Optimization of Tissue Digestion Methods for Characterization of Photoaged Skin by Single Cell RNA Sequencing Reveals Preferential Enrichment of T Cell Subsets.

Healthy human skin tissue is often used as a control for comparison to diseased skin in patients with skin pathologies, including skin cancers or other inflammatory conditions such as atopic dermatitis or psoriasis. Although non-affected skin from these patients is a more appropriate choice for comparison, there is a paucity of studies examining such tissue. This lack is exacerbated by the difficulty of processing skin tissue for experimental analysis. In addition, choosing a processing protocol for skin tissue which preserves cell viability and identity while sufficiently dissociating cells for single-cell analysis is not a trivial task. Here, we compare three digestion methods for human skin tissue, evaluating the cell yield and viability for each protocol. We find that the use of a sequential dissociation method with multiple enzymatic digestion steps produces the highest cell viability. Using single-cell sequencing, we show this method results in a relative increase in the proportion of non-antigen-presenting mast cells and CD8 T cells as well as a relative decrease in the proportion of antigen-presenting mast cells and KYNU+ CD4 T cells. Overall, our findings support the use of this sequential digestion method on freshly processed human skin samples for optimal cell yield and viability.

EBV-induced T-cell responses in EBV-specific and nonspecific cancers.

Epstein-Barr virus (EBV) is a ubiquitous human tumor virus associated with various malignancies, including B-lymphoma, NK and T-lymphoma, and epithelial carcinoma. It infects B lymphocytes and epithelial cells within the oropharynx and establishes persistent infection in memory B cells. With a balanced virus-host interaction, most individuals carry EBV asymptomatically because of the lifelong surveillance by T cell immunity against EBV. A stable anti-EBV T cell repertoire is maintained in memory at high frequency in the blood throughout persistent EBV infection. Patients with impaired T cell immunity are more likely to develop life-threatening lymphoproliferative disorders, highlighting the critical role of T cells in achieving the EBV-host balance. Recent studies reveal that the EBV protein, LMP1, triggers robust T-cell responses against multiple tumor-associated antigens (TAAs) in B cells. Additionally, EBV-specific T cells have been identified in EBV-unrelated cancers, raising questions about their role in antitumor immunity. Herein, we summarize T-cell responses in EBV-related cancers, considering latency patterns, host immune status, and factors like human leukocyte antigen (HLA) susceptibility, which may affect immune outcomes. We discuss EBV-induced TAA-specific T cell responses and explore the potential roles of EBV-specific T cell subsets in tumor microenvironments. We also describe T-cell immunotherapy strategies that harness EBV antigens, ranging from EBV-specific T cells to T cell receptor-engineered T cells. Lastly, we discuss the involvement of γδ T-cells in EBV infection and associated diseases, aiming to elucidate the comprehensive interplay between EBV and T-cell immunity.

Immunohistochemical expression of T-cell subsets (CD4 and CD8) in oral lichen planus.

Introduction: oral lichen planus (OLP) is a common oral mucosal disease with various clinical manifestations. The most predominant types are reticular and erosive. Despite extensive research on the causes of OLP, the exact etiology remains unclear. However, it is believed that a T-cell-mediated response, which triggers the apoptosis of oral epithelial cells, may contribute to the development of this disorder. This study aims to investigate the different types of T-cells (specifically CD4 and CD8) present in OLP tissue samples. By using immunohistochemistry, the expressions of cluster of differentiation 4 (CD4) and cluster of differentiation 8 (CD8) will be evaluated in biopsy samples taken from OLP patients who exhibit various clinical presentations. Methods: this study was a retrospective analysis study. Oral lichen planus was established histologically in forty paraffin-embedded tissue samples. Blocks of OLP were diagnosed and characterized as reticular or erosive. Immunohistochemical staining was conducted using a monoclonal antibody for (CD4) and a polyclonal antibody for CD8. Semi-quantitative techniques were used to analyze the patterns of positively stained cells. Results: forty biopsies of OLP cases were obtained from 24 females and 16 males. The mean age was (49.15±11.39) years. Using an immunohistochemical method, the proportion of CD4 expression: CD8 expression among the epithelial-connective tissue interface was shown to be 24 (60%) cases with a predominance of CD8, 9 (22.5%) cases with no difference, and only 7 (17.5%) cases with a predominance of CD4. The proportion of CD4: CD8 among perivascular parts was shown to be 8 (20%) cases with a predominance of CD8, 20 (50%) cases with no difference, while only 12 (30%) cases had a predominance of CD4. The CD4 perivascular expression was significantly stronger in (71.4%) of erosive OLP than in reticular cases. Conclusion: T-cell subsets (CD4 and CD8) were found in the OLP infiltrates. The correlation may have contributed to the pathogenesis of OLP.

