12-week combined strength and endurance exercise attenuates CD8+ T-cell differentiation and affects the kynurenine pathway in the elderly: a randomized controlled trial.

BACKGROUND: Age-related accumulation of highly differentiated CD8+ effector memory re-expressing CD45RA (EMRA) T-cells and disruption of the kynurenine (KYN) pathway are associated with chronic inflammation and the development of insulin resistance. In this study the aim was to investigate the effects of 12-week combined strength and endurance exercise on CD8+ T-cell differentiation and KYN pathway metabolites. Ninety-six elderly subjects (f/m, aged 50-70) were randomized to a control (CON) or exercise (EX) group. The EX group completed combined strength and endurance training twice weekly for one hour each time at an intensity of 60% of the one-repetition maximum for strength exercises and a perceived exertion of 15/20 for endurance exercises. The EX group was also randomly subdivided into two groups with or without a concomitant balanced diet intervention in order to examine additional effects besides exercise alone. Before and after the intervention phase, the proportions of CD8+ T-cell subsets and levels of KYN pathway metabolites in peripheral blood were determined. RESULTS: The CD8+ EMRA T-cell subsets increased in the CON group but remained almost unchanged in the EX group (p = .02). Plasma levels of kynurenic acid (KA) increased in the EX group and decreased in the CON group (p = .03). Concomitant nutritional intervention resulted in lower levels of quinolinic acid (QA) compared with exercise alone (p = .03). Overall, there was a slight increase in the QA/KA ratio in the CON group, whereas it decreased in the EX group (p > .05). CONCLUSIONS: Combined strength and endurance training seems to be a suitable approach to attenuate CD8+ T-cell differentiation in the elderly and to redirect the KYN pathway towards KA. The clinical relevance of these effects needs further investigation.

Hyperinflammatory State and Low T1 Adaptive Immune Response in Severe and Critical Acute COVID-19 Patients.

Background: A better understanding of COVID-19 immunopathology is needed to identify the most vulnerable patients and improve treatment options. Objective: We aimed to identify immune system cell populations, cytokines, and inflammatory markers related to severity in COVID-19. Methods: 139 hospitalized patients with COVID-19-58 mild/moderate and 81 severe/critical-and 74 recovered patients were included in a prospective longitudinal study. Clinical data and blood samples were obtained on admission for laboratory markers, cytokines, and lymphocyte subsets study. In the recovered patients, lymphocyte subsets were analyzed 8-12 weeks after discharge. Results: A National Early Warning Score 2 >2 (OR:41.4; CI:10.38-167.0), ferritin >583 pg/mL (OR:16.3; CI: 3.88-69.9), neutrophil/lymphocyte ratio >3 (OR: 3.5; CI: 1.08-12.0), sIL-2rα (sCD25) >512 pg/mL (OR: 3.3; CI: 1.48-7.9), IL-1Ra >94 pg/mL (OR: 3.2; IC: 1.4-7.3), and IL-18 >125 pg/mL (OR: 2.4; CI: 1.1-5.0) were associated with severe/critical COVID-19 in the multivariate models used. Lower absolute values of CD3, CD4, CD8, and CD19 lymphocytes together with higher frequencies of NK cells, a CD4 and CD8 activated (CD38+HLA-DR+) memory T cell and effector memory CD45RA+ (EMRA) phenotype, and lower T regulatory cell frequencies were found in severe/critical patients relative to mild/moderate and recovered COVID-19 patients. A significant reduction in Th1, Tfh1, and Tc1 with higher Th2, Tfh2, Tc2, and plasma cell frequencies was found in the most severe cases. Conclusion: A characteristic hyperinflammatory state with significantly elevated neutrophil/lymphocyte ratio and ferritin, IL-1Ra, sIL-2rα, and IL-18 levels together with a "low T1 lymphocyte signature" was found in severe/critical COVID-19 patients.

Cytomegalovirus Latent Infection is Associated with an Increased Risk of COVID-19-Related Hospitalization.

