Ganoderma lucidum dry extract supplementation modulates T lymphocyte function in older women.

Ganoderma lucidum (a mushroom used in traditional Chinese medicine) compounds may attenuate ageing-related physiological changes and restore normal immunity. However, studies on the physiological effects of Ganoderma lucidum dry extract food supplements are few. Therefore, here, we aimed to investigate the effects of Ganoderma lucidum dry extract food supplement on the lymphocyte function of older women. This was a double-blind clinical trial (n 60) with a final 39 older volunteers, divided into two groups Ganoderma lucidum (n 23) and placebo (n 16). The Ganoderma lucidum group received 2000 mg/d of Ganoderma lucidum dry extract for 8 weeks. We used flow cytometry to determine the lymphocyte profile. CD4+ lymphocyte gene expression was evaluated by real-time polymerase chain reaction. We observed that in the Ganoderma lucidum group, concanavalin A stimulation increased lymphocyte proliferation. Further, we observed an increase in expression of Forkhead box P3, transforming growth factor-beta, IL-10, IL-6, retinoic acid receptor-related orphan receptor gamma, GATA-binding protein 3 and interferon gamma genes in the Ganoderma lucidum group. Furthermore, in the Ganoderma lucidum group, ionomycin and phorbol 12-myristate 13-acetate stimulation led to decrease in Th17+ cells and increase in Th2+ cells. Thus, in older women, Ganoderma lucidum regulates T lymphocyte function leading to a predominant anti-inflammatory action but does not induce T lymphocyte proliferation through CD28 signalling pathway.

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.

Molecular and cellular mechanisms involved in tissue-specific metabolic modulation by SARS-CoV-2.

Coronavirus disease 2019 (COVID-19) is triggered by the SARS-CoV-2, which is able to infect and cause dysfunction not only in lungs, but also in multiple organs, including central nervous system, skeletal muscle, kidneys, heart, liver, and intestine. Several metabolic disturbances are associated with cell damage or tissue injury, but the mechanisms involved are not yet fully elucidated. Some potential mechanisms involved in the COVID-19-induced tissue dysfunction are proposed, such as: (a) High expression and levels of proinflammatory cytokines, including TNF-α IL-6, IL-1ß, INF-α and INF-ß, increasing the systemic and tissue inflammatory state; (b) Induction of oxidative stress due to redox imbalance, resulting in cell injury or death induced by elevated production of reactive oxygen species; and (c) Deregulation of the renin-angiotensin-aldosterone system, exacerbating the inflammatory and oxidative stress responses. In this review, we discuss the main metabolic disturbances observed in different target tissues of SARS-CoV-2 and the potential mechanisms involved in these changes associated with the tissue dysfunction.

Leukocyte metabolism in obese type 2 diabetic individuals associated with COVID-19 severity.

Recent studies show that the metabolic characteristics of different leukocytes, such as, lymphocytes, neutrophils, and macrophages, undergo changes both in the face of infection with SARS-CoV-2 and in obesity and type 2 diabetes mellitus (DM2) condition. Thus, the objective of this review is to establish a correlation between the metabolic changes caused in leukocytes in DM2 and obesity that may favor a worse prognosis during SARS-Cov-2 infection. Chronic inflammation and hyperglycemia, specific and usual characteristics of obesity and DM2, contributes for the SARS-CoV-2 replication and metabolic disturbances in different leukocytes, favoring the proinflammatory response of these cells. Thus, obesity and DM2 are important risk factors for pro-inflammatory response and metabolic dysregulation that can favor the occurrence of the cytokine storm, implicated in the severity and high mortality risk of the COVID-19 in these patients.

Evidence for Monocyte Reprogramming in a Long-Term Postsepsis Study.