Disturbance in the reconstitution of distinct T-cell subsets and the incidence of GvHD following allo-HSCT in pediatric patients with non-malignant hematological disorders.

BACKGROUND: The reconstitution of different T-cell subsets following allogeneic hematopoietic stem cell transplantation (allo-HSCT) is critical for efficient pathogen protection and the prevention of graft-versus-host disease (GvHD). In particular, studies have highlighted the importance of balanced ratios of regulatory T-cells (Tregs) and distinct functionally T-cells in preventing acute and chronic GvHD. METHODS: We evaluated the regeneration of CD4 and CD8 T-cell subpopulations in nine pediatric patients with non-malignant disorders following allo-HSCT from a fully HLA-identical donor. RESULTS: CD4 and CD8 T-cells were higher 12 months after the transplant but still lower than in healthy controls and pre-transplant. However, we found after allo-HSCT, central memory and effector memory cell subsets were the predominant phenotypes in the CD4 and CD8 T-cell populations, respectively. In patients who had developed acute GvHD, there was an increase in the frequency of TEMRA (effector memory T cells that re-express CD45RA) cells within the CD4 T-cell population. Meanwhile, in patients with chronic GvHD, we observed a decrease in Th1 cells in CD4 T-cells and effector memory cells within the CD8 T-cell population. In addition, we found decreased TEMRA cell subsets in CD4 and CD8 T-cell populations in chronic GvHD. CONCLUSION: Our findings suggest a possible relationship between the influence of acute GvHD and its prevention on delayed CD4 T-cell reconstitution and, reciprocally, unbalanced regeneration of CD4 and CD8 T-cell subsets in the development of chronic GvHD.

Spatial distribution of tumor-infiltrating T cells indicated immune response status under chemoradiotherapy plus PD-1 blockade in esophageal cancer.

Background: The spatial distribution of tumor-infiltrating T cells and its dynamics during chemoradiotherapy combined with PD-1 blockade is little known in esophageal squamous cell carcinoma (ESCC). Methods: We applied the multiplex immunofluorescence method to identify T cells (CD4+, CD8+ T cells, and their PD-1- or PD-1+ subsets) and myeloid-derived cells (CD11c+ dendritic cells, CD68+ macrophages, and their PD-L1+ subpopulations) in paired tumor biopsies (n = 36) collected at baseline and during combination (40 Gy of radiation) from a phase Ib trial (NCT03671265) of ESCC patients treated with first-line chemoradiotherapy plus anti-PD-1 antibody camrelizumab. We used the FoundationOne CDx assay to evaluate tumor mutational burden (TMB) in baseline tumor biopsies (n = 14). We dynamically assessed the nearest distance and proximity of T-cell subsets to tumor cells under combination and estimated the association between T-cell spatial distribution and combination outcome, myeloid-derived subsets, TMB, and patient baseline characteristics. Findings: We found that the tumor compartment had lower T-cell subsets than the stromal compartment but maintained a comparable level under combination. Both before and under combination, PD-1- T cells were located closer than PD-1+ T cells to tumor cells; T cells, dendritic cells, and macrophages showed the highest accumulation in the 5-10-µm distance. Higher CD4+ T cells in the tumor compartment and a shorter nearest distance of T-cell subsets at baseline predicted poor OS. Higher baseline CD4+ T cells, dendritic cells, and macrophages were associated with worse OS in less than 10-µm distance to tumor cells, but related with better OS in the farther distance. Higher on-treatment PD-1-positive-expressed CD4+ and CD8+ T cells within the 100-µm distance to tumor cells predicted longer OS. T cells, dendritic cells, and macrophages showed a positive spatial correlation. Both high TMB and smoking history were associated with a closer location of T cells to tumor cells at baseline. Conclusions: We firstly illustrated the T-cell spatial distribution in ESCC. Combining chemoradiotherapy with PD-1 blockade could improve the antitumor immune microenvironment, which benefits the treatment outcome. Further understanding the precision spatiality of tumor-infiltrating T cells would provide new evidence for the tumor immune microenvironment and for the combination treatment with immunotherapy.

