The onset of rare earth metallosis begins with renal gadolinium-rich nanoparticles from magnetic resonance imaging contrast agent exposure.

The leitmotifs of magnetic resonance imaging (MRI) contrast agent-induced complications range from acute kidney injury, symptoms associated with gadolinium exposure (SAGE)/gadolinium deposition disease, potentially fatal gadolinium encephalopathy, and irreversible systemic fibrosis. Gadolinium is the active ingredient of these contrast agents, a non-physiologic lanthanide metal. The mechanisms of MRI contrast agent-induced diseases are unknown. Mice were treated with a MRI contrast agent. Human kidney tissues from contrast-naïve and MRI contrast agent-treated patients were obtained and analyzed. Kidneys (human and mouse) were assessed with transmission electron microscopy and scanning transmission electron microscopy with X-ray energy-dispersive spectroscopy. MRI contrast agent treatment resulted in unilamellar vesicles and mitochondriopathy in renal epithelium. Electron-dense intracellular precipitates and the outer rim of lipid droplets were rich in gadolinium and phosphorus. We conclude that MRI contrast agents are not physiologically inert. The long-term safety of these synthetic metal-ligand complexes, especially with repeated use, should be studied further.

Autophagic response to exercise in peripheral blood mononuclear cells from young men is intensity-dependent and is altered by exposure to environmental heat.

Autophagy is essential to maintaining cellular homeostasis in all eukaryotic cells and to tolerance of acute stressors such as starvation, heat, and recovery after exercise. Limited information exists regarding the exercise intensity-dependent autophagic response in humans, and it is unknown how environmental heat stress may modulate this response. Therefore, we evaluated autophagy and accompanying pathways of cellular stress [the heat-shock response (HSR), apoptosis, and acute inflammation] in peripheral blood mononuclear cells (PBMCs) from 10 young men (mean [SD]; 22 [2] years) before, immediately after and up to 6-h postexercise recovery from 30 min of low-, moderate-, and high-intensity semirecumbent cycling [40%, 55%, and 70% of maximal oxygen consumption (V̇o2max), respectively] in a temperate environment (25°C) and at 70% of V̇o2max in a hot environment (40°C). Changes in protein content were analyzed via Western blot. Each increase in exercise intensity was associated with elevations in mean body temperature. LC3-II increased after moderate-intensity exercise, with further increases after high-intensity exercise (P < 0.05). However, an increase in beclin-2 and ULK1, with a decrease in p62 was only observed after high-intensity exercise, which was paralleled by elevated TNF-α and cleaved-caspase-3, with the HSR peaking at 6 h after exercise (P < 0.05). When exercise was performed in the heat, greater LC3-II and cleaved-caspase-3 accumulation were observed; however, beclin-2 declined in recovery (P < 0.05). Therefore, our findings indicate that autophagy in PBMCs during exercise may be associated with greater heat strain exhibited during increasing exercise intensities, which is modulated by exposure to heat.

Substrate-selective positive allosteric modulation of PTPRD's phosphatase by flavonols.

The receptor type protein tyrosine phosphatase D (PTPRD) is expressed by neurons and implicated in interesting phenotypes that include reward from addictive substances, restless leg syndrome and neurofibrillary tangle densities in Alzheimer's disease (AD-NFTs). However, the brain phosphotyrosine phosphoprotein (PTPP) substrates for PTPRD's phosphatase have not been clearly defined. Although we have identified small molecule inhibitors of PTPRD's phosphatase that are candidates for reducing reward from addictive substances, no positive allosteric modulators of this phosphatase that might be candidates for reducing AD-NFTs have been reported. We now report identification of candidate brain substrates for PTPRD based on their increased phosphorylation in knockout vs wildtype animals, coexpression with PTPRD in neuronal subtypes and brisk dephosphorylation by recombinant human PTPRD phosphatase. We also report discovery that quercetin and other flavonols, though not closely-related flavones, enhance rates of PTPRD's dephosphorylation of a group of these candidate substrate PTPPs but not others. This substrate-selective positive allosteric modulation provides a novel pharmacological action. Flavonol-mediated increases in PTPRD's dephosphorylation of the GSK3 ß and α kinases that hyperphosphorylate tau, the major component of AD-NFTs, could help to explain recent data concerning genetic and dietary impacts on Alzheimer's disease.

