Tissue-resident Memory Cells Research Articles

Estradiol Enhances Antiviral CD4+Tissue-Resident Memory T Cell Responses following Mucosal Herpes Simplex Virus 2 Vaccination through an IL-17-Mediated Pathway

Estradiol (E2) is a sex hormone which has been shown to be protective against sexually transmitted infections such as herpes simplex virus 2 (HSV-2). However, few studies have examined the underlying mechanisms by which this occurs. Here, we investigated the effect of E2 on the establishment of memory T cells post-intranasal immunization with HSV-2. CD4+ T cell responses first appeared in the upper respiratory tract (URT) within 3 days postimmunization before being detected in the female reproductive tract (FRT) at 7 days. E2 treatment resulted in greater and earlier Th17 responses, which preceded augmented Th1 responses at these sites. The CD4+ T cells persisted in the URT for up to 28 days, and E2 treatment resulted in higher frequencies of memory T cells. Intranasal immunization also led to the establishment of CD4+ tissue-resident memory T cells (TRM cells) in the FRT, and E2 treatment resulted in increased Th1 and Th17 TRM cells. When the migration of circulating T cells into the FRT was blocked by FTY720, immunized E2-treated mice remained completely protected against subsequent genital HSV-2 challenge compared to non-E2 controls, confirming that TRM cells alone are adequate for protection in these mice. Finally, the enhanced vaginal Th1 TRM cells present in E2-treated mice were found to be modulated through an interleukin 17 (IL-17)-mediated pathway, as E2-treated IL-17A-deficient mice had impaired establishment of Th1 TRM cells. This study describes a novel role for E2 in enhancing CD4+ memory T cells and provides insight on potential strategies for generating optimal immunity during vaccination.IMPORTANCE Herpes simplex virus 2 (HSV-2) is a highly prevalent sexually transmitted infection for which there is currently no vaccine available. Interestingly, the female sex hormone estradiol has been shown to be protective against HSV-2. However, the underlying mechanisms by which this occurs remains relatively unknown. Our study demonstrates that under the influence of estradiol treatment, intranasal immunization with an attenuated strain of HSV-2 leads to enhanced establishment of antiviral memory T cell responses in the upper respiratory tract and female reproductive tract. In these sites, estradiol treatment leads to greater Th17 memory cells, which precede enhanced Th1 memory responses. Consequently, the T cell responses mounted by tissue-resident memory cells in the female reproductive tract of estradiol-treated mice are sufficient to protect mice against vaginal HSV-2 challenge. This study offers important insights regarding the regulation of mucosal immunity by hormones and on potential strategies for generating optimal immunity during vaccination.

Brain tissue-resident Immune Memory Cells are Required for long-term Protection Against CNS Infection With Rabies Virus

Immune memory cells residing in previously infected, nonlymphoid tissues play a role in immune surveillance. In the event that circulating antibodies fail to prevent virus spread to the tissues in a secondary infection, these memory cells provide an essential defense against tissue reinfection. CNS tissues are isolated from circulating immune cells and antibodies by the blood-brain barrier, making the presence of tissue-resident immune memory cells particularly needed to combat recurrent infection by neurotropic viruses. Wild-type and laboratory-engineered rabies virusesare neurotropic, differ in pathogenicity, and have varying effects on BBB functions. These viruses have proven invaluable tools in demonstrating the importance of tissue-resident immune memory cells in the reinfection of CNS tissues. Only Type 1 immune memory is effective at therapeutically clearing a secondary infection with wild-type rabies viruses from the CNS and does so despite the maintenance of blood-brain barrierintegrity.

IL-21 from high-affinity CD4 T cells drives differentiation of brain-resident CD8 T cells during persistent viral infection.

