Comprehensive analysis of T-cell regulatory factors and tumor immune microenvironment in stomach adenocarcinoma.

BACKGROUND: Gastric cancer (GC) is one of the most prevalent malignant tumors worldwide and is associated with high morbidity and mortality rates. However, the specific biomarkers used to predict the postoperative prognosis of patients with gastric cancer remain unknown. Recent research has shown that the tumor microenvironment (TME) has an increasingly positive effect on anti-tumor activity. This study aims to build signatures to study the effect of certain genes on gastric cancer. METHODS: Expression profiles of 37 T cell-related genes and their TME characteristics were comprehensively analyzed. A risk signature was constructed and validated based on the screened T cell-related genes, and the roles of hub genes in GC were experimentally validated. RESULTS: A novel T cell-related gene signature was constructed based on CD5, ABCA8, SERPINE2, ESM1, SERPINA5, and NMU. The high-risk group indicated lower overall survival (OS), poorer immune efficacy, and higher drug resistance, with SERPINE2 promoting GC cell proliferation, according to experiments. SERPINE2 and CXCL12 were significantly correlated, indicating poor OS via the Youjiang cohort. CONCLUSIONS: This study identified T cell-related genes in patients with stomach adenocarcinoma (STAD) for prognosis estimation and proposed potential immunotherapeutic targets for STAD.

Glucose competition between endothelial cells in the blood-spinal cord barrier and infiltrating regulatory T cells is linked to sleep restriction-induced hyperalgesia.

BACKGROUND: Sleep loss is a common public health problem that causes hyperalgesia, especially that after surgery, which reduces the quality of life seriously. METHODS: The 48-h sleep restriction (SR) mouse model was created using restriction chambers. In vivo imaging, transmission electron microscopy (TEM), immunofluorescence staining and Western blot were performed to detect the status of the blood-spinal cord barrier (BSCB). Paw withdrawal mechanical threshold (PWMT) was measured to track mouse pain behavior. The role of infiltrating regulatory T cells (Tregs) and endothelial cells (ECs) in mouse glycolysis and BSCB damage were analyzed using flow cytometry, Western blot, CCK-8 assay, colorimetric method and lactate administration. RESULTS: The 48-h SR made mice in sleep disruption status and caused an acute damage to the BSCB, resulting in hyperalgesia and neuroinflammation in the spinal cord. In SR mice, the levels of glycolysis and glycolysis enzymes of ECs in the BSCB were found significantly decreased [CON group vs. SR group: CD31+Glut1+ cells: p < 0.001], which could cause dysfunction of ECs and this was confirmed in vitro. Increased numbers of infiltrating T cells [p < 0.0001] and Treg population [p < 0.05] were detected in the mouse spinal cord after 48-h SR. In the co-cultured system of ECs and Tregs in vitro, the competition of Tregs for glucose resulted in the glycolysis disorder of ECs [Glut1: p < 0.01, ENO1: p < 0.05, LDHα: p < 0.05; complete tubular structures formed: p < 0.0001; CCK8 assay: p < 0.001 on 24h, p < 0.0001 on 48h; glycolysis level: p < 0.0001]. An administration of sodium lactate partially rescued the function of ECs and relieved SR-induced hyperalgesia. Furthermore, the mTOR signaling pathway was excessively activated in ECs after SR in vivo and those under the inhibition of glycolysis or co-cultured with Tregs in vitro. CONCLUSIONS: Affected by glycolysis disorders of ECs due to glucose competition with infiltrating Tregs through regulating the mTOR signaling pathway, hyperalgesia induced by 48-h SR is attributed to neuroinflammation and damages to the barriers, which can be relieved by lactate supplementation.

Increased proportions of circulating PD-1+ CD4+ memory T cells and PD-1+ regulatory T cells associate with good response to prednisone in pulmonary sarcoidosis.