Effect of extracellular hyperosmolality during normothermia and hyperthermia on the autophagic response in peripheral blood mononuclear cells from young men.

Heat-stress-induced dehydration is associated with extracellular hyperosmolality. To counteract the associated stress, cells employ cytoprotective mechanisms, including autophagy; however, the autophagic response to hyperosmotic stress has yet to be evaluated in humans. Thus, we investigated autophagy and associated cellular stress pathways [the heat shock response (HSR), apoptosis, and the acute inflammatory response] to isosmotic and hyperosmotic conditions with and without hyperthermia in 12 young men (mean [SD]; 25 [5] yr). Participants received a 90-min intravenous infusion of either isosmotic (ISO; 0.9% NaCl; serum osmolality of 293 [4] mosmol/kgH2O) or hyperosmotic (HYP; 3.0% NaCl; 300 [6] mosmol/kgH2O) saline, followed by passive whole body heating using water perfused suit to increase esophageal temperature by ∼0.8°C. Peripheral blood mononuclear cells were harvested at baseline (preinfusion), postinfusion, and after heating, and changes in protein content were analyzed via Western blotting. Post infusion, the LC3-II/I ratio was higher in HYP compared with ISO infusion (P < 0.001), although no other protein changes were observed (all P > 0.050). Following passive heating, autophagy increased in HYP, as demonstrated by an increase in LC3-II from baseline (P = 0.004) and an elevated LC3-II/I ratio compared with ISO (P = 0.035), and a decrease in p62 when compared with the ISO condition (P = 0.019). This was accompanied by an elevation in cleaved caspase-3 following heating in the HYP condition (P < 0.010); however, the HSR and acute inflammatory response did not change under any condition (all P > 0.050). Taken together, our findings indicate that serum hyperosmolality induces autophagy and apoptotic signaling during mild hyperthermia with minimal autophagic activation during normothermia.NEW & NOTEWORTHY We demonstrate that a physiologically relevant increase in serum osmolality causes minimal activation of the autophagic response. However, the combined stressors of serum hyperosmolality and mild hyperthermia causes activation of both autophagy and apoptotic signaling. Thus, changes in osmotic homeostasis appear to influence the cell's cytoprotective ability during periods of heat stress, highlighting the importance of considering osmotic status when examining autophagic responses in vivo.

Cross-Talk between Probiotic Nissle 1917 and Human Colonic Epithelium Affects the Metabolite Composition and Demonstrates Host Antibacterial Effect.

Colonic epithelium-commensal interactions play a very important role in human health and disease development. Colonic mucus serves as an ecologic niche for a myriad of commensals and provides a physical barrier between the epithelium and luminal content, suggesting that communication between the host and microbes occurs mainly by soluble factors. However, the composition of epithelia-derived metabolites and how the commensal flora influences them is less characterized. Here, we used mucus-producing human adult stem cell-derived colonoid monolayers exposed apically to probiotic E. coli strain Nissle 1917 to characterize the host-microbial communication via small molecules. We measured the metabolites in the media from host and bacterial monocultures and from bacteria-colonoid co-cultures. We found that colonoids secrete amino acids, organic acids, nucleosides, and polyamines, apically and basolaterally. The metabolites from host-bacteria co-cultures markedly differ from those of host cells grown alone or bacteria grown alone. Nissle 1917 affects the composition of apical and basolateral metabolites. Importantly, spermine, secreted apically by colonoids, shows antibacterial properties, and inhibits the growth of several bacterial strains. Our data demonstrate the existence of a cross-talk between luminal bacteria and human intestinal epithelium via metabolites, which might affect the numbers of physiologic processes including the composition of commensal flora via bactericidal effects.

SARS-COV-2 induced Diarrhea is inflammatory, Ca 2+ Dependent and involves activation of calcium activated Cl channels.

