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Adoptive immunotherapy with Tregs is a promising approach for preventing or treating type 1 diabetes. Islet Antigen-specific Tregs have more potent therapeutic effects than polyclonal cells, but their low frequency is a barrier for clinical application. To generate Tregs that recognize islet antigens, we engineered a chimeric antigen receptor (CAR) derived from a monoclonal antibody with specificity for the insulin B chain 10-23 peptide presented in the context of the I[A.sup.g7] MHC class II allele present in NOD mice. Peptide specificity of the resulting InsB-g7 CAR was confirmed by tetramer staining and T cell proliferation in response to recombinant or islet-derived peptide. The InsB-g7 CAR redirected NOD Treg specificity such that insulin B 10-23-peptide stimulation enhanced suppressive function, measured via reduction of proliferation and IL-2 production by BDC2.5 T cells and CD80 and CD86 expression on dendritic cells. Cotransfer of InsB-g7 CAR Tregs prevented adoptive transfer diabetes by BDC2.5 T cells in immunodeficient NOD mice. In WT NOD mice, InsB-g7 CAR Tregs prevented spontaneous diabetes. These results show that engineering Treg specificity for islet antigens using a T cell receptor-like CAR is a promising therapeutic approach for the prevention of autoimmune diabetes., Introduction Autoimmune type 1 diabetes (T1D) is caused by T cell-mediated destruction of the insulin-producing [beta] cells of the islets of Langerhans in the pancreas. Tregs, defined by expression of [...]

Non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis (NASH) are prevalent liver conditions that underlie the development of life-threatening cirrhosis, liver failure and liver cancer. Chronic necro-inflammation is a critical factor in development of NASH, yet the cellular and molecular mechanisms of immune dysregulation in this disease are poorly understood. Here, using single-cell transcriptomic analysis, we comprehensively profiled the immune composition of the mouse liver during NASH. We identified a significant pathology-associated increase in hepatic conventional dendritic cells (cDCs) and further defined their source as NASH-induced boost in cycling of cDC progenitors in the bone marrow. Analysis of blood and liver from patients on the NAFLD/NASH spectrum showed that type 1 cDCs (cDC1) were more abundant and activated in disease. Sequencing of physically interacting cDC-T cell pairs from liver-draining lymph nodes revealed that cDCs in NASH promote inflammatory T cell reprogramming, previously associated with NASH worsening. Finally, depletion of cDC1 in XCR1.sup.DTA mice or using anti-XCL1-blocking antibody attenuated liver pathology in NASH mouse models. Overall, our study provides a comprehensive characterization of cDC biology in NASH and identifies XCR1.sup.+ cDC1 as an important driver of liver pathology. Single-cell analyses reveal cDC1 as conserved immunological drivers of non-alcoholic steatohepatitis in mice and humans, Author(s): Aleksandra Deczkowska [sup.1] [sup.14] , Eyal David [sup.1] , Pierluigi Ramadori [sup.2] , Dominik Pfister [sup.2] , Michael Safran [sup.3] , Baoguo At the [sup.1] , Amir Giladi [sup.1] [...]

PE07.01 COVID-19 and associated disruptions may indirectly affect HIV outcomes in two capital cities in Western/Central Africa: a modelling study R. Silhol (1) ; L. Geidelberg (1) ; K. Mitchell [...]

