Changes in Skeletal Muscle Protein Metabolism Signaling Induced by Glutamine Supplementation and Exercise.

AIM: To evaluate the effects of resistance exercise training (RET) and/or glutamine supplementation (GS) on signaling protein synthesis in adult rat skeletal muscles. METHODS: The following groups were studied: (1) control, no exercise (C); (2) exercise, hypertrophy resistance exercise training protocol (T); (3) no exercise, supplemented with glutamine (G); and (4) exercise and supplemented with glutamine (GT). The rats performed hypertrophic training, climbing a vertical ladder with a height of 1.1 m at an 80° incline relative to the horizontal with extra weights tied to their tails. The RET was performed three days a week for five weeks. Each training session consisted of six ladder climbs. The extra weight load was progressively increased for each animal during each training session. The G groups received daily L-glutamine by gavage (one g per kilogram of body weight per day) for five weeks. The C group received the same volume of water during the same period. The rats were euthanized, and the extensor digitorum longus (EDL) muscles from both hind limbs were removed and immediately weighed. Glutamine and glutamate concentrations were measured, and histological, signaling protein contents, and mRNA expression analyses were performed. RESULTS: Supplementation with free L-glutamine increased the glutamine concentration in the EDL muscle in the C group. The glutamate concentration was augmented in the EDL muscles from T rats. The EDL muscle mass did not change, but a significant rise was reported in the cross-sectional area (CSA) of the fibers in the three experimental groups. The levels of the phosphorylated proteins (pAkt/Akt, pp70S6K/p70S6K, p4E-BP1/4E-BP1, and pS6/S6 ratios) were significantly increased in EDL muscles of G rats, and the activation of p4E-BP1 was present in T rats. The fiber CSAs of the EDL muscles in T, G, and GT rats were increased compared to the C group. These changes were accompanied by a reduction in the 26 proteasome activity of EDL muscles from T rats. CONCLUSION: Five weeks of GS and/or RET induced muscle hypertrophy, as indicated by the increased CSAs of the EDL muscle fibers. The increase in CSA was mediated via the upregulated phosphorylation of Akt, 4E-BP1, p70S6k, and S6 in G animals and 4E-BP1 in T animals. In the EDL muscles from T animals, a decrease in proteasome activity, favoring a further increase in the CSA of the muscle fibers, was reported.

Fish Oil Supplementation Improves the Repeated-Bout Effect and Redox Balance in 20-30-Year-Old Men Submitted to Strength Training.

Herein, we investigated the effect of fish oil supplementation combined with a strength-training protocol, for 6 weeks, on muscle damage induced by a single bout of strength exercise in untrained young men. Sixteen men were divided into two groups, supplemented or not with fish oil, and they were evaluated at the pre-training period and post-training period. We investigated changes before and 0, 24, and 48 h after a single hypertrophic exercise session. Creatine kinase (CK) and lactate dehydrogenase (LDH) activities, plasma interleukin-6 (IL-6) and C-reactive protein (CRP) levels, and the redox imbalance were increased in response to the single-bout session of hypertrophic exercises at baseline (pre-training period) and decreased during the post-training period in the control group due to the repeated-bout effect (RBE). The fish oil supplementation exacerbated this reduction and improved the redox state. In summary, our findings demonstrate that, in untrained young men submitted to a strength-training protocol, fish oil supplementation is ideal for alleviating the muscle injury, inflammation, and redox imbalance induced by a single session of intense strength exercises, highlighting this supplementation as a beneficial strategy for young men that intend to engage in strength-training programs.

Green tea beneficial effects involve changes in the profile of immune cells in the adipose tissue of obese mice.

