Differential expression patterns of purinergic ectoenzymes and the antioxidative role of IL-6 in hospitalized COVID-19 patient recovery.

Introduction: We have acquired significant knowledge regarding the pathogenesis of severe acute respiratory syndrome caused by coronavirus 2 (SARS-CoV-2). However, the underlying mechanisms responsible for disease recovery still need to be fully understood. Methods: To gain insights into critical immune markers involved in COVID-19 etiopathogenesis, we studied the evolution of the immune profile of peripheral blood samples from patients who had recovered from COVID-19 and compared them to subjects with severe acute respiratory illness but negative for SARS-CoV-2 detection (controls). In addition, linear and clustered correlations between different parameters were determined. Results: The data obtained revealed a significant reduction in the frequency of inflammatory monocytes (CD14+CD16+) at hospital discharge vs. admission. Remarkably, nitric oxide (NO) production by the monocyte compartment was significantly reduced at discharge. Furthermore, interleukin (IL)-6 plasma levels were negatively correlated with the frequency of NO+CD14+CD16+ monocytes at hospital admission. However, at the time of hospital release, circulating IL-6 directly correlated with the NO production rate by monocytes. In line with these observations, we found that concomitant with NO diminution, the level of nitrotyrosine (NT) on CD8 T-cells significantly diminished at the time of hospital release. Considering that purinergic signaling constitutes another regulatory system, we analyzed the kinetics of CD39 and CD73 ectoenzyme expression in CD8 T-cells. We found that the frequency of CD39+CD8+ T-cells significantly diminished while the percentage of CD73+ cells increased at hospital discharge. In vitro, IL-6 stimulation of PBMCs from COVID-19 patients diminished the NT levels on CD8 T-cells. A clear differential expression pattern of CD39 and CD73 was observed in the NT+ vs. NT-CD8+ T-cell populations. Discussion: The results suggest that early after infection, IL-6 controls the production of NO, which regulates the levels of NT on CD8 T-cells modifying their effector functions. Intriguingly, in this cytotoxic cell population, the expression of purinergic ectoenzymes is tightly associated with the presence of nitrated surface molecules. Overall, the data obtained contribute to a better understanding of pathogenic mechanisms associated with COVID-19 outcomes.

Omega-3-Supplemented Fat Diet Drives Immune Metabolic Response in Visceral Adipose Tissue by Modulating Gut Microbiota in a Mouse Model of Obesity.

Obesity is a chronic, relapsing, and multifactorial disease characterized by excessive accumulation of adipose tissue (AT), and is associated with inflammation mainly in white adipose tissue (WAT) and an increase in pro-inflammatory M1 macrophages and other immune cells. This milieu favors the secretion of cytokines and adipokines, contributing to AT dysfunction (ATD) and metabolic dysregulation. Numerous articles link specific changes in the gut microbiota (GM) to the development of obesity and its associated disorders, highlighting the role of diet, particularly fatty acid composition, in modulating the taxonomic profile. The aim of this study was to analyze the effect of a medium-fat-content diet (11%) supplemented with omega-3 fatty acids (D2) on the development of obesity, and on the composition of the GM compared with a control diet with a low fat content (4%) (D1) over a 6-month period. The effect of omega-3 supplementation on metabolic parameters and the modulation of the immunological microenvironment in visceral adipose tissue (VAT) was also evaluated. Six-weeks-old mice were adapted for two weeks and then divided into two groups of eight mice each: a control group D1 and the experimental group D2. Their body weight was recorded at 0, 4, 12, and 24 weeks post-differential feeding and stool samples were simultaneously collected to determine the GM composition. Four mice per group were sacrificed on week 24 and their VAT was taken to determine the immune cells phenotypes (M1 or M2 macrophages) and inflammatory biomarkers. Blood samples were used to determine the glucose, total LDL and HDL cholesterol LDL, HDL and total cholesterol, triglycerides, liver enzymes, leptin, and adiponectin. Body weight measurement showed significant differences at 4 (D1 = 32.0 ± 2.0 g vs. D2 = 36.2 ± 4.5 g, p-value = 0.0339), 12 (D1 = 35.7 ± 4.1 g vs. D2 = 45.3 ± 4.9 g, p-value = 0.0009), and 24 weeks (D1 = 37.5 ± 4.7 g vs. D2 = 47.9 ± 4.7, p-value = 0.0009). The effects of diet on the GM composition changed over time: in the first 12 weeks, α and ß diversity differed considerably according to diet and weight increase. In contrast, at 24 weeks, the composition, although still different between groups D1 and D2, showed changes compared with previous samples, suggesting the beneficial effects of omega-3 fatty acids in D2. With regard to metabolic analysis, the results did not reveal relevant changes in biomarkers in accordance with AT studies showing an anti-inflammatory environment and conserved structure and function, which is in contrast to reported findings for pathogenic obesity. In conclusion, the results suggest that the constant and sustained administration of omega-3 fatty acids induced specific changes in GM composition, mainly with increases in Lactobacillus and Ligilactobacillus species, which, in turn, modulated the immune metabolic response of AT in this mouse model of obesity.

