Abstract The causal link between the aging microenvironment and T cell aging remains elusive. Here, we demonstrate that adenosine within aging tissues actively reprograms CD8+ T cells into a pro-aging Granzyme K + (Gzmk+) population. Mechanistically, senescent cells create an adenosine-rich niche via p16-dependent CD39 upregulation, triggering A2aR signaling to induce Gzmk + T cell differentiation. Once released, Gzmk promotes systemic inflammaging through PAR1 and complement activation. Crucially, targeting this axis—either via genetic Gzmk ablation or pharmacological A2aR blockade—reverses multi-organ aging phenotypes and significantly extends healthy lifespan in mice. Human analysis reveals age-dependent Gzmk + T cell accumulation in multi organs, while coffee intake (an A2aR antagonist) inversely correlates with plasma Gzmk levels. Our findings uncover how metabolic niche changes drive T cell aging and establish the adenosine-Gzmk axis as a pivotal therapeutic target for combating age-related diseases.

T cell senescence causes T cell dysfunction in tumors, but its drivers are unclear. Here we found that protein overload in the tumor microenvironment (TME) induces T cell differentiation into effector memory T cells re-expressing CD45RA (TEMRA). TEMRA cells exhibit senescent-like features, including reduced proliferative capacity and expression of senescence-associated markers. Both CD4+ and CD8+ T cells activated under high protein-to-amino-acid-ratio conditions in vitro or within TME niches underwent enhanced TEMRA differentiation. Single-cell transcriptomics showed that protein overload co-activates terminal effector and senescence programs. Mechanistically, it disrupts proteostasis by inhibiting lysosomal and proteasomal degradation, triggering a maladaptive unfolded protein response (UPR) that drives TEMRA commitment. Exogenous amino acid supplementation prevented UPR activation and TEMRA differentiation. In humanized tumors, intra-tumoral amino-acid administration reduced TEMRA accumulation, boosted T cell proliferation, and improved tumor control. Our work reveals managing proteostatic stress as a strategy to counter TEMRA generation and restore anti-tumor immunity.

Abstract Despite the pivotal role of mRNA isoform diversity in governing immune cell function, current investigations into peripheral immune aging predominantly focused on gene-level expression, obscuring deeper regulatory layers of transcriptome complexity. Here, we leveraged a 5′ scRNA-seq atlas comprising approximately 2.5 million PBMCs from 378 healthy donors. We demonstrate that immune aging is characterized by profound, non-linear transcriptional reprogramming that extends beyond gene-level shifts to include fine-tuned regulation of alternative transcription initiation and splice site selection. By quantifying the transcriptional activity of cis-regulatory elements, we resolved their contributions to age-related expression dynamics. Notably, we identified a subset of endogenous retroviruses that are reactivated in older individuals, some of which served as alternative promoters driving the production of chimeric transcripts. Furthermore, our analysis revealed EDA as a top-ranked gene consistently upregulated with age across multiple independent cohorts. Increasing EDA expression in in vitro-stimulated naïve CD4 + T cells from young individuals recapitulated aged phenotypes. This comprehensive resource elucidates the multi-layered transcriptomic landscape of the aging immune system and facilitates the identification of novel drivers of immune aging.

Proteins are the most abundant source of amino acids in body fluids. However, the potential contribution of extracellular protein catabolism to the regulation of T cell immunity remains poorly understood. In this study, we show that endocytosed extracellular proteins function as an amino acid source in activated T cells, maintaining mTORC1 activity and sustaining cytokine production following T cell activation. Genetic ablation of Tfe3 impairs the activation-induced upregulation of lysosomal genes and disrupts extracellular protein catabolism, resulting in attenuated mTORC1 signaling and compromised anti-viral and anti-tumor T cell responses. The TFE3-protein-mTORC1 signaling axis demonstrates clinical relevance. CD8+PD-1+ tumor-infiltrating T cells from older patients with lung cancer display reduced lysosomal degradation capacity and impaired cytokine secretion compared to their middle-aged counterparts. This functional defect is rescued by treatment with Vismodegib, a TFE3-inducing drug. Our findings reveal lysosome-mediated extracellular protein catabolism as an important metabolic pathway supporting T cell immunity.

