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Members of the Protein kinases D (PKD) family are described as regulators of T cell responses. From the two T cell‐expressed isoforms PKD2 and PKD3, so far mainly the former was thoroughly investigated and is well understood. Recently, we have investigated also PKD3 using conventional as well as conditional T cell‐specific knockout models. These studies suggested PKD3 to be a T cell‐extrinsic regulator of the cells' fate. However, these former model systems did not take into account possible redundancies with the highly homologous PKD2. To overcome this issue and thus properly unravel PKD3's T cell‐intrinsic functions, here we additionally used a mouse model overexpressing a constitutively active isoform of PKD3 specifically in the T cell compartment. These transgenic mice showed a slightly higher proportion of central memory T cells in secondary lymphoid organs and blood. This effect could not be explained via differences upon polyclonal stimulation in vitro, however, may be connected to the observed developmental aberrances in the CD8 single positive compartment during thymic development. Lastly, the observed alterations in the CD8+ T cell compartment did not impact proper immune response upon immunization with ovalbumin or in a subcutaneous tumour model suggesting only a small to absent biological relevance. Taking together the knowledge of all our published studies on PKD3 in the T cell compartment, we now conclude that T cell‐intrinsic PKD3 is a fine‐tuner of central memory T cell as well as CD8 single positive thymocyte development. [ABSTRACT FROM AUTHOR]

Human immunodeficiency virus (HIV) infects CD4+ cells and causes progressive immune function failure, and CD8+ cells lyse infected CD4+ cell via recognising peptide presented by human leukocyte antigens (HLA). Variations in HLA allele lead to observed different HIV infection outcomes. Within-host HIV dynamics involves virus replication within infected cells and lysing of infected cells by CD8+ cells, but how variations in HLA alleles determine different infection outcomes was far from clear. Here, we used mathematical modelling and parameter inference with a new analysis of published virus inhibition assay data to estimate CD8+ cell lysing efficiency, and found that lysing efficiency fall in the gap between low bound (0.1–0.2 day−1 (Elemans et al. in PLoS Comput Biol 8(2):e1002381, 2012)) and upper boundary (6.5–8.4 day−1 (Wick et al. in J Virol 79(21):13579–13586, 2005)). Our outcomes indicate that both lysing efficiency and viral inoculum size jointly determine observed different infection outcomes. Low lysing rate associated with non-protective HLA alleles leads to monostable viral kinetic to high viral titre and oscillatory viral kinetics. High lysing rate associated with protective HLA alleles leads monostable viral kinetic to low viral titre and bistable viral kinetics; at a specific interval of CD8+ cell counts, small viral inoculum sizes are inhibited but not large viral inoculum sizes remain infectious. Further, with CD8+ cell recruitment, HIV kinetics always exhibit oscillatory kinetics, but lysing rate is negatively correlated with range of CD8+ cell count. Our finding highlights role of HLA allele determining different infection outcomes, thereby providing a potential mechanistic explanation for observed good and bad HIV infection outcomes induced by protective HLA allele. [ABSTRACT FROM AUTHOR]

Background: To evaluate the efficacy and safety of allogenic CD8 + natural killer T (CD8+ NKT) immunotherapy combined with gefitinib in the treatment of advanced or metastatic EGFR mutant non-small cell lung cancer (NSCLC). Methods: This study is prospective. The NSCLC patients with exon 19 (Ex19del) or exon 21 L858R point mutations, and response to gefitinib treatment were enrolled into the trial to be randomly assigned into the gefitinib arm and the gefitinib/NKT arm. Allogenic CD8+ NKT cells were cultured in vitro and adaptive transferred into the patients via vein in the gefitinib/NKT arm. The primary endpoint was progression-free survival (PFS). Secondary endpoint analysis included time to disease progression (TTP), overall survival (OS), levels of serum tumour markers for carcinoembryonic antigen (CEA) and alanine aminotransferase (ALT) in the blood, the response rate and safety. From July 2017 to June 2021, 19 patients were randomly assigned to the gefitinib arm (n = 8) and the gefitinib/NKT arm (n = 11). Results: The estimated median survival PFS in the gefitinib/NKT arm was significantly longer than that of the gefitinib arm (12 months vs 7 months). Similar results were also observed for the median TTP. Moreover, the gefitinib/NKT arm had better CEA control than the gefitinib arm. Clinical grade 3 adverse reactions occurred in 64% and 39% of patients in the gefitinib/NKT arm and the gefitinib arm, respectively. The most common grade 3 adverse events in the gefitinib/NKT arm included abnormal liver function in 8 cases (73%) and diarrhoea in 1 case (9%), both of which resolved after drug intervention. Conclusion: The PFS of EGFR-mutated advanced NSCLC treated with allogenic CD8+ NKT cells combined with gefitinib was longer than that of gefitinib alone. No obvious serious adverse reactions occurred, and the patients compliance and survival status were good. [ABSTRACT FROM AUTHOR]

Although proteomics is extensively used in immune research, there is currently no publicly accessible spectral assay library for the comprehensive proteome of immune cells. This study generated spectral assay libraries for five human immune cell lines and four primary immune cells: CD4 T, CD8 T, natural killer (NK) cells, and B cells. This was achieved by utilizing data-dependent acquisition (DDA) and employing fractionated samples from over 100 µg of proteins, which was applied to acquire the highest-quality MS/MS spectral data. In addition, Data-indedendent acquisition (DIA) was used to obtain sufficient data points for analyzing proteins from 10,000 primary CD4 T, CD8 T, NK, and B cells. The immune cell spectral assay library generated included 10,544 protein groups and 127,106 peptides. The proteomic profiles of 10,000 primary human immune cells obtained from 15 healthy volunteers analyzed using DIA revealed the highest heterogeneity of B cells among other immune cell types and the similarity between CD4 T and CD8 T cells. All data and spectral library are deposited in ProteomeXchange (PXD047742). [ABSTRACT FROM AUTHOR]

This research was conducted to examine the impact of Wei Qi Booster (WQB) on immune parameters and anti-oxidative function in aged mice. Fifty aged mice were randomly assigned to five different groups. Group A was designated as the control group. Mice in Group B were receiving Levamisole at 10  mg/kg body weight. Each mouse in groups C, D and E received 0.1, 1, and 2% WQB, respectively. Another ten young mice, designated as group F, were fed regularly. The mice were fed according to the above methods for 28  days. Results showed that relative to the control group, the body weight and immune organs indexes experienced a substantial rise in the group with 1% WQB. In addition, 1% WQB could improve the activity of SOD and reduce the MDA levels. Expressions of CD4 and sIgA increased while CD8 decreased in the jejunum of aged mice treated with WQB. IL2 and IFN-γ levels increased in the 1% WQB group, showing no notable difference compared to the young mice group. The results demonstrated that WQB can elevate immune levels and enhance anti-oxidative functions in aged mice. [ABSTRACT FROM AUTHOR]

Objective: This study aimed to investigate the expression of GPR56 in the T cells of early-stage lung adenocarcinoma (LUAD) patients and clarify its diagnostic significance. Methods: Blood samples were collected from 32 patients with stage IA LUAD and 31 healthy controls. GPR56 and perforin were analysed in circulating T-cell subsets by flow cytometry. In addition, a correlation between perforin and GPR56 expression was detected. Changes in GPR56+ cells in early LUAD patients were analysed, and the diagnostic significance of GPR56+ T cells for early LUAD was studied by receiver operating characteristic (ROC) curve analysis. Results: The expression of GPR56 in CD8+ T cells from early-stage LUAD patients was significantly greater than that in CD4+ T cells. The percentage of perforin-positive GPR56+ cells in early-stage LUAD patients was high. GPR56 levels in the T cells of LUAD patients were significantly lower than those in healthy controls. ROC analysis revealed that the area under the curve for the percentage of GPR56-positive CD8+ TEMRA cells to distinguish early-stage LUAD patients from healthy individuals- reached 0.7978. Conclusion: The decreased expression of GPR56 in the peripheral blood of early-stage LUAD patients correlated with perforin levels, reflecting compromised antitumor immunity and aiding early-stage LUAD screening. [ABSTRACT FROM AUTHOR]

Despite the numerous studies on the clinical aspects of early-onset preeclampsia, our understanding of the immunological consequences of inadequate placenta development remains incomplete. The Th1-predominance characteristic of early-onset preeclampsia significantly impacts maternal immunotolerance, and the role of immune checkpoint molecules in these mechanisms is yet to be fully elucidated. Our study aims to fill these crucial knowledge gaps. A total of 34 pregnant women diagnosed with early-onset preeclampsia and 34 healthy pregnant women were enrolled in this study. A mononuclear cell fragment from the venous blood was separated and frozen. The CD8+ and CD8− NK cell subpopulations were identified and compared to their immune checkpoint molecule expressions using multicolor flow cytometry. The serum CD226 levels were measured by ELISA. Based on our measures, the frequency of the CD8− subpopulation was significantly higher than that of the CD8+ counterpart in both the NKdim and NKbright subsets. Significantly lower CD226 surface expressions were detected in the preeclamptic group compared to healthy women in all the investigated subpopulations. However, while no difference was observed in the level of the soluble CD226 molecule between the two groups, the CD112 and CD155 surface expressions were significantly different. Our study's findings underscore the significant role of the CD8+ and CD8− NK subpopulations in the Th1-dominated immune environment. This deepens our understanding of early-onset preeclampsia and suggests that each subpopulation could contribute to the compensation mechanisms and the restoration of the immunological balance in this condition, a crucial step toward developing effective interventions. [ABSTRACT FROM AUTHOR]

