Senescent CD8 Research Articles

Lesional senescent CD4+ T cells mediate bystander cytolysis and contribute to the skin pathology of human cutaneous leishmaniasis.

Cytotoxic activity is a hallmark of the immunopathogenesis in human cutaneous leishmaniasis (CL). In this study, we identified accumulation of CD4+ granzyme B producing T cells with increased cytotoxic capacity in CL lesions. These cells showed enhanced expression of activating NK receptors (NKG2D and NKG2C), diminished expression of inhibitory NKG2A, along with the upregulation of the senescence marker CD57. Notably, CD4+ T cells freshly isolated from CL lesions demonstrated remarkable capacity to mediate NL-like bystander cytolysis. Phenotypic analyses revealed that lesional CD4+ T cells are mainly composed of late-differentiated effector (CD27-CD45RA-) and terminally differentiated (senescent) TEMRA (CD27-CD45RA+) subsets. Interestingly, the TEMRA CD4+ T cells exhibited higher expression of granzyme B and CD107a. Collectively, our results provide the first evidence that senescent cytotoxic CD4+ T cells may support the skin pathology of human cutaneous leishmaniasis and, together with our previous findings, support the notion that multiple subsets of cytotoxic senescent cells may be involved in inducing the skin lesions in these patients.

Quercetin ameliorates lupus symptoms by promoting the apoptosis of senescent Tfh cells via the Bcl-2 pathway

Systemic lupus erythematosus (SLE) is an autoimmune disorder that commonly affects the skin, kidneys, joints, and various other systemic tissues, with its development intricately linked to the process of immunosenescence. Quercetin (QC), a phytochemical that occurs naturally, demonstrates many different biological capabilities, such as antibacterial, antioxidant, and anti-inflammatory activities. Our investigation found that QC effectively reduced kidney damage and relieved mesenteric lymph nodes (mLNs) swelling in MRL/lpr lupus mice. Moreover, QC has been found to decrease the number of senescent follicular helper T (Tfh) cells, a pivotal kind of T cells that contribute to the progression of SLE. In vitro, QC exhibited the capacity to modulate mRNA expression levels, with the downregulation of IL-6, IL21-AS1, IL-27, BCL6, and BCL2L12, and the upregulation of FOXP1 and BIM. This modulation resulted in the suppression of Tfh cells differentiation and the enhancement of apoptosis in senescent CD4+ T cells. In addition, the HuProtTM Human Proteome Microarray revealed that QC can directly bind to BCL-2 protein and therefore promote the apoptosis of senescent CD4+ T cell. As a result, our investigative elucidate the potent inhibitory action of QC on the ontogeny of Tfh cells, along with its capacity to abrogate the immunosenescent phenotype. This positions QC as a promising therapeutic strategy for treating SLE.

Senescent CD8+ T cells: a novel risk factor in atrial fibrillation.

Immune cell alterations may play a role in the development of atrial fibrillation (AF). Our objective was to comprehensively characterize immune cells in AF, and investigate the potential mechanisms. Single-cell RNA sequencing and multicolor flow cytometry revealed that T cells constituted the most significant subset alterations in AF, and senescent CD8+ T cells were AF-associated subset. Senescent CD8+ T cells increased in both peripheral veins (p < 0.0001) and the left atria (p < 0.05) in patients with AF compared to non-AF control. Senescent CD8+ T cells were independently associated with AF prevalence (odds ratio = 2.876, p < 0.05) and postprocedural recurrence (hazard ratio = 22.955, p < 0.0001) using a cross-sectional study and a subsequent prospective cohort study. Senescent CD8+ T cells secreted an increased amount of interferon (IFN)-γ, which induces Ca2+ handling abnormalities in human induced pluripotent stem cell-derived atrial cardiomyocytes, and translated into an increased susceptibility to AF assessed by heart optical mapping. An increased amount of senescent CD8+ T cells may be a hallmark of the immune senescence phenotype in AF and potentially serve as a valid biomarker for assessing prevalence and postprocedural recurrence of AF. By connecting immune senescence with electrophysiological disturbances in AF, this research provides a potential mechanism for the involvement of senescent CD8+ T cells in proarrhythmic calcium disorders and suggests novel avenues for developing new immune-modulatory and senolytic therapies for AF.