BACKGROUND: The treatment response to corticosteroids in patients with sarcoidosis is highly variable. CD4+ T cells are central in sarcoid pathogenesis and their phenotype in peripheral blood (PB) associates with disease course. We hypothesized that the phenotype of circulating T cells in patients with sarcoidosis may correlate with the response to prednisone treatment. Therefore, we aimed to correlate frequencies and phenotypes of circulating T cells at baseline with the pulmonary function response at 3 and 12 months during prednisone treatment in patients with pulmonary sarcoidosis. METHODS: We used multi-color flow cytometry to quantify activation marker expression on PB T cell populations in 22 treatment-naïve patients and 21 healthy controls (HCs). Pulmonary function tests at baseline, 3 and 12 months were used to measure treatment effect. RESULTS: Patients with sarcoidosis showed an absolute forced vital capacity (FVC) increase of 14.2% predicted (± 10.6, p < 0.0001) between baseline and 3 months. Good response to prednisone (defined as absolute FVC increase of ≥ 10% predicted) was observed in 12 patients. CD4+ memory T cells and regulatory T cells from patients with sarcoidosis displayed an aberrant phenotype at baseline, compared to HCs. Good responders at 3 months had significantly increased baseline proportions of PD-1+CD4+ memory T cells and PD-1+ regulatory T cells, compared to poor responders and HCs. Moreover, decreased fractions of CD25+ cells and increased fractions of PD-1+ cells within the CD4+ memory T cell population correlated with ≥ 10% FVC increase at 12 months. During treatment, the aberrantly activated phenotype of memory and regulatory T cells reversed. CONCLUSIONS: Increased proportions of circulating PD-1+CD4+ memory T cells and PD-1+ regulatory T cells and decreased proportions of CD25+CD4+ memory T cells associate with good FVC response to prednisone in pulmonary sarcoidosis, representing promising new blood biomarkers for prednisone efficacy. TRIAL REGISTRATION: NL44805.078.13.

Tracking in situ checkpoint inhibitor-bound target T cells in patients with checkpoint-induced colitis.

The success of checkpoint inhibitors (CPIs) for cancer has been tempered by immune-related adverse effects including colitis. CPI-induced colitis is hallmarked by expansion of resident mucosal IFNγ cytotoxic CD8+ T cells, but how these arise is unclear. Here, we track CPI-bound T cells in intestinal tissue using multimodal single-cell and subcellular spatial transcriptomics (ST). Target occupancy was increased in inflamed tissue, with drug-bound T cells located in distinct microdomains distinguished by specific intercellular signaling and transcriptional gradients. CPI-bound cells were largely CD4+ T cells, including enrichment in CPI-bound peripheral helper, follicular helper, and regulatory T cells. IFNγ CD8+ T cells emerged from both tissue-resident memory (TRM) and peripheral populations, displayed more restricted target occupancy profiles, and co-localized with damaged epithelial microdomains lacking effective regulatory cues. Our multimodal analysis identifies causal pathways and constitutes a resource to inform novel preventive strategies.

Thymic gene expression analysis reveals a potential link between HIF-1A and Th17/Treg imbalance in thymoma associated myasthenia gravis.

Myasthenia gravis (MG) is an immune-mediated disease frequently associated with thymic changes. Increased T helper 17 (Th17) cell activity and dysfunctional regulatory T (Treg) cells have been demonstrated in subgroups of MG. On the other hand, hypoxia-inducible factor 1 (HIF-1) has been shown to regulate the Th17/Treg balance by inducing Th17 differentiation while attenuating Treg development. To identify the underlying mechanisms of different thymic pathologies in MG development, we evaluated thymic samples from thymoma-associated myasthenia gravis (TAMG), MG with hyperplasia (TFH-MG) and thymoma without MG (TOMA) patients. Differential gene expression analysis revealed that TAMG and TFH-MG cells are associated with different functional pathways. A higher RORC/FOXP3 ratio provided evidence for Th17/Treg imbalance in TAMG potentially related to increased HIF1A. The hypoxic microenvironment in thymoma may be a driver of TAMG by increasing HIF1A. These findings may lead to new therapeutic approaches targeting HIF1A in the development of TAMG.