Development of tissue-resident memory (TRM) CD8 T cells depends on CD4 T cells. In polyomavirus central nervous system infection, brain CXCR5hi PD-1hi CD4 T cells produce interleukin-21 (IL-21), and CD8 T cells lacking IL-21 receptors (IL21R-/-) fail to become bTRM IL-21+ CD4 T cells exhibit elevated T cell receptor (TCR) affinity and higher TCR density. IL21R-/- brain CD8 T cells do not express CD103, depend on vascular CD8 T cells for maintenance, are antigen recall defective, and lack TRM core signature genes. CD4 T cell-deficient and IL21R-/- brain CD8 T cells show similar deficiencies in expression of genes for oxidative metabolism, and intrathecal delivery of IL-21 to CD4 T cell-depleted mice restores expression of electron transport genes in CD8 T cells to wild-type levels. Thus, high-affinity CXCR5hi PD-1hi CD4 T cells in the brain produce IL-21, which drives CD8 bTRM differentiation in response to a persistent viral infection.

Peripherally induced brain tissue-resident memory CD8+ T cells mediate protection against CNS infection.

The central nervous system (CNS) is classically viewed as immune-privileged; however, recent advances highlight interactions between the peripheral immune system and CNS in controlling infections and tissue homeostasis. Tissue-resident memory (Trm) CD8+ T cells in the CNS are generated after brain infections, but it is unknown whether CNS infection is required to generate brain Trm cells. We show that peripheral infections generate antigen-specific CD8+ memory T cells in the brain that adopt a unique Trm signature. Upon depletion of circulating and perivascular memory T cells, this brain signature was enriched and the surveilling properties of brain Trm cells revealed by intravital imaging. Importantly, peripherally induced brain Trm cells showed evidence of rapid activation, enhanced cytokine production and mediated protection after brain infections. These data reveal that peripheral immunizations can generate brain Trm cells and will guide potential use of T cells as therapeutic strategies against CNS infections and neurological diseases.

Human T cells interacting with HNSCC‐derived mesenchymal stromal cells acquire tissue‐resident memory like properties

Tissue-resident memory (Trm) cells are specialized components of both CD4+ and CD8+ T cell subsets that persist in peripheral nonlymphoid tissues following infections and provide fast response in case of a secondary invasion by the same pathogen. Trm cells express the surface markers CD69, CD103, and the immune checkpoint molecule PD-1. Trm cells develop not only in the context of infections but also in tumors, where they can provide a line of defense as suggested by the positive correlation between the frequency of tumor-infiltrating Trm cells and patients' survival. Trm cells persistence in peripheral tissues depends on their adaptation to the local microenvironment and the presence of survival factors, mainly IL-7, IL-15, and Notch ligands. However, the cell sources of these factors are largely unknown, especially in the context of tumors. Here, we show that head-neck squamous cell carcinoma (HNSCC) is enriched in CD4+ and CD8+ T cells with a Trm phenotype. Moreover, we show that mesenchymal stromal cells that accumulate in HNSCC are a source of survival factors and allow proper expression of Trm-typical markers in a VCAM1-dependent manner.

Decidual CD8+T cells exhibit both residency and tolerance signatures modulated by decidual stromal cells

BackgroundDuring early pregnancy, tolerance of the semi-allogeneic fetus necessitates comprehensive modifications of the maternal immune system. How decidual CD8+T (CD8+dT) cells balance maternal tolerance of the fetus with defense from invading pathogens remains undefined.MethodsWe investigated the distribution patterns of CD8+T cells and their heterogeneity in paired peripheral blood and decidual tissue in the first trimester of pregnancy using flow cytometry and mRNA-Seq. Gene Set Enrichment Analysis was utilized to determine the transcriptional features of CD8+dT cells. Moreover, we examined activation of T cells when they were cocultured with trophoblasts, in addition to the effect of the fetal–maternal environment on peripheral CD8+T (CD8+pT) cells.ResultsWe found that, compared with CD8+pT cells, CD8+dT cells consisted mainly of effector memory cells (TEM) and terminally differentiated effector memory cells (TEMRA). Both TEM and TEMRA subsets contained increased numbers of CD27+CD28− cells, which have been shown to possess only partial effector functions. In-depth analysis of the gene-expression profiles of CD8+dT cells revealed significant enrichment in T cell exhaustion-related genes and core tissue residency signature genes that have been found recently to be shared by tissue resident memory cells and tumor−infiltrating lymphocytes (TILs). In accordance with gene expression, protein levels of the exhaustion-related molecules PD-1 and CD39 and the tissue resident molecules CD103 and CXCR3 were increased significantly with almost no perforin secretion in CD8+dT cells compared with CD8+pT cells. However, the levels of granzyme B, IFN-γ, and IL-4 in CD8+dT cells were increased significantly compared with those in CD8+pT cells. Both CD8+dT and CD8+pT cells were not activated after being cocultured with autologous trophoblast cells. Moreover, the production of granzyme B in CD103+CD8+dT cells decreased significantly compared with that in their CD103− counterparts. Coculture with decidual stromal cells and trophoblasts upregulated CD103 expression significantly in CD8+pT cells.ConclusionsOur findings indicate that the selective silencing of effector functions of resident CD8+dT cells may favor maternal–fetal tolerance and that the decidual microenvironment plays an important role in promoting the residency of CD8+T cells and their tolerance–defense balance.