HIV reservoir and premature aging: risk factors for aging-associated illnesses in adolescents and young adults with perinatally acquired HIV.

Despite receiving antiretroviral therapy (ART), an increasing number of adolescents and young adults with perinatally acquired HIV (PHIVAYA) are at risk of developing premature senescence and aging-associated illnesses, including cancer. Given this concern, it is crucial to assess aging biomarkers and their correlation with the HIV reservoir in order to comprehensively characterize and monitor these individuals. Fifty-five PHIVAYA (median age: 23, interquartile range [IQR]: 20-27 years, and 21 [18-23] years on ART at the time of study sampling) were studied along with 23 age-matched healthy controls. The PHIVAYA exhibited significantly higher percentages of activated, senescent, exhausted CD4 and CD8 T cells, shorter telomeres, reduced thymic output, and higher levels of circulating inflammatory markers (PAMPs, DAMPs, and pro-inflammatory cytokines IL-6, IL-8, and TNFα) as well as denervation biomarkers (neural cell adhesion molecule 1 [NCAM1] and C-terminal Agrin fragment [CAF]), compared to controls. HIV-DNA levels positively correlated with activated, senescent, exhausted CD4 and CD8 T cells, circulating biomarkers levels, and inversely with regulatory T and B cells and telomere length. According to their viremia over time, PHIVAYA were subgrouped into 14 Not Suppressed (NS)-PHIVAYA and 41 Suppressed (S)-PHIVAYA, of whom 6 who initiated ART within one year of age and maintained sustained viral suppression overtime were defined as Early Suppressed (ES)-PHIVAYA and the other 35 as Late Suppressed (LS)-PHIVAYA. ES-PHIVAYA exhibited significantly lower HIV-DNA reservoir, decreased percentages of senescent and exhausted CD4 and CD8 T cells, reduced levels of circulating inflammatory and denervation biomarkers, but longer telomere compared to LS- and NS-PHIVAYA. They differed significantly from healthy controls only in a few markers, including higher percentages of regulatory T and B cells, and higher levels of DAMPs. Overall, these results underscore the importance of initiating ART early and maintaining viral suppression to limit the establishment of the viral reservoir and to counteract immune and cellular premature aging. These findings also suggest new approaches for minimally invasive monitoring of individuals at high risk of developing premature aging and age-related illnesses.

Implications of accumulation of clonally expanded and senescent CD4+GNLY+ T cells in immunological non-responders of HIV-1 infection

ABSTRACT Increased CD4+GNLY+ T cells have been confirmed to be inversely associated with CD4+ T cell count in immunological non-responders (INRs), however, the underlying mechanisms are unknown. This study aimed to elucidate the characteristics of CD4+GNLY+ T cells and their relationship with immune restoration. Single-cell RNA sequencing, single-cell TCR sequencing, and flow cytometry were used to analyze the frequency, phenotypes, and function of CD4+GNLY+ T cells. Moreover, Enzyme linked immunosorbent assay was performed to detect plasma cytokines production in patients. CD4+GNLY+ T cells were found to be highly clonally expanded, characterized by higher levels of cytotoxicity, senescence, P24, and HIV-1 DNA than CD4+GNLY− T cells. Additionally, the frequency of CD4+GNLY+ T cells increased after ART, and further increased in INRs, and were positively associated with the antiretroviral therapy duration in INR. Furthermore, increased IL-15 levels in INRs positively correlated with the frequency and senescence of CD4+GNLY+ T cells, suggesting that CD4+GNLY+ T cells may provide new insights for understanding the poor immune reconstitution of INRs. In conclusion, increased, highly clonally expanded, and senescent CD4+GNLY+ T cells may contribute to poor immune reconstitution in HIV-1 infection.

Reverse aging to treat lupus.

Systemic lupus erythematosus (SLE) is a complex autoimmune disease with multifaceted pathogenetic processes, including abnormalities of T-cell subset distribution and function. Accumulation of senescent CD4+ T cells has been found to contribute to the development of the disease. In this issue, Jiang etal. provide compelling evidence that links an expanded pool of CD4+CD57+ senescent T cells in patients with SLE to disease activity favored by interleukin-15. Importantly, treatment of lupus-prone mice with a senolytic drug resulted in decreased autoimmune pathology. The findings of this study suggest possible novel therapeutics to treat patients with SLE.