Diarrhea occurs in 2-50% of cases of COVID-19 (∼8% is average across series). The diarrhea does not appear to account for the disease mortality and its contribution to the morbidity has not been defined, even though it is a component of Long Covid or post-infectious aspects of the disease. Even less is known about the pathophysiologic mechanism of the diarrhea. To begin to understand the pathophysiology of COVID-19 diarrhea, we exposed human enteroid monolayers obtained from five healthy subjects and made from duodenum, jejunum, and proximal colon to live SARS-CoV-2 and virus like particles (VLPs) made from exosomes expressing SARS-CoV-2 structural proteins (Spike, Nucleocapsid, Membrane and Envelope). Results: 1) Live virus was exposed apically for 90 min, then washed out and studied 2 and 5 days later. SARS-Cov-2 was taken up by enteroids and live virus was present in lysates and in the apical>>basolateral media of polarized enteroids 48 h after exposure. This is the first demonstration of basolateral appearance of live virus after apical exposure. High vRNA concentration was detected in cell lysates and in the apical and basolateral media up to 5 days after exposure. 2) Two days after viral exposure, cytokine measurements of media showed significantly increased levels of IL-6, IL-8 and MCP-1. 3) Two days after viral exposure, mRNA levels of ACE2, NHE3 and DRA were reduced but there was no change in mRNA of CFTR. NHE3 protein was also decreased. 4) Live viral studies were mimicked by some studies with VLP exposure for 48 h. VLPs with Spike-D614G bound to the enteroid apical surface and was taken up; this resulted in decreased mRNA levels of ACE2, NHE3, DRA and CFTR. 4) VLP effects were determined on active anion secretion measured with the Ussing chamber/voltage clamp technique. S-D614G acutely exposed to apical surface of human ileal enteroids did not alter the short-circuit current (Isc). However, VLPS-D614G exposure to enteroids that were pretreated for ∼24 h with IL-6 plus IL-8 induced a concentration dependent increase in Isc indicating stimulated anion secretion, that was delayed in onset by ∼8 min. The anion secretion was inhibited by apical exposure to a specific calcium activated Cl channel (CaCC) inhibitor (AO1) but not by a specific CFTR inhibitor (BP027); was inhibited by basolateral exposure to the K channel inhibit clortimazole; and was prevented by pretreatment with the calcium buffer BAPTA-AM. 5) The calcium dependence of the VLP-induced increase in Isc was studied in Caco-2/BBe cells stably expressing the genetically encoded Ca2+ sensor GCaMP6s. 24 h pretreatment with IL-6/IL-8 did not alter intracellular Ca2+. However, in IL-6/IL-8 pretreated cells, VLP S-D614G caused appearance of Ca 2+ waves and an overall increase in intracellular Ca 2+ with a delay of ∼10 min after VLP addition. We conclude that the diarrhea of COVID-19 appears to an example of a calcium dependent inflammatory diarrhea that involves both acutely stimulated Ca2+ dependent anion secretion (stimulated Isc) that involves CaCC and likely inhibition of neutral NaCl absorption (decreased NHE3 protein and mRNA and decreased DRA mRNA).

Human Epithelial Stem Cell-Derived Colonoid Monolayers as a Model to Study Shiga Toxin-Producing Escherichia coli-Host Interactions.

Human intestinal organoid cultures established from crypt-derived stem cells truly revolutionized our approach to study intestinal epithelial physiology and pathologies as they can be propagated indefinitely and preserve the genetic signature of the donor and the gut segment specificity in culture. Here we describe human stem cell-derived colonoid monolayers as a reliable and reproducible model to study Shiga toxin-producing Escherichia coli (STEC) infection and STEC-caused pathologies of the whole colonic epithelium comprising a mixture of colonocytes, goblet, enteroendocrine, and other rare cells present in human colonic epithelial tissue.

Autophagy and heat: a potential role for heat therapy to improve autophagic function in health and disease.

Autophagy is a crucial cell survival mechanism that involves the degradation and recycling of old or damaged organelles and proteins to maintain cellular homeostasis. Impairments in autophagy are central to the pathogenesis of many conditions including metabolic and neurodegenerative disorders, cardiovascular and pulmonary diseases, diabetes, and aging. Although various pharmacological agents may be able to stimulate autophagic function, to our knowledge, few interventions exist that have been deemed safe and effective in humans. An emerging body of evidence suggests that targeting the autophagic pathway via passive heating (heat therapy) may stimulate autophagic function. Therefore, the primary focus of the present review is to analyze the mechanisms in which passive heating induces autophagy as defined by in vitro and in vivo (animal and human) models. Our secondary focus is to examine the implications of utilizing passive heating to restore dysfunctional autophagy in chronic disease and aging. Finally, we discuss potential therapeutic strategies to implement passive heating to stimulate autophagic function in humans.