Interferon gamma promotes long term retention of tissue resident memory T cells in the brain following alphavirus clearance in mice

Abstract The prototypic alphavirus Sindbis virus (SINV) targets neurons in mice, providing a well characterized model of alphavirus encephalomyelitis. SINV clearance from the brain requires a noncytolytic mechanism to avoid neurological damage and dysfunction and is facilitated by synergistic cooperation between anti-SINV antibody and interferon gamma (IFN-γ). While infectious virus is cleared from the brain by 7 to 10 days post infection (DPI), SINV RNA levels decrease slowly and remain detectable at low levels for at least a year following infection. Because alphaviral RNA is infectious, strict control by the immune system is required to prevent virus reactivation and relapse of disease. To characterize the long-term T cell response following clearance of infectious virus, C57BL/6 mice with intact or impaired IFN-γ signaling were infected with the nonfatal TE strain of SINV, and the frequency and functionality of different T cell populations were evaluated in the brain by flow cytometry. While highly proliferative short-lived T effector cells made up the majority of CD4+ and CD8+ T cells during active infection at 7 DPI, long-lived effector memory T cells became the predominant T cell type by 14 DPI. Following the period of virus clearance, both CD4+ and CD8+ T cell polyfunctionality increased, with more cells expressing multiple effector proteins at higher intensities, and PD-1 and TIM-3 exhaustion marker expression decreased. The percentage of CD8+ T cells characterized as CD103+ tissue resident memory (TRM) cells was decreased in mice with impaired IFN-γ signaling. This indicates IFN-γ signaling promotes the development of TRM cells in the brain during SINV infection and suggests IFN-γ supports long term control of SINV reactivation.

Immune pressure against ovarian cancer depends on antigen-specific CD69+CD103+TRM T cells

Abstract The relationship between stem-like, tissue resident memory (TRM) and exhausted T cells at tumor beds is incompletely understood. We found that more than 50% of the CD8+ T cells in human ovarian carcinomas are TRM cells. RNA seq of TRM and their re-circulating counterparts infiltrating multiple human ovarian carcinomas showed a very distinctive phenotype, characterized by co-upregulation of effector and exhaustion markers, along with increased clonality and marked proliferative and lipid metabolism signatures. Interestingly, both populations showed very little overlap in TCR repertoire. Single cell analysis of TRM population showed that this is a heterogeneous population composed of different clusters defined by gene expression and TCR repertoire. A distinctive cluster shows higher expression of cytotoxic mediators and exhaustion markers, along with higher clonality and higher proliferation rate, with a TCR repertoire shared by CD103+TCF7+ stem-like cells. Single cell ATACseq showed that TRM and their counterpart have different chromatin structure and, within both populations also appear variables epigenetic landscapes. In addition, the presence of TRM is correlated with better prognosis, while tumor antigen-specific TRM T cells transferred into syngeneic tumor-bearing mice were more effective at delaying tumor growth than their tumor antigen-specific re-circulating counterparts. Together, our data indicate that productive immune pressure against malignant progression depends on a subset of CD8+ stem-like T cells differentiating into a narrow cluster of TRM T cells of ~300 TCR clones, which represent true tumor antigen-specific effector lymphocytes.