Transplantation of human-induced pluripotent stem cell-derived cardiomyocytes (hiPS-CMs) has emerged as a promising therapy to treat end-stage heart failure. However, the immunogenicity of hiPS-CMs in transplanted patients has not been fully elucidated. Thus, in vivo models are required to estimate immune responses against hiPS-CMs in transplant recipients. We transferred human peripheral blood mononuclear cells (hPBMCs) into NOD/Shi- scid IL-2rg null (NOG) MHC class I/II double knockout (NOG-ΔMHC) mice, which were reported to accept hPBMCs without xenogeneic-graft- versus -host disease (xeno-GVHD). Then, hiPS-CM sheets generated from the hiPS cell line 201B7 harboring a luciferase transgene were transplanted into the subcutaneous space of NOG-ΔMHC mice. Graft survival was monitored by bioluminescent images using a Xenogen In Vivo Imaging System. The human immune cells were engrafted for more than 3 months in NOG-ΔMHC mice without lethal xeno-GVHD. The hiPS-CMs expressed a moderate level of human leukocyte antigen (HLA)-class I, but not HLA-class II, molecules even after interferon-gamma (IFN-γ) stimulation. Consistently, the allogenic IFN-γ-treated hiPS-CMs induced weak CD8+ but not CD4+ T cell responses in vitro. hiPS-CM sheets disappeared approximately 17 to 24 days after transplantation in hPBMC-transferred NOG-ΔMHC mice, and CD8+ T cell depletion significantly prolonged graft survival, similar to what was observed following tacrolimus treatment. hiPS-CMs are less immunogenic in vitro but induce sufficient CD8+ T cell-mediated immune responses for graft rejection in vivo. [ABSTRACT FROM AUTHOR]

Background: T lymphocytes in tumor microenvironment play a pivotal role in the anti-tumor immunity, and the memory of T cells contributes to the longterm protection against tumor antigens. Compared to solid tumors, studies focusing on the T-cell differentiation in the acute myeloid leukemia (AML) bone marrow (BM) microenvironment remain limited. Patients and methods: Fresh BM specimens collected from 103 adult AML patients at diagnosis and 12 healthy donors (HDs) were tested T-cell differentiation subsets by multi-parameter flow cytometry. Results: CD4 and CD8 T-cell compartments had different constituted profiles of T-cell differentiated subsets, which was similar between AML patients and HDs. Compared to HDs, AML patients as a whole had a significantly higher proportion of CD8 effector T cells (Teff, P = 0.048). Moreover, the T-cell compartment of AML patients with no DNMT3A mutations skewed toward terminal differentiation at the expense of memory T cells (CD4 Teff: P = 0.034; CD8 Teff: P = 0.030; CD8 memory T: P = 0.017), whereas those with mutated DNMT3A had a decrease in CD8 naïve T (Tn) and CD4 effector memory T cells (Tem) as well as an increase in CD4 central memory T cells (Tcm) (P = 0.037, 0.053 and 0.053). Adverse ELN genetic risk correlated with a lower proportion of CD8 Tn. In addition, the low proportions of CD4 Tem and CD8 Tn independently predicted poorer relapsefree survival (RFS, HR [95%CI]: 5.7 (1.4-22.2), P = 0.017 and 4.8 [1.3-17.4], P = 0.013) and event-free survival (EFS, HR [95% CI]: 3.3 (1.1-9.5), P = 0.029; 4.0 (1.4-11.5), P = 0.010), respectively. Conclusions: AML patients had abnormal profiles of BM T-cell differentiation subsets at diagnosis, which was related to DNMT3A mutations. The low proportions of CD4 Tem and CD8 Tn predicted poor outcomes. [ABSTRACT FROM AUTHOR]

Background: T cells play a pivotal role in chemotherapy-triggered anti-tumor effects. Emerging evidence underscores the link between impaired anti-tumor immune responses and resistance to paclitaxel therapy in triple-negative breast cancer (TNBC). Tumor-related endothelial cells (ECs) have potential immunoregulatory activity. However, how ECs regulate T cell activity during TNBC chemotherapy remains poorly understood. Methods: Single-cell analysis of ECs in patients with TNBC receiving paclitaxel therapy was performed using an accessible single-cell RNA sequencing (scRNA-seq) dataset to identify key EC subtypes and their immune characteristics. An integrated analysis of a tumor-bearing mouse model, immunofluorescence, and a spatial transcriptome dataset revealed the spatial relationship between ECs, especially Tumor necrosis factor receptor (TNFR) 2+ ECs, and CD8+ T cells. RNA sequencing, CD8+ T cell proliferation assays, flow cytometry, and bioinformatic analyses were performed to explore the immunosuppressive function of TNFR2 in ECs. The downstream metabolic mechanism of TNFR2 was further investigated using RNA sequencing, cellular glycolysis assays, and western blotting. Results: In this study, we identified an immunoregulatory EC subtype, characterized by enhanced TNFR2 expression in non-responders. By a mouse model of TNBC, we revealed a dynamic reduction in the proportion of the CD8+ T cell-contacting tumor vessels that could co-localize spatially with CD8+ T cells during chemotherapy and an increased expression of TNFR2 by ECs. TNFR2 suppresses glycolytic activity in ECs by activating NF-κB signaling in vitro. Tuning endothelial glycolysis enhances programmed death-ligand (PD-L) 1-dependent inhibitory capacity, thereby inducing CD8+ T cell suppression. In addition, TNFR2+ ECs showed a greater spatial affinity for exhausted CD8+ T cells than for non-exhausted CD8+ T cells. TNFR2 blockade restores impaired anti-tumor immunity in vivo, leading to the loss of PD-L1 expression by ECs and enhancement of CD8+ T cell infiltration into the tumors. Conclusions: These findings reveal the suppression of CD8+ T cells by ECs in chemoresistance and indicate the critical role of TNFR2 in driving the immunosuppressive capacity of ECs via tuning glycolysis. Targeting endothelial TNFR2 may serve as a potent strategy for treating TNBC with paclitaxel. [ABSTRACT FROM AUTHOR]

Introduction: Older recipient age is associated with a significant decreased risk for rejection after kidney transplantation which is incompletely understood. Methods: In a longitudinal study, circulating alloreactive T cells were assessed of young (≤45 years) and older (≥55 years) stable kidney transplant recipients. Alloreactive T-cells were identified by CD137-expression and phenotype, cytokine producing and proliferative capacity, were evaluated using multiparameter flowcytometry. Results: The results show that before transplantation frequencies of alloreactive CD4+ and CD8+ T-cells in older KT-recipients are significantly higher and shifted towards an effector memory-phenotype. However, the frequency of polyfunctional (≥2 pro-inflammatory cytokines) CD4+ T-cells was significantly lower and less IL2 was produced. The frequency of polyfunctional alloreactive CD4 + T-cells and proliferation of alloreactive T-cells donor-specifically declined after transplantation reaching a nadir at 12 months after transplantation, irrespective of age. A striking difference was observed for the proliferative response of alloreactive CD8+ T-cells. Thiswas not only lower in older compared to younger recipients but could also not be restored by exogenous IL2 or IL15 in the majority of older recipients while the response to polyclonal stimulation was unaffected. Conclusion: In conclusion, older age is associated with a distinct and marked reduction of functionality of both alloreactive CD4+ and CD8+ T-cells. [ABSTRACT FROM AUTHOR]

Antigen-specific cytotoxic CD8 T cells are extremely effective in controlling tumor growth and have been the focus of immunotherapy approaches. We leverage in silico tools to investigate whether the occurrence of mutations in proteins previously described as immunogenic and highly expressed by glioblastoma multiforme (GBM), such as Epidermal Growth Factor Receptor (EGFR), Isocitrate Dehydrogenase 1 (IDH1), Phosphatase and Tensin homolog (PTEN) and Tumor Protein 53 (TP53), may be contributing to the differential presentation of immunogenic epitopes. We recovered Class I MHC binding information from wild-type and mutated proteins using the Immune Epitope Database (IEDB). After that, we built peptide-MHC (pMHC-I) models in HLA-arena, followed by hierarchical clustering analysis based on electrostatic surface features from each complex. We identified point mutations that are determinants for the presentation of a set of peptides from TP53 protein. We point to structural features in the pMHC-I complexes of wild-type and mutated peptides, which may play a role in the recognition of CD8 T cells. To further explore these features, we performed 100 ns molecular dynamics simulations for the peptide pairs (wt/mut) selected. In pursuit of novel therapeutic targets for GBM treatment, we selected peptides where our predictive results indicated that mutations would not disrupt epitope presentation, thereby maintaining a specific CD8 T cell immune response. These peptides hold potential for future GBM interventions, including peptide-based or mRNA vaccine development applications. [ABSTRACT FROM AUTHOR]