Osteomyelitis is associated with increased anti-inflammatory response and immune exhaustion.

Introduction: Osteomyelitis (OMS) is a bone infection causing bone pain and severe complications. A balanced immune response is critical to eradicate infection without harming the host, yet pathogens manipulate immunity to establish a chronic infection. Understanding OMS-driven inflammation is essential for disease management, but comprehensive data on immune profiles and immune cell activation during OMS are lacking. Methods: Using high-dimensional flow cytometry, we investigated the detailed innate and adaptive systemic immune cell populations in OMS and age- and sex-matched controls. Results: Our study revealed that OMS is associated with increased levels of immune regulatory cells, namely T regulatory cells, B regulatory cells, and T follicular regulatory cells. In addition, the expression of immune activation markers HLA-DR and CD86 was decreased in OMS, while the expression of immune exhaustion markers TIM-3, PD-1, PD-L1, and VISTA was increased. Members of the T follicular helper (Tfh) cell family as well as classical and typical memory B cells were significantly increased in OMS individuals. We also found a strong correlation between memory B cells and Tfh cells. Discussion: We conclude that OMS skews the host immune system towards the immunomodulatory arm and that the Tfh memory B cell axis is evident in OMS. Therefore, immune-directed therapies may be a promising alternative for eradication and recurrence of infection in OMS, particularly in individuals and areas where antibiotic resistance is a major concern.

Preparation and application of fluorescent monoclonal antibodies recognizing goat CD4+CD25+ regulatory T cells.

Regulatory T cells (Tregs) are a subset of T cells participating in a variety of diseases including mycoplasmal pneumonia, contagious ecthyma, and so on. The role of Tregs in goat contagious ecthyma is not completely understood due to the lack of species-specific antibodies. Here, we developed a combination of CD4 and CD25 fluorescence monoclonal antibodies (mAb) to recognize goat Tregs and assessed its utility in flow cytometry, immunofluorescence staining. Using immunofluorescence staining, we found that the frequency of Treg cells was positively correlated with the viral load during orf virus infection. These antibodies could serve as important tools to monitor Tregs during orf virus infection in goats. KEY POINTS: • A combination of fluorescent mAbs (C11 and D12) was prepared for the detection of goat Tregs. • C11 and D12 are effective in flow cytometry, immunofluorescence staining, and C11 has excellent species specificity. • The frequency of Treg cells was positively correlated with the viral load during orf virus infection.

Identification of master regulator genes controlling pathogenic CD4+ T cell fate in inflammatory bowel disease through transcriptional network analysis.

Inflammatory bowel diseases (IBD) are a group of chronic inflammatory conditions of the gastrointestinal tract associated with multiple pathogenic factors, including dysregulation of the immune response. Effector CD4+ T cells and regulatory CD4+ T cells (Treg) are central players in maintaining the balance between tolerance and inflammation. Interestingly, genetic modifications in these cells have been implicated in regulating the commitment of specific phenotypes and immune functions. However, the transcriptional program controlling the pathogenic behavior of T helper cells in IBD progression is still unknown. In this study, we aimed to find master transcription regulators controlling the pathogenic behavior of effector CD4+ T cells upon gut inflammation. To achieve this goal, we used an animal model of IBD induced by the transfer of naïve CD4+ T cells into recombination-activating gene 1 (Rag1) deficient mice, which are devoid of lymphocytes. As a control, a group of Rag1-/- mice received the transfer of the whole CD4+ T cells population, which includes both effector T cells and Treg. When gut inflammation progressed, we isolated CD4+ T cells from the colonic lamina propria and spleen tissue, and performed bulk RNA-seq. We identified differentially up- and down-regulated genes by comparing samples from both experimental groups. We found 532 differentially expressed genes (DEGs) in the colon and 30 DEGs in the spleen, mostly related to Th1 response, leukocyte migration, and response to cytokines in lamina propria T-cells. We integrated these data into Gene Regulatory Networks to identify Master Regulators, identifying four up-regulated master gene regulators (Lef1, Dnmt1, Mybl2, and Jup) and only one down-regulated master regulator (Foxo3). The altered expression of master regulators observed in the transcriptomic analysis was confirmed by qRT-PCR analysis and found an up-regulation of Lef1 and Mybl2, but without differences on Dnmt1, Jup, and Foxo3. These two master regulators have been involved in T cells function and cell cycle progression, respectively. We identified two master regulator genes associated with the pathogenic behavior of effector CD4+ T cells in an animal model of IBD. These findings provide two new potential molecular targets for treating IBD.