This study sought to identify monocyte alterations from septic patients after hospital discharge by evaluating gene expression of inflammatory mediators and monocyte polarization markers. It was hypothesized that sepsis reprograms the inflammatory state of monocytes, causing effects that persist after hospital discharge and influencing patient outcomes. DESIGN: The gene expression patterns of inflammatory receptors, M1 and M2 macrophage polarization markers, NLRP3 inflammasome components, and pro- and anti-inflammatory cytokines in monocytes were assessed. PATIENTS: Thirty-four patients from the University of São Paulo Hospital, during the acute sepsis phase (phase A), immediately after ICU discharge (phase B), and 3 months (phase C), 6 months (phase D), 1 year (phase E), and 3 years (phase F) after discharge, were included. Patients that died during phases A and B were grouped separately, and the remaining patients were collectively termed the survivor group. MEASUREMENTS AND MAIN RESULTS: The gene expression of toll-like receptor (TLR)2 and TLR4 (inflammatory receptors), NLRP3, NFκB1, adaptor molecule apoptosis-associated speck-like protein containing a CARD, caspase 1, caspase 11, and caspase 12 (NLRP3 inflammasome components), interleukin-1α, interleukin-1ß, interleukin-18, and high-mobility group box 1 protein (proinflammatory cytokines), interleukin-10 (anti-inflammatory cytokine), C-X-C motif chemokine ligand 10, C-X-C motif chemokine ligand 11, and interleukin-12p35 (M1 inflammatory polarization markers), and C-C motif chemokine ligand 14, C-C motif chemokine ligand 22, transforming growth factor-beta (TGF-ß), SR-B1, and peroxisome proliferator-activated receptor γ (M2 anti-inflammatory polarization and tissue repair markers) was upregulated in monocytes from phase A until phase E compared with the control group. CONCLUSIONS: Sepsis reprograms the inflammatory state of monocytes, probably contributing to postsepsis syndrome development and mortality.

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.

Features of Neutrophils From Atopic and Non-Atopic Elite Endurance Runners.

We collected peripheral blood from thirty-nine elite male endurance runners at rest (24 hours after the last exercise session) and used the Allergy Questionnaire for Athletes score and plasma specific IgE level to separate them into atopic and non-atopic athletes. Neutrophils obtained from atopic and non-atopic athletes were subsequently stimulated in vitro with fMLP (N-formyl-methionyl-leucyl-phenylalanine), LPS (lipopolysaccharide), or PMA (phorbol 12-myristate 13-acetate). Neutrophils from non-atopic runners responded appropriately to LPS, as evidenced by the production of pro (IL-8, TNF-α, and IL-6) and anti-inflammatory (IL-10) cytokines. Neutrophils from atopic elite runners exhibited lower responses to LPS stimulus as indicated by no increase in IL-1ß, TNF-α, and IL-6 production. Neutrophils from non-atopic and atopic runners responded similarly to fMLP stimulation, indicating that migration function remained unaltered. Both groups were unresponsive to PMA induced reactive oxygen species (ROS) production. Training hours and training volume were not associated with neutrophil IgE receptor gene expression or any evaluated neutrophil function. Since non-atopic runners normally responded to LPS stimulation, the reduced neutrophil response to the stimuli was most likely due to the atopic state and not exercise training. The findings reported are of clinical relevance because atopic runners exhibit a constant decline in competition performance and are more susceptible to invading microorganisms.

Host cell glutamine metabolism as a potential antiviral target.

A virus minimally contains a nucleic acid genome packaged by a protein coat. The genome and capsid together are known as the nucleocapsid, which has an envelope containing a lipid bilayer (mainly phospholipids) originating from host cell membranes. The viral envelope has transmembrane proteins that are usually glycoproteins. The proteins in the envelope bind to host cell receptors, promoting membrane fusion and viral entry into the cell. Virus-infected host cells exhibit marked increases in glutamine utilization and metabolism. Glutamine metabolism generates ATP and precursors for the synthesis of macromolecules to assemble progeny viruses. Some compounds derived from glutamine are used in the synthesis of purines and pyrimidines. These latter compounds are precursors for the synthesis of nucleotides. Inhibitors of glutamine transport and metabolism are potential candidate antiviral drugs. Glutamine is also an essential nutrient for the functions of leukocytes (lymphocyte, macrophage, and neutrophil), including those in virus-infected patients. The increased glutamine requirement for immune cell functions occurs concomitantly with the high glutamine utilization by host cells in virus-infected patients. The development of antiviral drugs that target glutamine metabolism must then be specifically directed at virus-infected host cells to avoid negative effects on immune functions. Therefore, the aim of this review was to describe the landscape of cellular glutamine metabolism to search for potential candidates to inhibit glutamine transport or glutamine metabolism.