T cell aging contributes to the lower vaccine efficacy in older adults, yet the molecular mechanism remains elusive. Here, we show the density of initially responding naïve CD4 + T cells is instructive in T follicular helper (TFH) cell fate decisions and declines with age. A lower number of initially responding cells did not affect TFH differentiation at peak responses after immunization but accounted for an increased contraction phase manifesting as a larger loss of CXCR5 expression. Mechanistically, cells activated at a lower initial density had more sustained mammalian target of rapamycin complex 1 (mTORC1) activities that impair CXCR5 maintenance. YAP-dependent regulation of SLC7A5 involved in the cell density–dependent regulation of mTORC1 activities and TFH loss. Old mice fed with a leucine-restricted diet after peak responses showed smaller TFH loss and improved humoral immune responses. Attenuating mTORC1 signaling after peak response is a strategy to boost vaccine responses in older individuals.

Naive CD4+ T cells are more resistant to age-related loss than naive CD8+ T cells, suggesting mechanisms that preferentially protect naive CD4+ T cells during aging. Here, we show that TRIB2 is more abundant in naive CD4+ than CD8+ T cells and counteracts quiescence exit by suppressing AKT activation. TRIB2 deficiency increases AKT activity and accelerates proliferation and differentiation in response to interleukin-7 (IL-7) in humans and during lymphopenia in mice. TRIB2 transcription is controlled by the lineage-determining transcription factors ThPOK and RUNX3. Ablation of Zbtb7b (encoding ThPOK) and Cbfb (obligatory RUNT cofactor) attenuates the difference in lymphopenia-induced proliferation between naive CD4+ and CD8+ cells. In older adults, ThPOK and TRIB2 expression wanes in naive CD4+ T cells, causing loss of naivety. These findings assign TRIB2 a key role in regulating T cell homeostasis and provide a model to explain the lesser resilience of CD8+ T cells to undergo changes with age.

Chronic systemic inflammation is one of the hallmarks of the aging immune system. Here we show that activated T cells from older adults contribute to inflammaging by releasing mitochondrial DNA (mtDNA) into their environment due to an increased expression of the cytokine-inducible SH2-containing protein (CISH). CISH targets ATP6V1A, an essential component of the proton pump V-ATPase, for proteasomal degradation, thereby impairing lysosomal function. Impaired lysosomal activity caused intracellular accumulation of multivesicular bodies and amphisomes and the export of their cargos, including mtDNA. CISH silencing in T cells from older adults restored lysosomal activity and prevented amphisomal release. In antigen-specific responses in vivo, CISH-deficient CD4 + T cells released less mtDNA and induced fewer inflammatory cytokines. Attenuating CISH expression may present a promising strategy to reduce inflammation in an immune response of older individuals.

This article describes the systematic design of an electronic recording peak expiratory flow (PEF) meter to provide accurate data for the diagnosis of occupational asthma. Traditional diagnosis of asthma relies on accurate data of PEF tests performed by the patients in their own homes and places of work. Unfortunately there are high error rates in data produced and recorded by the patient, most of these are transcription errors and some patients falsify their records. The PEF measurement itself is not effort independent, the data produced depending on the way in which the patient performs the test. Patients are taught how to perform the test giving maximal effort to the expiration being measured. If the measurement is performed incorrectly then errors will occur. Accurate data can be produced if an electronically recording PEF instrument is developed, thus freeing the patient from the task of recording the test data. It should also be capable of determining whether the PEF measurement has been correctly performed. A requirement specification for a recording PEF meter was produced. A commercially available electronic PEF meter was modified to provide the functions required for accurate serial recording of the measurements produced by the patients. This is now being used in three hospitals in the West Midlands for investigations into the diagnosis of occupational asthma. In investigating current methods of measuring PEF and other pulmonary quantities a greater understanding was obtained of the limitations of current methods of measurement, and quantities being measured. It is now possible to define the quantities that need to be measured for accurate pulmonary diagnosis and the requirements of future instrumentation to make these necessary measurements. We are now in the process of developing this new instrumentation.