Understanding the immunosuppressive microenvironment of glioma: mechanistic insights and clinical perspectives.

Glioblastoma (GBM), the predominant and primary malignant intracranial tumor, poses a formidable challenge due to its immunosuppressive microenvironment, thereby confounding conventional therapeutic interventions. Despite the established treatment regimen comprising surgical intervention, radiotherapy, temozolomide administration, and the exploration of emerging modalities such as immunotherapy and integration of medicine and engineering technology therapy, the efficacy of these approaches remains constrained, resulting in suboptimal prognostic outcomes. In recent years, intensive scrutiny of the inhibitory and immunosuppressive milieu within GBM has underscored the significance of cellular constituents of the GBM microenvironment and their interactions with malignant cells and neurons. Novel immune and targeted therapy strategies have emerged, offering promising avenues for advancing GBM treatment. One pivotal mechanism orchestrating immunosuppression in GBM involves the aggregation of myeloid-derived suppressor cells (MDSCs), glioma-associated macrophage/microglia (GAM), and regulatory T cells (Tregs). Among these, MDSCs, though constituting a minority (4-8%) of CD45+ cells in GBM, play a central component in fostering immune evasion and propelling tumor progression, angiogenesis, invasion, and metastasis. MDSCs deploy intricate immunosuppressive mechanisms that adapt to the dynamic tumor microenvironment (TME). Understanding the interplay between GBM and MDSCs provides a compelling basis for therapeutic interventions. This review seeks to elucidate the immune regulatory mechanisms inherent in the GBM microenvironment, explore existing therapeutic targets, and consolidate recent insights into MDSC induction and their contribution to GBM immunosuppression. Additionally, the review comprehensively surveys ongoing clinical trials and potential treatment strategies, envisioning a future where targeting MDSCs could reshape the immune landscape of GBM. Through the synergistic integration of immunotherapy with other therapeutic modalities, this approach can establish a multidisciplinary, multi-target paradigm, ultimately improving the prognosis and quality of life in patients with GBM.

Multi-Omics profiling identifies aldehyde dehydrogenase 2 as a critical mediator in the crosstalk between Treg-mediated immunosuppression microenvironment and hepatocellular carcinoma.

Dysregulation of the aldehyde dehydrogenase (ALDH) family has been implicated in various pathological conditions, including cancer. However, a systematic evaluation of ALDH alterations and their therapeutic relevance in hepatocellular carcinoma (HCC) remains lacking. Herein, we found that 15 of 19 ALDHs were transcriptionally dysregulated in HCC tissues compared to normal liver tissues. A four gene signature, including ALDH2, ALDH5A1, ALDH6A1, and ALDH8A1, robustly predicted prognosis and defined a high-risk subgroup exhibiting immunosuppressive features like regulatory T cell (Tregs) infiltration. Single-cell profiling revealed selective overexpression of tumor necrosis factor receptor superfamily member 18 (TNFRSF18) on Tregs, upregulated in high-risk HCC patients. We identified ALDH2 as a tumor suppressor in HCC, with three novel phosphorylation sites mediated by protein kinase C zeta that enhanced enzymatic activity. Mechanistically, ALDH2 suppressed Tregs differentiation by inhibiting ß-catenin/TGF-ß1 signaling in HCC. Collectively, our integrated multi-omics analysis defines an ALDH-Tregs-TNFRSF18 axis that contributes to HCC pathogenesis and represents potential therapeutic targets for this aggressive malignancy.