Interleukin-27 potentiates CD8+ T-cell-mediated antitumor immunity in chronic lymphocytic leukemia.

Chronic lymphocytic leukemia (CLL) cells are highly dependent on interactions with the immunosuppressive tumor microenvironment (TME) for survival and proliferation. In the search for novel treatments, pro-inflammatory cytokines have emerged as candidates to reactivate the immune system. Among those, interleukin 27 (IL-27) has recently gained attention, but its effects differ among malignancies. Here, we utilized the Eµ-TCL1 and EBI3 knock-out mouse models as well as clinical samples from patients to investigate the role of IL-27 in CLL. Characterization of murine leukemic spleens revealed that the absence of IL-27 leads to enhanced CLL development and a more immunosuppressive TME in transgenic mice. Gene-profiling of T-cell subsets from EBI3 knock-out highlighted transcriptional changes in the CD8+ T-cell population associated with T-cell activation, proliferation, and cytotoxicity. We also observed an increased anti-tumor activity of CD8+ T cells in the presence of IL-27 ex vivo with murine and clinical samples. Notably, IL-27 treatment led to the reactivation of autologous T cells from CLL patients. Finally, we detected a decrease in IL-27 serum levels during CLL development in both pre-clinical and patient samples. Altogether, we demonstrated that IL-27 has a strong anti-tumorigenic role in CLL and postulate this cytokine as a promising treatment or adjuvant for this malignancy.

OX40 shapes an inflamed tumor immune microenvironment and predicts response to immunochemotherapy in diffuse large B-cell lymphoma.

OX40 enhances the T-cell activation via costimulatory signaling. However, its molecular characteristics and value in predicting response to immunochemotherapy in DLBCL remain largely unexplored. Here, we performed an integrative analysis of sequencing and multiplex immunofluorescence staining, and discovered abnormally higher expression of OX40 in DLBCL patients. Elevated OX40 could activate T cells leading to a higher immune score for tumor immune microenvironment (TiME). OX40 upregulation simultaneously happened with immune-related genes including PD-1, CTLA4 and TIGIT et,al. Patients with high OX40 expression exhibited a lower Ann Arbor stage and IPI score and more easily achieved a complete response/partial response. The analysis of infiltrated T-cell subset revealed that patients with a greater number of CD4+/OX40+ or CD8+/OX40+ T cells had a longer OS. Our findings indicated that OX40 shapes an inflamed tumor immune microenvironment and predicts response to immunochemotherapy, providing insights for the application of OX40 agonist in DLBCL patients.

[Evaluation of peripheral blood T-lymphocyte subpopulations features in patients with hepatitis B virus-related acute-on-chronic liver failure based on single-cell sequencing technology].

Objective: T lymphocyte exhaustion is an important component of immune dysfunction. Therefore, exploring peripheral blood-exhausted T lymphocyte features in patients with hepatitis B virus-related acute-on-chronic liver failure may provide potential therapeutic target molecules for ACLF immune dysfunction. Methods: Six cases with HBV-ACLF and three healthy controls were selected for T-cell heterogeneity detection using the single-cell RNA sequencing method. In addition, exhausted T lymphocyte subpopulations were screened to analyze their gene expression features, and their developmental trajectories quasi-timing. An independent sample t-test was used to compare the samples between the two groups. Results: Peripheral blood T lymphocytes in HBV-ACLF patients had different differentiation trajectories with different features distinct into eight subpopulations. Among them, the CD4(+)TIGIT(+) subsets (P = 0.007) and CD8(+)LAG3(+) (P = 0.010) subsets with highly exhausted genes were significantly higher than those in healthy controls. Quasi-time analysis showed that CD4(+)TIGIT(+) and CD8(+)LAG3(+) subsets appeared in the late stage of T lymphocyte differentiation, suggesting the transition of T lymphocyte from naïve-effector-exhausted during ACLF pathogenesis. Conclusion: There is heterogeneity in peripheral blood T lymphocyte differentiation in patients with HBV-ACLF, and the number of exhausted T cells featured by CD4(+)TIGIT(+)T cell and CD8(+)LAG3(+) T cell subsets increases significantly, suggesting that T lymphocyte immune exhaustion is involved in the immune dysfunction of HBV-ACLF, thereby identifying potential effective target molecules for improving ACLF patients' immune function.