SARS-CoV-2 is responsible for the development of coronavirus disease 2019 (COVID-19) in infected individuals, who can either exhibit mild symptoms or progress toward a life-threatening acute respiratory distress syndrome (ARDS). Exacerbated inflammation and dysregulated immune responses involving T and myeloid cells occur in COVID-19 patients with severe clinical progression. However, the differential contribution of specific subsets of dendritic cells and monocytes to ARDS is still poorly understood. In addition, the role of [CD8.sup.+] T cells present in the lung of COVID-19 patients and relevant for viral control has not been characterized. Here, we have studied the frequencies and activation profiles of dendritic cells and monocytes present in the blood and lung of COVID-19 patients with different clinical severity in comparison with healthy individuals. Furthermore, these subpopulations and their association with antiviral effector [CD8.sup.+] T cell subsets were also characterized in lung infiltrates from critical COVID-19 patients. Our results indicate that inflammatory transitional and nonclassical monocytes and [CD1c.sup.+] conventional dendritic cells preferentially migrate from blood to lungs in patients with severe COVID-19. Thus, this study increases the knowledge of specific myeloid subsets involved in the pathogenesis of COVID-19 disease and could be useful for the design of therapeutic strategies for fighting SARS-CoV-2 infection., Introduction The new SARS-CoV-2 virus causing coronavirus disease 2019 (COVID-19) has unleashed the current pandemic. Individuals infected with this pathogen can either remain asymptomatic or progress from mild to severe [...]

COVID-19 spans a wide range of symptoms, sometimes with profound immune system involvement. How immune cell subsets change during the disease course and with disease severity needs further study. While myeloid cells have been shown to initiate and maintain responses to pneumonia and lung inflammation, often playing a role in resolution, their involvement with COVID-19 remains unknown. In this issue of the JCI, Sanchez-Cerrillo and Landete et al. investigated DCs and monocytes from blood and bronchial secretions of patients with varying COVID-19 severity and with healthy controls. The authors conclude that circulating monocytes and DCs migrate from the blood into the inflamed lungs. While sampling differences in sex, collection timing, bacteria/fungal infection, and corticosteroid treatment limit interpretation, we believe that reprogramming monocyte or macrophages by targeting immunometabolism, epigenetics, or the cytokine milieu holds promise in resolving lung inflammation associated with COVID-19., Introduction COVID-19 disease morbidity and mortality have swept the globe since early 2020. The role of the immune response in SARS-CoV-2 pathogenesis and its determinants for the clinical course and [...]

Graft-versus-host disease (GVHD) remains an important cause of morbidity and mortality after allogeneic hematopoietic cell transplantation (allo-HCT). For decades, GVHD prophylaxis has included calcineurin inhibitors, despite their incomplete efficacy and impairment of graft-versus-leukemia (GVL). Distinct from pharmacologic immune suppression, we have developed what we believe is a novel, human CD83-targeted chimeric antigen receptor (CAR) T cell for GVHD prevention. CD83 is expressed on allo-activated conventional [CD4.sup.+] T cells (Tconvs) and proinflammatory dendritic cells (DCs), which are both implicated in GVHD pathogenesis. Human CD83 CAR T cells eradicate pathogenic [CD83.sup.+] target cells, substantially increase the ratio of regulatory T cells (Tregs) to allo-activated Tconvs, and provide durable prevention of xenogeneic GVHD. CD83 CAR T cells are also capable of treating xenogeneic GVHD. We show that human acute myeloid leukemia (AML) expresses CD83 and that myeloid leukemia cell lines are readily killed by CD83 CAR T cells. Human CD83 CAR T cells are a promising cell-based approach to preventing 2 critical complications of allo-HCT--GVHD and relapse. Thus, the use of human CD83 CAR T cells for GVHD prevention and treatment, as well as for targeting [CD83.sup.+] AML, warrants clinical investigation., Introduction Allogeneic hematopoietic cell transplantation (allo-HCT) is a procedure performed with curative intent for high-risk hematologic malignancies and bone marrow failure syndromes. Annually, thousands of patients receive allo-HCT worldwide, and [...]

