In Vivo CD4+ T Cell Differentiation and Function: Revisiting the Th1/Th2 Paradigm.

The discovery of CD4+ T cell subset-defining master transcription factors and framing of the Th1/Th2 paradigm ignited the CD4+ T cell field. Advances in in vivo experimental systems, however, have revealed that more complex lineage-defining transcriptional networks direct CD4+ T cell differentiation in the lymphoid organs and tissues. This review focuses on the layers of fate decisions that inform CD4+ T cell differentiation in vivo. Cytokine production by antigen-presenting cells and other innate cells influences the CD4+ T cell effector program [e.g., T helper type 1 (Th1), Th2, Th17]. Signals downstream of the T cell receptor influence whether individual clones bearing hallmarks of this effector program become T follicular helper cells, supporting development of B cells expressing specific antibody isotypes, or T effector cells, which activate microbicidal innate cells in tissues. These bifurcated, parallel axes allow CD4+ T cells to augment their particular effector program and prevent disease.

Imprinted SARS-CoV-2-specific memory lymphocytes define hybrid immunity.

Immune memory is tailored by cues that lymphocytes perceive during priming. The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic created a situation in which nascent memory could be tracked through additional antigen exposures. Both SARS-CoV-2 infection and vaccination induce multifaceted, functional immune memory, but together, they engender improved protection from disease, termed hybrid immunity. We therefore investigated how vaccine-induced memory is shaped by previous infection. We found that following vaccination, previously infected individuals generated more SARS-CoV-2 RBD-specific memory B cells and variant-neutralizing antibodies and a distinct population of IFN-γ and IL-10-expressing memory SARS-CoV-2 spike-specific CD4+ T cells than previously naive individuals. Although additional vaccination could increase humoral memory in previously naive individuals, it did not recapitulate the distinct CD4+ T cell cytokine profile observed in previously infected subjects. Thus, imprinted features of SARS-CoV-2-specific memory lymphocytes define hybrid immunity.

Functional SARS-CoV-2-Specific Immune Memory Persists after Mild COVID-19.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus is causing a global pandemic, and cases continue to rise. Most infected individuals experience mildly symptomatic coronavirus disease 2019 (COVID-19), but it is unknown whether this can induce persistent immune memory that could contribute to immunity. We performed a longitudinal assessment of individuals recovered from mild COVID-19 to determine whether they develop and sustain multifaceted SARS-CoV-2-specific immunological memory. Recovered individuals developed SARS-CoV-2-specific immunoglobulin (IgG) antibodies, neutralizing plasma, and memory B and memory T cells that persisted for at least 3 months. Our data further reveal that SARS-CoV-2-specific IgG memory B cells increased over time. Additionally, SARS-CoV-2-specific memory lymphocytes exhibited characteristics associated with potent antiviral function: memory T cells secreted cytokines and expanded upon antigen re-encounter, whereas memory B cells expressed receptors capable of neutralizing virus when expressed as monoclonal antibodies. Therefore, mild COVID-19 elicits memory lymphocytes that persist and display functional hallmarks of antiviral immunity.

Canonical BAF complex activity shapes the enhancer landscape that licenses CD8+ T cell effector and memory fates.

CD8+ T cells provide host protection against pathogens by differentiating into distinct effector and memory cell subsets, but how chromatin is site-specifically remodeled during their differentiation is unclear. Due to its critical role in regulating chromatin and enhancer accessibility through its nucleosome remodeling activities, we investigated the role of the canonical BAF (cBAF) chromatin remodeling complex in antiviral CD8+ T cells during infection. ARID1A, a subunit of cBAF, was recruited early after activation and established de novo open chromatin regions (OCRs) at enhancers. Arid1a deficiency impaired the opening of thousands of activation-induced enhancers, leading to loss of TF binding, dysregulated proliferation and gene expression, and failure to undergo terminal effector differentiation. Although Arid1a was dispensable for circulating memory cell formation, tissue-resident memory (Trm) formation was strongly impaired. Thus, cBAF governs the enhancer landscape of activated CD8+ T cells that orchestrates TF recruitment and activity and the acquisition of specific effector and memory differentiation states.

PD-1-Expressing SARS-CoV-2-Specific CD8+ T Cells Are Not Exhausted, but Functional in Patients with COVID-19.