How to Improve the Antioxidant Defense in Asphyxiated Newborns-Lessons from Animal Models.

Oxygen free radicals have been implicated in brain damage after neonatal asphyxia. In the early phase of asphyxia/reoxygenation, changes in antioxidant enzyme activity play a pivotal role in switching on and off the cascade of events that can kill the neurons. Hypoxia/ischemia (H/I) forces the brain to activate endogenous mechanisms (e.g., antioxidant enzymes) to compensate for the lost or broken neural circuits. It is important to evaluate therapies to enhance the self-protective capacity of the brain. In animal models, decreased body temperature during neonatal asphyxia has been shown to increase cerebral antioxidant capacity. However, in preterm or severely asphyxiated newborns this therapy, rather than beneficial seems to be harmful. Thus, seeking new therapeutic approaches to prevent anoxia-induced complications is crucial. Pharmacotherapy with deferoxamine (DFO) is commonly recognized as a beneficial regimen for H/I insult. DFO, via iron chelation, reduces oxidative stress. It also assures an optimal antioxidant protection minimizing depletion of the antioxidant enzymes as well as low molecular antioxidants. In the present review, some aspects of recently acquired insight into the therapeutic effects of hypothermia and DFO in promoting neuronal survival after H/I are discussed.

Effect of Acute Aerobic Exercise and Rapamycin Treatment on Autophagy in Peripheral Blood Mononuclear Cells of Adults With Prediabetes.

OBJECTIVES: Recently, a malfunction of the autophagic pathway has been implicated with impaired glucose metabolism and progression from prediabetes to type 2 diabetes. The aims of this study were to investigate the effect of exercise and rapamycin (RAPA) treatment on the autophagic process in peripheral blood mononuclear cells (PBMCs) from people with prediabetes compared with control subjects. METHODS: Two groups matched for age and sex served as participants and included 6 participants with prediabetes (42.4±11.7 years) and 6 control subjects (44.4±11.9 years). Participants exercised at 50% of maximal oxygen consumption for 60 min with 5 min of rest interspersed every 20 min. PBMCs were isolated pre-exercise, immediately postexercise and 4 h after exercise recovery. Additional PBMCs were incubated for 24 h and either exposed to bafilomycin, rapamycin with bafilomycin (RAPA), or no treatment with vehicle (dimethyl sulfoxide). Proteins and mRNA were analyzed via western blot and quantitative real-time polymerase chain reaction, respectively. RESULTS: Exercise increased autophagy immediately postexercise and recovered 4 h after exercise in control participants but not in participants with prediabetes. Autophagy increased in PBMCs from people with prediabetes and control participants after RAPA treatment; however, a significantly impaired autophagic response was observed in people with prediabetes when compared with control subjects. CONCLUSIONS: Our results indicate an impairment in autophagic flux in PBMCs from people with prediabetes when compared with control subjects in response to both exercise and RAPA treatment. Future methods of autophagic upregulation should be investigated to spare malfunctions in autophagy in people with prediabetes.

MMP-9-induced increase in intestinal epithelial tight permeability is mediated by p38 kinase signaling pathway activation of MLCK gene.