PURPOSE: During obesity, the adipose tissue is usually infiltrated by immune cells which are related to hallmarks of obesity such as systemic inflammation and insulin resistance (IR). Green tea (GT) has been widely studied for its anti-inflammatory actions, including the modulation in the proliferation and activity of immune cells, in addition to preventing cardiovascular and metabolic diseases. METHODS: The aim of the present study was to analyze the population of immune cells present in the subcutaneous and epididymal white adipose tissue (WAT) of mice kept at thermoneutrality (TN) and fed with a high-fat diet (HFD) for 16 weeks, supplemented or not with GT extract (500 mg/kg/12 weeks). RESULTS: The HFD in association with TN has induced chronic inflammation, and IR in parallel with changes in the profile of immune cells in the subcutaneous and epidydimal WAT, increasing pro-inflammatory cytokines release, inflammatory cells infiltration, and fibrotic aspects in WAT. On the other hand, GT prevented body weight gain, in addition to avoiding IR and inflammation, and the consequent tissue fibrosis, maintaining a lower concentration of cytokines and a profile of immune cells similar to the control mice, preventing the harmful modulations induced by both HFD and TN. CONCLUSIONS: GT beneficial effects in WAT abrogated the deleterious effects triggered by HFD and TN, maintaining all immune cells and fibrotic markers at the same level as in lean mice. These results place WAT immune cells population as a potential target of GT action, also highlighting the positive effects of GT in obese mice housed at TN.

Changes in skeletal muscle protein metabolism signaling induced by glutamine supplementation and esxercise

Aim: To evaluate the effects of resistance exercise training (RET) and/or glutamine supplementation (GS) on signaling protein synthesis in adult rat skeletal muscles. Methods: The following groups were studied: (1) control, no exercise (C); (2) exercise, hypertrophy resistance exercise training protocol (T); (3) no exercise, supplemented with glutamine (G); and (4) exercise and supplemented with glutamine (GT). The rats performed hypertrophic training, climbing a vertical ladder with a height of 1.1 m at an 80° incline relative to the horizontal with extra weights tied to their tails. The RET was performed three days a week for five weeks. Each training session consisted of six ladder climbs. The extra weight load was progressively increased for each animal during each training session. The G groups received daily L-glutamine by gavage (one g per kilogram of body weight per day) for five weeks. The C group received the same volume of water during the same period. The rats were euthanized, and the extensor digitorum longus (EDL) muscles from both hind limbs were removed and immediately weighed. Glutamine and glutamate concentrations were measured, and histological, signaling protein contents, and mRNA expression analyses were performed. Results: Supplementation with free L-glutamine increased the glutamine concentration in the EDL muscle in the C group. The glutamate concentration was augmented in the EDL muscles from T rats. The EDL muscle mass did not change, but a significant rise was reported in the cross-sectional area (CSA) of the fibers in the three experimental groups. The levels of the phosphorylated proteins (pAkt/Akt, pp70S6K/p70S6K, p4E-BP1/4E-BP1, and pS6/S6 ratios) were significantly increased in EDL muscles of G rats, and the activation of p4E-BP1 was present in T rats. The fiber CSAs of the EDL muscles in T, G, and GT rats were increased compared to the C group. These changes were accompanied by a reduction in the 26 proteasome activity of EDL muscles from T rats. Conclusion: Five weeks of GS and/or RET induced muscle hypertrophy, as indicated by the increased CSAs of the EDL muscle fibers. The increase in CSA was mediated via the upregulated phosphorylation of Akt, 4E-BP1, p70S6k, and S6 in G animals and 4E-BP1 in T animals. In the EDL muscles from T animals, a decrease in proteasome activity, favoring a further increase in the CSA of the muscle fibers, was reported.

Early Signs of Inflammation With Mild Oxidative Stress in Mixed Martial Arts Athletes After Simulated Combat.