SUV39H1 is a prognosis and immune microenvironment-related biomarker in diffuse large B-cell lymphoma.

BACKGROUND: The tumor microenvironment plays a crucial role in the oncogenesis and treatment of diffuse large B-cell lymphoma (DLBCL). The H3K9me3-specific histone methyltransferase Suppressor of variegation 3-9 homolog 1 (SUV39H1) is a significant gene that promotes the progression of various malignancies. However, the specific expression of SUV39H1 in DLBCL remains unclear. METHODS: By retrieving data from GEPIA, UCSC XENA and TCGA public databases, we observed the high expression of SUV39H1 in DLBCL. Combined with an immunohistochemical validation assay, we analyzed our hospital's clinical characteristics and prognosis of 67 DLBCL patients. The results showed that high SUV39H1 expression was closely associated with age over 50 years (P = 0.014) and low albumin levels (P = 0.023) of patients. Furthermore, the experiments in vitro were deployed to evaluate the regulation of SUV39H1 on the DLBCL immune microenvironment. RESULTS: The results showed that high SUV39H1 expression was closely associated with age over 50 years (P = 0.014) and low albumin levels (P = 0.023) of patients. The prognostic analysis showed that the high SUV39H1 expression group had a lower disease-free survival (DFS) rate than the low SUV39H1 expression group (P < 0.05). We further discovered that SUV39H1 upregulated the expression of CD86+ and CD163+ tumor-associated macrophages by DLBCL patients' tissues and cell experiments in vitro (P < 0.05). And SUV39H1-associated T lymphocyte subsets and cytokines IL-6/CCL-2 were downregulated in DLBCL (P < 0.05). CONCLUSIONS: In summary, SUV39H1 might be not only a potential target for treating DLBCL but also a clinical indicator for doctors to evaluate the trend of disease development.

Changes in T lymphocyte subsets in peripheral blood of patients with middle-advanced cervical cancer before and after nimotuzumab combined with concurrent chemoradiotherapy.

The current study sought to investigate the effect of nimotuzumab combined with concurrent chemoradiotherapy (CCRT + Nim) on T lymphocyte subsets in middle-advanced CC. Firstly, patients with middle-advanced CC were administered CCRT or CCRT + Nim. Next, levels of T lymphocytes in peripheral blood of CC patients pre- or post-treatment and healthy females were determined by flow cytometry. The short-term efficacy was evaluated, and overall survival (OS) and progression-free survival (PFS) of patients were recorded. In addition, the correlation of T lymphocyte subsets post-treatment with OS/PFS was assessed with Pearson analysis. CC patients exhibited decreased total T cells/T helper cells/CD4+/CD8+ ratio and increased T suppressor cells/Tregs in peripheral blood. Meanwhile, CCRT and CCRT + Nim improved T lymphocyte subset imbalance, with CCRT + Nim exhibiting better efficacy. CCRT + Nim exhibited better short-term efficacy and higher PFS than CCRT, with no evident difference in OS. The levels of total T cells/T helper cells/T suppressor cells/Tregs were not significantly-correlated with OS/PFS, and the CD4+/CD8+ ratio was correlated with PFS but not OS. Collectively, CCRT + nimotuzumab ameliorate the imbalance of T lymphocyte subsets in peripheral blood of middle-advanced CC patients, and the CD4+/CD8+ ratio after therapy is correlated with PFS.IMPACT STATEMENTWhat is already known on this subject? The utilisation of Nimotuzumab targeting epidermal growth factor receptor (EGFR) combined with concurrent chemoradiotherapy (CCRT) as an efficient treatment for middle-advanced cervical cancer (CC) has garnered the attention of numerous researchers over the years. T cells represent a major immune cell type in the tumour microenvironment and serve as the basis for maintaining cellular immune functions.What do the results of this study add? Our findings revealed that nimotuzumab combined with CCRT improves the abnormality of T lymphocyte subsets in peripheral blood of patients with middle-advanced CC, such that the CD4+/CD8+ ratio after treatment was significantly correlated with progression-free survival (PFS).What are the implications of these findings for clinical practice and/or further research? CCRT of CC may have a short-term negative impact on the peripheral T-cell immune micro-environment, and the combination of nimotuzumab, cisplatin-based chemotherapy, and radiotherapy enhances the frequency of Tregs in peripheral blood. Our findings illustrated that nimotuzumab combined with CCRT can improve the imbalance of T lymphocyte subsets in peripheral blood of patients with middle-advanced CC. A better understanding of the mechanisms of these therapies will optimise the selection of patients most likely to benefit from treatment, serving as a reference for further research on the relationship between EGFR-specific T cells and clinical benefit in patients treated with nimotuzumab in combination with CCRT.