Topical application of a BCL-2 inhibitor ameliorates imiquimod-induced psoriasiform dermatitis by eliminating senescent cells

BackgroundPsoriasis is an inflammatory skin disease with unclear pathogenesis and unmet therapeutic needs. ObjectiveTo investigate the role of senescent CD4+ T cells in psoriatic lesion formation and explore the application of senolytics in treating psoriasis. MethodsWe explored the expression levels of p16INK4a and p21, classical markers of cellular senescence, in CD4+ T cells from human psoriatic lesions and imiquimod (IMQ)-induced psoriatic lesions. We prepared a senolytic gel using B-cell lymphoma 2 (BCL-2) inhibitor ABT-737 and evaluated its therapeutic efficacy in treating psoriasis. ResultsUsing multispectrum immunohistochemistry (mIHC) staining, we detected increased expression levels of p16INK4a and p21 in CD4+ T cells from psoriatic lesions. After topical application of ABT-737 gel, significant alleviation of IMQ-induced psoriatic lesions was observed, with milder pathological alterations. Mechanistically, ABT-737 gel significantly decreased the percentage of senescent cells, expression of T cell receptor (TCR) α and β chains, and expression of Tet methylcytosine dioxygenase 2 (Tet2) in IMQ-induced psoriatic lesions, as determined by mIHC, high-throughput sequencing of the TCR repertoire, and RT-qPCR, respectively. Furthermore, the severity of psoriatic lesions in CD4creTet2f/f mice was milder than that in Tet2f/f mice in the IMQ-induced psoriasis model. ConclusionWe revealed the roles of senescent CD4+ T cells in developing psoriasis and highlighted the therapeutic potential of topical ABT-737 gel in treating psoriasis through the elimination of senescent cells, modulation of the TCR αβ repertoire, and regulation of the TET2-Th17 cell pathway.

Metabolic shifts during coffee consumption refresh the immune response: insight from comprehensive multiomics analysis.

Coffee, a widely consumed beverage, has shown benefits for human health but lacks sufficient basic and clinical evidence to fully understand its impacts and mechanisms. Here, we conducted a cross-sectional observational study of coffee consumption and a 1-month clinical trial in humans. We found that coffee consumption significantly reshaped the immune system and metabolism, including reduced levels of inflammatory factors and a reduced frequency of senescent T cells. The frequency of senescent T cells and the levels of the senescence-associated secretory phenotype were lower in both long-term coffee consumers and new coffee consumers than in coffee nondrinking subjects, suggesting that coffee has anti-immunosenescence effects. Moreover, coffee consumption downregulated the activities of the The Janus kinase/signal transduction and activator of transcription (JAK/STAT) and mitogen-activated protein kinases (MAPK) signaling pathways and reduced systemic proinflammatory cytokine levels. Mechanistically, coffee-associated metabolites, such as 1-methylxanthine, 3-methylxanthine, paraxanthine, and ceramide, reduced the frequency of senescent CD4+CD57+ T cells in vitro. Finally, in vivo, coffee intake alleviated inflammation and immunosenescence in imiquimod-induced psoriasis-like mice. Our results provide novel evidence of the anti-inflammatory and anti-immunosenescence effects of coffee, suggesting that coffee consumption could be considered a healthy habit.

Association of circulating T lymphocytes and their functional states with pathologic complete response (pCR) in breast cancer (BC). A prospective Italian study.