The Critical Role of Cell Metabolism for Essential Neutrophil Functions.

Neutrophils were traditionally considered as short-lived cells with abundant secretory and protein synthetic activity. Recent studies, however, indicate neutrophils are in reality a heterogeneous population of cells. Neutrophils differentiate from pluripotent stem cells in the bone marrow, and can further mature in the blood stream and can have different phenotypes in health and disease conditions. Neutrophils undergo primary functions such as phagocytosis, production of reactive oxygen species (ROS), release of lipid mediators and inflammatory proteins (mainly cytokines), and apoptosis. Neutrophils stimulate other neutrophils and trigger a cascade of immune and inflammatory responses. The underpinning intracellular metabolisms that support these neutrophil functions are herein reported. It has been known for many decades that neutrophils utilize glucose as a primary fuel and produce lactate as an end product of glycolysis. Neutrophils metabolize glucose through glycolysis and the pentose- phosphate pathway (PPP). Mitochondrial glucose oxidation is very low. The PPP provides the reduced nicotinamide adenine dinucleotide phosphate (NADPH) for the NADPH-oxidase (NOX) complex activity to produce superoxide from oxygen. These cells also utilize glutamine and fatty acids to produce the required adenosine triphosphate (ATP) and precursors for the synthesis of molecules that trigger functional outcomes. Neutrophils obtained from rat intraperitoneal cavity and incubate for 1 hour at 37°C metabolize glutamine at higher rate than that of glucose. Glutamine delays neutrophil apoptosis and maintains optimal NOX activity for superoxide production. Under limited glucose provision, neutrophils move to fatty acid oxidation (FAO) to obtain the required energy for the cell function. FAO is mainly associated with neutrophil differentiation and maturation. Hypoxia, hormonal dysfunction, and physical exercise markedly change neutrophil metabolism. It is now become clear that neutrophil metabolism underlies the heterogeneity of neutrophil phenotypes and should be intense focus of investigation.

Moderate physical exercise improves lymphocyte function in melanoma-bearing mice on a high-fat diet.

BACKGROUND: Obesity can lead to a chronic systemic inflammatory state that increases the risk of cancer development. Therefore, this study aimed to evaluate the alterations in tumor non-infiltrated lymphocytes function and melanoma growth in animals maintained on a high-fat diet and/or moderate physical exercise program in a murine model of melanoma. METHODS: Female mice were randomly divided into eight groups: 1) normolipidic control (N), 2) normolipidic + melanoma (NM), 3) high-fat control (H), 4) high-fat + melanoma (HM), 5) normolipidic control + physical exercise (NE), 6) normolipidic melanoma + physical exercise (NEM), 7) high-fat control + physical exercise (HE), and 8) high-fat melanoma + physical exercise (HEM). After 8 weeks of diet treatment and/or moderate physical exercise protocol, melanoma was initiated by explanting B16F10 cells into one-half of the animals. RESULTS: Animals fed a high-fat diet presented high-energy consumption (30%) and body weight gain (H and HE vs N and NE, 37%; HM and HEM vs NM and NEM, 73%, respectively), whether or not they carried melanoma explants. Although the tumor growth rate was higher in animals from the HM group than in animals from any other sedentary group, it was reduced by the addition of a physical exercise regimen. We also observed an increase in stimulated peripheral lymphocyte proliferation and a decrease in the T-helper 1 response in the HEM group. CONCLUSIONS: The results of the present study support the hypothesis that altering function of tumor non-infiltrated lymphocytes via exercise-related mechanisms can slow melanoma progression, indicating that the incorporation of a regular practice of moderate-intensity exercises can be a potential strategy for current therapeutic regimens in treating advanced melanoma.

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.

Marathon Race Affects Neutrophil Surface Molecules: Role of Inflammatory Mediators.