Thought of as a metastasis‐associated gene, however, NME/NM23 nucleoside diphosphate kinase 4 (NME4) has rarely been described in the context of the tumour microenvironment. To understand the immunological implications of NME4 in oesophageal squamous cell carcinoma (ESCC), we used multiplex immunohistochemistry to analyse the clinicopathological and prognostic importance of NME4 expression. Then, after establishing a syngeneic tumour model with a C57BL/6 mouse strain that can recapitulate the tumour microenvironment of humans, we examined the immunological involvement of NME4 expression. To explore the underlying molecular mechanism, via quantitative proteomics and protein microarray screening, we investigated the potential signalling pathways involved. The clinicopathological and prognostic importance of NME4 expression is limited in ESCC patients. In vivo, single‐cell RNA sequencing showed that NME4 strikingly prevented CD8+ T cells from infiltrating the tumour microenvironment in murine ESCC. Mechanistically, we mapped out the NFκB2‐CCL5 axis that was negatively controlled by NME4 in the murine ESCC cell line AKR. Collectively, these data demonstrated that regulation of NFκB2‐CCL5 axis by NME4 prevents CD8+ T cells infiltration in ESCC. [ABSTRACT FROM AUTHOR]

Introduction: Alveolar epithelial regeneration depends on the activity of resident quiescent progenitor cells. Alveolar epithelial type II (AT2) cells are known as the alveolar epithelial progenitor cells. They exit quiescent state, proliferate rapidly in response to injury and differentiate into alveolar epithelial type I (AT1) cells to regenerate the damaged alveolar epithelium. Although AT2 cell plasticity has been a very intense field of research, the role of CD8 T cell response and their released cytokine IFN-γ, in regulating AT2 cell plasticity and alveolar epithelial repair and regeneration after injury remains largely unknown. Methods: We used flow cytometry to quantify the amount of CD8 T cells in mouse lungs after bacterial pneumonia caused by Streptococcus pneumoniae. To determine whether CD8 T cells and their released cytokine IFN-γ are necessary for AT2 cell activity during alveolar epithelial regeneration, we performed loss of function studies using anti-CD8 or anti-IFN-γ monoclonal antibody (mAb) treatment in vivo. We assessed the effects of CD8 T cells and cytokine IFN-γ on AT2 cell differentiation capacity using the AT2- CD8 T cell co-culture system in vitro. Results: We detected a transient wave of accumulation of CD8 T cells in mouse lungs, which coincided with the burst of AT2 cell proliferation during alveolar epithelial repair and regeneration in mice following bacterial pneumonia caused by Streptococcus pneumoniae. Depletion of CD8 T cells or neutralization of cytokine IFN-γ using anti-CD8 or anti-IFN-γ monoclonal antibody significantly reduced AT2 cell proliferation and differentiation into AT1 cells in mice after bacterial pneumonia. Furthermore, co-culture of CD8 T cells or cytokine IFN-γ with AT2 cells promoted AT2-to-AT1 cell differentiation in both murine and human systems. Conversely, blockade of IFN-γ signaling abrogated the increase in AT2-to-AT1 cell differentiation in the AT2- CD8 T cell co-culture system. Discussion: Our data demonstrate that CD8 T-cell response and cytokine IFN-γ are necessary for promoting AT2 cell activity during alveolar epithelial repair and regeneration after acute lung injury caused by bacterial pneumonia. [ABSTRACT FROM AUTHOR]

Background: Study of expression of lymphocyte subsets could help to understand the pathophysiology of COVID-19 infection, development of novel treatment protocols and reducing rate of mortality among COVID-19 infected patients. The aim of this work was to correlate expression of lymphocyte subsets with the inflammatory markers and clinical status of patients with COVID-19 infection. Methods: This prospective study was performed at Zagazig University Isolation Hospitals and clinical pathology department, Zagazig, Egypt on 80 persons who were enrolled in this study; 40 COVID-19 patients with pneumonia made up group I, and the following individuals made up group II: 40 COVID-19 patients without pneumonia. Immunophenotyping of CD4 CD8 were measured by Flow Cytometry in both groups. Results: Patients had a considerable drop in CD4 CD8 levels with pneumonia than those without pneumonia Conclusions: Low levels of CD4 CD8 in COVID-19 patients are associated with a higher risk of pneumonia. [ABSTRACT FROM AUTHOR]

Staphylococcal toxic shock syndrome (STSS) is a rare, yet potentially fatal disease caused by Staphylococcus aureus (S. aureus) enterotoxins, known as superantigens, which trigger an intense immune response. Our previous study demonstrated the protective effect of tofacitinib against murine toxin-induced shock and a beneficial effect against S. aureus sepsis. In the current study, we examined the effects of tofacitinib on T-cell response in peripheral blood using a mouse model of enterotoxin-induced shock. Our data revealed that tofacitinib suppresses the activation of both CD4+ and CD8+ T cells in peripheral blood. Furthermore, both gene and protein levels of Th1 cytokines were downregulated by tofacitinib treatment in mice with enterotoxin-induced shock. Importantly, we demonstrated that CD4+ cells, but not CD8+ cells, are pathogenic in mice with enterotoxin-induced shock. In conclusion, our findings suggest that tofacitinib treatment suppresses CD4+ T-cell activation and Th1 response, thereby aiding in protection against staphylococcal toxic shock in mice. This insight may guide the future development of novel therapies for STSS. [ABSTRACT FROM AUTHOR]

Background: Infants with frequent viral and bacterial respiratory infections exhibit compromised immunity to routine immunizations. They are also more likely to develop chronic respiratory diseases in later childhood. This study investigated the feasibility of epigenetic profiling to reveal endotype-specific molecular pathways with potential for early identification and immuno-modulation. Peripheral blood mononuclear cells from respiratory infection allergy/asthma-prone (IAP) infants and non-infection allergy/asthma prone (NIAP) were retrospectively selected for genome-wide DNA methylation and single nucleotide polymorphism analysis. The IAP infants were enriched for the low vaccine responsiveness (LVR) phenotype (Fisher's exact p-value = 0.02). Results: An endotype signature of 813 differentially methylated regions (DMRs) comprising 238 lead CpG associations (FDR < 0.05) emerged, implicating pathways related to asthma, mucin production, antigen presentation and inflammasome activation. Allelic variation explained only a minor portion of this signature. Stimulation of mononuclear cells with monophosphoryl lipid A (MPL), a TLR agonist, partially reversed this signature at a subset of CpGs, suggesting the potential for epigenetic remodeling. Conclusions: This proof-of-concept study establishes a foundation for precision endotyping of IAP children and highlights the potential for immune modulation strategies using adjuvants for future investigation. [ABSTRACT FROM AUTHOR]

Targeting intracellular inhibiting proteins has been revealed to be a promising strategy to improve CD8+ T cell anti-tumor efficacy. Here, we are focusing on intracellular inhibiting proteins specific to TCR signaling: DOK1 and DOK2 expressed in T cells. We hypothesized that depletion of intracellular inhibition checkpoint DOK1 and DOK2 could improve CD8+ T-cell based cancer therapies. To evaluate the role of DOK1 and DOK2 depletion in physiology and effector function of CD8+ T lymphocytes and in cancer progression, we established a transgenic T cell receptor mouse model specific to melanoma antigen hgp100 (pmel-1 TCR Tg) in WT and Dok1/Dok2 DKO (double KO) mice. We showed that both DOK1 and DOK2 depletion in CD8+ T cells after an in vitro pre-stimulation induced a higher percentage of effector memory T cells as well as an up regulation of TCR signaling cascade- induced by CD3 mAbs, including the increased levels of pAKT and pERK, two major phosphoproteins involved in T cell functions. Interestingly, this improved TCR signaling was not observed in naïve CD8+ T cells. Despite this enhanced TCR signaling essentially shown upon stimulation via CD3 mAbs, pre-stimulated Dok1/Dok2 DKO CD8+ T cells did not show any increase in their activation or cytotoxic capacities against melanoma cell line expressing hgp100 in vitro. Altogether we demonstrate here a novel aspect of the negative regulation by DOK1 and DOK2 proteins in CD8+ T cells. Indeed, our results allow us to conclude that DOK1 and DOK2 have an inhibitory role following long term T cell stimulations. [ABSTRACT FROM AUTHOR]

Acute lung allograft dysfunction (ALAD) is an imprecise syndrome denoting concern for the onset of chronic lung allograft dysfunction (CLAD). Mechanistic biomarkers are needed that stratify risk of ALAD progression to CLAD. We hypothesized that single cell investigation of bronchoalveolar lavage (BAL) cells at the time of ALAD would identify immune cells linked to progressive graft dysfunction. We prospectively collected BAL from consenting lung transplant recipients for single cell RNA sequencing. ALAD was defined by a ≥10% decrease in FEV 1 not caused by infection or acute rejection and samples were matched to BAL from recipients with stable lung function. We examined cell compositional and transcriptional differences across control, ALAD with decline, and ALAD with recovery groups. We also assessed cell-cell communication. BAL was assessed for 17 ALAD cases with subsequent decline (ALAD declined), 13 ALAD cases that resolved (ALAD recovered), and 15 cases with stable lung function. We observed broad differences in frequencies of the 26 unique cell populations across groups (p = 0.02). A CD8 T cell (p = 0.04) and a macrophage cluster (p = 0.01) best identified ALAD declined from the ALAD recovered and stable groups. This macrophage cluster was distinguished by an anti-inflammatory signature and the CD8 T cell cluster resembled a Tissue Resident Memory subset. Anti-inflammatory macrophages signaled to activated CD8 T cells via class I HLA, fibronectin, and galectin pathways (p < 0.05 for each). Recipients with discordance between these cells had a nearly 5-fold increased risk of severe graft dysfunction or death (HR 4.6, 95% CI 1.1–19.2, adjusted p = 0.03). We validated these key findings in 2 public lung transplant genomic datasets. BAL anti-inflammatory macrophages may protect against CLAD by suppressing CD8 T cells. These populations merit functional and longitudinal assessment in additional cohorts. [ABSTRACT FROM AUTHOR]