559 Background: In BC, pCR is associated with substantial survival benefit. Hence, escalated neoadjuvant treatments (NATs) are being increasingly employed to improve pCR rates, warranting biomarkers for better patients’ selection. T-cell exhaustion and senescence are dysfunctional states that coexist in patients with cancer, impairing antitumor immunity and sustaining a suppressive immune microenvironment. Here, we assessed the impact of such states on pCR across different BC subtypes and NATs. Methods: This is a prospective study on patients undergoing NAT for BC at the Maggiore Hospital of Novara, Italy. Blood was collected at baseline and after NAT. Circulating T lymphocytes were characterized with exhausted (CD28, PD1, LAG3) and senescent (CD57, KLRG-1) biomarkers utilizing flow cytometry. Neutrophiles/lymphocytes ratio (NLR) and lymphocytes/monocytes ratio (LMR) were also retrieved. Subpopulations with frequency &lt;1% were normalized for total CD3+ cells. Results: From October 2020 to January 2024, 134 patients were enrolled and 115 completed NAT (85.8%). Fifty-one (38,1%) had adequate samples for analysis, 16 (31,4%) with hormone receptor positive HER2 negative BC (HR+HER2-), 15 (29,4%) with HER2+, and 20 (39,2%) with triple negative BC (TN). At baseline, patients with TNBC had the lowest NLR (p=0.018) and lowest senescent CD4+CD28- (p=0.042). Associations with pCR are shown in the Table. Overall, LMR associated with increased pCR (OR 1.34 [95% CIs 1.00-1.89]), and NLR with decreased pCR (OR 0.67 [0.39-1.04]). Both associations were driven by HR+ status (interaction p = 0.061 and 0.060). Senescent CD4+ predicted lower pCR in both HR-HER2+ (OR 0.10 [0.01-0.36]) and TNBC (OR 0.10 [0.01-0.5]). In TNBC, exhausted CD8+ predicted higher pCR (OR 1.78 [1.08-3.79]). When assessed separately in patients who received NAT with or without pembrolizumab, exhausted CD8+ remained predictor of pCR in chemotherapy + pembrolizumab (OR 1.84 [1.03-5.00]), but not with chemotherapy alone (p=0.203). The dynamics of paired pre-NAT and post-NAT TNBC samples revealed significant differences. Senescent CD4+ increased after chemotherapy, but not with chemotherapy + pembrolizumab (p=0.007). Pembrolizumab was associated with depletion of PD1-positive CD8+, while chemotherapy alone was not (p=0.020). Conclusions: The characterization of circulating T lymphocytes could help identify patients who benefit more in terms of pCR from specific NATs, including chemoimmunotherapy. [Table: see text]

The Role of Senescent CD8+T Cells in the Pathogenesis of Disseminated Leishmaniasis.

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.

Decreased but persistent epigenetic age acceleration is associated with changes in T-cell subsets after initiation of highly active antiretroviral therapy in persons living with HIV.

Persons living with HIV (PLWH) experience the early onset of age-related illnesses, even in the setting of successful human immunodeficiency virus (HIV) suppression with highly active antiretroviral therapy (HAART). HIV infection is associated with accelerated epigenetic aging as measured using DNA methylation (DNAm)-based estimates of biological age and of telomere length (TL). DNAm levels (Infinium MethylationEPIC BeadChip) from peripheral blood mononuclear cells from 200 PLWH and 199 HIV-seronegative (SN) participants matched on chronologic age, hepatitis C virus, and time intervals were used to calculate epigenetic age acceleration, expressed as age-adjusted acceleration residuals from 4 epigenetic clocks [Horvath's pan-tissue age acceleration residual (AAR), extrinsic epigenetic age acceleration (EEAA), phenotypic epigenetic age acceleration (PEAA), and grim epigenetic age acceleration (GEAA)] plus age-adjusted DNAm-based TL (aaDNAmTL). Epigenetic age acceleration was compared for PLWH and SN participants at two visits: up to 1.5years prior and 2-3years after HAART (or equivalent visits). Flow cytometry was performed in PLWH and SN participants at both visits to evaluate T-cell subsets. Epigenetic age acceleration in PLWH decreased after the initiation of HAART but remained greater post-HAART than that in age-matched SN participants, with differences in medians of 6.6, 9.1, and 7.7 years for AAR, EEAA, and PEAA, respectively, and 0.39 units of aaDNAmTL shortening (all p < 0.001). Cumulative HIV viral load after HAART initiation was associated with some epigenetic acceleration (EEAA, PEAA, and aaDNAmTL), but even PLWH with undetectable HIV post-HAART showed persistent epigenetic age acceleration compared to SN participants (p < 0.001). AAR, EEAA, and aaDNAmTL showed significant associations with total, naïve, and senescent CD8 T-cell counts; the total CD4 T-cell counts were associated with AAR, EEAA, and PEAA (p = 0.04 to <0.001). In an epigenome-wide analysis using weighted gene co-methylation network analyses, 11 modules demonstrated significant DNAm differences pre- to post-HAART initiation. Of these, nine were previously identified as significantly different from pre- to post-HIV infection but in the opposite direction. In this large longitudinal study, we demonstrated that, although the magnitude of the difference decreases with HAART is associated with the cumulative viral load, PLWH are persistently epigenetically older than age-matched SN participants even after the successful initiation of HAART, and these changes are associated with changes in T-cell subsets.

