlying mechanisms. CSS was induced by LPS administration in mice, which were concurrently administered leucine orally. In vitro, bone marrow derived macrophages (BMDMs) were polarized to M1 and M2 phenotypes with LPS and interleukin-4 (IL-4), respectively, and treated with leucine. Leucine decreased mortality in mice treated with lethal doses of LPS. Specifically, leucine decreased M1 polarization and promoted M2 polarization, thus diminishing pro-inflammatory cytokine levels and ameliorating CSS in mice. Further studies revealed that leucine-induced macrophage polarization through the mechanistic target of rapamycin complex 1 (mTORC1)/liver X receptor α (LXRα) pathway, which synergistically enhanced the expression of the IL-4-induced M2 marker Arg1 and subsequent M2 polarization. In summary, this study revealed that leucine ameliorates CSS in LPS mice by promoting M2 polarization through the mTORC1/LXRα/Arg1 signaling pathway. Our findings indicate that a fundamental link between metabolism and immunity contributes to the resolution of inflammation and the repair of damaged tissues.

Engineering Enhanced Antimicrobial Properties in α-Conotoxin RgIA through D-Type Amino Acid Substitution and Incorporation of Lysine and Leucine Residues.

Antimicrobial peptides (AMPs), acknowledged as host defense peptides, constitute a category of predominant cationic peptides prevalent in diverse life forms. This study explored the antibacterial activity of α-conotoxin RgIA, and to enhance its stability and efficacy, D-amino acid substitution was employed, resulting in the synthesis of nine RgIA mutant analogs. Results revealed that several modified RgIA mutants displayed inhibitory efficacy against various pathogenic bacteria and fungi, including Candida tropicalis and Escherichia coli. Mechanistic investigations elucidated that these polypeptides achieved antibacterial effects through the disruption of bacterial cell membranes. The study further assessed the designed peptides' hemolytic activity, cytotoxicity, and safety. Mutants with antibacterial activity exhibited lower hemolytic activity and cytotoxicity, with Pep 8 demonstrating favorable safety in mice. RgIA mutants incorporating D-amino acids exhibited notable stability and adaptability, sustaining antibacterial properties across diverse environmental conditions. This research underscores the potential of the peptide to advance innovative oral antibiotics, offering a novel approach to address bacterial infections.

Treatment with bestatin (the exogenous synthetic inhibitor of metalloproteinases) reduces the activity of metalloproteinase 2 and 12 in the spleen and lung tissues of rats in a model of lipopolysaccharide-induced sepsis.

Sepsis is caused by an inadequate or dysregulated host response to infection. Enzymes causing cellular degradation are matrix metalloproteinases (MMPs). Lipopolysaccharide (LPS) is used in models of sepsis in laboratory settings The aim of the study was to measure MMP 2 and 12 concentrations in spleen and lungs in rats in which septic shock was induced by LPS. The experiment was carried out on 40 male Wistar rats (5 groups of 8): 0. controls 1. administered LPS 2. administered bestatin 3. LPS and bestatin 4.bestatin and after 6 hours LPS Animals were decapitated. Lungs and spleens were collected. Concentrations of MMP-2 and MMP-12 were determined using immunoenzymatic methods. Mean (±SD) MMP-2 in the controls was 43.57 ± 20.53 ng/ml in the lungs and 1.7 ± 0.72 ng/ml in the spleen; Group 1: 31.28 ± 13.13 ng/ml, 0.83 ± 0.8 ng/ml; Group 2: 44.24 ± 22.75 ng /ml, 1.01 ± 0.32 ng/ml; Group 3: 35.94 ± 15.13 ng/ml, 0.41 ± 0.03 ng/ml; Group 4:79.42 ± 44.70 ng/ml, 0.45 ± 0.15, respectively. Mean MMP-12 in controls was 19.79 ± 10.01 ng/ml in lungs and 41.13 ± 15.99 ng/ml in the spleen; Group 1:27.97 ± 15.1 ng/ml; 40.44 ± 11.2 ng/ml; Group 2: 37.93 ± 25.38 ng/ml 41.05 ± 18.08 ng/ml; Group 3: 40.59 ± 11.46 ng/ml, 35.16 ± 12.89 ng/ml; Group 4: 39.4 ± 17.83 ng/ml, 42.04 ± 12.35 ng/ml, respectively. CONCLUSIONS: 1. Bestatin reduces MMP 2 and 12 levels in spleen and lungs. 2. Treatment with bestatin minimizes the effect of LPS.