Advances in understanding the pathological mechanisms of breast cancer have resulted in the emergence of novel therapeutic strategies. However, triple-negative breast cancer (TNBC), a molecular subtype of breast cancer with a poor prognosis, lacks classical and general therapeutic targets, hindering the clinical application of several therapies to breast cancer. As insights into the unique immunity and molecular mechanisms of TNBC have become more extensive, immunotherapy has gradually become a valuable complementary approach to classical radiotherapy and chemotherapy. CD8+ cells are significant actors in the tumor immunity cycle; thus, research on TNBC immunotherapy is increasingly focused in this direction. Recently, CD8+ tissue-resident memory (TRM) cells, a subpopulation of CD8+ cells, have been explored in relation to breast cancer and found to seemingly play an undeniably important role in tumor surveillance and lymphocytic infiltration. In this review, we summarize the recent advances in the mechanisms and relative targets of CD8+ T cells, and discuss the features and potential applications of CD8+ TRM cells in non-luminal breast cancer immunotherapy. [ABSTRACT FROM AUTHOR]

Purpose: Hepatocellular carcinoma has become one of the severe diseases threatening human health. T cell exhaustion is deemed as a reason for immunotherapy resistance. However, little is known about the roles of CD8 Tex-related lncRNAs in HCC. Materials and Methods: We processed single-cell RNA sequencing to identify CD8 Tex-related genes. CD8 Tex-related lncRNAs were identified based on their correlations with mRNAs. Unsupervised clustering approach was used to identify molecular clusters of CD8 Tex-related lncRNAs. Differences in prognosis and immune infiltration between the clusters were explored. Machine learning algorithms were used to construct a prognostic signature. Samples were classified as low- and high-risk groups based on their risk scores. We identified prognosis-related lncRNAs and constructed a ceRNA network. In vitro experiments were conducted to investigate the impacts of CD8 Tex-related lncRNAs on proliferation and apoptosis of HCC cells. Results: We clarified cell types within two HCC single-cell datasets. We identified specific markers of CD8 Tex cells and analyzed their potential functions. Twenty-eight lncRNAs were identified as CD8 Tex-related. Based on CD8 Tex-related lncRNAs, samples were categorized into two distinct clusters, which exhibited significant differences in survival rates and immune infiltration. Ninety-six algorithm combinations were employed to establish a prognostic signature. RSF emerged as the one with the highest C-index. Patients in high- and low-risk groups exhibited marked differences in prognosis, enriched pathways, mutations and drug sensitivities. MCM3AP-AS1, MAPKAPK5-AS1 and PART1 were regarded as prognosis-related lncRNAs. A ceRNA network was constructed based on CD8 Tex-related lncRNAs and mRNAs. Experiments on cell lines and organoids indicated that downregulation of MCM3AP-AS1, MAPKAPK5-AS1 and PART1 suppressed cell proliferation and induced apoptosis. Conclusion: CD8 Tex-related lncRNAs played crucial roles in HCC progression. Our findings provided new insights into the regulatory mechanisms of CD8 Tex-related lncRNAs in HCC. [ABSTRACT FROM AUTHOR]

Background/Aim: The activation of phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway has been implicated in canine soft tissue sarcoma (STS) and may serve as a prognostic marker. This study investigated the correlation between PI3K/Akt activation in tumor cells and tumorinfiltrating lymphocytes (TILs). Materials and Methods: A total of 59 STS samples were labeled via immunohistochemistry to calculate the density of TILs, including CD3+ T cells, CD8+ T cells, CD20+ B cells, and FOXP3+ regulatory T cells. Results: Forty-eight samples (81.3%) had intra-tumoral TILs with a high density of CD3+ T cells (mean: 283.3 cells/mm²) and CD8+ T cells (mean: 134.8 cells/mm²). Conversely, CD20+ B cells (mean: 73.6 cells/mm²) and FOXP3+ regulatory T cells (mean: 9.2 cells/mm²) were scarce. The abundance of CD3+/CD8+, CD3+/CD20+, and CD8+/CD20+ TILs were highly correlated in multivariate analyses (r=0.895, 0.946, and 0.856, respectively). Nonetheless, TIL density was unrelated to clinicopathological parameters (sex, age, tumor location, breed) and tumor grade. The abundance of CD8+ T cells was positively correlated with the activation of PI3K/Akt, indicating that samples with high levels of phospho-Akt and phospho-S6 tend to have a higher CD8+ T cell density (p=0.0032 and 0.0218, respectively). Furthermore, TIL density was correlated with the Ki-67 index, a tumor proliferation and growth marker. Samples with a high Ki-67 index had a significantly higher abundance of CD3+ T cells, CD8+ T cells, and CD20+ B cells (p=0.0392, 0.0254, 0.0380, respectively). Conclusion: PI3K/Akt pathway activation may influence the infiltration of CD8+ T cells within the tumor microenvironment in canine STS. Prospective studies involving a higher number of cases are warranted to confirm these findings. [ABSTRACT FROM AUTHOR]

Human cytomegalovirus (hCMV) is a ubiquitous latent persistent herpesvirus infecting 60–90% of the population worldwide. hCMV carriage in immunocompetent people is asymptomatic; thus, hCMV can be considered a component of normative aging. However, hCMV powerfully modulates many features of the immune, and likely other, systems and organs. Questions remain as to how hCMV carriage affects the human host. We used anti-CMV antibody titers as a stratifying criterion to examine the impact of "intensity" of hCMV infection as a potential biomarker of aging, inflammation, and immune homeostasis in a cohort of 247 participants stratified into younger (21–40 years) and older (> 65 years of age) groups. We showed that anti-CMV antibody titers increased with age and directly correlated to increased levels of soluble tumor necrosis factor (sTNFR) I in younger but not older participants. CD8 + cell numbers were reduced in the older group due to the loss in CD8 + T naïve (Tn) cells. In CMV carriers and, in particular, in anti-CMV Ab-high participants, this loss was mitigated or reversed by an increase in the numbers of CD8 + T effector memory (Tem) and T effector memory reexpressing CD45RA (Temra) cells. Analysis of CD38, HLA-DR, and CD57 expression revealed subset (CD4 or CD8)-specific changes that correlated with anti-CMV Ab levels. In addition, anti-CMV Ab levels predicted anti-CMV CD8 T cell responsiveness to different CMV open reading frames (ORFs) selectively in older participants, which correlated to the transcriptional order of expression of specific CMV ORFs. Implications of these results for the potential predictive value of anti-CMV Ab titers during aging are discussed. [ABSTRACT FROM AUTHOR]

The explicit identification of CD8+ T cell subpopulation is important for deciphering the role of CD8+ T cells for protecting our body against invading pathogens and cancer. Our generated monoclonal antibody (mAb), named FE‐1H10, recognized two novel subpopulations of peripheral blood CD8+ T cells, FE‐1H10+ and FE‐1H10− CD8+ T cells. The molecule recognized by mAb FE‐1H10 (FE‐1H10 molecules) had a higher distribution on effector memory CD8+ T cell subsets. The functions of FE‐1H10− and FE‐1H10+ CD8+ T cells were investigated. T cell proliferation assays revealed that FE‐1H10− CD8+ T cells exhibited a higher proliferation rate than FE‐1H10+ CD8+ T cells, whereas FE‐1H10+ CD8+ T cells produced higher levels of IFN‐γ and TNF‐α than FE‐1H10− CD8+ T cells. In T cell cytotoxicity assays, FE‐1H10+ CD8+ T cells were able to kill target cells better than FE‐1H10− CD8+ T cells. RNA‐sequencing analysis confirmed that these subpopulations were distinct: FE‐1H10+ CD8+ T cells have higher expression of genes involved in effector functions (IFNG, TNF, GZMB, PRF1, GNLY, FASL, CX3CR1) while FE‐1H10− CD8+ T cells have greater expression of genes related to memory CD8+ T cell populations (CCR7, SELL, TCF7, CD40LG). The results suggested that mAb FE‐1H10 identifies two novel distinctive CD8+ T cell subpopulations. The FE‐1H10+ CD8+ T cells carried a superior functionality in response to tumour cells. The uncover of these novel CD8+ T cell subpopulations may be the basis knowledge of an optional immunotherapy for the selection of potential CD8+ T cells in cancer treatment. [ABSTRACT FROM AUTHOR]

Psoriasis is a chronic inflammatory skin disease, the etiology of which has not been fully elucidated, in which CD8+ T cells play an important role in the pathogenesis of psoriasis. However, there is a lack of in-depth studies on the molecular characterization of different CD8+ T cell subtypes and their role in the pathogenesis of psoriasis. This study aims to further expound the pathogenesy of psoriasis at the single-cell level and to explore new ideas for clinical diagnosis and new therapeutic targets. Our study identified a unique subpopulation of CD8+ T cells highly infiltrated in psoriasis lesions. Subsequently, we analyzed the hub genes of the psoriasis-specific CD8+ T cell subpopulation using hdWGCNA and constructed a machine-learning prediction model, which demonstrated good efficacy. The model interpretation showed the influence of each independent variable in the model decision. Finally, we deployed the machine learning model to an online website to facilitate its clinical transformation. [ABSTRACT FROM AUTHOR]