CD4+CD57+ senescent T cells as promoters of systemic lupus erythematosus pathogenesis and the therapeutic potential of senolytic BCL-2 inhibitor.

Systemic lupus erythematosus (SLE) is a complex autoimmune disease characterized by persistent activation of immune cells and overproduction of autoantibodies. The accumulation of senescent T and B cells has been observed in SLE and other immune-mediated diseases. However, the exact mechanistic pathways contributing to this process in SLE remain incompletely understood. In this study, we found that in SLE patients: (1) the frequency of CD4+CD57+ senescent T cells was significantly elevated and positively correlated with disease activity; (2) the expression levels of B-lymphoma-2 (BCL-2) family and interferon-induced genes (ISGs) were significantly upregulated; and (3) in vitro, the cytokine IL-15 stimulation increased the frequency of senescent CD4+ T cells and upregulated the expression of BCL-2 family and ISGs. Further, treatment with ABT-263 (a senolytic BCL-2 inhibitor) in MRL/lpr mice resulted in decreased: (1) frequency of CD4+CD44hiCD62L-PD-1+CD153+ senescent CD4+ T cells; (2) frequency of CD19+CD11c+T-bet+ age-related B cells; (3) level of serum antinuclear antibody; (4) proteinuria; (5) frequency of Tfh cells; and (6) renal histopathological abnormalities. Collectively, these results indicated a dominant role for CD4+CD57+ senescent CD4+ T cells in the pathogenesis of SLE and senolytic BCL-2 inhibitor ABT-263 may be the potential treatment in ameliorating lupus phenotypes.

HLA-C*07 is associated with symptomatic HIV-1-associated neurocognitive disorders (HAND) and immune dysregulation

Background HIV-1-associated neurocognitive disorders (HAND) in stable patients undergoing antiretroviral therapy (ART) may result from ongoing immune dysregulation and chronic inflammation. A contributing factor may result from the unstable HLA class I allele, HLA-C*07. Objective To assess the genetic profile of killer-cell immunoglobulin-like receptors (KIR), human leukocyte antigens (HLA), and immune activation or senescence markers and their association with HAND in stable HIV-1 patients receiving ART. Methods An observational cross-sectional study was carried out with 96 patients with asymptomatic or symptomatic HAND. HLA and KIR as well as immune activation/senescence biomarkers in peripheral blood cells were assessed by SSO-Luminex typing and flow cytometry, respectively. Results HLA-C*07 is associated with symptomatic HAND. The frequency of two copies of HLA-C*07 was higher in patients with symptomatic than with asymptomatic HAND (12.0 vs. 2.2%, ρ < 0.001). The percentage of senescent CD8+CD28- T-cells was higher in patients with two copies of HLA-C*07 (ρ < 0.05). In patients with symptomatic HAND, the percentages of non-senescent CD8+CD28+ T cells were inversely proportional to the number of copies of the HLA-C*07 (ρ < 0.05). Conclusion Patients with symptomatic HAND showed a higher frequency of the homozygotic unstable HLA-C*07 allotype, which could be associated with neurocognitive complications. Two copies of HLA-C*07 were associated with immune senescent T lymphocyte profiles characterized by the loss of CD28 expression.

