The effect of diminished metabolic acidosis on thermoregulatory response during exercise.

It was reported that metabolic acidosis inhibits the activity of warm-sensitive hypothalamic neurons. The present study was designed to test the hypothesis that metabolic alkalosis may improve heat loss during intensive exercise in men. Fifteen male subjects aged 22-24 years were submitted to incremental exercise on two randomized occasions one week apart. During the bicarbonate trial exercise was preceded by ingestion of NaHCO3 at a dose 250 mg/kg whilst during the placebo trial lactose was administered. Exercise load was increased every 3 min by 30 W until volitional exhaustion. Ambient temperature was kept at 23-24°C and humidity 50-60%. Tympanic and skin temperatures were recorded and the rate of sweating was assayed by humidity measurement of nitrogen flowing through a capsule attached to the mid posterior chest. Total sweat loss was determined by the changes in body mass. Venous blood samples were taken before exercise and at the end of each workload for determination of acid-base parameters. The subjects attained similar maximal workload in the two tests (260 ± 6 W) with heart rate 185 ± 6 beats/min. Blood concentration of hydrogen ions was lower (p < 0.001) in the bicarbonate than in the placebo trial throughout the whole exercise period. There were no significant differences between these tests in tympanic and mean skin temperatures, sweating rate and total sweat loss. The present data showed that in men attenuation of metabolic acidosis by bicarbonate ingestion did not influence thermoregulation during incremental exercise performed until volitional exhaustion, possibly due to too short duration of exertional uncompensated metabolic acidosis.

Psychomotor Performance after 30 h of Sleep Deprivation Combined with Exercise.

Sleep deprivation (SD) usually impairs psychomotor performance, but most experiments are usually focused on sedentary conditions. The purpose of this study was to evaluate the influence of 30 h of complete SD combined with prolonged, moderate exercise (SDE) on human psychomotor performance. Eleven endurance-trained men accustomed to overnight exertion were tested twice: in well-slept and non-fatigued conditions (Control) and immediately after 30 h of SDE. They performed a multiple-choice reaction time test (MCRT) at rest and during each workload of the graded exercise test to volitional exhaustion. At rest, the MCRT was shorter after SDE than in the Control (300 ± 13 ms vs. 339 ± 11 ms, respectively, p < 0.05). During graded exercise, there were no significant differences in MCRT between groups, but the fastest reaction was observed at lower workloads after SDE (158 ± 7 W vs. 187 ± 11 W in Control, p < 0.05). The total number of missed reactions tended to be higher after SDE (8.4 ± 0.7 vs. 6.3 ± 0.8 in Control, p = 0.06). In conclusion, SDE is different from SD alone; however, well-trained men, accustomed to overnight exertion can maintain psychomotor abilities independently of the extent of central fatigue. Exercise can be used to enhance psychomotor performance in sleep-deprived subjects in whom special caution is required in order to avoid overload.

ANGPTL4, IL-6 and TNF-α as regulators of lipid metabolism during a marathon run.

The aim of the study was to reveal whether marathon running influences regulators of lipid metabolism i.e. angiopoietin-like protein 4 (ANGPTL4), interleukin 6 (IL-6) and tumor necrosis factor-α (TNF-α). Plasma concentration of ANGPTL4, IL-6, TNF-α and lipids were determined in samples collected from 11 male runners before the marathon, immediately after the run and at 90 min of recovery. Plasma ANGPTL4 increased during exercise from 55.5 ± 13.4 to 78.1 ± 15.0 ng/ml (P < 0.001). This was accompanied by a significant increase in IL-6, TNF-α, free fatty acids (FFA) and glycerol (Gly) and a decrease in triacylglycerols (TG). After 90 min of recovery ANGPTL4 and TG did not differ from the exercise values, while plasma IL-6, TNF-α, FFA and Gly concentration were significantly lower. The exercise-induced increase in plasma concentration of ANGPTL4 correlated positively with the rise in plasma IL-6, TNF-α, FFA and Gly and negatively with the duration of the run. The increase in plasma IL-6 and TNF-α correlated positively with the rise in Gly. Summarizing, marathon running induced an increase in plasma ANGPTL4 and the value was higher in faster runners. The increase in plasma FFA, IL-6 and TNF-α concentration during a marathon run may be involved in plasma ANGPTL4 release, which could be a compensatory mechanism against FFA-induced lipotoxicity and oxidative stress. All of the analyzed cytokines may stimulate lipolysis during exercise.