Background: Immune-positron emission tomography (PET) imaging with tracers that target CD8 and granzyme B has shown promise in predicting the therapeutic response following immune checkpoint blockade (ICB) in immunologically "hot" tumors. However, immune dynamics in the low T-cell infiltrating "cold" tumor immune microenvironment during ICB remain poorly understood. This study uses molecular imaging to evaluate changes in CD4 + T cells and CD8 + T cells during ICB in breast cancer models and examines biomarkers of response. Methods: [89Zr]Zr-DFO-CD4 and [89Zr]Zr-DFO-CD8 radiotracers were used to quantify changes in intratumoral and splenic CD4 T cells and CD8 T cells in response to ICB treatment in 4T1 and MMTV-HER2 mouse models, which represent immunologically "cold" tumors. A correlation between PET quantification metrics and long-term anti-tumor response was observed. Further biological validation was obtained by autoradiography and immunofluorescence. Results: Following ICB treatment, an increase in the CD8-specific PET signal was observed within 6 days, and an increase in the CD4-specific PET signal was observed within 2 days in tumors that eventually responded to immunotherapy, while no significant differences in CD4 or CD8 were found at the baseline of treatment that differentiated responders from nonresponders. Furthermore, mice whose tumors responded to ICB had a lower CD8 PET signal in the spleen and a higher CD4 PET signal in the spleen compared to non-responders. Intratumoral spatial heterogeneity of the CD8 and CD4-specific PET signals was lower in responders compared to non-responders. Finally, PET imaging, autoradiography, and immunofluorescence signals were correlated when comparing in vivo imaging to ex vivo validations. Conclusions: CD4- and CD8-specific immuno-PET imaging can be used to characterize the in vivo distribution of CD4 + and CD8 + T cells in response to immune checkpoint blockade. Imaging metrics that describe the overall levels and distribution of CD8 + T cells and CD4 + T cells can provide insight into immunological alterations, predict biomarkers of response to immunotherapy, and guide clinical decision-making in those tumors where the kinetics of the response differ. [ABSTRACT FROM AUTHOR]

CD8 + T cells exert a critical role in eliminating cancers and chronic infections, and can provide long-term protective immunity. However, under the exposure of persistent antigen, CD8 + T cells can differentiate into terminally exhausted CD8 + T cells and lose the ability of immune surveillance and disease clearance. New insights into the molecular mechanisms of T-cell exhaustion suggest that it is a potential way to improve the efficacy of immunotherapy by restoring the function of exhausted CD8 + T cells. Transforming growth factor-β (TGF-β) is an important executor of immune homeostasis and tolerance, inhibiting the expansion and function of many components of the immune system. Recent studies have shown that TGF-β is one of the drivers for the development of exhausted CD8 + T cells. In this review, we summarized the role and mechanisms of TGF-β in the formation of exhausted CD8 + T cells and discussed ways to target those to ultimately enhance the efficacy of immunotherapy. [ABSTRACT FROM AUTHOR]

CD8+ T lymphocytes are critical in the immune response against neoplasms, yet the prognostic relevance of CD8+ T cell-associated genes in hepatocellular carcinoma (HCC) is not fully understood. We sourced single-cell RNA-sequencing (scRNA-seq) and bulk RNA-seq data for HCC from the GSE98638 dataset and The Cancer Genome Atlas (TCGA) repository. We utilized Weighted Gene Correlation Network Analysis (WGCNA) to identify CD8+ T cell-related genes. A clinical prognostic model for risk stratification was then constructed via Cox-Lasso regression analysis. The Immunophenotypic Score (IPS) was utilized to evaluate the potential of immunotherapeutic interventions in the categorized cohorts. Validation of the expression of CD8+ T cell-associated risk genes was performed using quantitative reverse transcription PCR (qRT-PCR). Integrating scRNA-seq with RNA-seq data, we identified five CD8+ T cell-related signature genes: IKBKE, ATP1B3, MSC, ADA, and BATF. Notably, HCC patients in the high-risk group had markedly decreased overall survival. Elevated infiltration levels of CD8+ T cells, B cells, and macrophages were observed in the high-risk group. Moreover, there was a positive correlation between the risk score and immune checkpoints (ICPs), including PDCD1, CD274, and CTLA4. Patients within the high-risk group subject to PD1 and CTLA4 blockade exhibited higher IPS levels. Additionally, the expression of the five risk genes was upregulated in HCC cell lines and tissues compared to normal cells and tissues. Our findings establish a prognostic signature based on CD8+ T cells, offering a potent predictive model for clinical outcomes and responsiveness to immunotherapy in HCC patients. [ABSTRACT FROM AUTHOR]

Antigen-driven human effector-memory CD8+ T cells expressing low levels of the CD8β chain have been previously described. However, little is known on a possible antigen-independent trigger. We have examined the impact that IL-15 has on the expression of CD8β on purified human naïve CD8+ T cells after CFSE labeling and culture with IL-15. As expected, IL-15 induced naïve CD8+ T cells to proliferate and differentiate. Remarkably, the process was associated with a cell-cycle dependent down-modulation of CD8β from the cell surface, leading to the generation of CD8αβlow and CD8αβ- (i.e., CD8αα) T cells. In contrast, expression of the CD8α chain remained steady or even increased. Neither IL-2 nor IL-7 reproduced the effect of IL-15. Determination of mRNA levels for CD8α and CD8β isoforms by qPCR revealed that IL-15 promoted a significant decrease in mRNA levels of the CD8β M-4 isoform, while levels of the M-1/M-2 isoforms and of CD8α increased. Noteworthy, CD8+ T cell blasts obtained after culture of CD8+ T cells with IL-15 showed a cell-cycle dependent increase in the level of the tyrosine kinase Lck, when compared to CD8+ T cells at day 0. This study has shown for the first time that IL-15 generates CD8αα+αβlow and CD8αα+αβ- T cells containing high levels of Lck, suggesting that they may be endowed with unique functional features. [ABSTRACT FROM AUTHOR]

Background: The impact of chronic hepatic infection on antigen non-specific immune cells in circulation remains poorly understood. We reported lasting global hyperfunction of peripheral CD8 T cells in HCV-infected individuals with cirrhosis. Whether gene expression patterns in bulk CD8 T cells are associated with the severity of liver fibrosis in HCV infection is not known. Methods: RNA sequencing of blood CD8 T cells from treatment naïve, HCV infected individuals with minimal (Metavir F0-1 ≤ 7.0 kPa) or advanced fibrosis or cirrhosis (F4 ≥ 12.5 kPa), before and after direct-acting antiviral therapy, was performed. CD8 T cell function was assessed by flow cytometry. Results: In CD8 T cells from pre-DAA patients with advanced compared to minimal fibrosis, Gene Ontology analysis and Gene Set Enrichment Analysis identified differential gene expression related to cellular function and metabolism, including upregulated Hedgehog (Hh) signaling, IFN-a, -g, TGF-b response genes, apoptosis, apical surface pathways, phospholipase signaling, phosphatidyl-choline/inositol activity, and second-messenger-mediated signaling. In contrast, genes in pathways associated with nuclear processes, RNA transport, cytoskeletal dynamics, cMyc/E2F regulation, oxidative phosphorylation, and mTOR signaling, were reduced. Hh signaling pathway was the top featured gene set upregulated in cirrhotics, wherein hallmark genes GLI1 and PTCH1 ranked highly. Inhibition of Smo-dependent Hh signaling ablated the expression of IFN-g and perforin in stimulated CD8 T cells from chronic HCV-infected patients with advanced compared to minimal fibrosis. CD8 T cell gene expression profiles post-DAA remained clustered with pre-DAA profiles and disparately between advanced and minimal fibrosis, suggesting a persistent perturbation of gene expression long after viral clearance. Conclusions: This analysis of bulk CD8 T cell gene expression in chronic HCV infection suggests considerable reprogramming of the CD8 T cell pool in the cirrhotic state. Increased Hh signaling in cirrhosis may contribute to generalized CD8 T cell hyperfunction observed in chronic HCV infection. Understanding the lasting nature of immune cell dysfunction may help mitigate remaining clinical challenges after HCV clearance and more generally, improve long term outcomes for individuals with severe liver disease. [ABSTRACT FROM AUTHOR]

Peripheral blood CD8+ T lymphocytes play a crucial role in cell-mediated immunity and tumor-related immune responses in breast cancer. In this study, label-free quantification analysis and gene set enrichment analysis (GSEA) of CD8+ T lymphocytes in the peripheral blood of benign patients and patients with different breast cancer (BC) subtypes, i.e., luminal A, luminal B, and triple-negative breast cancer (TNBC), were performed using nano-UHPLC and Orbitrap mass spectrometry. Differential protein expression in CD8+ T lymphocytes revealed significant downregulation (log2 FC ≥ 0.38 or ≤−0.38, adj. p < 0.05), particularly in proteins involved in cytotoxicity, cytolysis, and proteolysis, such as granzymes (GZMs) and perforin 1 (PRF1). This downregulation was observed in the benign group (GZMH, GZMM, and PRF1) and luminal B (GZMA, GZMH) subtypes, whereas granzyme K (GZMK) was upregulated in TNBC in comparison to healthy controls. The RNA degradation pathway was significantly downregulated (p < 0.05, normalized enrichment score (NES) from −1.47 to −1.80) across all BC subtypes, suggesting a potential mechanism for regulating gene expression during T cell activation. Also, the Sm-like proteins (LSM2, LSM3, and LSM5) were significantly downregulated in the RNA degradation pathway. Proteomic analysis of CD8+ T lymphocytes in peripheral blood across different breast cancer subtypes provides a comprehensive view of the molecular mechanisms of the systemic immune response that can significantly contribute to advancements in the diagnosis, treatment, and prognosis of this disease. [ABSTRACT FROM AUTHOR]