Abstract PO2-13-04: Single cell characterization of senescent CD8+ T cell promotes immunotherapy resistance in early-stage triple-negative breast cancer

Abstract Background: Immune checkpoint blockade (ICB) has been explored in triple-negative breast cancer (TNBC) and has been proven to improve patient survival in advanced TNBCs (IMpassion130) or increase the pathological complete response rate in early TNBCs (KEYNOTE522). However, ICB could only benefit a subset of patients, highlighting the urgent need to optimize patient selection. Unlike in advanced TNBCs, a predictive biomarker for ICB in early TNBCs is still lacking. The main reason might be that previous studies neglected the heterogeneity of tumor ecosystems between early and advanced TNBCs. There is an urgent need to understand the unique ecosystem of early-stage TNBC and develop biomarkers for precision immunotherapy. Objective: This study aims to characterize the micro-loop associated with the immune efficacy of early TNBC, so as to explain the mechanisms behind the differences in early and late efficacy markers, identify efficacy markers in early TNBC immunotherapy, and search for clinical targeting strategies to sensitize TNBC for neoadjuvant therapy. Methods: We conducted an in-depth analysis of 32 single-cell transcriptomes, comprising 26 early-stage TNBC enrolled in prospective clinical trials (NCT04418154 and NCT04613674), receiving neoadjuvant chemotherapy alone (n=8) or combined with anti-PD1 (n=18), and 6 advanced TNBC. We analyzed the microenvironmental differences between early and advanced tumors, and between different treatment response groups in early stages, and identified a subpopulation of cells specific to early stages and associated with immunotherapeutic efficacy. We used mIHC, flow-through, and validated the presence of this cell in clinical samples. We also explored its predictive performance on four independent cohorts of TNBCs with ICB treatment (GSE194040, NCT04129996, GSE169246, GSE124821). Furthermore, to understand the impact of ISG+ T cells on immunotherapy efficacy and explore targeting strategies, we employed a multi-faceted approach to assess their tumor-killing ability, response to Anti-PD1 treatment, and sensitization effect of drugs targeting ISG+ T cells. ISG+ T cells were sorted using flow cytometry for further experiment. In vitro, we established a co-culture model of primary TNBC patients-derived organoid (PDO) and tumor infiltrated CD8+T cells and evaluated the tumor cell viability as well as T cell cytotoxicity. In vivo, we established TS/A tumor cell bearing BalB/c mouse and E0771 tumor cell-bearing C57BL mouse and measured tumor size and tumor microenvironment by flow cytometry. Results: We obtained 236,017 high-quality cells and identified cell subsets across 32 samples with different stages, treatments and responses. We then uncovered an enriched subset of interferon-induced (ISG+) CD8+ T cells specific to early TNBC, serving as a predictive factor for ICB resistance. Mechanistically, the upregulation of the IFN pathway in early-stage TNBC triggers a transition in ISG+ CD8+ T cells, resulting in cellular senescence, diminished cytotoxicity, and impaired response to ICB. Encouragingly, we made a noteworthy discovery that Nicotinamide mononucleotide, an anti-aging drug, effectively restores the functionality of ISG+ CD8+ T cells, amplifying immunotherapy sensitivity. Conclusions: This study provided a comprehensive single-cell transcriptome atlas of early-stage TNBC and depicted stage-related ISG+CD8+ T cell for predicting ICB response in early TNBC. We emphasize the critical role of T cell dysfunction prompted by cellular senescence in driving ICB resistance during the early stages of TNBC and propose an approach for sensitizing immunotherapy with aging drugs. Citation Format: Tong Fu, Yi-Yu Chen, Xi Jin, Yi-Zhou Jiang, Zhi-Ming Shao. Single cell characterization of senescent CD8+ T cell promotes immunotherapy resistance in early-stage triple-negative breast cancer [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO2-13-04.

Abstract 1030: Zeb2 Regulates Senescence And Cytotoxic Phenotype In Atherosclerotic Plaque CD8 T Cells