The fatigue induced by marathon races was observed in terms of inflammatory and immunological outcomes. Neutrophil survival and activation are essential for inflammation resolution and contributes directly to the pathogenesis of many infectious and inflammatory conditions. The aim of this study was to investigate the effect of marathon races on surface molecules related to neutrophil adhesion and extrinsic apoptosis pathway and its association with inflammatory markers. We evaluated 23 trained male runners at the São Paulo International Marathon 2013. The following components were measured: hematological and inflammatory mediators, muscle damage markers, and neutrophil function. The marathon race induced an increased leukocyte and neutrophil counts; creatine kinase (CK), lactate dehydrogenase (LDH), CK-MB, interleukin (IL)-6, IL-10, and IL-8 levels. C-reactive protein (CRP), IL-12, and tumor necrosis factor (TNF)-α plasma concentrations were significantly higher 24 h and 72 h after the marathon race. Hemoglobin and hematocrit levels decreased 72 h after the marathon race. We also observed an increased intercellular adhesion molecule-1 (ICAM-1) expression and decreasedTNF receptor-1 (TNFR1) expression immediately after and 24 h after the marathon race. We observed an increased DNA fragmentation and L-selectin and Fas receptor expressions in the recovery period, indicating a possible slow rolling phase and delayed neutrophil activation and apoptosis. Marathon racing affects neutrophils adhesion and survival in the course of inflammation, supporting the "open-window" post-exercise hypothesis.

Regulation of Gene Expression by Exercise-Related Micrornas.

Gene expression control by microRNAs (miRs) is an important mechanism for maintenance of cellular homeostasis in physiological and pathological conditions as well as in response to different stimuli including nutritional factors and exercise. MiRs are involved in regulation of several processes such as growth and development, fuel metabolism, insulin secretion, immune function, miocardium remodeling, cell proliferation, differenciation, survival, and death. These molecules have also been proposed to be potential biomarkers and/or therapeutical targets in obesity, type 2 diabetes mellitus, cardiovascular diseases, metabolic syndrome, and cancer. MiRs are released by most cells and potentially act on intercellular communication to borderer or distant cells. Various studies have been performed to elucidate the involvement of miRs in exercise-induced effects. The aims of this review are: 1) to bring up the main advances for the comprehension of the mechanisms of action of miRs; 2) to present the main results on miR involvement in physical exercise; 3) to discuss the physiological effects of miRs modified by exercise. The state of the art and the perspectives on miRs associated with physical exercise will be presented. Thus, this review is important for updating recent advances and driving further strategies and studies on the exercise-related miR research.

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.

Changes in lymphocyte and neutrophil function induced by a marathon race.

The aim of this study was to investigate the changes in lymphocyte and neutrophil selected functions before and after a marathon race. Fifteen professional athletes were recruited, and the following parameters were measured: plasma concentrations of IL-1ra, IL-6, IL-8, IL-10, TNF-α and C-reactive protein (CRP); neutrophil phagocytic capacity; cytokine production by neutrophils and lymphocytes and signs of neutrophil and lymphocyte death. The marathon race had no effect on CRP levels, but plasma concentrations of IL-6 and IL-1ra were increased. Although no effect was observed on the production of IL-6, IL1-ra, TNF-α, IL-1ß and IL-8 by unstimulated or stimulated neutrophils, a decrease in neutrophil phagocytic activity was observed immediately following the marathon. A high percentage of neutrophils undergoing apoptosis was observed due to the intense training regimen, whereas the percentages of apoptotic neutrophils were reduced after the race. The production of IL-2, TNF-α, IL-1ß and IL-10 by lymphocytes was decreased by 50%-80%, and the percentage of apoptotic and necrotic lymphocytes was increased by 42% and fourfold, respectively, as a result of the race. In conclusion, the increase in plasma levels of IL-6, IL-8, IL-1ra and IL-10 after the race was not due to the production of the cytokines by neutrophils or lymphocytes. In fact, the marathon led to a decrease in lymphocyte and neutrophil function, and the diminished function was more pronounced in lymphocytes, indicating an impairment in acquired immunity.

Effects of DHA-rich fish oil supplementation on lymphocyte function before and after a marathon race.