Chronic Toxoplasma gondii infection induces brain-resident CD8+ T cells (bTr), but the protective functions and differentiation cues of these cells remain undefined. Here, we used a mouse model of latent infection by T. gondii leading to effective CD8+ T cell-mediated parasite control. Thanks to antibody depletion approaches, we found that peripheral circulating CD8+ T cells are dispensable for brain parasite control during chronic stage, indicating that CD8+ bTr are able to prevent brain parasite reactivation. We observed that the retention markers CD69, CD49a, and CD103 are sequentially acquired by brain parasite-specific CD8+ T cells throughout infection and that a majority of CD69/CD49a/CD103 triple-positive (TP) CD8+ T cells also express Hobit, a transcription factor associated with tissue residency. This TP subset develops in a CD4+ T cell-dependent manner and is associated with effective parasite control during chronic stage. Conditional invalidation of Transporter associated with Antigen Processing (TAP)-mediated major histocompatibility complex (MHC) class I presentation showed that presentation of parasite antigens by glutamatergic neurons and microglia regulates the differentiation of CD8+ bTr into TP cells. Single-cell transcriptomic analyses revealed that resistance to encephalitis is associated with the expansion of stem-like subsets of CD8+ bTr. In summary, parasite-specific brain-resident CD8+ T cells are a functionally heterogeneous compartment which autonomously ensure parasite control during T. gondii latent infection and which differentiation is shaped by neuronal and microglial MHC I presentation. A more detailed understanding of local T cell-mediated immune surveillance of this common parasite is needed for harnessing brain-resident CD8+ T cells in order to enhance control of chronic brain infections. [ABSTRACT FROM AUTHOR]

Multiple sclerosis (MS) and glioblastoma (GBM) are CNS diseases in whose development and progression immune privilege is intimately important, but in a relatively opposite manner. Maintenance and strengthening of immune privilege have been shown to be an important mechanism in glioblastoma immune evasion, while the breakdown of immune privilege leads to MS initiation and exacerbation. We hypothesize that molecular signaling pathways can be oppositely regulated in peripheral blood CD8+ T cells of MS and glioblastoma patients at a transcriptional level. We analyzed publicly available data of the peripheral blood CD8+ T cell MS vs. control (MSvsCTRL) and GBM vs. control (GBMvsCTRL) differentially expressed gene (DEG) contrasts with Qiagen's Ingenuity pathway analysis software (IPA). We have identified sphingolipid signaling pathway which was significantly downregulated in the GBMvsCTRL and upregulated in the MSvsCTRL. As the pathway is important for the CD8+ T lymphocytes CNS infiltration, this result is in line with our previously stated hypothesis. Comparing publicly available lists of differentially expressed serum exosomal miRNAs from MSvsCTRL and GBMvsCTRL contrasts, we have identified that hsa-miR-182-5p has the greatest potential effect on sphingolipid signaling regarding the number of regulated DEGs in the GBMvsCTRL contrast, while not being able to find any relevant potential sphingolipid signaling target transcripts in the MSvsCTRL contrast. We conclude that the sphingolipid signaling pathway is a top oppositely regulated pathway in peripheral blood CD8+ T cells from GBM and MS, and might be crucial for the differences in CNS immune privilege maintenance of investigated diseases, but further experimental research is necessary. [ABSTRACT FROM AUTHOR]

Background: Despite the success of antiretroviral therapy (ART), people living with HIV (PLWH) suffer from a high burden of pulmonary diseases, even after accounting for their smoking status. Cytotoxic CD8 T-cells are likely implicated in this phenomenon and may act as a double-edged sword. While being essential in viral infection control, their hyperactivation can also contribute to lung mucosal tissue damage. The effects of HIV and smoking on pulmonary mucosal CD8 T-cell dynamics has been a neglected area of research, which we address herein. Methods: Bronchoalveolar lavage (BAL) fluid were obtained from ART-treated PLWH (median duration of supressed viral load: 9 years; smokers: n = 14; non-smokers: n = 21) and HIV-uninfected controls (smokers: n = 11; non-smokers: n = 20) without any respiratory symptoms or active infection. Lymphocytes were isolated and CD8 T-cell subsets and homing markers were characterized by multiparametric flow cytometry. Results: Both smoking and HIV infection were independently associated with a significant increase in frequencies of total pulmonary mucosal CD8 T-cell. BAL CD8 T-cells were primarily CD69 + expressing CD103 and/or CD49a, at least one of the two granzymes (GzmA/GzmB), and little Perforin. Higher expression levels of CD103, CD69, and GzmB were observed in smokers versus non-smokers. The ex vivo phenotype of GzmA + and GzmB + cells revealed increased expression of CD103 and CXCR6 in smokers, while PLWH displayed elevated levels of CX3CR1 compared to controls. Conclusion: Smoking and HIV could promote cytotoxic CD8 T-cell retention in small airways through different mechanisms. Smoking likely increases recruitment and retention of GzmB + CD8 Trm via CXCR6 and CD103. Heightened CX3CR1 expression could be associated with CD8 non-Trm recruitment from the periphery in PLWH. [ABSTRACT FROM AUTHOR]

Cancer immunotherapy is a pillar of clinical oncology but only achieves long-term remissions in a minority of cases. In this issue, van Elsas et al. show that effective immunotherapy requires a series of processes orchestrated by CD8+ T cells that result in the recruitment and local activation of M1-like macrophages. [ABSTRACT FROM AUTHOR]

Background: Allergic rhinitis (AR), a prevalent chronic inflammatory condition triggered by immunoglobulin E (IgE), involves pivotal roles of immune and metabolic factors in its onset and progression. However, the intricacies and uncertainties in clinical research render current investigations into their interplay somewhat inadequate. Objective: To elucidate the causal relationships between immune cells, metabolites, and AR, we conducted a mediation Mendelian randomization (MR) analysis. Methods: Leveraging comprehensive publicly accessible summary-level data from genome-wide association studies (GWAS), this study employed the two-sample MR research method to investigate causal relationships among 731 immune cell phenotypes, 1400 metabolite levels, and AR. Additionally, employing the mediation MR approach, the study analyzed potential mediated effect of metabolites in the relationships between immune cells and AR. Various sensitivity analysis methods were systematically employed to ensure the robustness of the results. Results: Following false discovery rate (FDR) correction, we identified three immune cell phenotypes as protective factors for AR: Naive CD8br %CD8br (odds ratio (OR): 0.978, 95% CI = 0.966-0.990, P = 4.5x10-4), CD3 on CD39+ activated Treg (OR: 0.947, 95% CI = 0.923-0.972, P = 3x10-5), HVEM on CD45RA-CD4+ (OR: 0.967, 95% CI = 0.948--0.986, P = 4x10-5). Additionally, three metabolite levels were identified as risk factors for AR: N-methylhydroxyproline levels (OR: 1.219, 95% CI = 1.104--1.346, P = 9x10-5), N-acetylneuraminate levels (OR: 1.133, 95% CI = 1.061--1.211, P = 1.7x10-4), 1-stearoyl-2-arachidonoyl-gpc (18:0/20:4) levels (OR: 1.058, 95% CI = 1.029-1.087, P = 5x10-5). Mediation MR analysis indicated a causal relationship between Naive CD8br %CD8br and N-methylhydroxyproline levels, acting as a protective factor (OR: 0.971, 95% CI = 0.950-0.992, P = 8.31x10-3). The mediated effect was -0.00574, accounting for 26.1% of the total effect, with a direct effect of -0.01626. Naive CD8+ T cells exert a protective effect on AR by reducing N-methylhydroxyproline levels. Conclusion: Our study, delving into genetic information, has substantiated the intricate connection between immune cell phenotypes and metabolite levels with AR. This reveals a potential pathway to prevent the onset of AR, providing guiding directions for future clinical investigations. [ABSTRACT FROM AUTHOR]