Circulating senescent CD8 + CD57 + T cells have been implicated in reduced post-myocardial infarction survival and impaired glucose homeostasis in type 2 diabetes (T2D) patients. Despite these associations, their specific role in human atherosclerotic plaques, particularly in the context of aggravated plaque pathology in T2D remains unknown. In this study, we analyzed single-cell transcriptomic data from 75,049 immune cells obtained from plaques from 12 T2D and 10 no diabetes patients enrolled in the ATHERO-IN study. We identified 14 clusters of CD8 + T cells clusters, revealing distinct subsets such cytotoxic CD8 + T cells with a terminally differentiated effector memory phenotype, cytotoxic/exhausted, naïve/CM and regulatory CD8 + T cells. These clusters expressed varying degrees of senescent genes, with a notable heightened senescent transcriptional signature in T2D patients. Fibrocalcific plaques, which were prevalent in T2D patients, showed significantly enrichment in cytotoxic, inflammatory, naïve/CM, and interferon response CD8 T cell clusters. In contrast, fibroatheroma samples were predominantly enriched in MAIT-like CD8 T cells and cytotoxic/exhausted clusters. Aged CD8 T cells expanded in T2D patients exhibited elevated senescence and cytotoxic score, manifested a senescence-associated secretory phenotype expressing cytotoxic, pro-inflammatory and proatherogenic genes. Trajectory analysis and regulatory network analysis, highlighted ZEB2— a GWAS gene associated to coronary artery disease and myocardial infarction— as a potential transcription factor driving the senescence phenotype. CRISPR-based studies corroborated this, demonstrating that ZEB2 KO CD8 T cells reduced senescence, cytotoxicity, and activation markers, underscoring the pivotal role of ZEB2 in driving the terminal differentiation and aging phenotype. In conclusion, our findings shed light on the intricate interplay between senescent CD8+ T cells and atherosclerotic plaque pathology in the context of T2D. The identification of ZEB2 as a key regulator of T cell senescence offers a potential target for therapeutic interventions aimed at mitigating the senescence-associated effects on plaque pathology and cardiovascular outcomes in T2D patients.

Frequency of exhausted and senescent CD4+ T cells in peripheral blood, specific to antigen-associated tumors (TPI1, PRDX2, A1AT, AHSG AND PPIA) and its impact and its association with the diagnosis of benign and malignant ovarian tumors

Frequency of exhausted and senescent CD4+ T cells in peripheral blood, specific to antigen-associated tumors (TPI1, PRDX2, A1AT, AHSG AND PPIA) and its impact and its association with the diagnosis of benign and malignant ovarian tumors

Immune cell phenotype and function patterns across the life course in individuals from rural Uganda.

To determine the pattern of immune cell subsets across the life span in rural sub-Saharan Africa (SSA), and to set a reference standard for cell subsets amongst Africans, we characterised the major immune cell subsets in peripheral blood including T cells, B cells, monocytes, NK cells, neutrophils and eosinophils, in individuals aged 3 to 89 years from Uganda. Immune phenotypes were measured using both conventional flow cytometry in 72 individuals, and full spectrum flow cytometry in 80 individuals. Epstein-Barr virus (EBV) IFN-γ T cell responses were quantified in 332 individuals using an ELISpot assay. Full blood counts of all study participants were also obtained. The percentages of central memory (TCM) and senescent CD4+ and CD8+ T cell subsets, effector memory (TEM) CD8+ T cells and neutrophils increased with increasing age. On the other hand, the percentages of naïve T (TN) and B (BN) cells, atypical B cells (BA), total lymphocytes, eosinophils and basophils decreased with increasing age. There was no change in CD4+ or CD8+ T effector memory RA (TEMRA) cells, exhausted T cells, NK cells and monocytes with age. Higher eosinophil and basophil percentages were observed in males compared to females. T cell function as measured by IFN-γ responses to EBV increased with increasing age, peaking at 31-55 years. The percentages of cell subsets differ between individuals from SSA compared to those elsewhere, perhaps reflecting a different antigenic milieu. These results serve as a reference for normal values in this population.

Microfluidic Capture Device for Simple, Cell Surface Marker-Based Quantification of Senescent CD8+ T Cells

Among human CD8+ T cells, senescent cells are marked by the expression of CD57. The frequency of senescent CD57+CD8+ T cells is significantly correlated with aging and age-associated disorders, and it can be measured by multi-color flow cytometry. However, multi-color flow cytometry presents challenges in terms of accessibility and requires significant resource allocation. Therefore, developing a rapid and straightforward method for quantifying CD57+CD8+ T cells remains a key challenge. This study introduces a microfluidic device composed of a PDMS microfluidic channel with a pre-modified glass substrate for anti-CD8 antibody immobilization. This design allows blood samples to flow through, enabling the selective capture of CD8+ T cells while minimizing the required blood sample volume. This technology enables accurate and reliable quantification of CD57+ cells among captured CD8+ T cells through fluorescence image analysis. The ability of the device to easily quantify senescent CD57+CD8+ T cells is anticipated to contribute significantly to both immunological research and clinical applications.