Pre-heating stress associated with acute oral leucine supplementation effects in rat gastrocnemius muscle: Implications for protein synthesis signaling pathways.

Skeletal muscle is a highly plastic tissue. The role of heat shock protein 72 (Hsp72) in heat stress-induced skeletal muscle hypertrophy has been well demonstrated; however, the precise mechanisms remain unclear. Essential amino acids, such as leucine, mainly mediate muscle protein synthesis. We investigated the effects of pre-heating and increased Hsp72 expression on the mechanistic target of rapamycin (mTOR) signaling and protein synthesis following leucine administration in rat gastrocnemius muscle. To ensure increased Hsp72 expression in both the red and white portions of the muscle, one leg of male Wistar rats (10-week-old, n = 23) was heat-stressed in 43 °C water for 30 min twice at a 48-h-interval (heat-stressed leg, HS leg). The contralateral leg served as a non-heated internal control (CT leg). After the recovery period (48 h), rats were divided into the pre-administration or oral leucine administration groups. We harvested the gastrocnemius muscle (red and white parts) prior to administration and 30 and 90 min after leucine treatment (n = 7-8 per group) and intramuscular signaling responses to leucine ingestion were determined using western blotting. Heat stress significantly upregulated the expression of Hsp72 and was not altered by leucine administration. Although the phosphorylation levels of mTOR/S6K1 and ERK were similar regardless of heating, 4E-BP1 was less phosphorylated in the HS legs than the CT legs after leucine administration in the red portion of the muscles (P < 0.05). Moreover, c-Myc expression differed significantly after leucine administration in both the red and white portions of the muscles. Our findings indicate that following oral leucine administration, pre-heating partially blunted the muscle protein synthesis signaling response in the rat gastrocnemius muscle.

Dietary tryptophan alleviates intestinal inflammation caused by long photoperiod via gut microbiota derived tryptophan metabolites-NLRP3 pathway in broiler chickens.

Light pollution is a potential risk factor for intestinal health. Tryptophan plays an important role in the inhibition of intestinal inflammation. However, the mechanism of tryptophan in alleviating intestinal inflammation caused by long photoperiod is still unclear. This study investigated the anti-inflammatory effect of dietary tryptophan on intestinal inflammatory damage induced by long photoperiod and its potential mechanism in broiler chickens. We found that dietary tryptophan mitigated long photoperiod-induced intestinal tissue inflammatory damage and inhibited the activation of Nucleotide-Binding Oligomerization Domain, Leucine-Rich Repeat and Pyrin Domain-Containing 3 inflammasome. Moreover, dietary tryptophan significantly increased the relative abundance of Faecalibacterium, Enterococcus, and Lachnospiraceae_NC2004_group were significantly decreased the relative abundance of Ruminococcus_torques_group and norank_f_UCG-010 under the condition of long photoperiod (P < 0.05). The results of tryptophan targeted metabolomics show that tryptophan significantly increased indole-3-acetic acid (IAA) and indole-3 lactic acid (ILA), and significantly decreased xanthurenic acid (XA) under long photoperiod (P < 0.05). In conclusion, the results indicated that dietary tryptophan alleviates intestinal inflammatory damage caused by long photoperiod via the inhibition of Nucleotide-Binding Oligomerization Domain, Leucine-Rich Repeat and Pyrin Domain-Containing 3 inflammasome activation, which was mediated by tryptophan metabolites. Therefore, tryptophan supplementation could be a promising way to protect the intestine health under the condition of long photoperiod.

Identification of a leucine-mediated threshold effect governing macrophage mTOR signalling and cardiovascular risk.