Background and Aim: Staphylococcus aureus, with its diverse virulence factors and immune response evasion mechanisms, presents a formidable challenge as an opportunistic pathogen. Developing an effective vaccine against S. aureus has proven elusive despite extensive efforts. Autologous Staphylococcus lysate (ASL) treatment has proven effective in triggering an immune response against bovine mastitis. Peptides that stimulate the immune response can be the subject of further research. The study aimed to use immunoinformatics tools to identify epitopes on S. aureus surface and secretory proteins that can bind to major histocompatibility complex class I (MHC I) and CD8+ T-cells. This method aids in discovering prospective vaccine candidates and elucidating the rationale behind ASL therapy's efficacy. Materials and Methods: Proteins were identified using both literature search and the National Center for Biotechnology Information search engine Entrez. Self and non-self peptides, allergenicity predictions, epitope locations, and physicochemical characteristics were determined using sequence alignment, AllerTOP, SVMTriP, and Protein-Sol tools. Hex was employed for simulating the docking interactions between S. aureus proteins and the MHC I + CD8+ T-cells complex. The binding sites of S. aureus proteins were assessed using Computer Atlas of Surface Topography of Proteins (CASTp) while docked with MHC I and CD8+ T-cells. Results: Nine potential S. aureus peptides and their corresponding epitopes were identified in this study, stimulating cytotoxic T-cell mediated immunity. The peptides were analyzed for similarity with self-antigens and allergenicity. 1d20, 2noj, 1n67, 1nu7, 1amx, and 2b71, non-self and stable, are potential elicitors of the cytotoxic T-cell response. The energy values from docking simulations of peptide-MHC I complexes with the CD8+ and T-cell receptor (TCR) indicate the stability and strength of the formed complexes. These peptides - 2noj, 1d20, 1n67, 2b71, 1nu7, 1yn3, 1amx, 2gi9, and 1edk - demonstrated robust MHC I binding, as evidenced by their low binding energies. Peptide 2gi9 exhibited the lowest energy value, followed by 2noj, 1nu7, 1n67, and 1d20, when docked with MHC I and CD8 + TCR, suggesting a highly stable complex. CASTp analysis indicated substantial binding pockets in the docked complexes, with peptide 1d20 showing the highest values for area and volume, suggesting its potential as an effective elicitor of immunological responses. These peptides - 2noj, 2gi9, 1d20, and 1n67 - stand out for vaccine development and T-cell activation against S. aureus. Conclusion: This study sheds light on the design and development of S. aureus vaccines, highlighting the significance of employing computational methods in conjunction with experimental verification. The significance of T-cell responses in combating S. aureus infections is emphasized by this study. More experiments are needed to confirm the effectiveness of these vaccine candidates and discover their possible medical uses. [ABSTRACT FROM AUTHOR]

Background: T cell stress response state (TSTR), as a novel immune concept previous studies have proposed, has not yet been explored in prostate cancer (PC). As a type of cellular efflux, exosomes play important roles in the occurrence and development of PC. Method: Here, we conducted a combined analysis on extracellular vesicle related genes (EVRGs) in PC using data from single-cell RNA (scRNA), spatial transcriptome (ST), and bulk RNA sequencing. Result: Preliminary findings have revealed that heat shock protein family H (Hsp110) member 1 (HSPH1) possesses two identities, one being EVRGs and the other being a member of the heat shock protein family involved in TSTR, which may promote the differentiation of conventional T cells towards Th1 or Th2 cells through the pathway of IL2-MYC-IL2RA, thereby promoting the increase of CD8 + T cells in the tumor area, especially in the invasive zone, and inhibiting the invasion of PCs. We also notice the negative response of HSPH1 + CD8 + T cell related genes in immune checkpoint blockade (ICB). Western blot (WB) and droplet digital Polymerase Chain Reaction (ddPCR) demonstrated that the mRNA and protein levels of HSPH1 in EVs of PCs were significantly higher than those in adjacent tissues. Conclusion: Results above indicate the potential of HSPH1 as a critical therapeutic target in PC. [ABSTRACT FROM AUTHOR]

Simple Summary: A widely used test to establish if a person has T-cells that react against an infection or against a cancer is the enzyme-linked ImmunoSpot (ELISPOT assay). The person's T-cells are mixed with short lengths of amino acids called peptides that span the protein targets (antigens) step-wise in an overlapping manner, in order to stimulate a measurable response in the form of countable spots on the membrane of the ELISPOT plate, which indicate cytokine secretion by individual T cells. Whilst research has shown that responsive peptides for certain individuals can be predicted by computer programs, we show that using large peptide pools representing several antigens reveal most effectively the majority of the T cell reactivity against viruses. Such an approach is also likely to be most effective in cancers. Synthetic peptides derived from antigen sequences are essential reagents for the detection of CD8+ cytotoxic T lymphocytes (CTLs), in assays such as ELISPOT/ImmunoSpot®. Indeed, the combination of peptides and ImmunoSpot® has been widely used for immune monitoring in numerous vaccine trials. Target antigens in pathogens or cancers may be large in size and multiple in number, often seemingly necessitating in silico peptide epitope predictions using algorithms and programs for certain HLA alleles to narrow down the numbers of required peptides. In this commentary, we discuss our data in the context of immune responses to viral and cancer antigens, concluding that systematic high-throughput immune monitoring of CD8+ T cells will provide more reliable insights on the host's response to cancer than the reliance on select CD8+ T cell epitopes, no matter whether these are in silico predicted or even if they had been empirically established. We show the feasibility of large scale, high-throughput systematic CD8+ T cell epitope testing towards this goal. [ABSTRACT FROM AUTHOR]

Trichloroethylene (TCE) is an industrial solvent and widespread environmental contaminant associated with CD4+ T-cell activation and autoimmune disease. Prior studies showed that exposure to TCE in the drinking water of autoimmune-prone mice expanded effector/memory CD4+ T cells with an interferon-γ (IFN-γ)-secreting Th1-like phenotype. However, very little is known how TCE exposure skews CD4+ T cells towards this pro-inflammatory Th1 subset. As observed previously, TCE exposure was associated with hypermethylation of regions of the genome related to transcriptional repression in purified effector/memory CD4 T cells. We hypothesized that TCE modulates transcriptional and/or epigenetic programming of CD4+ T cells as they differentiate from a naive to effector phenotype. In the current study, purified naive CD4 T cells from both male and female autoimmune-prone MRL/MpJ mice were activated ex vivo and polarized towards a Th1 subset for 4 days in the presence or absence of the oxidative metabolite of TCE, trichloroacetaldehyde hydrate (TCAH) in vitro. An RNA-seq assessment and reduced representation bisulfite sequencing for DNA methylation were conducted on Th1 cells or activated, non-polarized cells. The results demonstrated TCAH's ability to regulate key genes involved in the immune response and autoimmunity, including Ifng, by altering the level of DNA methylation at the gene promoter. Intriguing sex differences were observed and for the most part, the effects were more robust in females compared to males. In conclusion, TCE via TCAH epigenetically regulates gene expression in CD4+ T cells. These results may have implications for mechanistic understanding or future therapeutics for autoimmunity. [ABSTRACT FROM AUTHOR]

CD8 downregulation is an overlooked component of self-reactive CD8+ T cell tolerance. CD8 downregulation is accompanied by T cell inhibition at the signaling and transcriptional levels. Loss of CD8 expression impairs the capacity of T cells to engage peptide-major histocompatibility complex complexes. CD8 downregulation may be a temporary event, as cells may regain CD8 expression and effector function. CD8 downregulation in mammals may be a part of a multilayered mechanism that inactivates self-reactive CD8+ T cells. Loss of CD8 may impair their capacity to engage target cells, while other simultaneous changes curb their activation and effector functions. Understanding this process represents a central element in our interpretation of CD8+ T cell physiology and its role in autoimmune diseases and cancer immunology. Activated CD8+ T cells directly kill target cells. Therefore, the regulation of their function is central to avoiding immunopathology. Mechanisms that curb effector functions in CD4+ and CD8+ T cells are mostly shared, yet important differences occur. Here, we focus on the control of CD8+ T cell activity and discuss the importance of a poorly understood aspect of tolerance that directly impairs engagement of target cells: the downregulation of CD8. We contextualize this process and propose that it represents a key element during CD8+ T cell modulation. [ABSTRACT FROM AUTHOR]

The proteasome generates the majority of peptides presented on MHC class I molecules. The cleavage pattern of the proteasome has been shown to be changed via the proteasome activator (PA)28 alpha beta (PA28αβ). In particular, several immunogenic peptides have been reported to be PA28αβ-dependent. In contrast, we did not observe a major impact of PA28αβ on the generation of different major histocompatibility complex (MHC) classI ligands. PA28αβ-knockout mice infected with the lymphocytic choriomeningitis virus (LCMV) or vaccinia virus showed a normal cluster of differentiation (CD) 8 response and viral clearance. However, we observed that the adoptive transfer of wild-type cells into PA28αβ-knockout mice led to graft rejection, but not vice versa. Depletion experiments showed that the observed rejection was mediated by CD8+ cytotoxic T cells. These data indicate that PA28αβ might be involved in the development of the CD8+ T cell repertoire in the thymus. Taken together, our data suggest that PA28αβ is a crucial factor determining T cell selection and, therefore, impacts graft acceptance. [ABSTRACT FROM AUTHOR]

Objectives: Gout is characterized by hyperuricemia and recurrent inflammatory episodes caused by intra-articular crystal deposition of monosodium urate (MSU). There is a clear relationship between gout and metabolic syndrome. Recent evidence indicates that perforin plays a role in regulating glucose homeostasis and provides protection in diet-induced non-alcoholic steatohepatitis models. However, the impact of perforin on immune inflammation in gout remains unclear. Methods: We induced acute gout models in both wild-type (WT) mice and Prf1null mice by administering intra-articular injections of MSU crystals. We compared the ankle joint swelling and the histological score between the two groups. Furthermore, we investigated underlying mechanisms through in vitro co-culture experiments involving CD8 T cells and macrophages. Results: In this study, Prf1null mice showed significantly more pronounced ankle swelling with increased inflammatory cell infiltrations compared with WT mice 24 h after local MSU injection. Moreover, MSU-induced Prf1null mice exhibited increased accumulation of CD8 T cells but not NK cells. Perforin-deficient CD8 T cells displayed reduced cytotoxicity towards bone marrow–derived M0 and M1 macrophages and promoted TNF-α secretion from macrophage. Conclusions: Perforin from CD8 T cells limits joint inflammation in mice with acute gout by downregulating macrophage-mediated inflammation. Key Points • Perforin deficiency increased swelling in the ankle joints of mice upon MSU injection. • Perforin deficiency is associated with increased immune cell recruitment and severe joint damage in gout. • Perforin regulated CD8 T cell accumulation in gout and promoted CD8 T cell cytotoxicity towards M0 and M1 macrophages. • CD8 T cell-derived perforin regulated pro-inflammatory cytokine secretion of macrophage. [ABSTRACT FROM AUTHOR]