DOP84 Colonic CD4+ T cell senescence is implicated in the progression of experimental colitis in aged mice

Abstract Background Evidence from recent epidemiological data suggests that the burden of IBD in the elderly is increasing globally. However, the mechanisms underlying the age-related IBD susceptibility remain elusive. Aging leads to a progressive functional decline of the immune system, rendering the elderly increasingly susceptible to autoimmune disease. Herein, we sought to determine the role of immunosenescence in age-dependent colitis and to explore the underlying mechanisms. Methods Young (2-3 months), middle-aged (10-12 months), and aged (20-24 months) mice were established models of colitis by 2% dextran sulfate sodium salt (DSS) treatment. Weight loss, colon length, disease activity indexes (DAI) and histological scores were evaluated to assess the severity of colitis. Using a second-generation fluorogenic substrate for β-galactosidase and multi-parameter flow cytometry, we detected the senescence-associated β-galactosidase (SA-βGal) activity in lymphocytes isolated from colon, including CD4+ T cells, CD8+ T cells, B cells, natural killer cells, microphages, neutrophils and dendritic cells. Based on the immunocytes that exhibit greatest age-related increases in SA-βGal activity, inflammatory cytokines such as TNF-α、IFN-γ、IL-17 were evaluated in senescent immunocytes through flow cytometry. Single-cell and bulk RNA-seq of colonic immune cells in mice with different ages were performed to further investigate the precise molecular mechanisms. Results Ageing increased the severity of DSS-induced colitis wherein aged mice exhibited worsened weight loss, shortened colon length, higher DAI and histological scores compared to young mice. The greatest age-associated increases of SA-βGal activity were observed in colonic CD4+ T cell populations. Additionally, aging resulted in systemic activation of CD4+ T cells, exhibiting an expansion of CD4+ effector memory T (TEM) cells in colon, as well as in spleen. Aged colonic CD4+ T cells generate higher levels of IFN-γ and IL-17 comparing to young colonic CD4+ T cells, especially in status of colitis. Moreover, the accumulated CD4+ TEM cells in the aged colon were the main source of IL-17 production. Single-cell analysis revealed that aging increased colonic CD4+ TEM cells, which were associated with T cell-mediated cytotoxicity and cytokine-mediated signaling pathway. Splenic CD4+ TEM cells isolated from aged colitis mice were confirmed to involve in cellular senescence, Th17 differentiation and inflammatory signaling pathway through bulk RNA-seq. Conclusion These results provide a significant insight into the contribution of senescent CD4+ T cells to age-dependent colitis and provide an attractive possibility that targeting T cell specifically might be a potential strategy to treat elderly IBD patients.

NAD+ supplementation prevents STING-induced senescence in CD8+ T cells by improving mitochondrial homeostasis.

Understanding the connection between senescence phenotypes and mitochondrial dysfunction is crucial in aging and premature aging diseases. Loss of mitochondrial function leads to a decline in T cell function, which plays a significant role in this process. However, more research is required to determine if improving mitochondrial homeostasis alleviates senescence phenotypes. Our research has shown an association between NAD+ and senescent T cells through the cGAS-STING pathway, which can lead to an inflammatory phenotype. Further research is needed to fully understand the role of NAD+ in T-cell aging and how it can be utilized to improve mitochondrial homeostasis and alleviate senescence phenotypes. We demonstrate here that mitochondrial dysfunction and cellular senescence with a senescence-associated secretory phenotype (SASP) occur in senescent T cells and tumor-bearing mice. Senescence is mediated by a stimulator of interferon genes (STING) and involves ectopic cytoplasmic DNA. We further show that boosting intracellular NAD+ levels with nicotinamide mononucleotide (NMN) prevents senescence and SASP by promoting mitophagy. NMN treatment also suppresses senescence and neuroinflammation and improves the survival cycle of mice. Encouraging mitophagy may be a useful strategy to prevent CD8+ T cells from senescence due to mitochondrial dysfunction. Additionally, supplementing with NMN to increase NAD+ levels could enhance survival rates in mice while also reducing senescence and inflammation, and enhancing mitophagy as a potential therapeutic intervention.