Machine learning on the road to unlocking microbiota's potential for boosting immune checkpoint therapy.

The intestinal microbiota is a complex and diverse ecological community that fulfills multiple functions and substantially impacts human health. Despite its plasticity, unfavorable conditions can cause perturbations leading to so-called dysbiosis, which have been connected to multiple diseases. Unfortunately, understanding the mechanisms underlying the crosstalk between those microorganisms and their host is proving to be difficult. Traditionally used bioinformatic tools have difficulties to fully exploit big data generated for this purpose by modern high throughput screens. Machine Learning (ML) may be a potential means of solving such problems, but it requires diligent application to allow for drawing valid conclusions. This is especially crucial as gaining insight into the mechanistic basis of microbial impact on human health is highly anticipated in numerous fields of study. This includes oncology, where growing amounts of studies implicate the gut ecosystems in both cancerogenesis and antineoplastic treatment outcomes. Based on these reports and first signs of clinical benefits related to microbiota modulation in human trials, hopes are rising for the development of microbiome-derived diagnostics and therapeutics. In this mini-review, we're inspecting analytical approaches used to uncover the role of gut microbiome in immune checkpoint therapy (ICT) with the use of shotgun metagenomic sequencing (SMS) data.

Effect of mountain ultra-marathon running on plasma angiopoietin-like protein 4 and lipid profile in healthy trained men.

PURPOSE: Angiopoietin-like protein 4 (ANGPTL4) regulates lipid metabolism by inhibiting lipoprotein lipase activity and stimulating lipolysis in adipose tissue. The aim of this study was to find out whether the mountain ultra-marathon running influences plasma ANGPTL4 and whether it is related to plasma lipid changes. METHODS: Ten healthy men (age 31 ± 1.1 years) completed a 100-km ultra-marathon running. Plasma ANGPTL4, free fatty acids (FFA), triacylglycerols (TG), glycerol (Gly), total cholesterol (TC), low (LDL-C) and high (HDL-C) density lipoprotein-cholesterol were determined before, immediately after the run and after 90 min of recovery. RESULTS: Plasma ANGPTL4 increased during exercise from 68.0 ± 16.5 to 101.2 ± 18.1 ng/ml (p < 0.001). This was accompanied by significant increases in plasma FFA, Gly, HDL-C and decreases in plasma TG concentrations (p < 0.01). After 90 min of recovery, plasma ANGPTL4 and TG did not differ significantly from the exercise values, while plasma FFA, Gly, TC and HDL-C were significantly lower than immediately after the run. TC/HDL-C and TG/HDL-C molar ratios were significantly reduced. The exercise-induced changes in plasma ANGPTL4 correlated positively with those of FFA (r = 0.73; p < 0.02), and HDL-C (r = 0.69; p < 0.05). Positive correlation was found also between plasma ANGPTL4 and FFA concentrations after 90 min of recovery (r = 0.77; p < 0.01). CONCLUSIONS: The present data suggest that increase in plasma FFA during mountain ultra-marathon run may be involved in plasma ANGPTL4 release and that increase in ANGPTL4 secretion may be a compensatory mechanism against fatty acid-induced oxidative stress. Increase in plasma HDL-C observed immediately after the run may be due to the protective effect of ANGPTL4 on HDL.

Inhibitors of Serine Proteases in Regulating the Production and Function of Neutrophil Extracellular Traps.

Neutrophil extracellular traps (NETs), DNA webs released into the extracellular environment by activated neutrophils, are thought to play a key role in the entrapment and eradication of microbes. However, NETs are highly cytotoxic and a likely source of autoantigens, suggesting that NET release is tightly regulated. NET formation involves the activity of neutrophil elastase (NE), which cleaves histones, leading to chromatin decondensation. We and others have recently demonstrated that inhibitors of NE, such as secretory leukocyte protease inhibitor (SLPI) and SerpinB1, restrict NET production in vitro and in vivo. SLPI was also identified as a NET component in the lesional skin of patients suffering from the autoinflammatory skin disease psoriasis. SLPI-competent NET-like structures (a mixture of SLPI with neutrophil DNA and NE) stimulated the synthesis of interferon type I (IFNI) in plasmacytoid dendritic cells (pDCs) in vitro. pDCs uniquely respond to viral or microbial DNA/RNA but also to nucleic acids of "self" origin with the production of IFNI. Although IFNIs are critical in activating the antiviral/antimicrobial functions of many cells, IFNIs also play a role in inducing autoimmunity. Thus, NETs decorated by SLPI may regulate skin immunity through enhancing IFNI production in pDCs. Here, we review key aspects of how SLPI and SerpinB1 can control NET production and immunogenic function.