High protein intake is common in western societies and is often promoted as part of a healthy lifestyle; however, amino-acid-mediated mammalian target of rapamycin (mTOR) signalling in macrophages has been implicated in the pathogenesis of ischaemic cardiovascular disease. In a series of clinical studies on male and female participants ( NCT03946774 and NCT03994367 ) that involved graded amounts of protein ingestion together with detailed plasma amino acid analysis and human monocyte/macrophage experiments, we identify leucine as the key activator of mTOR signalling in macrophages. We describe a threshold effect of high protein intake and circulating leucine on monocytes/macrophages wherein only protein in excess of ∼25 g per meal induces mTOR activation and functional effects. By designing specific diets modified in protein and leucine content representative of the intake in the general population, we confirm this threshold effect in mouse models and find ingestion of protein in excess of ∼22% of dietary energy requirements drives atherosclerosis in male mice. These data demonstrate a mechanistic basis for the adverse impact of excessive dietary protein on cardiovascular risk.

Cardioprotective Effects of Leucine Supplementation against Doxorubicin-Induced Cardiotoxicit.

Doxorubicin is one of the most important antitumor drugs used in oncology; however, its cardiotoxic effect limits the therapeutic use and raises concerns regarding patient prognosis. Leucine is a branched-chain amino acid used in dietary supplementation and has been studied to attenuate the toxic effects of doxorubicin in animals, which increases oxidative stress. Oxidative stress in different organs can be estimated using several methods, including catalase expression analysis. This study aimed to analyze the effect of leucine on catalase levels in rat hearts after doxorubicin administration. Adult male Wistar rats were separated into two groups: Standard diet (SD) and 5% Leucine-Enriched Diet (LED). The animals had free access to diet from D0 to D28. At D14, the groups were subdivided in animals injected with Doxorubicin and animals injected with vehicle, until D28, and the groups were SD, SD + Dox, LED and LED + Dox. At D28, the animals were submitted do Transthoracic Echocardiography and euthanized. Despite Dox groups had impaired body weight gain, raw heart weight was not different between the groups. No substantial alterations were observed in macroscopic evaluation of the heart. Although, Doxorubicin treatment increased total interstitial collagen in the heart, which in addition to Type I collagen, is lower in LED groups. Western blot analysis showed that catalase expression in the heart of LED groups was lower than that in SD groups. In conclusion, leucine supplementation reduced both the precocious Dox-induced cardiac remodeling and catalase levels in the heart.

Leucine improves the growth performance, carcass traits, and lipid nutritional quality of pork in Shaziling pigs.

Leucine is involved in promoting fatty acid oxidation and lipolysis, mediating lipid metabolism and energy homeostasis, thus it has been widely used in livestock production. However, the effects of leucine on fat deposition and nutrition in Shaziling pigs remain unclear. A total of 72 Shaziling pigs (150 days old, weight 35.00 ± 1.00 kg) were randomly divided into 2 groups and fed with basal diet (control group) or basal diet containing 1% leucine (leucine group) for 60 days. The results showed that leucine significantly increased the average daily feed intake but decreased the ratio of feed to gain (P < 0.05), increased the loin muscle area and serum glucose content (P < 0.05) of Shaziling pigs. Besides, leucine regulated the re-distribution of fatty acids from adipose tissue to muscle as it significantly increased the contents of C18:1n-9 and C22:6n-3 (DHA) in the longissimus thoracis while decreased the contents of C22:5n-3 (DPA), C20:5n-3 (EPA), and DHA in the adipose tissue of Shaziling pigs (P < 0.05). Lipidomic analysis showed that the contents of phosphatidylethanolamines (PEs), cardiolipins (CLs), and phosphatidylglycerols (PGs) in the longissimus thoracis and the contents of lysophosphatidylethanolamines (LPEs), ceramides (Cers), phosphatidylinositols (PIs) in adipose tissue of Shaziling pigs were decreased in leucine group (P < 0.05). Collectively, this study clarified that dietary addition of 1% leucine have a better effect on growth performance and the deposition of beneficial fatty acids in the muscle of Shaziling pigs, which is conductive to the production of high quality and healthy pork. In addition, leucine altered the lipid composition of muscle and fat in Shaziling pigs. The related results provide a theoretical basis and application guidance for regulating fat deposition in Shaziling pigs, which is important for the healthy breeding of Shaziling pigs.

Developing a comprehensive solution aimed to disrupt LARS1/RagD protein-protein interaction.