Matrix metalloproteinase-9 (MMP-9) has been implicated as being an important pathogenic factor in inflammatory bowel disease (IBD). MMP-9 is markedly elevated in intestinal tissue of patients with IBD, and IBD patients have a defective intestinal tight-junction (TJ) barrier manifested by an increase in intestinal permeability. The loss of intestinal epithelial barrier function is an important contributing factor in the development and prolongation of intestinal inflammation; however, the role of MMP-9 in intestinal barrier function remains unclear. The purpose of this study was to investigate the effect of MMP-9 on the intestinal epithelial TJ barrier and to delineate the intracellular mechanisms involved by using in vitro (filter-grown Caco-2 monolayers) and in vivo (mouse small intestine recycling perfusion) systems. MMP-9 caused a time- and dose-dependent increase in Caco-2 TJ permeability. MMP-9 also caused an increase in myosin light-chain kinase (MLCK) gene activity, protein expression, and enzymatic activity. The pharmacological MLCK inhibition and siRNA-induced knockdown of MLCK inhibited the MMP-9-induced increase in Caco-2 TJ permeability. MMP-9 caused a rapid activation of the p38 kinase signaling pathway and inhibition of p38 kinase activity prevented the MMP-9-induced increase in MLCK gene activity and the increase in Caco-2 TJ permeability. MMP-9 also caused an increase in mouse intestinal permeability in vivo, which was accompanied by an increase in MLCK expression. The MMP-9-induced increase in mouse intestinal permeability was inhibited in MLCK-deficient mice. These data show for the first time that the MMP-9-induced increase in intestinal TJ permeability in vitro and in vivo was mediated by the p38 kinase signal transduction pathway upregulation of MLCK gene activity and that therapeutic targeting of these pathways can prevent the MMP-9-induced increase in intestinal TJ permeability. NEW & NOTEWORTHY MMP-9 is highly elevated in patients with IBD. IBD patients have compromised intestinal TJ barrier function manifested by an increase in intestinal permeability and intestinal inflammation. This study shows that MMP-9, at clinically achievable concentrations, causes an increase in intestinal TJ permeability in vitro and in vivo. In addition, a MMP-9-induced increase in intestinal TJ permeability was mediated by an increase in MLCK gene and protein expression via the p38 kinase pathway.

Effect of Branched-Chain Amino Acid Supplementation on Recovery Following Acute Eccentric Exercise.

This study investigated the effect of branched-chain amino acid (BCAA) supplementation on recovery from eccentric exercise. Twenty males ingested either a BCAA supplement or placebo (PLCB) prior to and following eccentric exercise. Creatine kinase (CK), vertical jump (VJ), maximal voluntary isometric contraction (MVIC), jump squat (JS) and perceived soreness were assessed. No significant (p > 0.05) group by time interaction effects were observed for CK, soreness, MVIC, VJ, or JS. CK concentrations were elevated above baseline (p < 0.001) in both groups at 4, 24, 48 and 72 hr, while CK was lower (p = 0.02) in the BCAA group at 48 hr compared to PLCB. Soreness increased significantly from baseline (p < 0.01) in both groups at all time-points; however, BCAA supplemented individuals reported less soreness (p < 0.01) at the 48 and 72 hr time-points. MVIC force output returned to baseline levels (p > 0.05) at 24, 48 and 72 hr for BCAA individuals. No significant difference between groups (p > 0.05) was detected for VJ or JS. BCAA supplementation may mitigate muscle soreness following muscle-damaging exercise. However, when consumed with a diet consisting of ~1.2 g/kg/day protein, the attenuation of muscular performance decrements or corresponding plasma CK levels are likely negligible.

Understanding the factors that effect maximal fat oxidation.

Lipids as a fuel source for energy supply during submaximal exercise originate from subcutaneous adipose tissue derived fatty acids (FA), intramuscular triacylglycerides (IMTG), cholesterol and dietary fat. These sources of fat contribute to fatty acid oxidation (FAox) in various ways. The regulation and utilization of FAs in a maximal capacity occur primarily at exercise intensities between 45 and 65% VO2max, is known as maximal fat oxidation (MFO), and is measured in g/min. Fatty acid oxidation occurs during submaximal exercise intensities, but is also complimentary to carbohydrate oxidation (CHOox). Due to limitations within FA transport across the cell and mitochondrial membranes, FAox is limited at higher exercise intensities. The point at which FAox reaches maximum and begins to decline is referred to as the crossover point. Exercise intensities that exceed the crossover point (~65% VO2max) utilize CHO as the predominant fuel source for energy supply. Training status, exercise intensity, exercise duration, sex differences, and nutrition have all been shown to affect cellular expression responsible for FAox rate. Each stimulus affects the process of FAox differently, resulting in specific adaptions that influence endurance exercise performance. Endurance training, specifically long duration (>2 h) facilitate adaptations that alter both the origin of FAs and FAox rate. Additionally, the influence of sex and nutrition on FAox are discussed. Finally, the role of FAox in the improvement of performance during endurance training is discussed.

Reduced Hsp70 and Glutamine in Pediatric Severe Malaria Anemia: Role of Hemozoin in Suppressing Hsp70 and NF-κB activation.