ABSTRACT: Gomes-Santos, JAF, Lambertucci, RH, Vardaris, CV, Passos, MEP, Silva-Junior, EP, Hatanaka, E, Gorjão, R, McAnulty, SR, Souza-Junior, TP, and Barros, MP. Early signs of inflammation with mild oxidative stress in Mixed Martial Arts athletes after simulated combat. J Strength Cond Res 36(1): 180-186, 2022-Combat sports involve a combination of strenuous physical activity, usually at the anaerobic threshold, followed by intermittent low-intensity recovery periods for energy re-establishment. Oxidative stress and inflammation are inevitable exercise-related processes that could drastically affect athletic performance and practitioners' health, unless efficiently controlled during and after physical activities. This study aims to measure oxidative stress and inflammation biomarkers in the plasma of 12 top ranked professional Mixed Martial Arts (MMAs) athletes before and after simulated combats under official rules (pre-post study). Our results show that the athletes exhibited mild oxidative imbalances in plasma, evidenced by significant (p < 0.01) higher contents of both reduced (+7.3%) and oxidized glutathione (+28%), uric acid (+21%), and "free" iron (+21%) after combat, whereas variation tendencies (0.05 < p < 0.01) were observed in the antioxidant capacity in plasma (-40%), and SOD (-27%) or GPX (+20%) antioxidant activities in erythrocytes. However, a clear pro-inflammatory state was detected by increases in circulating cytokines IL-6 (+6,020%), IL-1ß (+4,357%), and tumor necrosis factor alpha (+63%), and by an abrupt drop of the anti-inflammatory cytokine IL-10 (-98%). A significant correlation was observed between pre-post variations of IL-6 and GSH/GSSG ratio in plasma (p < 0.0001), which reinforces the integration between oxidative stress and inflammation during MMA combats. Considering metabolic and mechanical stresses (imposed by combat techniques, e.g., punches and joint locks), this study indicates pre-existing inflammation, although minor oxidative stress, in MMA professionals after combat.

Hydrolyzed whey protein enriched with glutamine dipeptide attenuates skeletal muscle damage and improves physical exhaustion test performance in triathletes.

Purpose: To investigate the effects of hydrolyzed whey protein enriched with glutamine dipeptide on the percentage of oxygen consumption, second ventilatory threshold, duration and total distance covered, and skeletal muscle damage during an exhaustion test in elite triathletes. Methods: The study was a randomized, double-blinded, placebo-controlled, crossover trial. Nine male triathletes performed a progressive incremental test on a treadmill ergometer (1.4 km h-1·3 min-1) 30 min after ingesting either 50 g of maltodextrin plus four tablets of 700 mg hydrolyzed whey protein enriched with 175 mg of glutamine dipeptide diluted in 250 ml of water (MGln) or four tablets of 700 mg maltodextrin plus 50 g maltodextrin diluted in 250 ml of water (M). Each athlete was submitted to the two dietary treatments and two corresponding exhaustive physical tests with an interval of one week between the interventions. The effects of the two treatments were then compared within the same athlete. Maximal oxygen consumption, percentage of maximal oxygen consumption, second ventilatory threshold, and duration and total distance covered were measured during the exhaustion test. Blood was collected before and immediately after the test for the determination of plasma lactate dehydrogenase (LDH) and creatine kinase (CK) activities and lactate concentration (also measured 6, 10, and 15 min after the test). Plasma cytokines (IL-6, IL-1ß, TNF-α, IL-8, IL-10, and IL-1ra) and C-reactive protein levels were also measured. Results: A single dose of MGln increased the percentage of maximal oxygen consumption, second ventilatory threshold duration, and total distance covered during the exhaustion test and augmented plasma lactate levels 6 and 15 min after the test. MGln also decreased plasma LDH and CK activities indicating muscle damage protection. Plasma cytokine and C-reactive protein levels did not change across the study periods. Conclusion: Conditions including overnight fasting and a single dose of MGln supplementation resulted in exercising at a higher percentage of maximal oxygen consumption, a higher second ventilatory threshold, blood lactate levels, and reductions in plasma markers of muscle damage during an exhaustion test in elite triathletes. These findings support oral glutamine supplementation's efficacy in triathletes, but further studies require.

L-Glutamine Supplementation Enhances Strength and Power of Knee Muscles and Improves Glycemia Control and Plasma Redox Balance in Exercising Elderly Women.