Aminoacyl-tRNA synthetases are crucial enzymes involved in protein synthesis and various cellular physiological reactions. Aside from their standard role in linking amino acids to the corresponding tRNAs, they also impact protein homeostasis by controlling the level of soluble amino acids within the cell. For instance, leucyl-tRNA synthetase (LARS1) acts as a leucine sensor for the mammalian target of rapamycin complex 1 (mTORC1), and may also function as a probable GTPase-activating protein (GAP) for the RagD subunit of the heteromeric activator of mTORC1. In turn, mTORC1 regulates cellular processes, such as protein synthesis, autophagy, and cell growth, and is implicated in various human diseases including cancer, obesity, diabetes, and neurodegeneration. Hence, inhibitors of mTORC1 or a deregulated mTORC1 pathway may offer potential cancer therapies. In this study, we investigated the structural requirements for preventing the sensing and signal transmission from LARS to mTORC1. Building upon recent studies on mTORC1 regulation activation by leucine, we lay the foundation for the development of chemotherapeutic agents against mTORC1 that can overcome resistance to rapamycin. Using a combination of in-silico approaches to develop and validate an alternative interaction model, discussing its benefits and advancements. Finally, we identified a set of compounds ready for testing to prevent LARS1/RagD protein-protein interactions. We establish a basis for creating chemotherapeutic drugs targeting mTORC1, which can conquer resistance to rapamycin. We utilize in-silico methods to generate and confirm an alternative interaction model, outlining its advantages and improvements, and pinpoint a group of novel substances that can prevent LARS1/RagD interactions.Communicated by Ramaswamy H. Sarma.

Effects of L-Leucine Supplementation and Resistance Training on Adipokine Markers in Untrained Perimenopausal and Postmenopausal Women.

ABSTRACT: Chapman-Lopez, TJ, Funderburk, LK, Heileson, JL, Wilburn, DT, Koutakis, P, Gallucci, AR, and Forsse, JS. Effects of L-leucine supplementation and resistance training on adipokine markers in untrained perimenopausal and postmenopausal women. J Strength Cond Res 38(3): 526-532, 2024-This study examined the effects of supplementing 5 g of leucine compared with a placebo during a 10-week resistance training program on body composition parameters and adipokine concentrations in untrained, perimenopausal and postmenopausal women. Thirty-five women were randomly assigned to 2 groups-leucine (LEU, n = 17) and placebo (PLC, n = 18)-in a double-blind, placebo-controlled trial. Each group consumed the supplement or placebo every day and completed a resistance training program for 10 weeks. Using 3-day food records, a diet was assessed before the intervention and after its cessation. Body composition was assessed preintervention and postintervention using dual-energy x-ray absorptiometry. Moreover, the concentrations of adipokines, such as adiponectin, visfatin, leptin, and monocyte chemoattractant protein-1 (MCP-1), were assessed preintervention and postintervention. Both groups showed an increase in visceral adipose tissue (VAT) area ( p = 0.030) and fat-free mass (FFM; p = 0.023). There were significant group differences in concentrations of visfatin ( p = 0.020) and leptin ( p = 0.038) between the PLC and LEU groups. Visfatin displayed higher concentrations in the PLC group and leptin displayed higher concentrations in the LEU group. In addition, there were significant decreases in adiponectin concentrations for both groups (LEU: 652 ± 513 to 292 ± 447 pg·ml -1 ; PLC: 584 ± 572 to 245 ± 356 pg·ml -1 , p = 0.002) and MCP-1 only decreased in the PLC group (253 ± 119 to 206 ± 106 pg·ml -1 , p = 0.004). There were significant decreases in adiponectin concentrations in both groups and a decrease in MCP-1 concentrations in the PLC group. These decreases may be due to both adipokines possible relationship with VAT area. However, it is not known whether leucine has underlying properties that hinder changes in MCP-1 concentrations.

Multi-Omics Analysis Reveals Sphingomyelin Accumulation, Glycerolipids Loss, and Disorders of Lipid Metabolism Regulated by Leucine Deprivation in the Liver of Mice.