Severe malarial anemia [SMA, hemoglobin (Hb) <5.0 g/dL] is a leading cause of global morbidity and mortality among children residing in Plasmodium falciparum transmission regions. Exploration of molecular pathways through global gene expression profiling revealed that SMA was characterized by decreased HSPA1A, a heat shock protein (Hsp) 70 coding gene. Hsp70 is a ubiquitous chaperone that regulates Nuclear Factor-kappa B (NF-κB) signaling and production of pro-inflammatory cytokines known to be important in malaria pathogenesis (e.g., IL-1ß, IL-6 and TNF-α). Since the role of host Hsp70 in malaria pathogenesis is unexplored, we investigated Hsp70 and molecular pathways in children with SMA. Validation experiments revealed that leukocytic HSP70 transcripts were reduced in SMA relative to non-severe malaria, and that intraleukocytic hemozoin (PfHz) was associated with lower HSP70. HSP70 was correlated with reticulocyte production and Hb. Since glutamine (Gln) up-regulates Hsp70, modulates NF-κB activation, and attenuates over-expression of pro-inflammatory cytokines, circulating Gln was measured in children with malaria. Reduced Gln was associated with increased risk of developing SMA. Treatment of cultured peripheral blood mononuclear cells (PBMCs) with PfHz caused a time-dependent decrease in Hsp70 transcripts/protein, and NF-κB activation. Gln treatment of PBMCs overcame PfHz-induced suppression of HSP70 transcripts/protein, reduced NF-κB activation, and suppressed over-expression of IL-1ß, IL-6 and TNF-α. Findings here demonstrate that SMA is characterized by reduced intraleukocytic HSP70 and circulating Gln, and that PfHz-induced suppression of HSP70 can be reversed by Gln. Thus, Gln supplementation may offer important immunotherapeutic options for futures studies in children with SMA.

Intestinal epithelial barrier function and tight junction proteins with heat and exercise.

A single layer of enterocytes and tight junctions (intercellular multiprotein complexes) form the intestinal epithelial barrier that controls transport of molecules through transcellular and paracellular pathways. A dysfunctional or "leaky" intestinal tight junction barrier allows augmented permeation of luminal antigens, endotoxins, and bacteria into the blood stream. Various substances and conditions have been shown to affect the maintenance of the intestinal epithelial tight junction barrier. The primary focus of the present review is to analyze the effects of exertional or nonexertional (passive hyperthermia) heat stress on tight junction barrier function in in vitro and in vivo (animals and humans) models. Our secondary focus is to review changes in tight junction proteins in response to exercise or hyperthermic conditions. Finally, we discuss some pharmacological or nutritional interventions that may affect the cellular mechanisms involved in maintaining homeostasis of the intestinal epithelial tight junction barrier during heat stress or exercise.

Heat shock response and autophagy--cooperation and control.

Protein quality control (proteostasis) depends on constant protein degradation and resynthesis, and is essential for proper homeostasis in systems from single cells to whole organisms. Cells possess several mechanisms and processes to maintain proteostasis. At one end of the spectrum, the heat shock proteins modulate protein folding and repair. At the other end, the proteasome and autophagy as well as other lysosome-dependent systems, function in the degradation of dysfunctional proteins. In this review, we examine how these systems interact to maintain proteostasis. Both the direct cellular data on heat shock control over autophagy and the time course of exercise-associated changes in humans support the model that heat shock response and autophagy are tightly linked. Studying the links between exercise stress and molecular control of proteostasis provides evidence that the heat shock response and autophagy coordinate and undergo sequential activation and downregulation, and that this is essential for proper proteostasis in eukaryotic systems.

The effects of acute oral glutamine supplementation on exercise-induced gastrointestinal permeability and heat shock protein expression in peripheral blood mononuclear cells.