Triple-negative breast cancer (TNBC) poses a significant challenge for treatment efficacy. CD8+ T cells, which are pivotal immune cells, can be effectively analyzed for differential gene expression across diverse cell populations owing to rapid advancements in sequencing technology. By leveraging these genes, our objective was to develop a prognostic model that accurately predicts the prognosis of patients with TNBC and their responsiveness to immunotherapy. Sample information and clinical data of TNBC were sourced from The Cancer Genome Atlas and METABRIC databases. In the initial stage, we identified 67 differentially expressed genes associated with immune response in CD8+ T cells. Subsequently, we narrowed our focus to three key genes, namely CXCL13 , GBP2 , and GZMB , which were used to construct a prognostic model. The accuracy of the model was assessed using the validation set data and receiver operating characteristic (ROC) curves. Furthermore, we employed various methods, including Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway, immune infiltration, and correlation analyses with CD274 (PD-L1) to explore the model's predictive efficacy in immunotherapeutic responses. Additionally, we investigated the potential underlying biological pathways that contribute to divergent treatment responses. We successfully developed a model capable of predicting the prognosis of patients with TNBC. The areas under the curve (AUC) values for the 1-, 3-, and 5-year survival predictions were 0.618, 0.652, and 0.826, respectively. Employing this risk model, we stratified the samples into high- and low-risk groups. Through KEGG enrichment analysis, we observed that the high-risk group predominantly exhibited enrichment in metabolism-related pathways such as drug and chlorophyll metabolism, whereas the low-risk group demonstrated significant enrichment in cytokine pathways. Furthermore, immune landscape analysis revealed noteworthy variations between (PD-L1) expression and risk scores, indicating that our model effectively predicted the response of patients to immune-based treatments. Our study demonstrates the potential of CXCL13, GBP2, and GZMB as prognostic indicators of clinical outcomes and immunotherapy responses in patients with TNBC. These findings provide valuable insights and novel avenues for developing immunotherapeutic approaches targeting TNBC. [ABSTRACT FROM AUTHOR]

Chagas disease, caused by the protozoan Trypanosoma cruzi, remains a major public health challenge affecting millions in Latin America and worldwide. Although significant progress has been made in vector control, no vaccine exists to prevent infection or mitigate disease pathogenesis. We developed a rationally designed chimeric protein vaccine, N-Tc52/TSkb20, incorporating immunodominant epitopes from two T. cruzi antigens, the amino-terminal portion of Tc52 and the TSkb20 epitope derived from trans-sialidase. The objectives of this study were to construct and characterize the antigen and evaluate its protective potential in an immunoprophylactic murine model of T. cruzi infection. The N-Tc52/TSkb20 protein was recombinantly expressed in E. coli and its identity was confirmed using mass spectrometry and Western blotting. Immunization with the chimeric protein significantly controlled parasitemia and reduced the heart, colon, and skeletal muscle parasite burdens compared to non-vaccinated mice. Protection was superior to vaccination with the individual parental antigen components. Mechanistically, the vaccine induced potent CD8+ T-cell and IFNγ responses against the incorporated epitopes and a protective IgG antibody profile. A relatively low IL-10 response favored early parasite control. These results validate the promising multi-epitope approach and support the continued development of this type of rational vaccine design strategy against Chagas disease. [ABSTRACT FROM AUTHOR]

Disseminated leishmaniasis (DL) caused by L. braziliensis is characterized by the presence of 10 to more than 1000 lesions spread on the body. While protection against Leishmania is mediated by macrophages upon activation by IFN-γ produced by CD4+T cells, the pathology of disseminated leishmaniasis (DL) could be mediated by macrophages, NK, and CD8+T cells. Herein, we evaluate the participation of senescent CD8+T cells in the pathogenesis of DL. Methods: Peripheral blood mononuclear cells (PBMCs), biopsies, co-cultures of CD8+T cells with uninfected and infected macrophages (MØ), and PBMC cultures stimulated with soluble L. braziliensis antigen (SLA) for 72 h from patients with cutaneous leishmaniasis (CL) and DL were used to characterize senescent CD8+T cells. Statistical analysis was performed using the Mann–Whitney and Kruskal–Wallis tests, followed by Dunn's. Results: Patients with DL have an increase in the frequency of circulating CD8+T cells that present a memory/senescent phenotype, while lesions from DL patients have an increase in the frequency of infiltrating CD8+T cells with a senescent/degranulation phenotype. In addition, after specific stimuli, DL patients' circulating CD8+T with memory/senescent profile, showing degranulation characteristics, increased upon SLA stimuli, and those specific CD8+T cells from DL patients had an increased degranulation phenotype, causing more apoptosis of infected target cells. Conclusions: DL patients show a higher frequency of cytotoxic senescent CD8+T cells compared to CL patients, and that could promote the lysis of infected cells, although without parasite killing, releasing Leishmania to the extracellular compartment, contributing to the spread of parasites. [ABSTRACT FROM AUTHOR]

Infusion of 13C-labeled metabolites provides a gold standard for understanding the metabolic processes used by T cells during immune responses in vivo. Through infusion of 13C-labeled metabolites (glucose, glutamine, and acetate) in Listeria monocytogenes-infected mice, we demonstrate that CD8 T effector (Teff) cells use metabolites for specific pathways during specific phases of activation. Highly proliferative early Teff cells in vivo shunt glucose primarily toward nucleotide synthesis and leverage glutamine anaplerosis in the tricarboxylic acid (TCA) cycle to support adenosine triphosphate and de novo pyrimidine synthesis. In addition, early Teff cells rely on glutamic-oxaloacetic transaminase 1 (Got1)--which regulates de novo aspartate synthesis--for effector cell expansion in vivo. CD8 Teff cells change fuel preference over the course of infection, switching from glutamine-to acetate-dependent TCA cycle metabolism late in infection. This study provides insights into the dynamics of Teff metabolism, illuminating distinct pathways of fuel consumption associated with CD8 Teff cell function in vivo. [ABSTRACT FROM AUTHOR]

Purpose: This study mainly focuses on the immune function and introduces CD4+, CD8+ T cells and their ratios based on the MuLBSTA score, a previous viral pneumonia mortality risk warning model, to construct an early warning model of severe viral pneumonia risk. Methods: A retrospective single-center observational study was operated from January 2021 to December 2022 at the People's Hospital of Liangjiang New Area, Chongqing, China. A total of 138 patients who met the criteria for viral pneumonia in hospital were selected and their data, including demographic data, comorbidities, laboratory results, CT scans, immunologic and pathogenic tests, treatment regimens, and clinical outcomes, were collected and statistically analyzed. Results: Forty-one patients (29.7%) developed severe or critical illness. A viral pneumonia severe risk warning model was successfully constructed, including eight parameters: age, bacterial coinfection, CD4+, CD4+/CD8+, multiple lung lobe infiltrations, smoking, hypertension, and hospital admission days. The risk score for severe illness in patients was set at 600 points. The model had good predictive performance (AUROC = 0.94397), better than the original MuLBSTA score (AUROC = 0.8241). Conclusion: A warning system constructed based on immune function has a good warning effect on the risk of severe conversion in patients with viral pneumonia. [ABSTRACT FROM AUTHOR]

Killer cell lectin-like receptor G1 (KLRG1) has traditionally been regarded as an inhibitory receptor of T cell exhaustion in chronic infection and inflammation. However, its exact role in hepatitis B virus (HBV) infection remains elusive. CD8+ T cells from 190 patients with chronic hepatitis B were analyzed ex vivo for checkpoint and apoptosis markers, transcription factors, cytokines and subtypes in 190 patients with chronic hepatitis B. KLRG1+ and KLRG1− CD8+ T cells were sorted for transcriptome analysis. The impact of the KLRG1-E-cadherin pathway on the suppression of HBV replication mediated by virus-specific T cells was validated in vitro. As expected, HBV-specific CD8+ T cells expressed higher levels of KLRG1 and showed an exhausted molecular phenotype and function. However, despite being enriched for the inhibitory molecules, thymocyte selection-associated high mobility group box protein (TOX), eomesodermin (EOMES), and Helios, CD8+ T cells expressing KLRG1 produced significant levels of tumour necrosis factor (TNF)-α, interferon (IFN)-γ, perforin, and granzyme B, demonstrating not exhausted but active function. Consistent with the in vitro phenotypic assay results, RNA sequencing (RNA-seq) data showed that signature effector T cell and exhausted T cell genes were enriched in KLRG1+ CD8+ T cells. Furthermore, in vitro testing confirmed that KLRG1−E-cadherin binding inhibits the antiviral efficacy of HBV-specific CD8+ T cells. Based on these findings, we concluded that KLRG1+ CD8+ T cells are not only a terminally exhausted subgroup but also exhibit functional diversity, despite inhibitory signs in HBV infection. [ABSTRACT FROM AUTHOR]