We investigated the effects of oral L-glutamine (Gln) supplementation, associated or not with physical exercises, in control of glycemia, oxidative stress, and strength/power of knee muscles in elderly women. Physically active (n = 21) and sedentary (n = 23) elderly women aged 60 to 80 years were enrolled in the study. Plasma levels of D-fructosamine, insulin, reduced (GSH) and oxidized (GSSG) glutathione, iron, uric acid, and thiobarbituric acid-reactive substances (TBARs) (lipoperoxidation product), as well as knee extensor/flexor muscle torque peak and average power (isokinetic test), were assessed pre- and post-supplementation with Gln or placebo (30 days). Higher plasma D-fructosamine, insulin, and iron levels, and lower strength/power of knee muscles were found pre-supplementation in the NPE group than in the PE group. Post-supplementation, Gln subgroups showed higher levels of GSH, GSSG, and torque peak, besides lower D-fructosamine than pre-supplementation values. Higher muscle average power and plasma uric acid levels were reported in the PE + Gln group, whereas lower insulin levels were found in the NPE + Gln than pre-supplementation values. TBARs levels were diminished post-supplementation in all groups. Gln supplementation, mainly when associated with physical exercises, improves strength and power of knee muscles and glycemia control, besides boosting plasma antioxidant capacity of elderly women.

Glutamine depletion disrupts mitochondrial integrity and impairs C2C12 myoblast proliferation, differentiation, and the heat-shock response.

Glutamine and glucose are both oxidized in the mitochondria to supply the majority of usable energy for processes of cellular function. Low levels of plasma and skeletal muscle glutamine are associated with severe illness. We hypothesized that glutamine deficiency would disrupt mitochondrial integrity and impair cell function. C2C12 mouse myoblasts were cultured in control media supplemented with 5.6 mmol/L glucose and 2 mmol/L glutamine, glutamine depletion (Gln-) or glucose depletion (Glc-) media. We compared mitochondrial morphology and function, as well as cell proliferation, myogenic differentiation, and heat-shock response in these cells. Glc- cells exhibited slightly elongated mitochondrial networks and increased mitochondrial mass, with normal membrane potential (ΔΨm). Mitochondria in Gln- cells became hyperfused and swollen, which were accompanied by severe disruption of cristae and decreases in ΔΨm, mitochondrial mass, the inner mitochondrial membrane remodeling protein OPA1, electron transport chain complex IV protein expression, and markers of mitochondrial biogenesis and bioenergetics. In addition, Gln- increased the autophagy marker LC3B-II on the mitochondrial membrane. Notably, basal mitochondrial respiration was increased in Glc- cells as compared to control cells, whereas maximal respiration remained unchanged. In contrast, basal respiration, maximal respiration and reserve capacity were all decreased in Gln- cells. Consistent with the aforementioned mitochondrial deficits, Gln- cells had lower growth rates and myogenic differentiation, as well as a higher rate of cell death under heat stress conditions than Glc- and control cells. We conclude that glutamine is essential for mitochondrial integrity and function; glutamine depletion impairs myoblast proliferation, differentiation, and the heat-shock response.

New insights on the regulation of cancer cachexia by N-3 polyunsaturated fatty acids.

Cancer cachexia is a multifactorial syndrome that develops during malignant tumor growth. Changes in plasma levels of several hormones and inflammatory factors result in an intense catabolic state, decreased activity of anabolic pathways, anorexia, and marked weight loss, leading to cachexia development and/or accentuation. Inflammatory mediators appear to be related to the control of a highly regulated process of muscle protein degradation that accelerates the process of cachexia. Several mediators have been postulated to participate in this process, including TNF-α, myostatin, and activated protein degradation pathways. Some interventional therapies have been proposed, including nutritional (dietary, omega-3 fatty acid supplementation), hormonal (insulin), pharmacological (clenbuterol), and nonpharmacological (physical exercise) therapies. Omega-3 (n-3) polyunsaturated fatty acids (PUFAs), especially eicosapentaenoic acid (EPA) and docosahexaenoic acid, are recognized for their anti-inflammatory properties and have been used in therapeutic approaches to treat or attenuate cancer cachexia. In this review, we discuss recent findings on cellular and molecular mechanisms involved in inflammation in the cancer cachexia syndrome and the effectiveness of n-3 PUFAs to attenuate or prevent cancer cachexia.