Memory T cell responses have been demonstrated in COVID-19 convalescents, but ex vivo phenotypes of SARS-CoV-2-specific T cells have been unclear. We detected SARS-CoV-2-specific CD8+ T cells by MHC class I multimer staining and examined their phenotypes and functions in acute and convalescent COVID-19. Multimer+ cells exhibited early differentiated effector-memory phenotypes in the early convalescent phase. The frequency of stem-like memory cells was increased among multimer+ cells in the late convalescent phase. Cytokine secretion assays combined with MHC class I multimer staining revealed that the proportion of interferon-γ (IFN-γ)-producing cells was significantly lower among SARS-CoV-2-specific CD8+ T cells than those specific to influenza A virus. Importantly, the proportion of IFN-γ-producing cells was higher in PD-1+ cells than PD-1- cells among multimer+ cells, indicating that PD-1-expressing, SARS-CoV-2-specific CD8+ T cells are not exhausted, but functional. Our current findings provide information for understanding of SARS-CoV-2-specific CD8+ T cells elicited by infection or vaccination.

Heterogenous Populations of Tissue-Resident CD8+ T Cells Are Generated in Response to Infection and Malignancy.

Tissue-resident memory CD8+ T cells (Trm) provide host protection through continuous surveillance of non-lymphoid tissues. Using single-cell RNA-sequencing (scRNA-seq) and genetic reporter mice, we identified discrete lineages of intestinal antigen-specific CD8+ T cells, including a Blimp1hiId3lo tissue-resident effector cell population most prominent in the early phase of acute viral and bacterial infections and a molecularly distinct Blimp1loId3hi tissue-resident memory population that subsequently accumulated at later infection time points. These Trm populations exhibited distinct cytokine production, secondary memory potential, and transcriptional programs including differential roles for transcriptional regulators Blimp1, T-bet, Id2, and Id3 in supporting and maintaining intestinal Trm. Extending our analysis to malignant tissue, we also identified discrete populations of effector-like and memory-like CD8+ T cell populations with tissue-resident gene-expression signatures that shared features of terminally exhausted and progenitor-exhausted T cells, respectively. Our findings provide insight into the development and functional heterogeneity of Trm cells, which has implications for enhancing vaccination and immunotherapy approaches.

The Transcription Factor Bhlhe40 Programs Mitochondrial Regulation of Resident CD8+ T Cell Fitness and Functionality.

Tissue-resident memory CD8+ T (Trm) cells share core residency gene programs with tumor-infiltrating lymphocytes (TILs). However, the transcriptional, metabolic, and epigenetic regulation of Trm cell and TIL development and function is largely undefined. Here, we found that the transcription factor Bhlhe40 was specifically required for Trm cell and TIL development and polyfunctionality. Local PD-1 signaling inhibited TIL Bhlhe40 expression, and Bhlhe40 was critical for TIL reinvigoration following anti-PD-L1 blockade. Mechanistically, Bhlhe40 sustained Trm cell and TIL mitochondrial fitness and a functional epigenetic state. Building on these findings, we identified an epigenetic and metabolic regimen that promoted Trm cell and TIL gene signatures associated with tissue residency and polyfunctionality. This regimen empowered the anti-tumor activity of CD8+ T cells and possessed therapeutic potential even at an advanced tumor stage in mouse models. Our results provide mechanistic insights into the local regulation of Trm cell and TIL function.

Insights into phenotypic and functional CD8+ TRM heterogeneity.

Cytotoxic CD8+ T cells recognize and eliminate infected or cancerous cells. A subset of CD8+ memory T cells called tissue-resident memory T cells (TRM ) resides in peripheral tissues, monitors the periphery for pathogen invasion, and offers a rapid and potent first line of defense at potential sites of re-infection. TRM cells are found in almost all tissues and are transcriptionally and epigenetically distinct from circulating memory populations, which shows their ability to acclimate to the tissue environment to allow for long-term survival. Recent work and the broader availability of single-cell profiling have highlighted TRM heterogeneity among different tissues, as well as identified specialized subsets within individual tissues, that are time and infection dependent. TRM cell phenotypic and transcriptional heterogeneity has implications for understanding TRM function and longevity. This review aims to summarize and discuss the latest findings on CD8+ TRM heterogeneity using single-cell molecular profiling and explore the potential implications for immune protection and the design of immune therapies.

Microbiota configuration determines nutritional immune optimization.