Purinergic modulation of the immune response to infections.

Infectious diseases are caused by the invasion of pathogenic microorganisms such as fungi, bacteria, viruses, and parasites. After infection, disease progression relies on the complex interplay between the host immune response and the microorganism evasion strategies. The host's survival depends on its ability to mount an efficient protective anti-microbial response to accomplish pathogen clearance while simultaneously preventing tissue injury by keeping under control the excessive inflammatory process. The purinergic system has the dual function of regulating the immune response and triggering effector antimicrobial mechanisms. This review provides an overview of the current knowledge of the modulation of innate and adaptive immunity driven by the purinergic system during parasitic, bacterial and viral infections.

Associations between objectively measured physical activity, sedentary time, and cardiorespiratory fitness with inflammatory and oxidative stress markers and heart rate variability.

Background: To assess the associations between physical activity (PA) and sedentary time (SEDT) with inflammatory and oxidative stress markers, heart rate variability (HRV) and post-exercise recovery (HRR) controlling for cardiorespiratory fitness (CRF) and potential confounders. Design and methods: The following data was collected from 44 participants during 2019 (age = 49.5 ± 6.4 years, 66% women): Plasma levels of C-reactive protein (CRP) and cytokines (IL-1ß, INF-γ, TNF-α, MCP-1, IL-6, IL-8, IL-10, IL-18, IL-23); catalase (CAT) and glutathione peroxidase (GPX) activities; resting heart (HR) rate for HRV analysis, anthropometric measures, a submaximal cycling test to evaluate CRF with active recovery to assess and HRR (absolute and ΔHR), and 7-day accelerometry. Results: Women spent significantly more SEDT (p = 0.035), had higher inflammatory markers (IL-6 and TNF) and lower HRV indices [SDNN, LF/HF, SD2 (p > 0.05)]. Significant associations were found between SEDT and markers of inflammation [CRP, B = 0.006, p = 0.001; MCP-1, B = 0.003, p = 0.038]. HRV indices were significantly associated with inflammatory/oxidative stress markers [IL-10 (p = 0.04), GPX (p = 0.014), ln-IL 23 (p = 0.036), CAT (p = 0.026)] while HRR was positively associated with light PA [Δ3 (B = 0.051, p = 0.043), Δ4 (B = 0.062, p = 0.021)] and inversely related to catalase [Δ3 (B = -54.7, p = 0.042), Δ4 (B = -54.1, p = 0.021] and CRP [Δ5 (B = -19.8, p = 0.033)]. Higher CRF showed lower values for TNF-α (p = 0.02) and IL-10 (p = 0.003) and better HRV/HRR indices [RMSSD, PNS, SampEn, SD1 (p < 0.05)]. Conclusions: SEDT had a higher impact on inflammation and autonomic balance, independently of PA levels with differences by sex and CRF. PA appears to be more important for a better HRR. Lower HRV and HRR could be indicative of inflammatory status.

HIF-1α and CD73 expression in cardiac leukocytes correlates with the severity of myocarditis in end-stage Chagas disease patients.