Quantifying NF-κB Activation by Flow Cytometry of IκBα Degradation.

Nuclear factor-κB (NF-κB) is a crucial pro-inflammatory transcription factor whose activation is of immense interest to immunology research. Estimation of NF-κB activation through flow cytometry is not possible due to the unavailability of robust flow cytometry antibodies that can bind to its phosphorylated, active, nuclear form. In this protocol, we describe a flow cytometry assay that measures the activation of the pro-inflammatory transcription factor NF-κB in stimulated immune cells by quantifying the degradation of its upstream regulator IκBα. We demonstrate the utility of this protocol by assessment of intracellular IκBα in human primary regulatory T cells experiencing TNFR2 agonism, a process previously reported to activate NF-κB in these cells. We also show that this assay may be applied to study NF-κB activation in other cell types, such as human primary T cells and THP-1 cell-derived macrophages, when induced by their corresponding inflammatory cues. Thus, this robust and reproducible protocol will be of interest to a wide range of scientists who aim to measure NF-κB activity in medium-to-high-throughput assays. © 2024 Wiley Periodicals LLC. Basic Protocol: Quantifying inflammatory activation by flow cytometry of IκBα degradation Support Protocol 1: Isolating and expanding human regulatory T cells Support Protocol 2: Calculating IC50 from flow cytometry data using Excel.

The Role of Regulatory T Cells and Their Therapeutic Potential in Hypertensive Disease of Pregnancy: A Literature Review.

Hypertensive disorders of pregnancy (HDP), including preeclampsia (PE) and gestational hypertension (GH), are major causes of maternal and foetal morbidity and mortality. This review elucidates the role of regulatory T cells (Tregs) in the immunological aspects of HDP and explores their therapeutic potential. Tregs, which play a critical role in maintaining immune homeostasis, are crucial in pregnancy to prevent immune-mediated rejection of the foetus. The review highlights that Tregs contribute to immunological adaptation in normal pregnancy, ensuring foetal acceptance. In contrast, HDP is associated with Treg dysfunction, which is marked by decreased numbers and impaired regulatory capacity, leading to inadequate immune tolerance and abnormal placental development. This dysfunction is particularly evident in PE, in which Tregs fail to adequately modulate the maternal immune response against foetal antigens, contributing to the pathophysiology of the disorder. Therapeutic interventions aiming to modulate Treg activity represent a promising avenue for HDP management. Studies in animal models and limited clinical trials suggest that enhancing Treg functionality could mitigate HDP symptoms and improve pregnancy outcomes. However, given the multifactorial nature of HDP and the intricate regulatory mechanisms of Tregs, the review explores the complexities of translating in vitro and animal model findings into effective clinical therapies. In conclusion, while the precise role of Tregs in HDP is still being unravelled, their central role in immune regulation during pregnancy is indisputable. Further research is needed to fully understand the mechanisms by which Tregs contribute to HDP and to develop targeted therapies that can safely and effectively harness their regulatory potential for treating hypertensive diseases of pregnancy.

The impact of regulatory T cells on the graft-versus-leukemia effect.

Allogeneic Hematopoietic Stem Cell Transplantation (allo-HSCT) is the only curative therapy for many hematologic malignancies, whereby the Graft-versus-Leukemia (GVL) effect plays a pivotal role in controlling relapse. However, the success of GVL is hindered by Graft-versus-Host Disease (GVHD), where donor T cells attack healthy tissues in the recipient. The ability of natural regulatory T cells (Treg) to suppress immune responses has been exploited as a therapeutical option against GVHD. Still, it is crucial to evaluate if the ability of Treg to suppress GVHD does not compromise the benefits of GVL. Initial studies in animal models suggest that Treg can attenuate GVHD while preserving GVL, but results vary according to tumor type. Human trials using Treg as GVHD prophylaxis or treatment show promising results, emphasizing the importance of infusion timing and Treg/Tcon ratios. In this review, we discuss strategies that can be used aiming to enhance GVL post-Treg infusion and the proposed mechanisms for the maintenance of the GVL effect upon the adoptive Treg transfer. In order to optimize the therapeutic outcomes of Treg administration in allo-HSCT, future efforts should focus on refining Treg sources for infusion and evaluating their specificity for antigens mediating GVHD while preserving GVL responses.