Multi-dimensional analysis of tumor-specific and bystander CD8+ tumor infiltrating lymphocytes with TetTCR-SeqHD

Abstract Objective Antigen specific CD8+ T cells were isolated from primary tumor tissue and analyzed for on the single cell level to determine tumor-specific signatures for further study. Methods Primary tumor, metastatic lesions, adjacent healthy tissue and peripheral blood were from collected from patients at Dell Seton Medical Center at the University of Texas. Matched tumor and healthy samples underwent exome and RNA sequencing to allow prediction of tumor-specific epitopes. We then synthesized DNA-barcoded tetramers for all predicted tumor associated-epitopes and an HLA-matched library of exogenous epitopes. Tetramer stained CD8+ T cells were then sorted and analyzed by TetTCR-SeqHD, a technology that we recently developed to link TCR antigen specificity, TCR sequence, gene expression, and phenotype of single T cells. Results We analyzed ~2900 tumor infiltrating lymphocytes (TILs) and T cells from adjacent healthy tissue and observed enrichment for cytotoxic effector and tissue resident memory cells within the tumor versus the healthy tissues. We also note an enrichment of bystander cells, specifically those reactive to Epstein Barr Virus epitopes, within the cytotoxic cluster. Finally, we analyze the distribution and differential expression between tumor-specific lymphocytes and bystander T cells, noting reduced expression of CD57 and CD49d in the tumor-specific TILs, as previously described. Conclusions We validate the use of TetTCR-SeqHD for gaining insight into phenotypic and gene expression differences between bystander and tumor-specific TILs. We identified markers for future exploration and demonstrate key differences between shared clonal populations within healthy and cancerous tissues.

A single cell approach to identify the role of IL-9 secreting tissue-resident CD4+ T cells in allergic airway recall responses

Abstract Allergic asthma is a chronic intermittent inflammatory lung disease where patients experience relatively few symptoms between exacerbations precipitated by allergen-challenge of allergic memory responses. Allergen-specific CD4+ T cells are believed to be a major mediator of allergic memory responses through the production of type 2 cytokines including IL-4, IL-5, IL-13 and IL-9. Recently, tissue-resident memory (Trm) cells have been identified in peripheral tissues that mediate mucosal barrier immunity. However, the precise role of Trm CD4+ T cells in allergic exacerbations is not clearly defined. In studies to examine the role of IL-9 in allergen-specific Trm responses to Aspergillus fumigatus we observed that all IL-9-secreting T cells had a resident memory phenotype. Inhibition of circulating T cells by administering FTY720 in the last month of rest, diminished the total CD4 T cells population in the lung, while the Th9 cells remained stable. Blockade of IL-9 prior to recall challenge phase reduced overall allergic lung inflammation as assessed both by flow cytometry and single cell RNA-seq. Within eight hours after challenge, IL-9 is one of the top genes identified by scRNA-seq that distinguishes ST2+ and ST2− CD4+ T cell populations in the lung. Using IL-9-reporter mice we also compared gene expression in isolated IL-9+ T cells and other type 2 T cells. These findings demonstrate that IL-9-producing Trms play an important role in mediating allergic memory responses. IL-9 could be a promising therapeutic target specifically in patients showing symptoms of intermittent allergic response.

Regulation of Respiratory CD8+ T cell Immunity by Suppressive Monocyte Derived Dendritic Cells (moDCs)

Abstract Active immune suppression can mediate the balance between protective cellular immunity and harmful immunopathology. This suppression can occur locally, at an infection site, or in regional draining lymph nodes (dLN). Immune regulation is of particular importance in sites such as the lung where aberrant immunopathology can result in loss of tissue function. We have recently identified a novel population of CD11b+CD103+CCR2+monocyte-derived dendritic cells (moDCs) which directly suppress CD8+ T cell proliferation in vitro. Respiratory infection with RNA viruses recruits these moDCs either exclusively to the dLN (after vesicular stomatitis virus infection) or both the dLN and site of viral replication (after influenza infection). Exclusive depletion of moDCs from the dLN using CCR2-DTR bone marrow chimeras results in enhanced respiratory CD8+ lung tissue resident memory cell formation. By contrast, depletion of moDCs from both the dLN and respiratory tract following influenza infection results in enhanced respiratory CD8+ T cell responses coupled with fatal immunopathology. Together, these data suggest that suppressive moDCs govern key aspects of respiratory immunity by regulating the function and development of long-lived CD8+ T cells, thereby balancing immunity and adverse pathology in the context of viral infection. Ongoing experiments seek to uncouple moDC interactions with CD8+ T cells in specific sites in order to better understand, and potentially beneficially modulate, CD8+ T cell immunity within the lung.