Attenuation of obesity and insulin resistance by fish oil supplementation is associated with improved skeletal muscle mitochondrial function in mice fed a high-fat diet.

Omega-3 polyunsaturated fatty acids (n-3 PUFAs) have been reported to improve insulin sensitivity and glucose homeostasis in animal models of insulin resistance, but the involved mechanisms still remain unresolved. In this study, we evaluated the effects of fish oil (FO), a source of n-3 PUFAs, on obesity, insulin resistance and muscle mitochondrial function in mice fed a high-fat diet (HFD). C57Bl/6 male mice, 8 weeks old, were divided into four groups: control diet (C), high-fat diet (H), C+FO (CFO) and H+FO (HFO). FO was administered by oral gavage (2 g/kg b.w.), three times a week, starting 4 weeks before diet administration until the end of the experimental protocol. HFD-induced obesity and insulin resistance associated with impaired skeletal muscle mitochondrial function, as indicated by decreased oxygen consumption, tricarboxylic acid cycle intermediate (TCAi) contents (citrate, α-ketoglutarate, malate and oxaloacetate), oxidative phosphorylation protein content and mitochondrial biogenesis. These effects were associated with elevated reactive oxygen species production, decreased PGC1-a transcription and reduced Akt phosphorylation. The changes induced by the HFD were partially attenuated by FO, which decreased obesity and insulin resistance and increased mitochondrial function. In the H group, FO supplementation also improved oxygen consumption; increased TCAi content, and Akt and AMPK phosphorylation; and up-regulated mRNA expression of Gpat1, Pepck, catalase and mitochondrial proteins (Pgc1α, Pparα, Cpt1 and Ucp3). These results suggest that dietary FO attenuates the deleterious effects of the HFD (obesity and insulin resistance) by improving skeletal muscle mitochondrial function.

Effects of DHA-rich fish oil supplementation on the lipid profile, markers of muscle damage, and neutrophil function in wheelchair basketball athletes before and after acute exercise.

We investigated the effects of docosahexaenoic acid (DHA)-rich fish oil (FO) supplementation on the lipid profile, levels of plasma inflammatory mediators, markers of muscle damage, and neutrophil function in wheelchair basketball players before and after acute exercise. We evaluated 8 male basketball wheelchair athletes before and after acute exercise both prior to (S0) and following (S1) FO supplementation. The subjects were supplemented with 3 g of FO daily for 30 days. The following components were measured: the plasma lipid profile (total cholesterol, high-density lipoprotein cholesterol, low-density lipoprotein cholesterol, and triglycerides), plasma inflammatory mediators (C-reactive protein, interleukin (IL)-1ß, IL-1ra, IL-4, IL-6, IL-8, and tumor necrosis factor-α), markers of muscle damage (creatine kinase and lactate dehydrogenase (LDH)), and neutrophil function (cytokine production, phagocytic capacity, loss of membrane integrity, mitochondrial membrane potential, neutral lipid accumulation, phosphatidylserine externalization, DNA fragmentation, and production of reactive oxygen species (ROS)). Acute exercise increased the plasma levels of total cholesterol, LDH, IL1ra, and IL-6, led to the loss of membrane integrity, ROS production, and a high mitochondrial membrane potential in neutrophils, and reduced the phagocytic capacity and IL-6 production by the neutrophils (S0). However, supplementation prevented the increases in the plasma levels of LDH and IL-6, the loss of membrane integrity, and the alterations in ROS production and mitochondrial membrane potential in the neutrophils that were induced by exercise (S1). In conclusion, DHA-rich FO supplementation reduces the markers of muscle damage, inflammatory disturbances, and neutrophil death induced by acute exercise in wheelchair athletes.

Medium-chain dicarboxylic acylcarnitines as markers of n-3 PUFA-induced peroxisomal oxidation of fatty acids.