CD103 and CD39 coexpression identifies neoantigen-specific cytotoxic T cells in colorectal cancers with low mutation burden.

BACKGROUND: Expression of CD103 and CD39 has been found to pinpoint tumor-reactive CD8+ T cells in a variety of solid cancers. We aimed to investigate whether these markers specifically identify neoantigen-specific T cells in colorectal cancers (CRCs) with low mutation burden. EXPERIMENTAL DESIGN: Whole-exome and RNA sequencing of 11 mismatch repair-proficient (MMR-proficient) CRCs and corresponding healthy tissues were performed to determine the presence of putative neoantigens. In parallel, tumor-infiltrating lymphocytes (TILs) were cultured from the tumor fragments and, in parallel, CD8+ T cells were flow-sorted from their respective tumor digests based on single or combined expression of CD103 and CD39. Each subset was expanded and subsequently interrogated for neoantigen-directed reactivity with synthetic peptides. Neoantigen-directed reactivity was determined by flow cytometric analyses of T cell activation markers and ELISA-based detection of IFN-γ and granzyme B release. Additionally, imaging mass cytometry was applied to investigate the localization of CD103+CD39+ cytotoxic T cells in tumors. RESULTS: Neoantigen-directed reactivity was only encountered in bulk TIL populations and CD103+CD39+ (double positive, DP) CD8+ T cell subsets but never in double-negative or single-positive subsets. Neoantigen-reactivity detected in bulk TIL but not in DP CD8+ T cells could be attributed to CD4+ T cells. CD8+ T cells that were located in direct contact with cancer cells in tumor tissues were enriched for CD103 and CD39 expression. CONCLUSION: Coexpression of CD103 and CD39 is characteristic of neoantigen-specific CD8+ T cells in MMR-proficient CRCs with low mutation burden. The exploitation of these subsets in the context of adoptive T cell transfer or engineered T cell receptor therapies is a promising avenue to extend the benefits of immunotherapy to an increasing number of CRC patients.

Age-related changes in CD4+ T and NK cell compartments may contribute to the occurrence of pregnancy loss in advanced maternal age.

A recent study characterized novel immune cell subsets (T, NK, and γδ T cell subsets) related to recurrent pregnancy loss (RPL). This study aims to assess whether these RPL-related immune cell subsets are affected by aging. The percentages of peripheral blood immunes cells from nulligravida women (NGW), women with a history of normal pregnancy (NP), and women with a history of pregnancy loss (PL) were detected by flow cytometry. The correlations between maternal age and cell percentages were assessed. We found a significant positive correlation between PL and maternal age. The percentages of effector memory CD4+ T (CD3+ CD4+ CD45RA¯ CCR7¯), terminally differentiated CD4+ T (CD3+ CD4+ CD45RA+ CCR7¯), and mature NK cells (CD3¯ CD56+lo) significantly increased with maternal age. A significant decrease in the percentage of Naïve CD4+ T cells (CD3+ CD4+ CD45RA+ CCR7+) with age was observed in women from the NP group. Women aged 35 or older had significantly higher percentages of effector memory CD4+ T cells, terminally differentiated CD4+ T cells, and mature NK cells than younger women. Maternal age positively correlates with terminally differentiated CD4+ T, effector memory CD4+ T, and mature NK cell percentages. In contrast, an inverse correlation was observed between Naïve CD4+ T cell and age among women from the NP group. Our findings indicate that age-related CD4+ T and NK cell dysregulation might be involved in the pathogenesis of PL in women with advanced maternal age. The underlying mechanism needs further investigation.