Age-related changes in the immune phenotype of human memory CD4 T cells differ between colon and peripheral blood

Abstract Gut lamina propria (LP) CD4 T cells are primarily tissue resident memory cells that are critical in maintaining gut homeostasis and responding to enteric pathogens. We established that aging altered the phenotypic and functional profiles of human small intestinal CD4 T cells. However, given site-specific differences in immune cell composition in the gut, it is difficult to extrapolate these findings to other locations. To probe the effect of aging on large intestine CD4 T cells, we used flow cytometry to analyze immune phenotypes of colon LP CD4 T cells from younger (34±2yrs mean±SEM; N=6) and older (80±2yrs, N=6) adults. Tissue-specificity of these profiles was determined by comparing to memory (CD45RA−) peripheral blood (mPB) CD4 T cells in a separate cohort of similarly aged persons (N=6/age group). Among younger samples, LP CD4 T cells expressed higher levels of co-inhibitory molecules CTLA4 and PD1, but had lower turnover (Ki67+) and IL2 receptor (CD25) expression relative to mPB CD4 T cells. Expression of activation (CD38, HLA-DR), senescence (CD57) and survival (Bcl2) markers were similar. Regarding age effects, frequencies of LP CD4 T cells (% of CD3 T cells) were similar between older and younger samples; mPB CD4 T cell frequencies were higher in older vs younger samples. Older LP CD4 T cells had lower CTLA4, PD1, CD38, CD25 and Ki67 and higher Bcl2 expression than younger LP CD4 T cells. Among mPB CD4 T cells, greater age was only associated with a higher CD57+ fraction. Thus, age effects differed between colon LP and mPB CD4 memory T cells. Loss of co-inhibitory markers, decreased turnover and increased survival of colon LP CD4 T cells may contribute to altered homeostasis, loss of protective immunity, and chronic inflammation in older people.

Circulating memory is sufficient for protective immunity to infection with Chlamydia muridarum

Abstract Chlamydia trachomatis is the most prevalent bacterial sexually transmitted infection worldwide and yet no vaccine currently exists to protect the public from this reproductive tract pathogen. Recent reports suggest that CD4 tissue resident memory cells (TRM) are essential for vaccine-induced protection against Chlamydia trachomatis in a mouse infection model. Additionally, CD4 TRM in the female reproductive tract (FRT) have been shown to co-localize with APCs in memory lymphocyte clusters (MLCs) to enhance secondary adaptive immunity to reproductive tract infections. Here, we examine a requirement for CD4 TRM and MLCs in protective immunity against infection with Chlamydia muridarum (Cm). Using parabiosis of naïve and Chlamydia-immune mice, we demonstrate that although CD4 TRM form in the FRT after primary Cm infection, they are not required for protection. Furthermore, histological analysis of FRT tissue from parabionts revealed that circulating memory is sufficient for protection against intravaginal challenge with Cm in the absence of CD4 TRM or MLCs in the reproductive tract. Moreover, this protective circulating memory can be induced through intravaginal or intranasal challenge with Chlamydia, indicating that a strong mucosal immunization can generate circulating protection in a vaccine setting. Ongoing work using adoptive transfers and antibody depletions of CD4 T cells and B cells will reveal which memory lymphocytes are responsible for the observed circulating protection.