SCOPE: Branched-chain amino acids, especially leucine, have been reported to play a role in regulating lipid metabolism. This study aims to examine the effects of leucine deprivation on hepatic lipid metabolism. METHODS AND RESULTS: C57BL/6 mice are fed with a chow diet (control group, n = 8) or a leucine-free diet (-Leu group, n = 8) for 7 days. Histology, lipidomics, targeted metabolomics, and transcriptomics are performed to analyze the liver tissue. Compared to control group, -Leu group exhibits a notably reduced liver weight, accompanied by hepatic injury, and disorders of lipid metabolism. The level of sphingomyelin (SM) is significantly increased in the liver of -Leu group, while the glycerolipids (GL) level is significantly decreased. The expression of sphingomyelin synthase 1 (SGMS1) is upregulated by leucine deprivation in a time-dependent manner, leading to hepatic SM accumulation. Moreover, leucine deprivation results in hepatic GL loss via suppressing fatty acid synthase (FASN) and acetyl-CoA carboxylase 1 (ACC1) expression. CONCLUSION: The findings demonstrate that leucine deprivation results in abnormal lipid metabolism in the liver, mainly manifested as SM accumulation and GL loss. These results provide insights into the role of leucine in regulating lipid metabolism.

Valine and nonessential amino acids affect bidirectional transport rates of leucine and isoleucine in bovine mammary epithelial cells.

A more complete understanding of the mechanisms controlling AA transport in mammary glands of dairy cattle will help identify solutions to increase nitrogen feeding efficiency on farms. It was hypothesized that Ala, Gln, and Gly (NEAAG), which are actively transported into cells and exchanged for all branched-chain AA (BCAA), may stimulate transport of BCAA, and that Val may antagonize transport of the other BCAA due to transporter competition. Thus, we evaluated the effects of varying concentrations of NEAAG and Val on transport and metabolism of the BCAA Ala, Met, Phe, and Thr by bovine mammary epithelial cells. Primary cultures of bovine mammary epithelial cells were assigned to treatments of low (70% of mean in vivo plasma concentrations of lactating dairy cows) and high (200%) concentrations of Val and NEAAG (LVal and LNEAAG, HVal and HNEAAG, respectively) in a 2 × 2 factorial design. Cells were preloaded with treatment media containing [15N]-labeled AA for 24 h. The [15N]-labeled media were replaced with treatment media containing [13C]-labeled AA. Media and cells were harvested from plates at 0, 0.5, 1, 5, 15, 30, 60, and 240 min after application of the [13C]-labeled AA and assessed for [15N]- and [13C]-AA label concentrations. The data were used to derive transport, transamination, irreversible loss, and protein-synthesis fluxes. All Val fluxes, except synthesis of rapidly exchanging tissue protein, increased with the HVal treatment. Interestingly, the rapidly exchanging tissue protein, transamination, and irreversible-loss rate constants decreased with HVal, indicating that the significant flux increases were primarily driven by mass action with the cells resisting the flux increases by downregulating activity. However, the decreases could also reflect saturation of processes that would drive down the mass-action rate constants. This is supported by decreases in the same rate constants for Ile and Leu with HVal. This could be due to either competition for shared transamination and oxidation reactions or a reduction in enzymatic activity. Also, NEAAG did not affect Val fluxes, but influx and efflux rate constants increased for both Val and Leu with HNEAAG, indicating an activating substrate effect. Overall, AA transport rates generally responded concordantly with extracellular concentrations, indicating the transporters are not substrate-saturated within the in vivo range. However, BCAA transamination and oxidation enzymes may be approaching saturation within in vivo ranges. In addition, System L transport activity appeared to be stimulated by as much as 75% with high intracellular concentrations of Ala, Gln, and Gly. High concentrations of Val antagonized transport activity of Ile and Leu by 68% and 15%, respectively, indicating competitive inhibition, but this was only observable at HNEAAG concentrations. The exchange transporters of System L transport 8 of the essential AA that make up approximately 40% of milk protein, so better understanding this transporter is an important step for increased efficiency.

The impact of leucine supplementation on body composition and glucose tolerance following energy restriction: an 8-week RCT in adults at risk of the metabolic syndrome.