Secretory leukocyte protease inhibitor (SLPI), a multifunctional protein in the host defense response.

Secretory leukocyte protease inhibitor (SLPI), a ∼12kDa nonglycosylated cationic protein, is emerging as an important regulator of innate and adaptive immunity and as a component of tissue regenerative programs. First described as an inhibitor of serine proteases such as neutrophil elastase, this protein is increasingly recognized as a molecule that benefits the host via its anti-proteolytic, anti-microbial and immunomodulatory activities. Here, we discuss the diverse functions of SLPI. Moreover, we review several novel layers of SLPI-mediated control that protect the host from excessive/dysregulated inflammation typical of infectious, allergic and autoinflammatory diseases and that support healing responses through affecting cell proliferation, differentiation and apoptosis.

The inhibitory effect of secretory leukocyte protease inhibitor (SLPI) on formation of neutrophil extracellular traps.

Neutrophil extracellular traps (NETs), web-like DNA structures, provide efficient means of eliminating invading microorganisms but can also present a potential threat to its host because it is a likely source of autoantigens or by promoting bystander tissue damage. Therefore, it is important to identify mechanisms that inhibit NET formation. Neutrophil elastase (NE)-dependent chromatin decondensation is a key event in the release of NETs release. We hypothesized that inhibitors of NE, secretory leukocyte protease inhibitor (SLPI) and α(1)-proteinase inhibitor (α(1)-PI), has a role in restricting NET generation. Here, we demonstrate that exogenous human SLPI, but not α(1)-PI markedly inhibited NET formation in human neutrophils. The ability of exogenous SLPI to attenuate NET formation correlated with an inhibition of a core histone, histone 4 (H4), cleavage, and partial dependence on SLPI-inhibitory activity against NE. Moreover, neutrophils from SLPI(-/-) mice were more efficient at generating NETs than were neutrophils from wild-type mice in vitro, and in experimental psoriasis in vivo. Finally, endogenous SLPI colocalized with NE in the nucleus of human neutrophils in vitro, as well as in vivo in inflamed skin of patients with psoriasis. Together, these findings support a controlling role for SLPI in NET generation, which is of potential relevance to infectious and autoinflammatory diseases.

The role of metalloproteinase ADAM17 in regulating ICOS ligand-mediated humoral immune responses.

Immune cells regulate cell surface receptor expression during their maturation, activation, and motility. Although many of these receptors are regulated largely at the level of expression, protease-mediated ectodomain shedding represents an alternative means of refashioning the surface of immune cells. Shedding is largely attributed to a family of a disintegrin and metalloprotease domain (ADAM) metalloproteases, including ADAM17. Although ADAM17 is well known to contribute to the innate immune response, mainly by releasing TNF-α, much less is known about whether/how this metalloprotease regulates adaptive immunity. To determine whether ADAM17 contributes to regulating adaptive immune responses, we took advantage of ADAM17 hypomorphic (ADAM17(ex/ex)) mice, in which ADAM17 expression is reduced by 90-95% compared with wild-type littermates. In this study, we show that that ADAM17 deficiency results in spleen and lymph node enlargement, as well as increased levels of Ag-specific class-switched Ig production following immunization with OVA together with anti-CD40 mAbs and polyinosinic-polycytidylic acid. Moreover, we demonstrate that the costimulatory ligand ICOS ligand (ICOSL) is selectively downregulated on the surface of B cells in an ADAM17-specific manner, although it is not proteolitically processed by recombinant ADAM17 in vitro. Finally, we show that higher cell surface levels of ICOSL in ADAM17(ex/ex) mice may contribute to the development of excessive Ab responses. Therefore, our data suggest a functional link between ADAM17 and ICOSL in controlling adaptive immune responses.