P19 Frequencies of IL-17+CD8+ T-cells and tissue resident memory T-cells in early inflammatory arthritis

Abstract Background Early inflammatory arthritis (EIA) is classically described as arthritis of ≤ 12 months duration. EIA can be sub-divided into psoriatic arthritis (PsA), other spondyloarthritides (SpA), rheumatoid arthritis (RA) or reactive arthritis (ReA). EIA is often difficult to classify. IL-17+CD8+ T-cells have previously been identified within the joints of patients with PsA. Many of these cells have properties of tissue resident memory (TRM) T-cells, a recently defined subset of CD8+ T cells which remain resident within tissues ready to respond rapidly. We sought to identify the presence and frequency of Tc17 and/ or TRM cells within the joints of patients with EIA to facilitate correct diagnosis and management. Methods 27 EIA patient samples were collected from patients in the Rheumatology clinic setting. Consent was obtained using established ethics. Samples underwent synovial fluid mononuclear cell (SFMC) isolation and cryopreservation. After thawing, 2 million cells from each sample were stimulated for 3 hours with PMA, Ionomycin and Golgistop. Samples were stained for CD-3, CD-14, CD-4, CD-8 and cytokine expression (IL-17, IFN- γ and TNF-α). A further 2 million cells from 21 of these samples were stained using an extracellular stain for the presence of TRM cells (CD69+CD103+ CD8+ T cells). Samples were all acquired on BD Canto. Analysis was completed using FlowJo and PRISM. Results Patients were classified into disease groups based on clinical features and laboratory results, blinded to the SF results. Seronegative RA was subdivided based on disease pattern into RA like or oligo-arthritis like. Patient numbers per group: seropositive RA = 5, seronegative RA (RA-like) =4, seronegative RA (oligo-like) =8, PsA=4, SpA=6. The frequency of Tc17 cells was significantly increased in SF of patients with PsA compared to all other groups (ordinary one-way ANOVA p < 0.0001). Specific comparisons were seropositive RA vs PsA (p = 0.003), seronegative RA vs PsA (RA like p = 0.0002, oligo IA p < 0.0001) and PsA vs SpA (p = 0.0013). The frequency of TRM cells was also significantly increased in PsA compared to all other groups (ordinary one-way ANOVA p = 0.0013), and specific groups: seropositive RA and PsA (p = 0.0031), seronegative RA and PsA (RA like p = 0.0086, oligo IA p = 0.0039) and PsA and SpA P = 0.0009. In contrast, Th17 cells and IFN-γ+ CD8+ T-cells (Tc1 cells) were found in all SF with no significant difference between sub-groups. Conclusion This preliminary study of 27 patients with EIA shows that the frequency of Tc17 and CD69+CD103+CD8+ TRMcells are increased in the SF of patients with early PsA compared to other patient groups. Our data also show that TRM cells are present at detectable frequencies in the joints of most patients with EIA. Further work is required to establish if these cells could function as a clinical biomarker to aid diagnosis. Disclosures C.D. Hughes None. K.J.A. Steel None. S. Ryan None. S. Lahnunlimi None. B. Menon None. L.S. Taams None. B.W. Kirkham Consultancies; BK has worked as a consultant for Eli Lilly, Gilead, Jannsen and Novartis. Grants/research support; BK has received research support from Eli Lilly and Novartis.

Donor T and NK Cells with a Special Tissue-Resident Memory Phenotype Migrate into the Periphery of Lung Transplant Recipients - A Potential Feature for Tolerance Development