Some risk factors for severe coronavirus disease 2019 (COVID-19) have been identified, including age, race, and obesity. However, 20%-50% of severe cases occur in the absence of these factors. Cytomegalovirus (CMV) is a herpesvirus that infects about 50% of all individuals worldwide and is among the most significant nongenetic determinants of immune system. We hypothesized that latent CMV infection might influence the severity of COVID-19. Our analyses demonstrate that CMV seropositivity is associated with more than twice the risk of hospitalization due to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. Immune profiling of blood and CMV DNA quantitative polymerase chain reaction in a subset of patients for whom respiratory tract samples were available revealed altered T-cell activation profiles in absence of extensive CMV replication in the upper respiratory tract. These data suggest a potential role for CMV-driven immune perturbations in affecting the outcome of SARS-CoV-2 infection and may have implications for the discrepancies in COVID-19 severity between different human populations.

Abdominal Obesity-Related Disturbance of Insulin Sensitivity Is Associated with CD8+ EMRA Cells in the Elderly.

Aging and overweight increase the risk of developing type 2 diabetes mellitus. In this cross-sectional study, we aimed to investigate the potential mediating role of T-EMRA cells and inflammatory markers in the development of a decreased insulin sensitivity. A total of 134 healthy older volunteers were recruited (age 59.2 (SD 5.6) years). T cell subpopulations were analyzed by flow cytometry. Furthermore, body composition, HOMA-IR, plasma tryptophan (Trp) metabolites, as well as cytokines and adipokines were determined. Using subgroup and covariance analyses, the influence of BMI on the parameters was evaluated. Moreover, correlation, multiple regression, and mediation analyses were performed. In the subgroup of participants with obesity, an increased proportion of CD8+EMRA cells and elevated concentrations of plasma kynurenine (KYN) were found compared to the lower-weight subgroups. Linear regression analysis revealed that an elevated HOMA-IR could be predicted by a higher proportion of CD8+EMRA cells and KYN levels. A mediation analysis showed a robust indirect effect of the Waist-to-hip ratio on HOMA-IR mediated by CD8+EMRA cells. Thus, the deleterious effects of abdominal obesity on glucose metabolism might be mediated by CD8+EMRA cells in the elderly. Longitudinal studies should validate this assumption and analyze the suitability of CD8+EMRA cells as early predictors of incipient prediabetes.

Experimental Evolution of Escherichia coli K-12 in the Presence of Proton Motive Force (PMF) Uncoupler Carbonyl Cyanide m-Chlorophenylhydrazone Selects for Mutations Affecting PMF-Driven Drug Efflux Pumps.

Experimental evolution of Escherichia coli K-12 with benzoate, a partial uncoupler of the proton motive force (PMF), selects for mutations that decrease antibiotic resistance. We conducted experimental evolution in the presence of carbonyl cyanide m-chlorophenylhydrazone (CCCP), a strong uncoupler. Cultures were serially diluted daily 1:100 in LBK medium containing 20 to 150 µM CCCP buffered at pH 6.5 or at pH 8.0. After 1,000 generations, the populations tolerated up to 150 µM CCCP. Sequenced isolates had mutations in mprA (emrR), which downregulates the EmrAB-TolC pump that exports CCCP. A mprA::kanR deletion conferred growth at 60 µM CCCP, though not at the higher levels resisted by evolved strains (150 µM). Some mprA mutant strains also had point mutations affecting emrA, but deletion of emrA abolished the CCCP resistance. Thus, CCCP-evolved isolates contained additional adaptations. One isolate lacked emrA or mprA mutations but had mutations in cecR (ybiH), whose product upregulates drug pumps YbhG and YbhFSR, and in gadE, which upregulates the multidrug pump MdtEF. A cecR::kanR deletion conferred partial resistance to CCCP. Other multidrug efflux genes that had mutations included ybhR and acrAB The acrB isolate was sensitive to the AcrAB substrates chloramphenicol and tetracycline. Other mutant genes in CCCP-evolved strains include rng (RNase G) and cyaA (adenylate cyclase). Overall, experimental evolution revealed a CCCP-dependent fitness advantage for mutations increasing CCCP efflux via EmrA and for mutations that may deactivate proton-driven pumps for drugs not present (cecR, gadE, acrAB, and ybhR). These results are consistent with our previous report of drug sensitivity associated with evolved benzoate tolerance.IMPORTANCE The genetic responses of bacteria to depletion of proton motive force (PMF), and their effects on drug resistance, are poorly understood. PMF drives export of many antibiotics, but the energy cost may decrease fitness when antibiotics are absent. Our evolution experiment reveals genetic mechanisms of adaptation to the PMF uncoupler CCCP, including selection for increased CCCP efflux but also against the expression of PMF-driven pumps for drugs not present. The results have implications for our understanding of the gut microbiome, which experiences high levels of organic acids that decrease PMF.