Immune aging combines cellular defects in adaptive immunity with the activation of pathways causing a low-inflammatory state. Here we examined the influence of age on the kinetic changes in the epigenomic and transcriptional landscape induced by T cell receptor (TCR) stimulation in naive CD4 + T cells. Despite attenuated TCR signaling in older adults, TCR activation accelerated remodeling of the epigenome and induced transcription factor networks favoring effector cell differentiation. We identified increased phosphorylation of STAT5, at least in part due to aberrant IL-2 receptor and lower HELIOS expression, as upstream regulators. Human HELIOS-deficient, naive CD4 + T cells, when transferred into human-synovium-mouse chimeras, infiltrated tissues more efficiently. Inhibition of IL-2 or STAT5 activity in T cell responses of older adults restored the epigenetic response pattern to the one seen in young adults. In summary, reduced HELIOS expression in non-regulatory naive CD4 + T cells in older adults directs T cell fate decisions toward inflammatory effector cells that infiltrate tissue.

With increasing age, individuals are more vulnerable to viral infections such as with influenza or the SARS-CoV-2 virus. One age-associated defect in human T cells is the reduced expression of miR-181a. miR-181ab1 deficiency in peripheral murine T cells causes delayed viral clearance after infection, resembling human immune aging. Here we show that naive T cells from older individuals as well as miR-181ab1-deficient murine T cells develop excessive replication stress after activation, due to reduced histone expression and delayed S-phase cell cycle progression. Reduced histone expression was caused by the miR-181a target SIRT1 that directly repressed transcription of histone genes by binding to their promoters and reducing histone acetylation. Inhibition of SIRT1 activity or SIRT1 silencing increased histone expression, restored cell cycle progression, diminished the replication-stress response, and reduced the production of inflammatory mediators in replicating T cells from old individuals. Correspondingly, treatment with SIRT1 inhibitors improved viral clearance in mice with miR-181a-deficient T cells after LCMV infection. In conclusion, SIRT1 inhibition may be beneficial to treat systemic viral infection in older individuals by targeting antigen-specific T cells that develop replication stress due to miR-181a deficiency.

The nutrient-sensing mammalian target of rapamycin (mTOR) is integral to cell fate decisions after T cell activation. Sustained mTORC1 activity favors the generation of terminally differentiated effector T cells instead of follicular helper and memory T cells. This is particularly pertinent for T cell responses of older adults who have sustained mTORC1 activation despite dysfunctional lysosomes. Here, we show that lysosome-deficient T cells rely on late endosomes rather than lysosomes as an mTORC1 activation platform, where mTORC1 is activated by sensing cytosolic amino acids. T cells from older adults have an increased expression of the plasma membrane leucine transporter SLC7A5 to provide a cytosolic amino acid source. Hence, SLC7A5 and VPS39 deficiency (a member of the HOPS complex promoting early to late endosome conversion) substantially reduced mTORC1 activities in T cells from older but not young individuals. Late endosomal mTORC1 is independent of the negative-feedback loop involving mTORC1-induced inactivation of the transcription factor TFEB that controls expression of lysosomal genes. The resulting sustained mTORC1 activation impaired lysosome function and prevented lysosomal degradation of PD-1 in CD4+ T cells from older adults, thereby inhibiting their proliferative responses. VPS39 silencing of human T cells improved their expansion to pertussis and to SARS-CoV-2 peptides in vitro. Furthermore, adoptive transfer of CD4+ Vps39-deficient LCMV-specific SMARTA cells improved germinal center responses, CD8+ memory T cell generation, and recall responses to infection. Thus, curtailing late endosomal mTORC1 activity is a promising strategy to enhance T cell immunity.

Lysosomes were initially recognized as degradation centers that regulate digestion and recycling of cellular waste. More recent studies document that the lysosome is an important signaling hub that regulates cell metabolism. Our knowledge of the role of lysosomes in immunity is mostly derived from innate immune cells, especially lysosomal degradation-specialized cells such as macrophages and dendritic cells. Their function in adaptive immunity is less understood. However, with the recent emphasis on metabolic regulation of T cell differentiation, lysosomes are entering center stage in T cell immunology. In this review, we will focus on the role of lysosomes in adaptive immunity and discuss recent findings on lysosomal regulation of T cell immune responses and lysosomal dysfunction in T cell aging.