Chronic glutamine supplementation reduces exercise-induced intestinal permeability and inhibits the NF-κB pro-inflammatory pathway in human peripheral blood mononuclear cells. These effects were correlated with activation of HSP70. The purpose of this paper is to test if an acute dose of oral glutamine prior to exercise reduces intestinal permeability along with activation of the heat shock response leading to inhibition of pro-inflammatory markers. Physically active subjects (N = 7) completed baseline and exercise intestinal permeability tests, determined by the percent ratio of urinary lactulose (5 g) to rhamnose (2 g). Exercise included two 60-min treadmill runs at 70 % of VO2max at 30 °C after ingestion of glutamine (Gln) or placebo (Pla). Plasma levels of endotoxin and TNF-α, along with peripheral blood mononuclear cell (PBMC) protein expression of HSP70 and IκBα, were measured pre- and post-exercise and 2 and 4 h post-exercise. Permeability increased in the Pla trial compared to that at rest (0.06 ± 0.01 vs. 0.02 ± 0.018) and did not increase in the Gln trial. Plasma endotoxin was lower at the 4-h time point in the Gln vs. 4 h in the Pla (6.715 ± 0.046 pg/ml vs. 7.952 ± 1.11 pg/ml). TNF-α was lower 4 h post-exercise in the Gln vs. Pla (1.64 ± 0.09 pg/ml vs. 1.87 ± 0.12 pg/ml). PBMC expression of IkBα was higher 4 h post-exercise in the Gln vs. 4 h in the Pla (1.29 ± 0.43 vs. 0.8892 ± 0.040). HSP70 was higher pre-exercise and 2 h post-exercise in the Gln vs. Pla (1.35 ± 0.21 vs. 1.000 ± 0.000 and 1.65 ± 0.21 vs. 1.27 ± 0.40). Acute oral glutamine supplementation prevents an exercise-induced rise in intestinal permeability and suppresses NF-κB activation in peripheral blood mononuclear cells.

Exercise regulation of intestinal tight junction proteins.

Gastrointestinal distress, such as diarrhoea, cramping, vomiting, nausea and gastric pain are common among athletes during training and competition. The mechanisms that cause these symptoms are not fully understood. The stress of heat and oxidative damage during exercise causes disruption to intestinal epithelial cell tight junction proteins resulting in increased permeability to luminal endotoxins. The endotoxin moves into the blood stream leading to a systemic immune response. Tight junction integrity is altered by the phosphoylation state of the proteins occludin and claudins, and may be regulated by the type of exercise performed. Prolonged exercise and high-intensity exercise lead to an increase in key phosphorylation enzymes that ultimately cause tight junction dysfunction, but the mechanisms are different. The purpose of this review is to (1) explain the function and physiology of tight junction regulation, (2) discuss the effects of prolonged and high-intensity exercise on tight junction permeability leading to gastrointestinal distress and (3) review agents that may increase or decrease tight junction integrity during exercise.

Effects of oral glutamine supplementation on exercise-induced gastrointestinal permeability and tight junction protein expression.

The objectives of this study are threefold: 1) to assess whether 7 days of oral glutamine (GLN) supplementation reduces exercise-induced intestinal permeability; 2) whether supplementation prevents the proinflammatory response; and 3) whether these changes are associated with upregulation of the heat shock response. On separate occasions, eight human subjects participated in baseline testing and in GLN and placebo (PLA) supplementation trials, followed by a 60-min treadmill run. Intestinal permeability was higher in the PLA trial compared with baseline and GLN trials (0.0604 ± 0.047 vs. 0.0218 ± 0.008 and 0.0272 ± 0.007, respectively; P < 0.05). IκBα expression in peripheral blood mononuclear cells was higher 240 min after exercise in the GLN trial compared with the PLA trial (1.411 ± 0.523 vs. 0.9839 ± 0.343, respectively; P < 0.05). In vitro using the intestinal epithelial cell line Caco-2, we measured effects of GLN supplementation (0, 4, and 6 mM) on heat-induced (37° or 41.8°C) heat shock protein 70 (HSP70), heat shock factor-1 (HSF-1), and occludin expression. HSF-1 and HSP70 levels increased in 6 mM supplementation at 41°C compared with 0 mM at 41°C (1.785 ± 0.495 vs. 0.6681 ± 0.290, and 1.973 ± 0.325 vs. 1.133 ± 0.129, respectively; P < 0.05). Occludin levels increased after 4 mM supplementation at 41°C and 6 mM at 41°C compared with 0 mM at 41°C (1.236 ± 0.219 and 1.849 ± 0.564 vs. 0.7434 ± 0.027, respectively; P < 0.001). GLN supplementation prevented exercise-induced permeability, possibly through HSF-1 activation.