Unbalanced T-cell subsets in pediatric patients with beta-thalassemia.

BACKGROUND: Beta-thalassemia major is an autosomal recessive disorder in hemoglobin synthesis. Ineffective erythropoiesis is the main characteristic of the disease, which results in anemia following the extensive destruction of red blood cells. Chronic antigenic stimulation following frequent blood transfusions lead to immune abnormalities, especially regarding T cells, which is one of the reasons for the high susceptibility to infection in beta-thalassemia. METHODS: Six pediatric patients and six age- and sex-matched healthy children were selected. Immunophenotyping of functional T-cells was performed using flow cytometry with staining for surface and intracellular markers. The proliferative response of T lymphocytes was also investigated after labeling with CFSE and following stimulation with anti-CD3 and anti-CD28. RESULTS: Examination of T lymphocyte subpopulations showed a significant increase in regulatory T cells (Tregs) in beta-thalassemia patients. Hence, the Treg:Tcons (conventional T cells) and Treg:CD8 ratios were significantly increased. In addition, a significant increase in CD8 T cell proliferation activity was observed. Multivariate analysis showed a significant association of central memory cells with serum ferritin levels and the duration of transfusion. In particular, patients with cytomegalovirus (CMV) infection exhibited a significant increase in CD4 central memory cells. CONCLUSION: Patients with beta-thalassemia have functionally distinct CD4 and CD8 T cell subsets imbalances, and this may contribute to their high susceptibility to infections and immune dysregulation.

Distinct T cell subsets in adipose tissue are associated with obesity.

Adipose tissue inflammation is a driving factor for the development of obesity-associated metabolic disturbances, and a role of adipose tissue T cells in initiating the pro-inflammatory signaling is emerging. However, data on human adipose tissue T cells in obesity are limited, reflected by the lack of phenotypic markers to define tissue-resident T cell subsets. In this study, we performed a deep characterization of T cells in blood and adipose tissue depots using multicolor flow cytometry and RNA sequencing. We identified distinct subsets of T cells associated with obesity expressing the activation markers, CD26 and CCR5, and obesity-specific genes that are potentially engaged in activating pro-inflammatory pathway, including ceramide signaling, autophagy, and IL-6 signaling. These findings increase our knowledge on the heterogeneity of T cells in adipose tissue and on subsets that may play a role in obesity-related pathogenesis.

Targeting T cell checkpoints 41BB and LAG3 and myeloid cell CXCR1/CXCR2 results in antitumor immunity and durable response in pancreatic cancer.

Pancreatic ductal adenocarcinoma (PDAC) is considered non-immunogenic, with trials showing its recalcitrance to PD1 and CTLA4 immune checkpoint therapies (ICTs). Here, we sought to systematically characterize the mechanisms underlying de novo ICT resistance and to identify effective therapeutic options for PDAC. We report that agonist 41BB and antagonist LAG3 ICT alone and in combination, increased survival and antitumor immunity, characterized by modulating T cell subsets with antitumor activity, increased T cell clonality and diversification, decreased immunosuppressive myeloid cells and increased antigen presentation/decreased immunosuppressive capability of myeloid cells. Translational analyses confirmed the expression of 41BB and LAG3 in human PDAC. Since single and dual ICTs were not curative, T cell-activating ICTs were combined with a CXCR1/2 inhibitor targeting immunosuppressive myeloid cells. Triple therapy resulted in durable complete responses. Given similar profiles in human PDAC and the availability of these agents for clinical testing, our findings provide a testable hypothesis for this lethal disease.