Mild or transient dietary restriction (DR) improves many aspects of health and aging. Emerging evidence from us and others has demonstrated that DR also optimizes the development and quality of immune responses. However, the factors and mechanisms involved remain to be elucidated. Here, we propose that DR-induced optimization of immunological memory requires a complex cascade of events involving memory T cells, the intestinal microbiota, and myeloid cells. Our findings suggest that DR enhances the ability of memory T cells to recruit and activate myeloid cells in the context of a secondary infection. Concomitantly, DR promotes the expansion of commensal Bifidobacteria within the large intestine, which produce the short-chain fatty acid acetate. Acetate conditioning of the myeloid compartment during DR enhances the capacity of these cells to kill pathogens. Enhanced host protection during DR is compromised when Bifidobacteria expansion is prevented, indicating that microbiota configuration and function play an important role in determining immune responsiveness to this dietary intervention. Altogether, our study supports the idea that DR induces both memory T cells and the gut microbiota to produce distinct factors that converge on myeloid cells to promote optimal pathogen control. These findings suggest that nutritional cues can promote adaptation and co-operation between multiple immune cells and the gut microbiota, which synergize to optimize immunity and protect the collective metaorganism.

Tet2 deletion in CD4+ T cells disrupts Th1 lineage commitment in memory cells and enhances T follicular helper cell recall responses to viral rechallenge.

Following viral clearance, antigen-specific CD4+ T cells contract and form a pool of distinct Th1 and Tfh memory cells that possess unique epigenetic programs, allowing them to rapidly recall their specific effector functions upon rechallenge. DNA methylation programing mediated by the methylcytosine dioxygenase Tet2 contributes to balancing Th1 and Tfh cell differentiation during acute viral infection; however, the role of Tet2 in CD4+ T cell memory formation and recall is unclear. Using adoptive transfer models of antigen-specific wild type and Tet2 knockout CD4+ T cells, we find that Tet2 is required for full commitment of CD4+ T cells to the Th1 lineage and that in the absence of Tet2, memory cells preferentially recall a Tfh like phenotype with enhanced expansion upon secondary challenge. These findings demonstrate an important role for Tet2 in enforcing lineage commitment and programing proliferation potential, and highlight the potential of targeting epigenetic programing to enhance adaptive immune responses.

TGF-ß broadly modifies rather than specifically suppresses reactivated memory CD8 T cells in a dose-dependent manner.

Transforming growth factor ß (TGF-ß) directly acts on naive, effector, and memory T cells to control cell fate decisions, which was shown using genetic abrogation of TGF-ß signaling. TGF-ß availability is altered by infections and cancer; however, the dose-dependent effects of TGF-ß on memory CD8 T cell (Tmem) reactivation are still poorly defined. We examined how activation and TGF-ß signals interact to shape the functional outcome of Tmem reactivation. We found that TGF-ß could suppress cytotoxicity in a manner that was inversely proportional to the strength of the activating TCR or proinflammatory signals. In contrast, even high doses of TGF-ß had a comparatively modest effect on IFN-γ expression in the context of weak and strong reactivation signals. Since CD8 Tmem may not always receive TGF-ß signals concurrently with reactivation, we also explored whether the temporal order of reactivation versus TGF-ß signals is of importance. We found that exposure to TGF-ß before or after an activation event were both sufficient to reduce cytotoxic effector function. Concurrent ATAC-seq and RNA-seq analysis revealed that TGF-ß altered ~10% of the regulatory elements induced by reactivation and also elicited transcriptional changes indicative of broadly modulated functional properties. We confirmed some changes on the protein level and found that TGF-ß-induced expression of CCR8 was inversely proportional to the strength of the reactivating TCR signal. Together, our data suggest that TGF-ß is not simply suppressing CD8 Tmem but modifies functional and chemotactic properties in context of their reactivation signals and in a dose-dependent manner.

High CD62L expression predicts the generation of chimeric antigen receptor T cells with potent effector functions.

The efficient generation of chimeric antigen receptor (CAR) T cells is highly influenced by the quality of apheresed T cells. Healthy donor-derived T cells usually proliferate better than patients-derived T cells and are precious resources to generate off-the-shelf CAR-T cells. However, relatively little is known about the determinants that affect the efficient generation of CAR-T cells from healthy donor-derived peripheral blood mononuclear cells (PBMCs) compared with those from the patients' own PBMCs. We here examined the efficiency of CAR-T cell generation from multiple healthy donor samples and analyzed its association with the phenotypic features of the starting peripheral blood T cells. We found that CD62L expression levels within CD8+ T cells were significantly correlated with CAR-T cell expansion. Moreover, high CD62L expression within naïve T cells was associated with the efficient expansion of T cells with a stem cell-like memory phenotype, an indicator of high-quality infusion products. Intriguingly, genetic disruption of CD62L significantly impaired CAR-T cell proliferation and cytokine production upon antigen stimulation. Conversely, ectopic expression of a shedding-resistant CD62L mutant augmented CAR-T cell effector functions compared to unmodified CAR-T cells, resulting in improved antitumor activity in vivo. Collectively, we identified the surface expression of CD62L as a concise indicator of potent T-cell proliferation. CD62L expression is also associated with the functional properties of CAR-T cells. These findings are potentially applicable to selecting optimal donors to massively generate CAR-T cell products.