After lung transplantation (LuTx), a transient chimerism of donor cells exists in the blood of recipients due to the migration of lymphocytes from the transplanted lung into the periphery. We aimed to characterize the phenotype of donor CD8+ and CD4+ T and NK cells and to investigate whether they might represent tissue-resident memory (TRM) cells. Lymphocyte dynamics in recipient blood were determined in 97 lung transplant patients directly (T0), 24 hours (T24) and 3 (wks) weeks after LuTx using flow cytometry with lineage-, tissue-, and donor HLA class I allele-specific mAb. The same makers were used to determine the phenotype of lymphocytes present in organ storage solution (perfusate, n=102), recipient explanted lung parenchyma (n=28) and donor trachea (n=17). In peripheral blood of all recipients, donor-derived CD4+ and CD8+ T and CD56dim NK cells were detected at T0, T24 and 3 wks after LuTx and had higher CD69 expression compared to recipient cells (p<0.05) and were mostly CD25- CCR7- memory cells. This phenotype was similar to T and NK cells in corresponding perfusates. In lung parenchyma and trachea, most CD69+ T and NK cells showed coexpression of other tissue residency markers such as CD103, CD49a and PD-1 with significant enrichment in trachea (p<0.05). These markers were not found in circulating donor lymphocytes and perfusates, indicating that they represent distinct memory T and NK subsets. Donor T and NK cells showed higher IFN-γ production with PMA/ionomycin stimulation compared to recipient cells (p<0.05). The presence of these particular TRM cells did not have an impact on the development of PGD 24h after LuTx. However, patients with high frequencies of donor T cells showed a trend towards a CLAD-free survival 2 years post LuTx, although no statistical significance was reached (p=0.15). Our results demonstrate that donor T and NK cells found in the blood of LuTx recipients are a distinct subset from circulating lymphocytes and TRM cells present in lung tissue, since they express CD69 but lack expression of other classical TRM markers. They display a higher functional capacity despite the onset of immunosuppression. Donor T cells might be clinically relevant for tolerance induction and long-term survival after transplantation due to their unique features.

Asialo GM1-positive liver-resident CD8 T cells that express CD44 and LFA-1 are essential for immune clearance of hepatitis B virus.

Persistent hepatitis B virus (HBV) infection results in chronic liver diseases that may progress to chronic hepatitis, liver cirrhosis, and subsequent hepatocellular carcinoma. Previous studies demonstrated that adaptive immunity, in particular CD8 T cells, is critical in HBV elimination. Recent studies have revealed a distinct tissue-localized T cell lineage, tissue-resident memory (TRM) cells, that is crucial for protective immunity in peripheral tissues. In this study, we showed that treatment with an anti-asialo GM1 (ASGM1) antibody (Ab), which depletes NK cells, led to impairment of HBV clearance in a mouse animal model. Unexpectedly, the ability to clear HBV was not significantly impaired in NFIL3 KO mice, which are deficient in NK cells, implying that other non-NK ASGM1-positive immune cells mediate HBV clearance. We isolated intrahepatic ASGM1-positive cells from NFIL3 KO mice and analyzed the immune phenotype of these cells. Our results demonstrated a distinct population of CD44+ LFA-1hi CD8 T cells that were the major intrahepatic ASGM1-positive immune cells in NFIL3 KO mice. Importantly, transcriptome analysis revealed that these ASGM1-positive CD8 T cells had distinct gene profiles and shared a similar core gene signature with TRM cells. In addition to both transcriptional and phenotypic liver residency characteristics, ASGM1-positive CD8 T cells were able to home to and be retained in the liver after adoptive transfer. Taken together, our study results indicate that these ASGM1-positive liver-resident CD8 T cells are the major effector immune cells mediating anti-HBV immunity.

Resident Memory B Cells.

In mammals, adaptive immunity is mediated by a broadly diverse repertoire of naive B and T lymphocytes that recirculate between secondary lymphoid organs. Initial antigen exposure promotes lymphocyte clonal expansion and differentiation, including the formation of memory cells. Antigen-specific memory cells are maintained at higher frequencies than their naive counterparts and have different functional and homing abilities. Importantly, a subset of memory cells, known as tissue-resident memory cells, is maintained without recirculating in nonlymphoid tissues, often at barrier surfaces, where they can be reactivated by antigen and rapidly perform effector functions that help protect the tissue in which they reside. Although antigen-experienced B cells are abundant at many barrier surfaces, their characterization as tissue-resident memory B (BRM) cells is not well developed. In this study, we describe the characteristics of memory B cells in various locations and discuss their possible contributions to immunity and homeostasis as bona fide BRM cells.

Immunological Memory of Psoriatic Lesions.