Exosomal miR-125a-5p regulates T lymphocyte subsets to promote silica-induced pulmonary fibrosis by targeting TRAF6.

Silicosis caused by long-term inhalation of crystalline silica during occupational activities seriously threatens the health of occupational populations. Imbalances in T helper 1(Th1), Th2, Th17, and regulatory T cells (Tregs) promote the development of pulmonary silicosis. Exosomes and their contents, especially microRNAs (miRNAs), represent a new type of intercellular signal transmission mediator related to various diseases including pulmonary fibrosis. However, whether exosomal miRNAs can affect the progression of silicosis by regulating T cell differentiation remains to be determined. To test this hypothesis, we established a miR-125a-5p antagomir mouse model and examined changes in miR-125a-5p levels and T cell subtypes. We found that miR-125a-5p levels were increased in lung tissues and serum exosomes in the silica group at 7 days and 28 days. Downregulation of miR-125a-5p attenuated α-smooth muscle actin (α-SMA), collagen I, fibronectin, p-p65, and p-inhibitor of nuclear factor kappa B (NF-κB) kinase (IKK) protein expression, while tumor necrosis factor receptor-associated factor 6 (TRAF6) and p-inhibitor of κBα (IKBα) expression were increased. MiR-125a-5p anta-miR treatment contributes to the maintenance of Th1/Th2 balance during the progression of pulmonary fibrosis. Our findings indicated that knockdown miR-125a-5p could regulate T lymphocyte subsets and significantly reduce pulmonary fibrosis by targeting TRAF6.

Evaluation of peripheral blood T-lymphocyte subpopulations features in patients with hepatitis B virus-related acute-on-chronic liver failure based on single-cell sequencing technology / 中华肝脏病杂志

Objective: T lymphocyte exhaustion is an important component of immune dysfunction. Therefore, exploring peripheral blood-exhausted T lymphocyte features in patients with hepatitis B virus-related acute-on-chronic liver failure may provide potential therapeutic target molecules for ACLF immune dysfunction. Methods: Six cases with HBV-ACLF and three healthy controls were selected for T-cell heterogeneity detection using the single-cell RNA sequencing method. In addition, exhausted T lymphocyte subpopulations were screened to analyze their gene expression features, and their developmental trajectories quasi-timing. An independent sample t-test was used to compare the samples between the two groups. Results: Peripheral blood T lymphocytes in HBV-ACLF patients had different differentiation trajectories with different features distinct into eight subpopulations. Among them, the CD4(+)TIGIT(+) subsets (P = 0.007) and CD8(+)LAG3(+) (P = 0.010) subsets with highly exhausted genes were significantly higher than those in healthy controls. Quasi-time analysis showed that CD4(+)TIGIT(+) and CD8(+)LAG3(+) subsets appeared in the late stage of T lymphocyte differentiation, suggesting the transition of T lymphocyte from naïve-effector-exhausted during ACLF pathogenesis. Conclusion: There is heterogeneity in peripheral blood T lymphocyte differentiation in patients with HBV-ACLF, and the number of exhausted T cells featured by CD4(+)TIGIT(+)T cell and CD8(+)LAG3(+) T cell subsets increases significantly, suggesting that T lymphocyte immune exhaustion is involved in the immune dysfunction of HBV-ACLF, thereby identifying potential effective target molecules for improving ACLF patients' immune function.

Delineation of T cell subsets in chronic rhinosinusitis with nasal polyps.