Keeping T cell memories in mind.

The mammalian central nervous system (CNS) contains a vibrant community of resident adaptive immune cells at homeostasis. Among these are memory CD8+ and CD4+ T cells, which reside in the CNS in the settings of health, aging, and neurological disease. These T cells commonly exhibit a tissue-resident memory (TRM) phenotype, suggesting that they are antigen-experienced and remain separate from the circulation. Despite these characterizations, T cell surveillance of the CNS has only recently been studied through the lens of TRM immunology. In this Review, we outline emerging concepts of CNS TRM generation, localization, maintenance, function, and specificity. In this way, we hope to highlight roles of CNS TRM in health and disease to inform future studies of adaptive neuroimmunity.

Inflammatory memory in psoriasis: From remission to recurrence.

The routine use of targeted systemic immunomodulatory therapies has transformed outcomes for people with severe psoriasis, with skin clearance (clinical remission) rates up to 60% at 1 year of biologic treatment. However, psoriasis may recur following drug withdrawal, and as a result, patients tend to continue receiving costly treatment indefinitely. Here, we review research into the "inflammatory memory" in resolved psoriasis skin and the potential mechanisms leading to psoriasis recurrence following drug withdrawal. Research has implicated immune cells such as tissue resident memory T cells, Langerhans cells, and dermal dendritic cells, and there is growing interest in keratinocytes and fibroblasts. A better understanding of the interactions between these cell populations, enabled by single cell technologies, will help to elucidate the events underpinning the shift from remission to recurrence. This may inform the development of personalized strategies for sustaining remission while reducing long-term drug burden.

Treponema pallidum Periplasmic and Membrane Proteins Are Recognized by Circulating and Skin CD4+ T Cells.

BACKGROUND: Histologic and serologic studies suggest the induction of local and systemic Treponema pallidum-specific CD4+ T-cell responses to T. pallidum infection. We hypothesized that T. pallidum-specific CD4+ T cells are detectable in blood and in the skin rash of secondary syphilis and persist in both compartments after treatment. METHODS: Peripheral blood mononuclear cells collected from 67 participants were screened by interferon-γ (IFN-γ) ELISPOT response to T. pallidum sonicate. T. pallidum-reactive T-cell lines from blood and skin were probed for responses to 89 recombinant T. pallidum antigens. Peptide epitopes and HLA class II restriction were defined for selected antigens. RESULTS: We detected CD4+ T-cell responses to T. pallidum sonicate ex vivo. Using T. pallidum-reactive T-cell lines we observed recognition of 14 discrete proteins, 13 of which localize to bacterial membranes or the periplasmic space. After therapy, T. pallidum-specific T cells persisted for at least 6 months in skin and 10 years in blood. CONCLUSIONS: T. pallidum infection elicits an antigen-specific CD4+ T-cell response in blood and skin. T. pallidum-specific CD4+ T cells persist as memory in both compartments long after curative therapy. The T. pallidum antigenic targets we identified may be high-priority vaccine candidates.

IL-17 and IFN-γ-producing Respiratory Tissue-Resident Memory CD4 T Cells Persist for Decades in Adults Immunized as Children With Whole-Cell Pertussis Vaccines.

The objective was to determine if antigen-specific tissue-resident memory T (TRM) cells persist in respiratory tissues of adults immunized as children with whole-cell pertussis (wP) or acellular pertussis (aP) vaccines. Mononuclear cells from tonsil or nasal tissue cells were cultured with Bordetella pertussis antigens and TRM cells quantified by flow cytometry. Adults immunized with wP vaccines as children had significantly more interleukin 17A (IL-17A) and interferon-γ (IFN-γ)-producing TRM cells that respond to B. pertussis antigens in respiratory tissues when compared with aP-primed donors. Our findings demonstrate that wP vaccines induce CD4 TRM cells that can persist in respiratory tissues for decades.