BACKGROUND: L-Leucine (Leu) supplementation may benefit fat-free mass (FFM) per se and glucose metabolism. OBJECTIVES: To determine whether Leu supplementation during energy restriction blunted the loss of FFM, enhanced the loss of fat mass (FM) and improved glucose tolerance. DESIGN: Thirty-seven adults, aged 20-65 years, with increased waist circumference and at least one other metabolic syndrome (MetS) component, were selected. We employed a two-arm parallel, double blind, randomized control trial (RCT) design. Participants were randomly assigned to an intervention group (leucine - 3 g/d) or placebo (lactose - 2.67 g/d), while following an individualised energy restricted diet for an 8-week period. Detailed body composition (DEXA), oral glucose tolerance test (OGTT), insulin and components of MetS were measured before and after the trial. Analysis of covariance (ANCOVA) assessed the effect of Leu on an intention-to-treat (ITT) principle. Bootstrapping method with 1000 bootstrap samples was used to derive parameter estimates, standard errors, p-values, and 95% confidence intervals for all outcomes. RESULTS: Adjusted for baseline values and other covariates, FFM (p = 0.045) and lean tissue mass (LTM) (p = 0.050) were significantly higher following Leu. These outcomes were modified by a significant treatment x sex interaction that indicated Leu had the greater effect in men. However, on adjustment for body composition changes, there was no difference in insulin sensitivity, oral glucose tolerance, or change in MetS components following Leu. CONCLUSION: Short-term leucine supplementation during energy restriction resulted in a greater preservation of FFM and LTM particularly in men, but did not impact glucose metabolism.

Dietary leucine supplementation restores T-cell mitochondrial respiration and regulates T-lineage differentiation in denervation-induced sarcopenic mice.

With the aim of offsetting immune dysfunction preceded by sarcopenia, the feasibility and efficiency of nutritional leucine supplementation were evaluated using a murine denervation-induced sarcopenia model. Sciatic nerve axotomy caused significant loss of skeletal muscle of the hind limbs and accelerated mitochondrial stress along with suppressed ATP production in spleen-derived T cells. Dietary leucine intake not only ameliorated muscle mass anabolism in a sarcopenic state, but also restored mitochondrial respiratory function, as indicated by elevated levels of basal respiration, maximal respiration, spare respiratory capacity, and ATP production, in T cells, which in turn led to downregulated expression of mTOR and downstream signals, as indicated by the findings of comprehensive transcriptome analysis. Consequentially, this finally resulted in amelioration of the sarcopenia-induced relative Th1/Th17-dominant proinflammatory microenvironment. These results highlight the importance of leucine-promoted metabolic cues in directing T cell fate in a sarcopenic microenvironment. The present study provides insights that particularly help rationalize the design and optimization of leucine supplementation for chronic sarcopenic patients with autoimmune diseases.

Leucine Supplementation Improves Diastolic Function in HFpEF by HDAC4 Inhibition.

Heart failure with preserved ejection fraction (HFpEF) is a complex syndrome associated with a high morbidity and mortality rate. Leucine supplementation has been demonstrated to attenuate cardiac dysfunction in animal models of cachexia and heart failure with reduced ejection fraction (HFrEF). So far, no data exist on leucine supplementation on cardiac function in HFpEF. Thus, the current study aimed to investigate the effect of leucine supplementation on myocardial function and key signaling pathways in an established HFpEF rat model. Female ZSF1 rats were randomized into three groups: Control (untreated lean rats), HFpEF (untreated obese rats), and HFpEF_Leu (obese rats receiving standard chow enriched with 3% leucine). Leucine supplementation started at 20 weeks of age after an established HFpEF was confirmed in obese rats. In all animals, cardiac function was assessed by echocardiography at baseline and throughout the experiment. At the age of 32 weeks, hemodynamics were measured invasively, and myocardial tissue was collected for assessment of mitochondrial function and for histological and molecular analyses. Leucine had already improved diastolic function after 4 weeks of treatment. This was accompanied by improved hemodynamics and reduced stiffness, as well as by reduced left ventricular fibrosis and hypertrophy. Cardiac mitochondrial respiratory function was improved by leucine without alteration of the cardiac mitochondrial content. Lastly, leucine supplementation suppressed the expression and nuclear localization of HDAC4 and was associated with Protein kinase A activation. Our data show that leucine supplementation improves diastolic function and decreases remodeling processes in a rat model of HFpEF. Beneficial effects were associated with HDAC4/TGF-ß1/Collagenase downregulation and indicate a potential use in the treatment of HFpEF.

Leucine Supplementation Exacerbates Morbidity in Male but Not Female Mice with Colorectal Cancer-Induced Cachexia.