MicroRNAs play an important role in the regulation of T cell development, activation, and differentiation. One of the most abundant microRNAs in lymphocytes is miR-181a, which controls T cell receptor (TCR) activation thresholds in thymic selection as well as in peripheral T cell responses. We previously found that miR-181a levels decline in T cells in the elderly. In this study, we identified TCF1 as a transcriptional regulator of pri-miR-181a. A decline in TCF1 levels in old individuals accounted for the reduced miR-181a expression impairing TCR signaling. Inhibition of GSK3ß restored expression of miR-181a by inducing TCF1 in T cells from old adults. GSK3ß inhibition enhanced TCR signaling to increase downstream expression of activation markers and production of IL-2. The effect involved the upregulation of miR-181a and the inhibition of DUSP6 expression. Thus, inhibition of GSK3ß can restore responses of old T cells by inducing miR-181a expression through TCF1.

SummaryT cell aging is a complex process combining the emergence of cellular defects with activation of adaptive mechanisms. Generation of T cell memory is impaired, while a low-inflammatory state is induced, in part due to effector T cells. To determine whether age-associated changes in T cell fate decisions occur early after T cell activation, we profiled the longitudinal transcriptional and epigenetic landscape induced by TCR stimulation comparing naïve CD4+T cells from young and older adults. In spite of attenuated TCR signaling, activation-induced remodeling of the epigenome increased with age, culminating in heightened BATF and BLIMP1 activity. Single cell studies, integrating ATAC-seq and RNA-seq data, identified increases in dysfunctional and in effector T cells and a decrease in BACH2-expressing memory cell precursors. STAT5 activation, in part due to a decline in HELIOS and aberrant IL-2 receptor expression, accounted for the induction of transcription factor networks favoring effector cell differentiation.

AbstractsBackgroundCancer patients are considered to be highly susceptible to viral infections, however, the comprehensive features of COVID-19 in these patients remained largely unknown. The present study aimed to assess the clinical characteristics and outcomes of COVID-19 in a large cohort of cancer patients.Design, Setting, and ParticipantsData of consecutive cancer patients admitted to 33 designated hospitals for COVID-19 in Hubei province, China from December 17, 2019 to March 18, 2020 were retrospectively collected. The follow-up cutoff date was April 02, 2020. The clinical course and survival status of the cancer patients with COVID-19 were measured, and the potential risk factors of severe events and death were assessed through univariable and multivariable analyses.ResultsA total of 283 laboratory confirmed COVID-19 patients (50% male; median age, 63.0 years [IQR, 55.0 to 70.0]) with more than 20 cancer types were included. The overall mortality rate was 18% (50/283), and the median hospitalization stay for the survivors was 26 days. Amongst all, 76 (27%) were former cancer patients with curative resections for over five years without recurrence. The current cancer patients exhibited worse outcomes versus former cancer patients (overall survival, HR=2.45, 95%CI 1.10 to 5.44, log-rank p=0.02; mortality rate, 21% vs 9%). Of the 207 current cancer patients, 95 (46%) have received recent anti-tumor treatment, and the highest mortality rate was observed in the patients receiving recent chemotherapy (33%), followed by surgery (26%), other anti-tumor treatments (19%), and no anti-tumor treatment (15%). In addition, a higher mortality rate was observed in patients with lymphohematopoietic malignancies (LHM) (53%, 9/17), and all seven LHM patients with recent chemotherapy died. Multivariable analysis indicated that LHM (p=0.001) was one of the independent factors associating with critical illness or death.ConclusionsThis is the first systematic study comprehensively depicting COVID-19 in a large cancer cohort. Patients with tumors, especially LHM, may have poorer prognosis of COVID-19. Additional cares are warranted and non-emergency anti-tumor treatment should be cautiously used for these patients under the pandemic.

The transcription factor T cell factor 1 (TCF1), a pioneer transcription factor as well as a downstream effector of WNT/β-catenin signaling, is indispensable for T cell development in the thymus. Recent studies have highlighted the additional critical role of TCF1 in peripheral T cell responses to acute and chronic infections as well as cancer. Here, we review the regulatory functions of TCF1 in the differentiation of T follicular helper cells, memory T cells and recently described stem-like exhausted T cells, where TCF1 promotes less differentiated stem-like cell states by controlling common gene-regulatory networks. These studies also provide insights into the mechanisms of defective T cell responses in older individuals. We discuss alterations in TCF1 expression and related regulatory networks with age and their consequences for T cell responses to infections and vaccination. The increasing understanding of the pathways regulating TCF1 expression and function in aged T cells holds the promise of enabling the design of therapeutic interventions aiming at improving T cell responses in older individuals.