PURPOSE: To investigate the effects of docosahexaenoic-(DHA)-rich fish oil (FO) supplementation on lymphocyte function before and after a marathon race. METHODS: Twenty-one athletes participated in this study. Eight marathon runners were supplemented with 3 g of FO daily for 60 d (FO group), and 13 athletes were not supplemented (C group). The following measures of lymphocytes were taken before and after the marathon: cell proliferation, cytokine production (IL-2, IL-10, TNF-α, and IL-4), and signs of cell death. RESULTS: In the C group, the marathon had no effect on lymphocyte proliferation, DNA fragmentation, or mitochondrial membrane polarization; however, the marathon increased phosphatidylserine externalization (by 2.5-fold), induced a loss of plasma membrane integrity (by 20%), and decreased IL-2, TNF-α, and IL-10 production (by 55%, 95%, and 50%, respectively). FO supplementation did not prevent lymphocyte death induced by the marathon, as indicated by cell viability, DNA fragmentation, and phosphatidylserine externalization. However, FO supplementation increased lymphocyte proliferation before and after the marathon, and before the race, FO supplementation decreased IL-2, TNF-α, and IL-10 production in concanavalin-A-stimulated lymphocytes (by 55%, 95%, and 58%, respectively) compared with cells from the C group. The production of cytokines was not altered before or after the race in the FO group. CONCLUSIONS: DHA-rich FO supplementation increased lymphocyte proliferation and prevented a decrease in cytokine production, but it did not prevent lymphocyte death induced by participation in the marathon. Overall, DHA rich-FO supplementation has beneficial effects in preventing some of the changes in lymphocyte function induced by marathon participation.

Neurotoxicity of anhydroecgonine methyl ester, a crack cocaine pyrolysis product.

Smoking crack cocaine involves the inhalation of cocaine and its pyrolysis product, anhydroecgonine methyl ester (AEME). Although there is evidence that cocaine is neurotoxic, the neurotoxicity of AEME has never been evaluated. AEME seems to have cholinergic agonist properties in the cardiovascular system; however, there are no reports on its effects in the central nervous system. The aim of this study was to investigate the neurotoxicity of AEME and its possible cholinergic effects in rat primary hippocampal cell cultures that were exposed to different concentrations of AEME, cocaine, and a cocaine-AEME combination. We also evaluated the involvement of muscarinic cholinergic receptors in the neuronal death induced by these treatments using concomitant incubation of the cells with atropine. Neuronal injury was assessed using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and lactate dehydrogenase (LDH) assays. The results of the viability assays showed that AEME is a neurotoxic agent that has greater neurotoxic potential than cocaine after 24 and 48 h of exposure. We also showed that incubation for 48 h with a combination of both compounds in equipotent concentrations had an additive neurotoxic effect. Although both substances decreased cell viability in the MTT assay, only cocaine increased LDH release. Caspase-3 activity was increased after 3 and 6 h of incubation with 1mM cocaine and after 6 h of 0.1 and 1.0mM AEME exposure. Atropine prevented the AEME-induced neurotoxicity, which suggests that muscarinic cholinergic receptors are involved in AEME's effects. In addition, binding experiments confirmed that AEME has an affinity for muscarinic cholinergic receptors. Nevertheless, atropine was not able to prevent the neurotoxicity produced by cocaine and the cocaine-AEME combination, suggesting that these treatments activated other neuronal death pathways. Our results suggest a higher risk for neurotoxicity after smoking crack cocaine than after cocaine use alone.

The effect of an adventure race on lymphocyte and neutrophil death.