Cancer cachexia (CC) is a multifactorial wasting syndrome characterized by a significant loss in lean and/or fat mass and represents a leading cause of mortality in cancer patients. Nutraceutical treatments have been proposed as a potential treatment strategy to mitigate cachexia-induced muscle wasting. However, contradictory findings warrant further investigation. The purpose of this study was to determine the effects of leucine supplementation on skeletal muscle in male and female ApcMin/+ mice (APC). APC mice and their wild-type (WT) littermates were given normal drinking water or 1.5% leucine-supplemented water (n = 4-10/group/sex). We measured the gene expression of regulators of inflammation, protein balance, and myogenesis. Leucine treatment lowered survival rates, body mass, and muscle mass in males, while in females, it had no effect on body or muscle mass. Leucine treatment altered inflammatory gene expression by lowering Il1b 87% in the APC group and decreasing Tnfa 92% in both WT and APC males, while it had no effect in females (p < 0.05). Leucine had no effect on regulators of protein balance and myogenesis in either sex. We demonstrated that leucine exacerbates moribundity in males and is not sufficient for mitigating muscle or fat loss during CC in either sex in the ApcMin/+ mouse.

Protein dose requirements to maximize skeletal muscle protein synthesis after repeated bouts of resistance exercise in young trained women.

Studies examining the effect of protein (PRO) feeding on post resistance exercise (RE) muscle protein synthesis (MPS) have primarily been performed in men, and little evidence is available regarding the quantity of PRO required to maximally stimulate MPS in trained women following repeated bouts of RE. We therefore quantified acute (4 h and 8 h) and extended (24 h) effects of two bouts of resistance exercise, alongside protein-feeding, in women, and the PRO requirement to maximize MPS. Twenty-four RE trained women (26.6 ± 0.7 years, mean ± SEM) performed two bouts of whole-body RE (3 × 8 repetitions/maneuver at 75% 1-repetition maximum) 4 h apart, with post-exercise ingestion of 15 g, 30 g, or 60 g whey PRO (n = 8/group). Saliva, venous blood, and a vastus lateralis muscle biopsy were taken at 0 h, 4 h, 8 h, and 24 h post-exercise. Plasma leucine and branched chain amino acids were quantified using gas chromatography mass spectrometry (GC-MS) after ingestion of D2 O. Fifteen grams PRO did not alter plasma leucine concentration or myofibrillar synthetic rate (MyoFSR). Thirty and sixty grams PRO increased plasma leucine concentration above baseline (105.5 ± 5.3 µM; 120.2 ± 7.4 µM, respectively) at 4 h (151.5 ± 8.2 µM, p < 0.01; 224.8 ± 16.0 µM, p < 0.001, respectively) and 8 h (176.0 ± 7.3 µM, p < 0.001; 281.7 ± 21.6 µM, p < 0.001, respectively). Ingestion of 30 g PRO increased MyoFSR above baseline (0.068 ± 0.005%/h) from 0 to 4 h (0.140 ± 0.021%/h, p < 0.05), 0 to 8 h (0.121 ± 0.012%/h, p < 0.001), and 0 to 24 h (0.099 ± 0.011%/h, p < 0.01). Ingestion of 60 g PRO increased MyoFSR above baseline (0.063 ± 0.003%/h) from 0 to 4 h (0.109 ± 0.011%/h, p < 0.01), 0 to 8 h (0.093 ± 0.008%/h, p < 0.01), and 0 to 24 h (0.086 ± 0.006%/h, p < 0.01). Post-exercise ingestion of 30 g or 60 g PRO, but not 15 g, acutely increased MyoFSR following two consecutive bouts of RE and extended the anabolic window over 24 h. There was no difference between the 30 g and 60 g responses.

Exosome enriched leucine-rich alpha-2-glycoprotein-1 and extracellular matrix protein 1 proteins induce abnormal placental angiogenesis in pregnant mice.