Dimethyl fumarate modulates the regulatory T cell response in the mesenteric lymph nodes of mice with experimental autoimmune encephalomyelitis.

Dimethyl fumarate (DMF, Tecfidera) is an oral drug utilized to treat relapsing-remitting multiple sclerosis (MS). DMF treatment reduces disease activity in MS. Gastrointestinal discomfort is a common adverse effect of the treatment with DMF. This study aimed to investigate the effect of DMF administration in the gut draining lymph nodes cells of C57BL6/J female mice with experimental autoimmune encephalomyelitis (EAE), an animal model of MS. We have demonstrated that the treatment with DMF (7.5 mg/kg) significantly reduces the severity of EAE. This reduction of the severity is accompanied by the increase of both proinflammatory and anti-inflammatory mechanisms at the beginning of the treatment. As the treatment progressed, we observed an increasing number of regulatory Foxp3 negative CD4 T cells (Tr1), and anti-inflammatory cytokines such as IL-27, as well as the reduction of PGE2 level in the mesenteric lymph nodes of mice with EAE. We provide evidence that DMF induces a gradual anti-inflammatory response in the gut draining lymph nodes, which might contribute to the reduction of both intestinal discomfort and the inflammatory response of EAE. These findings indicate that the gut is the first microenvironment of action of DMF, which may contribute to its effects of reducing disease severity in MS patients.

Treg-derived TGF-ß1 dampens cGAS-STING signaling to downregulate the expression of class I MHC complex in multiple myeloma.

Multiple myeloma (MM) progression involves diminished tumor antigen presentation and an immunosuppressive microenvironment, characterized by diminished expression of major histocompatibility complexes (MHC) class I molecule and elevated programmed death ligand 1 (PDL1) in MM cells, along with an enriched population of regulatory T cells (Tregs). To investigate Treg's influence on MM cells, we established a co-culture system using Tregs from MM patients and the MM cell lines (MM.1S and SK-MM-1) in vitro and assessed the effects of intervening in the relevant pathways connecting Tregs and MM cells in vivo. In vitro, Tregs induced transforming growth factor beta-1 (TGF-ß1) production, downregulated MHC I members, and increased PDL1 expression in MM cells. Treg-derived TGF-ß1 suppressed the cGAS-STING pathway, contributing to the loss of MHC I molecule expression and PDL1 upregulation. Correspondingly, neutralizing TGF-ß1 or activating the cGAS-STING pathway restored MHC I and PDL1 expression, effectively countering the pro-tumorigenic effect of Tregs on MM cells in vivo. These data elucidated how Tregs influence tumor antigen presentation and immunosuppressive signal in MM cells, potentially providing therapeutic strategies, such as neutralizing TGF-ß1 or activating the cGAS-STING pathway, to address the immune escape and immunosuppressive dynamics in MM.

Adaptive plasticity of natural interleukin-35-induced regulatory T cells (Tr35) that are required for T-cell immune regulation.