Residual Immunity from Smallpox Vaccination and Possible Protection from Mpox, China.

Among persons born in China before 1980 and tested for vaccinia virus Tiantan strain (VVT), 28.7% (137/478) had neutralizing antibodies, 71.4% (25/35) had memory B-cell responses, and 65.7% (23/35) had memory T-cell responses to VVT. Because of cross-immunity between the viruses, these findings can help guide mpox vaccination strategies in China.

IL-2/IL-7-inducible factors pioneer the path to T cell differentiation in advance of lineage-defining factors.

When dormant naïve T cells first become activated by antigen-presenting cells, they express the autocrine growth factor IL-2 which transforms them into rapidly dividing effector T cells. During this process, hundreds of genes undergo epigenetic reprogramming for efficient activation, and also for potential reactivation after they return to quiescence as memory T cells. However, the relative contributions of IL-2 and T cell receptor signaling to this process are unknown. Here, we show that IL-2 signaling is required to maintain open chromatin at hundreds of gene regulatory elements, many of which control subsequent stimulus-dependent alternative pathways of T cell differentiation. We demonstrate that IL-2 activates binding of AP-1 and STAT5 at sites that can subsequently bind lineage-determining transcription factors, depending upon what other external factors exist in the local T cell environment. Once established, priming can also be maintained by the stroma-derived homeostatic cytokine IL-7, and priming diminishes if Il7r is subsequently deleted in vivo. Hence, IL-2 is not just a growth factor; it lays the foundation for T cell differentiation and immunological memory.

IL-21, not IL-17A, exacerbates murine primary biliary cholangitis.

Primary biliary cholangitis (PBC) is a chronic autoimmune liver disease caused by intrahepatic bile duct injuries, resulting in fibrosis, cirrhosis, and eventually liver failure. T helper (Th) 17 cells are proposed to involve in the pathogenesis of PBC. However, how and which Th17 cell-derived cytokines affect PBC remains unclear. In this study, we investigated the effects of Th17 effector cytokines, including interleukin (IL)-17A, IL-17F, and IL-21 in PBC using a xenobiotic-induced mouse model of autoimmune cholangitis (inducible chemical xenobiotic models of PBC) treated with cytokine-expressing adeno-associated virus. Our results showed that administration of IL-17A, the well-known main cytokine produced by Th17 cells, did not augment liver inflammation or fibrosis. In contrast, we noted IL-17A-treated mice had lower hepatic Th1 cell numbers and higher hepatic CD11b+Ly6G+ polymorphonuclear myeloid-derived suppressor cell numbers. IL-17F did not alter liver inflammation or fibrosis. However, the administration of IL-21 exacerbated liver inflammatory responses and portal cell infiltration. IL-21 markedly increased the numbers of activated CD8+ T cells and liver tissue-resident memory CD8+ T cells. Moreover, IL-21 aggravates liver fibrosis in mice with autoimmune cholangitis. These results emphasized that not IL-17A but IL-21 in Th17 cell-derived cytokines affected the pathogenesis of PBC. IL-21 enhanced liver inflammation and progression to fibrosis by enhancing the numbers and effector activities of CD8+ T cells. Delineation of the effects of different Th17 effector cytokines in PBC offers clues for developing new therapeutic approaches.

NK-like CD8 T cell: one potential evolutionary continuum between adaptive memory and innate immunity.

CD8 T cells are crucial adaptive immune cells with cytotoxicity to fight against pathogens or abnormal self-cells via major histocompatibility complex class I-dependent priming pathways. The composition of the memory CD8 T-cell pool is influenced by various factors. Physiological aging, chronic viral infection, and autoimmune diseases promote the accumulation of CD8 T cells with highly differentiated memory phenotypes. Accumulating studies have shown that some of these memory CD8 T cells also exhibit innate-like cytotoxicity and upregulate the expression of receptors associated with natural killer (NK) cells. Further analysis shows that these NK-like CD8 T cells have transcriptional profiles of both NK and CD8 T cells, suggesting the transformation of CD8 T cells into NK cells. However, the specific induction mechanism underlying NK-like transformation and the implications of this process for CD8 T cells are still unclear. This review aimed to deduce the possible differentiation model of NK-like CD8 T cells, summarize the functions of major NK-cell receptors expressed on these cells, and provide a new perspective for exploring the role of these CD8 T cells in health and disease.