INTRODUCTION: Gestational Diabetes Mellitus (GDM) is characterized by a high risk of fetal macrosomia and placenta hypervascularization. Exosomes has been known participating in various physiological and pathological processes, including pro-angiogenic function. However, the effects of umbilical cord blood derived exosomes from cases of GDM (GDM-exo) on placental vascular network formation remain unclear. METHODS: In the current study, we isolated and identified exosomes in umbilical cord blood from both normal (N-exo) and GDM pregnancies. Meanwhile, we investigated the effects of umbilical cord blood derived exosomes on placental angiogenesis both in vitro and in vivo. RESULTS: Our data indicated that in a mouse model, the placenta and fetus weight were significantly higher in the ones administrated with GDM-exo when compared with N-exo. Meanwhile, GDM-exo significantly enhanced placental endothelial cells functions in both HUVEC and HPMEC endothelial cell models. Importantly, we explored two up-regulated proteins in GDM-exo, namely leucine-rich alpha-2-glycoprotein-1 (LRG1) and extracellular matrix protein 1 (ECM1) by proteome analysis, which performed largely pro-angiogenic function and probably resulted in hypervascularization in GDM placenta. DISCUSSION: Thus, we proposed that abundant LRG1 and ECM1 enriched GDM-exo may take important roles in regulating pathological placental angiogenesis.

Dietary supplementation with L-leucine reduces nitric oxide synthesis by endothelial cells of rats.

This study tested the hypothesis that elevated L-leucine concentrations in plasma reduce nitric oxide (NO) synthesis by endothelial cells (ECs) and affect adiposity in obese rats. Beginning at four weeks of age, male Sprague-Dawley rats were fed a casein-based low-fat (LF) or high-fat (HF) diet for 15 weeks. Thereafter, rats in the LF and HF groups were assigned randomly into one of two subgroups (n = 8/subgroup) and received drinking water containing either 1.02% L-alanine (isonitrogenous control) or 1.5% L-leucine for 12 weeks. The energy expenditure of the rats was determined at weeks 0, 6, and 11 of the supplementation period. At the end of the study, an oral glucose tolerance test was performed on all the rats immediately before being euthanized for the collection of tissues. HF feeding reduced (P < 0.001) NO synthesis in ECs by 21% and whole-body insulin sensitivity by 19% but increased (P < 0.001) glutamine:fructose-6-phosphate transaminase (GFAT) activity in ECs by 42%. Oral administration of L-leucine decreased (P < 0.05) NO synthesis in ECs by 14%, increased (P < 0.05) GFAT activity in ECs by 35%, and reduced (P < 0.05) whole-body insulin sensitivity by 14% in rats fed the LF diet but had no effect (P > 0.05) on these variables in rats fed the HF diet. L-Leucine supplementation did not affect (P > 0.05) weight gain, tissue masses (including white adipose tissue, brown adipose tissue, and skeletal muscle), or antioxidative capacity (indicated by ratios of glutathione/glutathione disulfide) in LF- or HF-fed rats and did not worsen (P > 0.05) adiposity, whole-body insulin sensitivity, or metabolic profiles in the plasma of obese rats. These results indicate that high concentrations of L-leucine promote glucosamine synthesis and impair NO production by ECs, possibly contributing to an increased risk of cardiovascular disease in diet-induced obese rats.

Enhancement of PPARα-Inhibited Leucine Metabolism-Stimulated ß-Casein Synthesis and Fatty Acid Synthesis in Primary Bovine Mammary Epithelial Cells.

Leucine, a kind of branched-chain amino acid, plays a regulatory role in the milk production of mammalian mammary glands, but its regulatory functions and underlying molecular mechanisms remain unknown. This work showed that a leucine-enriched mixture (LEUem) supplementation increased the levels of milk protein and milk fat synthesis in primary bovine mammary epithelial cells (BMECs). RNA-seq of leucine-treated BMECs indicated alterations in lipid metabolism, translation, ribosomal structure and biogenesis, and inflammatory response signaling pathways. Meanwhile, the supplementation of leucine resulted in mTOR activation and increased the expression of BCKDHA, FASN, ACC, and SCD1. Interestingly, the expression of PPARα was independently correlated with the leucine-supplemented dose. PPARα activated by WY-14643 caused significant suppression of lipogenic genes expression. Furthermore, WY-14643 attenuated leucine-induced ß-casein synthesis and enhanced the level of BCKDHA expression. Moreover, promoter analysis revealed a peroxisome-proliferator-response element (PPRE) site in the bovine BCKDHA promoter, and WY-14643 promoted the recruitment of PPARα onto the BCKDHA promoter. Together, the present data indicate that leucine promotes the synthesis of ß-casein and fatty acid and that PPARα-involved leucine catabolism is the key target.