Thermogenic effect of glucose in hypothyroid subjects.

The importance of thyroid hormone, catecholamines, and insulin in modification of the thermogenic effect of glucose (TEG) was examined in 34 healthy and 32 hypothyroid subjects. We calculated the energy expenditure at rest and during oral glucose tolerance test. Blood samples for determinations of glucose, plasma insulin, adrenaline (A), and noradrenaline (NA) were collected. It was found that TEG was lower in hypothyroid than in control group (19.68 ± 3.90 versus 55.40 ± 7.32 kJ, resp., P < 0.0004). Mean values of glucose and insulin areas under the curve were higher in women with hypothyroidism than in control group (286.79 ± 23.65 versus 188.41 ± 15.84 mmol/L·min, P < 0.003 and 7563.27 ± 863.65 versus 4987.72 ± 583.88 mU/L·min, P < 0.03 resp.). Maximal levels of catecholamines after glucose ingestion were higher in hypothyroid patients than in control subjects (Amax-0.69 ± 0.08 versus 0.30 ± 0.07 nmol/L, P < 0.0001, and NAmax-6.42 ± 0.86 versus 2.54 ± 0.30 nmol/L, P < 0.0002). It can be concluded that in hypothyroidism TEG and glucose tolerance are decreased while the adrenergic response to glucose administration is enhanced. Presumably, these changes are related to decreased insulin sensitivity and responsiveness to catecholamine action.

Molecular pathway profiling of T lymphocyte signal transduction pathways; Th1 and Th2 genomic fingerprints are defined by TCR and CD28-mediated signaling.

BACKGROUND: T lymphocytes are orchestrators of adaptive immunity. Naïve T cells may differentiate into Th1, Th2, Th17 or iTreg phenotypes, depending on environmental co-stimulatory signals. To identify genes and pathways involved in differentiation of Jurkat T cells towards Th1 and Th2 subtypes we performed comprehensive transcriptome analyses of Jurkat T cells stimulated with various stimuli and pathway inhibitors. Results from these experiments were validated in a human experimental setting using whole blood and purified CD4+ Tcells. RESULTS: Calcium-dependent activation of T cells using CD3/CD28 and PMA/CD3 stimulation induced a Th1 expression profile reflected by increased expression of T-bet, RUNX3, IL-2, and IFNγ, whereas calcium-independent activation via PMA/CD28 induced a Th2 expression profile which included GATA3, RXRA, CCL1 and Itk. Knock down with siRNA and gene expression profiling in the presence of selective kinase inhibitors showed that proximal kinases Lck and PKCθ are crucial signaling hubs during T helper cell activation, revealing a clear role for Lck in Th1 development and for PKCθ in both Th1 and Th2 development. Medial signaling via MAPkinases appeared to be less important in these pathways, since specific inhibitors of these kinases displayed a minor effect on gene expression. Translation towards a primary, whole blood setting and purified human CD4+ T cells revealed that PMA/CD3 stimulation induced a more pronounced Th1 specific, Lck and PKCθ dependent IFNγ production, whereas PMA/CD28 induced Th2 specific IL-5 and IL-13 production, independent of Lck activation. PMA/CD3-mediated skewing towards a Th1 phenotype was also reflected in mRNA expression of the master transcription factor Tbet, whereas PMA/CD28-mediated stimulation enhanced GATA3 mRNA expression in primary human CD4+ Tcells. CONCLUSIONS: This study identifies stimulatory pathways and gene expression profiles for in vitro skewing of T helper cell activation. PMA/CD3 stimulation enhances a Th1-like response in an Lck and PKCθ dependent fashion, whereas PMA/CD28 stimulation results in a Th2-like phenotype independent of the proximal TCR-tyrosine kinase Lck. This approach offers a robust and fast translational in vitro system for skewed T helper cell responses in Jurkat T cells, primary human CD4+ Tcells and in a more complex matrix such as human whole blood.

PD-1 blockade augments Th1 and Th17 and suppresses Th2 responses in peripheral blood from patients with prostate and advanced melanoma cancer.