Chronic Chagas cardiomyopathy is the main infectious myocarditis worldwide. Almost 30% of Trypanosoma cruzi infected individuals develop slow and progressive myocarditis that leads to ventricular dilation and heart failure. Heart transplantation is an established, valuable therapeutic option for end-stage Chagas disease patients. Although the pathophysiology of Chagas disease has been addressed for decades by numerous groups, the cardiac immunologic mechanisms involved in the progression of clinical manifestation are still unknown. Growing evidence demonstrates that hypoxia-inducible factor (HIF)-1α plays indispensable roles in driving immune response by triggering the expression of CD73 purinergic ecto-enzyme. Purinergic system controls the duration and magnitude of purine signals directed to modulate immune cells through the conversion of extracellular ATP (microbicide/proinflammatory) to the immunoregulatory metabolite adenosine. In the present work, we described that infiltrating leukocytes within cardiac explants from patients with end-stage Chagas cardiomyopathy up-regulated HIF-1α and CD73 expression. Moreover, the number of HIF-1α+ and CD73+ leukocytes positively correlated with the myocarditis severity and the local parasite load. Furthermore, we demonstrated a direct relationship between tissue parasite persistence and the influx of immune cells to the infected hearts, which ultimately determine the severity of the myocarditis. These findings provide evidence that CD73-dependent regulatory pathways are locally triggered in the myocardium of patients with end-stage Chagas disease.

Deficiency of CD73 activity promotes protective cardiac immunity against Trypanosoma cruzi infection but permissive environment in visceral adipose tissue.

Damaged cells release the pro-inflammatory signal ATP, which is degraded by the ectonucleotidases CD39 and CD73 to the anti-inflammatory mediator adenosine (ADO). The balance between ATP/ADO is known to determine the outcome of inflammation/infection. However, modulation of the local immune response in different tissues due to changes in the balance of purinergic metabolites has yet to be investigated. Here, we explored the contribution of CD73-derived ADO on the acute immune response against Trypanosoma cruzi parasite, which invades and proliferates within different target tissues. Deficiency of CD73 activity led to an enhanced cardiac microbicidal immune response with an augmented frequency of macrophages with inflammatory phenotype and increased CD8+ T cell effector functions. The increment of local inducible nitric oxide (NO) synthase (iNOS)+ macrophages and the consequent rise of myocardial NO production in association with reduced ADO levels induced protection against T. cruzi infection as observed by the diminished cardiac parasite burden compared to their wild-type (WT) counterpart. Unexpectedly, parasitemia was substantially raised in CD73KO mice in comparison with WT mice, suggesting the existence of tissue reservoir/s outside myocardium. Indeed, CD73KO liver and visceral adipose tissue (VAT) showed increased parasite burden associated with a reduced ATP/ADO ratio and the lack of substantial microbicidal immune response. These data reveal that the purinergic system has a tissue-dependent impact on the host immune response against T. cruzi infection.

Monocyte glycolysis determines CD8+ T cell functionality in human Chagas disease.

Chagas disease is a lifelong pathology resulting from Trypanosoma cruzi infection. It represents one of the most frequent causes of heart failure and sudden death in Latin America. Herein, we provide evidence that aerobic glycolytic pathway activation in monocytes drives nitric oxide (NO) production, triggering tyrosine nitration (TN) on CD8+ T cells and dysfunction in patients with chronic Chagas disease. Monocytes from patients exhibited a higher frequency of hypoxia-inducible factor 1α and increased expression of its target genes/proteins. Nonclassical monocytes are expanded in patients' peripheral blood and represent an important source of NO. Monocytes entail CD8+ T cell surface nitration because both the frequency of nonclassical monocytes and that of NO-producing monocytes positively correlated with the percentage of TN+ lymphocytes. Inhibition of glycolysis in in vitro-infected peripheral blood mononuclear cells decreased the inflammatory properties of monocytes/macrophages, diminishing the frequency of IL-1ß- and NO-producing cells. In agreement, glycolysis inhibition reduced the percentage of TN+CD8+ T cells, improving their functionality. Altogether, these results clearly show that glycolysis governs oxidative stress on monocytes and modulates monocyte-T cell interplay in human chronic Chagas disease. Understanding the pathological immune mechanisms that sustain an inflammatory environment in human pathology is key to designing improved therapies.