Correlation between the number of Pro-Ala repeats in the EmrA homologue of Acinetobacter baumannii and resistance to netilmicin, tobramycin, imipenem and ceftazidime.

Acinetobacter baumannii coccobacilli are dangerous to patients in intensive care units because of their multidrug resistance to antibiotics, developed mainly in the past decade. This study aimed to examine whether there is a significant correlation between the number of Pro-Ala repeats in the CAP01997 protein, the EmrA homologue of A. baumannii, and resistance to antibiotics. A total of 79 multidrug-resistant A. baumannii strains isolated from patients were analysed. Resistance to antibiotics was determined on Mueller-Hinton agar plates using the Kirby-Bauer disk diffusion method. The number of CCTGCA repeats encoding Pro-Ala repeats in CAP01997 was determined by PCR and capillary electrophoresis. The 3D models of CAP01997 containing Pro-Ala repeats were initially generated using RaptorX Structure Prediction server and were assembled with EasyModeller 4.0. The models were embedded in a model bacterial membrane based on structural information from homologous proteins and were refined using 100-ns molecular dynamics simulations. The results of this research show significant correlation between susceptibility to netilmicin, tobramycin and imipenem and the number of repeated Pro-Ala sequences in the CAP01997 protein, a homologue of the Escherichia coli transporter EmrA. Predicted structures suggest potential mechanisms that confer drug resistance by reshaping the cytoplasmic interface between CAP01997 protein and the critical component of the multidrug efflux pump homologous to EmrB. Based on these results, we can conclude that the CAP01997 protein, an EmrA homologue of A. baumannii, confers resistance to netilmicin, tobramycin and imipenem, depending on the number of Pro-Ala repeats.

Signal transducer and activator of transcription 3 (STAT3) mutations underlying autosomal dominant hyper-IgE syndrome impair human CD8(+) T-cell memory formation and function.

BACKGROUND: The capacity of CD8(+) T cells to control infections and mediate antitumor immunity requires the development and survival of effector and memory cells. IL-21 has emerged as a potent inducer of CD8(+) T-cell effector function and memory development in mouse models of infectious disease. However, the role of IL-21 and associated signaling pathways in protective CD8(+) T-cell immunity in human subjects is unknown. OBJECTIVE: We sought to determine which signaling pathways mediate the effects of IL-21 on human CD8(+) T cells and whether defects in these pathways contribute to disease pathogenesis in patients with primary immunodeficiencies caused by mutations in components of the IL-21 signaling cascade. METHODS: Human primary immunodeficiencies resulting from monogenic mutations provide a unique opportunity to assess the requirement for particular molecules in regulating human lymphocyte function. Lymphocytes from patients with loss-of-function mutations in signal transducer and activator of transcription 1 (STAT1), STAT3, or IL-21 receptor (IL21R) were used to assess the respective roles of these genes in human CD8(+) T-cell differentiation in vivo and in vitro. RESULTS: Mutations in STAT3 and IL21R, but not STAT1, led to a decrease in multiple memory CD8(+) T-cell subsets in vivo, indicating that STAT3 signaling, possibly downstream of IL-21R, regulates the memory cell pool. Furthermore, STAT3 was important for inducing the lytic machinery in IL-21-stimulated naive CD8(+) T cells. However, this defect was overcome by T-cell receptor engagement. CONCLUSION: The IL-21R/STAT3 pathway is required for many aspects of human CD8(+) T-cell behavior but in some cases can be compensated by other signals. This helps explain the relatively mild susceptibility to viral disease observed in STAT3- and IL-21R-deficient subjects.