SCOPE: Omega-3 polyunsaturated fatty acids (n-3 PUFA) found in fish oil activate PPAR-α, stimulate peroxisomal fatty acid (FA) ß-oxidation and prevent impairments on glucose homeostasis. METHODS AND RESULTS: Glucose metabolism and FA oxidation were studied in C57/Bl6 mice fed with diets containing either 3.6 and 31.5% fish oil or lard. To assess the effects of peroxisomal proliferation on FA oxidation independent of n-3 PUFA intake, mice were treated with the PPAR-α agonist WY-14643. n-3 PUFA-fed mice were protected from glucose intolerance and dyslipidemia compared to animals fed a lard-based high-fat diet. Most importantly, mice fed on the hyperlipidic diet based on fish oil as well as the WY-14643 treated mice showed twofold increase of odd, medium-chain, dicarboxylic acylcarnitines in the liver suggesting that not only ß-oxidation, but also α- and ω-oxidation of FA were increased. Finally, an oxidation assay using liver homogenates and palmitic acid as substrate revealed an over tenfold increased production of similar acylcarnitines, indicating that FA are their precursors. CONCLUSION: This study shows at the metabolite level that peroxisome proliferation induced either by fish oil or WY-14643 is associated with increased α- and ω-oxidation of FA producing specific acylcarnitines that can be utilized as biomarkers of peroxisomal FA oxidation.

Omega-3 fatty acids protect from diet-induced obesity, glucose intolerance, and adipose tissue inflammation through PPARγ-dependent and PPARγ-independent actions.

SCOPE: We tested herein the hypothesis that peroxisome proliferator activated receptor γ (PPARγ) is a major mediator of omega-3 (n-3) protective actions against high-fat diet (HFD) induced obesity, glucose intolerance, and adipose tissue inflammation. METHODS AND RESULTS: C57BL6 wild-type and fat-1 transgenic (fat-1) mice were fed a low-fat diet (LFD) or HFD, treated or not with PPARγ antagonist, and evaluated for energy balance, adiposity, glucose tolerance, and adipose tissue inflammation. Fat-1 mice were protected from obesity, fasting hyperglycemia, glucose intolerance, and adipose tissue inflammation. PPARγ inhibition completely abolished fat-1 protection against HFD-induced glucose intolerance, but not obesity or adipose tissue inflammation. To investigate the role of myeloid cell as mediator of n-3 beneficial metabolic actions, mice with deletion (LyzM-PPARγ(KO)) or nondeletion (LyzM-PPARγ(WT)) of PPARγ in myeloid cells were fed either LFD or HFD (lard) or an HFD rich in n-3 (fish oil). Our findings indicate that myeloid cell associated PPARγ is not involved in the attenuation of HFD-induced glucose intolerance and adipose tissue inflammation induced by n-3. CONCLUSION: High endogenous n-3 fatty acid levels protect from HFD obesity, glucose intolerance, and adipose tissue inflammation. Among these, only protection against glucose intolerance is mediated by non-myeloid cell PPARγ.

Fish oil supplementation for two generations increases insulin sensitivity in rats.

We investigated the effect of fish oil supplementation for two consecutive generations on insulin sensitivity in rats. After the nursing period (21 days), female rats from the same prole were divided into two groups: (a) control group and (b) fish oil group. Female rats were supplemented with water (control) or fish oil at 1 g/kg body weight as a single bolus for 3 months. After this period, female rats were mated with male Wistar rats fed on a balanced chow diet (not supplemented). Female rats continued to receive supplementation throughout gestation and lactation periods. The same treatment was performed for the next two generations (G1 and G2). At 75 days of age, male offspring from G1 and G2 generations from both groups were used in the experiments. G1 rats did not present any difference with control rats. However, G2 rats presented reduction in glycemia and lipidemia and improvement in in vivo insulin sensitivity (model assessment of insulin resistance, insulin tolerance test) as well as in vitro insulin sensitivity in soleus muscle (glucose uptake and metabolism). This effect was associated with increased insulin-stimulated p38 MAP kinase phosphorylation and lower n-6/n-3 fatty acid ratio, but not with activation of proteins from insulin signaling (IR, IRS-1 and Akt). Global DNA methylation was decreased in liver but not in soleus muscle. These results suggest that long-term fish oil supplementation improves insulin sensitivity in association with increased insulin-stimulated p38 activation and decreased n-6:n-3 ratio in skeletal muscle and decreased global DNA methylation in liver.