Objectives: Detailed characterisation of CD45RA+ and CD45RO+ T cell subsets in both CD4+ and CD8+ cells in blood and tissue of patients with chronic rhinosinusitis with nasal polyps (CRSwNP) is lacking. We aimed to investigate T cell differentiation levels in blood and tissue and correlate this with CT score and white blood cell count. Methods: This case-control study was conducted on 21 patients with CRSwNP and 20 healthy controls. Lund-Mackay score was used for radiologic staging of chronic rhinosinusitis. T cell subsets were characterised in blood and nasal polyp tissue using CD4, CD8, CCR7, CD45RA, CD45RO, CD27, and CD95 monoclonal antibodies. Results: Most variations in T cell subsets were detected in tissue rather than blood. A higher level of CD8+ TCM, CD4+ TSCMs and lower level of CD8+ TEM were detected in the blood of CRSwNP patients vs controls. In CRSwNP patients, substantial decrease of tissue CD8+ TNs, CD8+ TCMs, CD4+ TNs, and CD4+TSCMs with marked increase in the percentages of TEMs and TEMRAs were detected among CD4+ and CD8+ T cells compared with levels in blood. Conclusions: The reduction of TNs, TSCMs, and TCMs and accumulation of TEMs and TEMRAs subsets that function as effector cells in sinonasal mucosa, especially CD8+ T cells, might indicate the role of immunomodulation of memory/effector T-cells in the pathogenesis of CRSwNP.

Differential distribution and prognostic value of CD4+ T cell subsets before and after radioactive iodine therapy in differentiated thyroid cancer with varied curative outcomes.

Differentiated thyroid cancer is the most frequently diagnosed endocrine tumor. While differentiated thyroid cancers often respond to initial treatment, little is known about the differences in circulating immune cells amongst patients who respond differently. A prospective study of 39 patients with differentiated thyroid cancer was conducted. Serum thyroglobulin levels and thyroid and immunological functions were tested before and after radioactive iodine treatment (RAIT). Efficacy assessments were performed 6 to 12 months after radioactive iodine treatment. Most patients showed an excellent response to radioactive iodine treatment. Before radioactive iodine treatment, the excellent response group had considerably fewer circulating CD4+ T cell subsets than the non-excellent response group. Both the excellent response and non-excellent response groups had considerably lower circulating CD4+ T lymphocyte subsets 30 days after radioactive iodine treatment, but those of the excellent response group were still lower than those of the non-excellent response group. All circulating CD4+ T cell subsets in the excellent response group rose by varying degrees by the 90th day, but only Treg cell amounts increased in the non-excellent response group. Interestingly, in the non-excellent response group, we noticed a steady drop in Th1 cells. However, the bulk of circulating CD4+ T cell subsets between the two groups did not differ appreciably by the 90th day. Finally, we discovered that CD4+ T cell subsets had strong predictive potential, and we thus developed high-predictive-performance models that deliver more dependable prognostic information. In conclusion, in individuals with differentiated thyroid cancer, there is great variation in circulating immune cells, resulting in distinct treatment outcomes. Low absolute CD4+ T cell counts is linked to improved clinical outcomes as well as stronger adaptive and resilience capacities.

Variable frequencies of peripheral T-lymphocyte subsets in the diabetes spectrum from type 1 diabetes through latent autoimmune diabetes in adults (LADA) to type 2 diabetes.

T lymphocytes are key players in the pathogenesis of autoimmune diabetes. We recruited subjects with T1D (n=81), LADA (n=82), T2D (n=95) and NGT (n=218) and analyzed the percentages of T-lymphocyte subsets, including T helper 1 (Th1), T helper 2 (Th2), T helper 17 (Th17), T cytotoxic 1 (Tc1), regulatory T cells (Tregs), effector T (Teff), naïve T, central memory T (Tcm), and effector memory T (Tem) cells by flow cytometry. LADA patients possessed similar frequencies of IFN-γ+CD4+ T (Th1), IFN-γ+CD8+ T and CD4+ Teff cells compared with T1D patients, but much lower than those of NGT subjects. Like T2D patients, LADA patients had increased frequencies of CD4+ Tem and CD8+ Tem cells with respect to T1D and NGT subjects. In LADA patients, Th2 cells were decreased while CD4+ Tcm cells were increased compared with NGT subjects. Notably, we observed significant negative correlations between the CD4+ Tcm cell frequency and C-peptide in LADA subjects. These data demonstrates that LADA patients possess T-cell subset changes resembling both T1D and T2D and represent the middle of the diabetes spectrum between T1D and T2D. Based on these T-cell subset alterations, we speculate that autoimmunity-induced ß-cell destruction and inflammation-induced insulin resistance might both be involved in the pathogenesis of LADA.