The effect of an adventure race (Ecomotion Pró), which lasted for 4-5 days, on neutrophil and lymphocyte death from elite athletes was investigated. Blood was collected from 11 athletes at rest and after the adventure race. The following parameters of cell death were measured in neutrophils and lymphocytes: cell membrane integrity, DNA fragmentation, mitochondrial transmembrane depolarization and reactive oxygen species (ROS) production. Phagocytosis capacity was also evaluated in neutrophils. The adventure race raised the proportion of cells with the loss of membrane integrity; lymphocytes by 14% and neutrophils by 16.4%. The proportion of lymphocytes with DNA fragmentation (2.9-fold) and mitochondrial transmembrane depolarization (1.5-fold) increased. However, these parameters did not change in neutrophils. ROS production remained unchanged in lymphocytes, whereas an increase by 2.2-fold was found in neutrophils due to the race. Despite these changes, the phagocytosis capacity did not change in neutrophils after the race. In conclusion, the Ecomotion Pró race-induced neutrophil death by necrosis (as indicated by the loss of membrane integrity) and led to lymphocyte death by apoptosis (as indicated by increase DNA fragmentation and depolarization of mitochondrial membrane).

Induction of lymphocyte death by short- and long-duration triathlon competitions.

PURPOSE: The effect of triathlon competitions on death of lymphocytes from elite athletes was investigated. MATERIAL AND METHODS: Blood was collected from sedentary volunteers and triathletes at rest and after a short-duration triathlon (SDT) and after a long-duration triathlon (LDT-half Ironman) competitions. RESULTS: The athletes had lowered lymphocyte proliferation capacity compared with sedentary volunteers either at rest or after the competitions. There was no difference in the parameters associated with lymphocyte death when sedentary volunteers were compared with triathletes at rest. Lymphocytes from triathletes after SDT competition showed an increase in DNA fragmentation, phosphatidylserine externalization, and mitochondrial transmembrane depolarization and did not alter membrane integrity when compared with cells from athletes at rest. In contrast, the LDT competition raised the proportion of lymphocytes with loss of membrane integrity when compared with cells from athletes at rest and did not change the apoptotic parameters. The LDT competition induced an increase of reactive oxygen species (ROS) production by lymphocytes compared with triathletes at rest. The SDT competition did not alter ROS production by lymphocytes when compared with cells from triathletes at rest. ROS production by lymphocytes after LDT competition was 60% higher than in SDT. CONCLUSIONS: Evidence is presented herein that an LDT competition caused lymphocyte death by necrosis, whereas an SDT induced lymphocyte apoptosis. The mechanism for lymphocyte death induced by the triathlon competitions may involve an increase in ROS production at different extents.

Neutrophil death induced by a triathlon competition in elite athletes.

INTRODUCTION/PURPOSE: The effect of a triathlon competition on death of neutrophils from elite athletes was investigated. METHODS: Blood was collected from 11 sedentary volunteers and 12 triathletes under rest and after a Half Ironman triathlon competition (2-km swimming, 80-km cycling, and 20-km running). RESULTS: The triathlon competition increased DNA fragmentation, phosphatidylserine externalization, and reactive oxygen species production in neutrophils when compared to the results at rest. The proportion of neutrophils with mitochondrial transmembrane depolarization was increased in the triathletes at rest and after competition as compared with sedentary volunteers. Plasma levels of thiobarbituric acid reactive substances were increased in triathletes after competition. Expression of bcl-xL (antiapoptotic) was decreased and that of bax (proapoptotic) was increased, whereas intracellular neutral lipid content was lowered in neutrophils after the triathlon. A positive correlation was found between the proportion of neutrophils with DNA fragmentation and the plasma free fatty acid levels (r = 0.688, P< 0.05), which was elevated by threefold after competition. Plasma levels of oleic, linoleic, and stearic acids were increased in triathletes after the competition when compared with sedentary volunteers. The plasma concentration of these three fatty acids, measured after the triathlon competition, was toxic for 3-h cultured neutrophils obtained from sedentary volunteers. The maximal tolerable (nontoxic) concentration of the fatty acids by 3-h cultured neutrophils was 100 micromol x L(-1) for oleic and linoleic acids and 200 micromol x L(-1) for stearic acid. CONCLUSION: The triathlon competition induced neutrophil death possibly by apoptosis as indicated by DNA fragmentation and phosphatidylserine externalization. The increase in plasma levels of oleic, linoleic, and stearic acids induced by the competition may be involved in the neutrophil death observed possibly by increasing the production of reactive oxygen species and by decreasing the accumulation of intracellular neutral lipid.