Comparative effects of DHA and EPA on cell function.

Fish oil supplementation has been reported to be generally beneficial in autoimmune, inflammatory and cardiovascular disorders. Most researchers have attributed these beneficial effects to the high content of omega-3 fatty acids in fish oil (FO). The effects of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) are not differentiated in most studies. In fact, up to 1990, purified DHA was not available for human use and there was no study regarding its effects on human immune response. In this review, the differences in the effects of these two fatty acids on cell function are discussed. Studies have shown that EPA and DHA have also different effects on leukocyte functions such as phagocytosis, chemotactic response and cytokine production. DHA and EPA modulate differently expression of genes in lymphocytes. Activation of intracellular signaling pathways involved with lymphocyte proliferation is also differently affected by these two fatty acids. In relation to insulin producing cell line RINm5F, DHA and EPA are cytotoxic at different concentrations and the proteins involved with cell death are differently modulated by these two fatty acids. Substantial improvement in the therapeutic usage of omega-3 fatty acid-rich FO will be possible with the discovery of the different mechanisms of actions of DHA and EPA.

Effects of exercise on leukocyte death: prevention by hydrolyzed whey protein enriched with glutamine dipeptide.

Lymphocyte and neutrophil death induced by exercise and the role of hydrolyzed whey protein enriched with glutamine dipeptide (Gln) supplementation was investigated. Nine triathletes performed two exhaustive exercise trials with a 1-week interval in a randomized, double blind, crossover protocol. Thirty minutes before treadmill exhaustive exercise at variable speeds in an inclination of 1% the subjects ingested 50 g of maltodextrin (placebo) or 50 g of maltodextrin plus 4 tablets of 700 mg of hydrolyzed whey protein enriched with 175 mg of glutamine dipeptide dissolved in 250 mL water. Cell viability, DNA fragmentation, mitochondrial transmembrane potential and production of reactive oxygen species (ROS) were determined in lymphocytes and neutrophils. Exhaustive exercise decreased viable lymphocytes but had no effect on neutrophils. A 2.2-fold increase in the proportion of lymphocytes and neutrophils with depolarized mitochondria was observed after exhaustive exercise. Supplementation of maltodextrin plus Gln (MGln) prevented the loss of lymphocyte membrane integrity and the mitochondrial membrane depolarization induced by exercise. Exercise caused an increase in ROS production by neutrophils, whereas supplementation of MGln had no additional effect. MGln supplementation partially prevented lymphocyte apoptosis induced by exhaustive exercise possibly by a protective effect on mitochondrial function.

Effect of olive oil-based emulsion on human lymphocyte and neutrophil death.

BACKGROUND: The incorporation of lipid emulsions in parenteral diets is a requirement for energy and essential fatty acid supply to critically ill patients. The most frequently used IV lipid emulsions (LE) are composed with long-chain triacylglycerols rich in omega-6 polyunsaturated fatty acids (PUFA) from soybean oil, but these LE promote lymphocyte and neutrophil death. A new emulsion containing 20% soybean oil and 80% olive oil rich in omega-9 monounsaturated fatty acids (MUFA) has been hypothesized not to cause impairment of immune function. In this study, the toxicity of an olive oil-based emulsion (OOE) on lymphocytes and neutrophils from healthy volunteers was investigated. METHODS: Twenty volunteers were recruited and blood was collected before a 6-hour infusion of an OOE, immediately after infusion, and again 18 hours postinfusion. Lymphocytes and neutrophils were isolated by gradient density. The cells were studied immediately after isolation and after 24 hours or 48 hours in culture. The following determinations were carried out: triacylglycerol levels and fatty acid composition and levels in plasma, lymphocyte proliferation, production of reactive oxygen species, and parameters of lymphocyte and neutrophil death (viability, DNA fragmentation, phosphatidylserine externalization, mitochondrial depolarization, and neutral lipid accumulation). RESULTS: OOE decreased lymphocyte proliferation, provoked lymphocyte necrosis, and had no effect on the proportion of viable neutrophils. The mechanism of cell death induced by OOE involved neutral lipid accumulation but had no effect on mitochondrial membrane depolarization. CONCLUSIONS: The OOE given as a single dose of 500 mL induced low toxicity to lymphocytes from healthy volunteers, probably by necrosis.