Purpose: With the proper insights, measurement of central venous pressure (CVP) can be a useful clinical aid. However, the formal utility of CVP measurement on mortality in septic patients has never been proved.Methods: The Medical Information Mart for Intensive Care III (MIMIC-III) was applied to identify septic patients who had and did not have CVP measured. The primary outcome was 28-day mortality. The statistical approaches including multivariate regression, propensity score matching (PSM) and an inverse probability of treatment weighing (IPTW) and causal mediation analysis (CMA) were utilized to elucidate the relationship between CVP measurement and 28-day mortality.Results: A total of 10275 patients were included in our study, of which 4516 patients (44%) had CVP measured within 24 h after ICU admission. A significant beneficial effect of CVP measurement in terms of 28-day mortality was observed (OR 0.60 (95% CI 0.51–0.70; p<0.001)). Patients in CVP group received more fluid on day 1, had a shorter duration of mechanical ventilation and vasopressor use, and the reduction of serum lactate was higher than that in the no CVP group. The mediation effect of serum lactate reduction was significant for the whole cohort (p=0.04 for average causal mediation effect (ACME)) and patients in the CVP group with an initial CVP level below 8mmHg (p=0.04 for ACME).Conclusion: CVP measurement is associated with a lower risk-adjusted 28-day mortality among patients with sepsis, which is proportionally mediated through serum lactate reduction.

T cell differentiation involves the dynamic regulation of FOXO1 expression, which rapidly declines after activation and is subsequently restored. Reexpression is impaired in naïve CD4+ T cell responses from older individuals. Here, we show that FOXO1 promotes lysosome function through the induction of the key transcription factor for lysosomal proteins, TFEB. Subdued FOXO1 reexpression in activated CD4+ T cells impairs lysosomal activity, causing an expansion of multivesicular bodies (MVBs). Expansion of the MVB compartment induces the sequestration of glycogen synthase kinase 3β (GSK3β), thereby suppressing protein turnover and enhancing glycolytic activity. As a consequence, older activated CD4+ T cells develop features reminiscent of senescent cells. They acquire an increased cell mass, preferentially differentiate into short-lived effector T cells, and secrete exosomes that harm cells in the local environment through the release of granzyme B.

Induction of protective vaccine responses, governed by the successful generation of antigen-specific antibodies and long-lived memory T cells, is increasingly impaired with age. Regulation of the T cell proteome by a dynamic network of microRNAs is crucial to T cell responses. Here, we show that activation-induced upregulation of miR-21 biases the transcriptome of differentiating T cells away from memory T cells and toward inflammatory effector T cells. Such a transcriptome bias is also characteristic of T cell responses in older individuals who have increased miR-21 expression and is reversed by antagonizing miR-21. miR-21 targets negative feedback circuits in several signaling pathways. The concerted, sustained activity of these signaling pathways in miR-21high T cells disfavors the induction of transcription factor networks involved in memory cell differentiation. Our data suggest that curbing miR-21 upregulation or activity in older individuals may improve their ability to mount effective vaccine responses.

One of the most prominent immunological changes during human aging is the alteration in CD8 T-cell subset distribution, predominated by a loss of naïve CD8 T cells. The molecular mechanisms that contribute to the loss of naïve CD8 T-cells during aging remain unclear. Considering that many CD8 T-cell functions are influenced by microRNAs (miRNAs), we explored miRNA expression profiling to identify novel dysfunctions that contribute to naïve CD8 T-cell loss during aging. Here, we describe age-dependent miRNA expression changes in naïve, central memory, and effector memory CD8 T-cell subsets. Changes in old naïve CD8 T-cells partially resembled those driven by an underlying shift in cellular differentiation toward a young central memory phenotype. Pathways enriched for targets of age-dependent miRNAs included FOXO1, NF-κB, and PI3K-AKT signaling. Transcriptome analysis of old naïve CD8 T-cells yielded corresponding patterns that correlated to those seen with reduced FOXO1 or altered NF-κB activities. Of particular interest, IL-7R expression, controlled by FOXO1 signaling, declines on naïve CD8 T cells with age and directly correlates with the frequencies of naïve CD8 T cells. Thus, age-associated changes in miRNA networks may ultimately contribute to the failure in CD8 T-cell homeostasis exemplified by the loss in naïve cells.