The natural course of psoriasis is the appearance of new lesions in the place of previous ones, which disappeared after a successful therapy. Recent studies of psoriasis etiopathogenesis showed that after psoriatic plaques have disappeared, in healthy skin we can still find a trace of inflammation in the form of tissue resident memory cells (TRM). They are originally responsible for protection against viral and bacterial infections in non-lymphatic tissues. In psoriatic inflammation, they are characterized by heterogeneity depending on their origin. CD8+ T cells TRM are abundantly present in psoriatic epidermis, while CD4+ TRM preferentially populate the dermis. In psoriasis, epidermal CD8+ TRM cells express CLA, CCR6, CD103 and IL-23R antigen and produce IL-17A during ex vivo stimulation. However, CD4+ CD103+ TRM can also colonize the epidermis and produce IL-22 during stimulation. Besides T cells, Th22 and epidermal DCs proved that epidermal cells in healed skin were still present and functioning after several years of disease remission. It explains the clinical phenomenon of the tendency of psoriatic lesions to relapse in the same location and it allows to develop new therapeutic strategies in the future.

CD8+ T-cell plasticity regulates vascular regeneration in type-2 diabetes.

In this study, we observe that the ischemic tissues of type-2 diabetic (T2D) patients and mice have significantly more CD8+ T-cells than that of their normoglycemic counterparts, respectively. However, the role of CD8+ T-cells in the pathogenesis of diabetic vascular complication has been less studied.Methods: We employed loss-of-function studies in mouse models using the non-lytic anti-CD8 antibody that blocks tissue infiltration of CD8+ T-cells into the injured tissue. We also performed genome-wide, single-cell RNA-sequencing of CD8+ T-cells to uncover their role in the pathogenesis of diabetic vascular diseases.Results: The vascular density is negatively correlated with the number of CD8+ T-cells in the ischemic tissues of patients and mice after injury. CD8+ T-cells or their supernatant can directly impair human and murine angiogenesis. Compared to normoglycemic mice that can regenerate their blood vessels after injury, T2D mice fail in this regeneration. Treatment with the CD8 checkpoint blocking antibody increases the proliferation and function of endothelial cells in both Leprdb/db mice and diet-induced diabetic Cdh5-Cre;Rosa-YFP lineage-tracing mice after ischemic injury. Furthermore, single-cell transcriptomic profiling reveals that CD8+ T-cells of T2D mice showed a de novo cell fate change from the angiogenic, tissue-resident memory cells towards the effector and effector memory cells after injury. Functional revascularization by CD8 checkpoint blockade is mediated through unleashing such a poised lineage commitment of CD8+ T-cells from T2D mice.Conclusion: Our results reveal that CD8+ T-cell plasticity regulates vascular regeneration; and give clinically relevant insights into the potential development of immunotherapy targeting vascular diseases associated with obesity and diabetes.

Triple Therapy with MerTK and PD1 Inhibition Plus Radiotherapy Promotes Abscopal Antitumor Immune Responses.

Radiotherapy (RT) traditionally has been used for local tumor control in the treatment of cancer. The recent discovery that radiotherapy can have anticancer effects on the immune system has led to recognition of its ability to sensitize the tumor microenvironment to immunotherapy. However, radiation can also prompt adverse immunosuppressive effects that block aspects of systemic response at other tumor sites. Our hypothesis was that inhibition of the MER proto-oncogene tyrosine kinase (MerTK) in combination with anti-programmed cell death-1 (α-PD1) checkpoint blockade will enhance immune-mediated responses to radiotherapy. We tested the efficacy of this triple therapy (Radiation + α-PD1 + α-MerTK mAbs) in 129Sv/Ev mice with bilateral lung adenocarcinoma xenografts. Primary tumors were treated with stereotactic radiotherapy (36 Gy in 3 12-Gy fractions), and tumors were monitored for response. The triple therapy significantly delayed abscopal tumor growth, improved survival rates, and reduced numbers of lung metastases. We further found that the triple therapy increased the activated CD8+ and NK cells populations measured by granzyme B expression with upregulation of CD8+CD103+ tissue-resident memory cells (TRM) within the abscopal tumor microenvironment relative to radiation only. The addition of α-PD1 + α-MerTK mAbs to radiotherapy could alter the cell death to be more immunogenic and generate adaptive immune response via increasing the retention of TRM cells in the tumor islets of the abscopal tumors which was proven to play a major role in survival of non-small cell lung cancer patients.