Background: IL-35 potently inhibits immune responses both in vivo and in vitro. However, the specific characteristics of IL-35-producing cells, including their developmental origin, cellular phenotype, and function, are unknown. Methods: By using a novel IL-35 reporter mouse (Ebi3-Dre-Thy1.1) and double transgenic fate-mapping reporter mice (35EbiT-Rosa26-rox-tdTomato reporter mice or Foxp3 fate-mapping system), we tracked and analyzed the differentiation and developmental trajectories of Tr35 cells in vivo. And then we investigated the therapeutic effects of OVA-specific Tr35 cells in an OVA-induced allergic airway disease model. Results: We identified a subset of cells, denoted Tr35 cells, that secrete IL-35 but do not express Foxp3. These cells have high expression of molecules associated with T-cell activation and can inhibit T-cell proliferation in vitro. Our analyses showed that Tr35 cells are a distinct subpopulation of cells that are independent of Tr1 cells. Tr35 cells exhibit a unique gene expression profile and tissue distribution. The presence of Thy1.1 (Ebi3) expression in Tr35 cells indicates their active secretion of IL-35. However, the proportion of ex-Tr35 cells (Thy1.1-) is significantly higher compared to Tr35 cells (Thy1.1+). This suggests that Tr35 cells possess the ability to regulate IL-35 expression rapidly in vivo. Tr35 cells downregulated the expression of the inflammatory cytokines IL-4, IFN-γ and IL-17A. However, once Tr35 cells lost IL-35 expression and became exTr35 cells, the expression of inflammatory cytokines was upregulated. Importantly, our findings indicate that Tr35 cells have therapeutic potential. In an OVA-induced allergic airway disease mouse model, Tr35 cell reinfusion significantly reduced airway hyperresponsiveness and histopathological airway and lung inflammation. Conclusions: We have identified a subset of Tregs, Tr35 cells, that are distinct from Tr1 cells. Tr35 cells can dynamically regulate the secretion of inflammatory cytokines by controlling IL-35 expression to regulate inflammatory immune responses.

CD4+ T cell heterogeneity in gestational age and preeclampsia using single-cell RNA sequencing.

A balance between pro-inflammatory decidual CD4+ T cells and FOXP3+ regulatory T cells (FOXP3+ Tregs) is important for maintaining fetomaternal tolerance. Using single-cell RNA-sequencing and T cell receptor repertoire analysis, we determined that diversity and clonality of decidual CD4+ T cell subsets depend on gestational age. Th1/Th2 intermediate and Th1 subsets of CD4+ T cells were clonally expanded in both early and late gestation, whereas FOXP3+ Tregs were clonally expanded in late gestation. Th1/Th2 intermediate and FOXP3+ Treg subsets showed altered gene expression in preeclampsia (PE) compared to healthy late gestation. The Th1/Th2 intermediate subset exhibited elevated levels of cytotoxicity-related gene expression in PE. Moreover, increased Treg exhaustion was observed in the PE group, and FOXP3+ Treg subcluster analysis revealed that the effector Treg like subset drove the Treg exhaustion signatures in PE. The Th1/Th2 intermediate and effector Treg like subsets are possible inflammation-driving subsets in PE.

Single-cell transcriptomic analyses reveal distinct immune cell contributions to epithelial barrier dysfunction in checkpoint inhibitor colitis.

Immune checkpoint inhibitor (ICI) therapy has revolutionized oncology, but treatments are limited by immune-related adverse events, including checkpoint inhibitor colitis (irColitis). Little is understood about the pathogenic mechanisms driving irColitis, which does not readily occur in model organisms, such as mice. To define molecular drivers of irColitis, we used single-cell multi-omics to profile approximately 300,000 cells from the colon mucosa and blood of 13 patients with cancer who developed irColitis (nine on anti-PD-1 or anti-CTLA-4 monotherapy and four on dual ICI therapy; most patients had skin or lung cancer), eight controls on ICI therapy and eight healthy controls. Patients with irColitis showed expanded mucosal Tregs, ITGAEHi CD8 tissue-resident memory T cells expressing CXCL13 and Th17 gene programs and recirculating ITGB2Hi CD8 T cells. Cytotoxic GNLYHi CD4 T cells, recirculating ITGB2Hi CD8 T cells and endothelial cells expressing hypoxia gene programs were further expanded in colitis associated with anti-PD-1/CTLA-4 therapy compared to anti-PD-1 therapy. Luminal epithelial cells in patients with irColitis expressed PCSK9, PD-L1 and interferon-induced signatures associated with apoptosis, increased cell turnover and malabsorption. Together, these data suggest roles for circulating T cells and epithelial-immune crosstalk critical to PD-1/CTLA-4-dependent tolerance and barrier function and identify potential therapeutic targets for irColitis.