Effect of docosahexaenoic acid-rich fish oil supplementation on human leukocyte function.

BACKGROUND: The effect of a docosahexaenoic acid (DHA)-rich fish oil (FO) supplementation on human leukocyte function was investigated. METHODS: Ten male volunteers were supplemented with 3g/day FO containing 26% eicosapentaenoic acid (EPA, 20:5, n-3) and 54% DHA (22:6, n-3) for 2 months. RESULTS: FO supplementation changed the fatty acid (FA) composition of leukocytes resulting in an increase of n-3/n-6 ratio from 0.18 to 0.62 in lymphocytes and from 0.15 to 0.70 in neutrophils. DHA-rich FO stimulated an increase in phagocytic activity by 62% and 145% in neutrophils and monocytes, respectively. Neutrophil chemotactic response was increased by 128%. The rate of production of reactive oxygen species by neutrophils was also increased, as it was with lymphocyte proliferation. These changes were partially reversed after a 2-month wash out period. With respect to cytokine production by lymphocytes, interleukin (IL)-4 release was not altered, whereas secretions of IL-10, interferon (IFN)-gamma and tumor necrosis factor (TNF)-alpha were raised. These results are in contrast to those described by others using EPA-rich FO supplementation. Lymphocyte pleiotropic gene expression was analyzed by a macroarray technique. Of the analyzed genes (588 in total), 77 were modified by the supplementation. FO supplementation resulted in up-regulation of 6 genes (GATA binding protein 2, IL-6 signal transducer, transforming growth factor alpha, TNF, heat shock 90kDa protein 1-alpha and heat shock protein 70kDa 1A) and a down regulation of 71 genes (92.2% of total genes changed). The largest functional group of altered genes was that related to signaling pathways (22% of the total modified genes). CONCLUSIONS: Therefore, although EPA and DHA are members of n-3 FA family, changes in the proportion of DHA and EPA exert different effects on neutrophil, monocyte and lymphocyte function, which may be a result of specific changes in gene expression.

Toxicity of a soybean oil emulsion on human lymphocytes and neutrophils.

BACKGROUND: The incorporation of lipid emulsions in parenteral diets is a requirement for energy and essential fatty acid supply to critically ill patients. In this study, the toxicity of a lipid emulsion rich (60%) in triacylglycerol of omega-6 polyunsaturated fatty acids on leukocytes from healthy volunteers was investigated. METHODS: Eleven volunteers were recruited, and blood samples were collected before infusion of a soybean oil emulsion, immediately afterwards, and 18 hours later. The cells were studied immediately after isolation and again after 24 hours or 48 hours in culture. The following determinations were made: composition and concentration of fatty acids in plasma, lymphocytes and neutrophils, lymphocyte proliferation, levels of cell viability, DNA fragmentation, phosphatidylserine externalization, mitochondrial depolarization, reactive oxygen species production, and neutral lipid accumulation. RESULTS: Soybean oil emulsion decreased lymphocyte proliferation and provoked neutrophil and lymphocyte apoptosis and necrosis. Evidence is presented herein that soybean oil emulsion is less toxic to neutrophils than to lymphocytes. The mechanism of cell death induced by this oil emulsion was characterized by mitochondrial membrane depolarization and neutral lipid accumulation but did not alter reactive oxygen species production. CONCLUSIONS: Soybean oil emulsion given as a single dose of 500 mL promotes lymphocyte and neutrophil death that may enhance the susceptibility of the patients to infections.