Negative costimulation on T cells is exploited by both prostate cancer and melanoma to evade antitumor immunity. Blocking such mechanisms restores antitumor immunity as was demonstrated by the improved survival of patients with metastatic melanoma after treatment with an antibody blocking the CTLA-4 inhibitory receptor (ipilimumab). Enhanced expression of another inhibitory immunoreceptor, programmed death-1 (PD-1), and its ligand, PD-L1, was found to correlate with a poor prognosis in prostate cancer and melanoma. PD-1-blocking antibodies are being developed to modulate antitumor immune responses. To support preclinical and clinical development of anti-PD-1 therapy, we sought to develop biomarker assays that can detect the effect of PD-1-blocking agents in whole blood and peripheral blood mononuclear cells. In this study, we assessed the effect of PD-1 blockade in modulating super antigen (staphylococcus enterotoxin B)-induced and recall antigen (tetanus toxoid)-induced T-cell reactivity in vitro using whole blood and peripheral blood mononuclear cells from patients with advanced melanoma, prostate cancer, and healthy controls. PD-1 blockade was found to shift antigen-induced cellular reactivity toward a proinflammatory Th1/Th17 response, as evidenced by enhanced production of interferon γ, interleukin (IL)-2, tumor necrosis factor α, IL-6, and IL-17 and reduced production of the Th2 cytokines IL-5 and IL-13. It is interesting to note that suppression of Th2 responsivity was seen with whole blood cells only from patients with cancer. Taken together, we identified novel biomarker assays that might be used to determine the functional consequences of PD-1 blockade in peripheral blood cells from patients with cancer. How these assays translate to the local antitumor response remains to be established in a clinical setting.

Therapeutic dosing of an orally active, selective cathepsin S inhibitor suppresses disease in models of autoimmunity.

The purpose of the study was to examine the potential of inhibition of cathepsin S as a treatment for autoimmune diseases. A highly selective cathepsin S inhibitor, CSI-75, was shown to upregulate levels of the cathepsin S substrate, invariant chain Lip10, in vitro as well as in vivo in C57Bl/6 mice after oral administration. Functional activity of the compound was shown by a reduction in the OVA-specific response of OVA-sensitized splenocytes from C57Bl/6 mice as well as from OVA-TCR transgenic mice (DO11.10). Since these studies revealed a selective suppression of the Th1 and Th17 cytokines causing a shift to Th2, CSI-75 was tested in the murine HC-gp39-immunization model. Indeed, CSI-75 specifically reduced the circulating HC-gp39-specific IgG2a in these mice indicating selective inhibition of the Th1 type of response in vivo. The importance of especially the Th1 and Th17 cell subsets in the pathology of autoimmune diseases, renders CatS inhibition a highly interesting potential therapeutic treatment of autoimmune diseases. Therefore, CSI-75 was tested in a murine model of multiple sclerosis (i.e. experimental autoimmune encephalomyelitis (EAE)) in a semi-therapeutic setting (ie. oral treatment after initial sensitization to antigen). Finally, in a murine model with features resembling rheumatoid arthritis (the collagen-induced arthritis (CIA) model), CSI-75 was tested in a therapeutic manner (after disease development). CSI-75 caused a significant reduction in disease score in both disease models, indicating a promising role for CatS inhibitors in the area of therapeutic treatments for autoimmune diseases.

Palmitic acid metabolism in the soleus muscle in vitro in hypo- and hyperthyroid rats.

The aim of this study was to establish whether the rate of fatty acid (FA) incorporation and its utilization by the isolated soleus muscle is modified under conditions of thyroid hormone deficit or excess. The rate of palmitic acid (PA) uptake, oxidation and incorporation into intramuscular lipids with increasing PA concentration (0.5-1.5 mM) in the incubation medium were determined. In hypothyroid rats intramuscular triacylglycerol (TG) synthesis was increased, while the rate of PA oxidation to CO2 and incorporation into mono- and diacylglycerols (MG/DG) and phospholipids (PL) remained unchanged. In rats with triiodothyronine (T3) excess the rate of all processes studied was enhanced, although the percentage incorporation of PA into different classes of intramuscular lipids was fairly constant and, independently of thyroid state and FA concentration in the medium, was 56-66% for TG, 9-14% for MG/DG and 24-32% for PL. Our results thus indicate that even short-term T3 excess accelerates the rate of FA uptake and metabolism in the oxidative soleus muscle, whereas in hypothyroid rats only intramuscular TG synthesis is affected.