T cell activation and deficits in T regulatory cells are associated with major depressive disorder and severity of depression.

Major depressive disorder (MDD) is associated with T cell activation, but no studies have examined the combined effects of T cell activation and deficits in T regulatory (Treg) cells on the severity of acute phase MDD. Using flow cytometry, we determined the percentage and median fluorescence intensity of CD69, CD71, CD40L, and HLADR-bearing CD3+, CD4+, and CD8+ cells, and cannabinoid type 1 receptor (CB1), CD152 and GARP (glycoprotein A repetitions predominant)-bearing CD25+ FoxP3 T regulatory (Treg) cells in 30 MDD patients and 20 healthy controls in unstimulated and stimulated (anti-CD3/CD28) conditions. Based on cytokine levels, we assessed M1 macrophage, T helper (Th)-1 cell, immune-inflammatory response system (IRS), T cell growth, and neurotoxicity immune profiles. We found that the immune profiles (including IRS and neurotoxicity) were significantly predicted by decreased numbers of CD152 or GARP-bearing CD25+ FoxP3 cells or CD152 and GARP expression in combination with increases in activated T cells (especially CD8+ CD40L+ percentage and expression). MDD patients showed significantly increased numbers of CD3+ CD71+, CD3+ CD40L+, CD4+ CD71+, CD4+ CD40L+, CD4+ HLADR+, and CD8+ HLADR+ T cells, increased CD3+ CD71+, CD4+ CD71+ and CD4+ HLADR+ expression, and lowered CD25+ FoxP3 expression and CD25+ FoxP+ CB1+ numbers as compared with controls. The Hamilton Depression Rating Scale score was strongly predicted (between 30 and 40% of its variance) by a lower number of CB1 or GARP-bearing Treg cells and one or more activated T cell subtypes (especially CD8+ CD40L+). In conclusion, increased T helper and cytotoxic cell activation along with decreased Treg homeostatic defenses are important parts of MDD that lead to enhanced immune responses and, as a result, neuroimmunotoxicity.

Quercetin exerts anti-tumor immune mechanism by regulating IL-6/JAK2/STAT3 signaling pathway to deplete Treg cells.

Breast cancer is still the leading cause of death among women worldwide. Due to the lack of effective drug targets, triple-negative breast cancer has a worse prognosis and higher mortality compared with other types of breast cancer, and chemotherapy is still the main treatment for triple-negative breast cancer at present. Quercetin (QUE) is a flavonoid compound found in a variety of fruits and vegetables. The mechanism of QUE has been extensively studied, such as prostate cancer, colon cancer, ovarian cancer, etc. However, the anti-tumor immune mechanism of QUE in triple-negative breast cancer remains unclear. Therefore, we assessed the anti-tumor immune effects of QUE on triple-negative breast cancer using both 4T1 cells and a xenograft mouse model of 4T1 cells. In vitro, we examined the inhibitory effects of QUE on 4T1 cells and its molecular mechanisms through MTT, Transwell, ELISA, and Western blotting. In vivo, by establishing a xenograft mouse model, we utilized flow cytometry, immunohistochemistry, ELISA, and Western blotting to evaluate the anti-tumor immune effects of QUE on triple-negative breast cancer. The results indicate that QUE inhibits the proliferation, migration, and invasion of 4T1 cells, concurrently significantly suppressing the IL-6/JAK2/STAT3 signaling pathway. Furthermore, it depletes Treg cell content in 4T1 xenograft mice, thereby improving the tumor immune microenvironment and promoting the cytotoxicity of relevant tumor immune cells. These findings suggest that QUE may serve as a potential adjuvant for immune therapy in triple-negative breast cancer.