Plasmacytoid dendritic cells (pDCs) are robust producers of IFN[alpha] and one of the first immune cells to respond to SIV infection. To elucidate responses to early HIV-1 replication, we studied blood pDCs in 29 HIV-infected participants who initiated antiretroviral therapy during acute infection and underwent analytic treatment interruption (ATI). We observed an increased frequency of partially activated pDCs in the blood before detection of HIV RNA. Concurrent with peak pDC frequency, we detected a transient decline in the ability of pDCs to produce IFN[alpha] in vitro, which correlated with decreased phosphorylation of IFN regulatory factory 7 (IRF7) and NF-[kappa]B. The levels of phosphorylated IRF7 and NF-[kappa]B inversely correlated with plasma IFNA2 levels, implying that pDCs were refractory to in vitro stimulation after IFN[alpha] production in vivo. After ATI, decreased expression of IFN genes in pDCs inversely correlated with the time to viral detection, suggesting that pDC IFN loss is part of an effective early immune response. These data from a limited cohort provide a critical first step in understanding the earliest immune response to HIV-1 and suggest that changes in blood pDC frequency and function can be used as an indicator of viral replication before detectable plasma viremia., Introduction Plasmacytoid dendritic cells (pDCs) are important mediators of the early innate immune response to viral infection through robust production of type I IFNs, particularly IFN[alpha] (1, 2). pDCs recognize [...]

Acute liver failure (ALF) is a life-threatening condition, and liver transplantation is the only therapeutic option. Although immune dysregulation is central to its pathogenesis, the precise mechanism remains unclear. Here, we show that the number of peripheral and hepatic plasmacytoid DCs (pDCs) decrease during acute liver injury in both humans and mice. Selective depletion of pDCs in [Siglech.sup.dtr/+] mice exacerbated concanavalin A-induced acute liver injury. In contrast, adoptively transferred BM-derived pDCs preferentially accumulated in the inflamed liver and protected against liver injury. This protective effect was independent of TLR7 and TLR9 signaling, since a similar effect occurred following transfer of MyD88-deficient pDCs. Alternatively, we found an unexpected immunosuppressive role of pDCs in an IL-35-dependent manner. Both Il12a and Ebi3, heterodimeric components of IL-35, were highly expressed in transferred pDCs and [CD4.sup.+][CD25.sup.+] Tregs. However, the protective effect of pDC transfer was completely lost in mice depleted of Tregs by anti-CD25 antibody. Moreover, pDCs derived from IL-35-deficient mice had less of a protective effect both in vivo and in vitro even in the presence of Tregs. These results highlight a unique aspect of pDCs in association with Tregs, serving as a guide for immunotherapeutic options in ALF., Introduction Acute liver failure (ALF) is a life-threatening condition characterized by progressive and extensive multilobular damage of the hepatocytes and massive intrahepatic infiltration of immune cells. However, there are no [...]

HIV-1 acquisition occurs most commonly after sexual contact. To establish infection, HIV-1 must infect cells that support high-level replication, namely [CD4.sup.+] T cells, which are absent from the outermost genital epithelium. Dendritic cells (DCs), present in mucosal epithelia, potentially facilitate HIV-1 acquisition. We show that vaginal epithelial DCs, termed [CD1a.sup.+] VEDCs, are unlike other blood- and tissue-derived DCs because they express langerin but not DC-SIGN, and unlike skin-based [langerin.sup.+] DC subset Langerhans cells (LCs), they do not harbor Birbeck granules. Individuals primarily acquire HIV-1 that utilizes the CCR5 receptor (termed either R5 or R5X4) during heterosexual transmission, and the mechanism for the block against variants that only use the CXCR4 receptor (classified as X4) remains unclear. We show that X4 as compared with R5 HIV-1 shows limited to no replication in [CD1a.sup.+] VEDCs. This differential replication occurs after fusion, suggesting that receptor usage influences postentry steps in the virus life cycle. Furthermore, [CD1a.sup.+] VEDCs isolated from HIV-1-infected virologically suppressed women harbor HIV-1 DNA. Thus, [CD1a.sup.+] VEDCs are potentially infected early during heterosexual transmission and also retain virus during treatment. Understanding the interplay between HIV-1 and [CD1a.sup.+] VEDCs is important for future prevention and cure strategies., Introduction The majority of new HIV-1 infections occur in women after heterosexual contact. To establish a systemic infection in a naive woman, HIV-1 must cross the genital epithelium and infect [...]

Background Anastomotic failure causes morbidity and mortality even in technically correct anastomoses. Initial leaks must be prevented by mucosal reapproximation across the anastomosis. Healing is a concerted effort between intestinal epithelial cells (IECs), immune cells, and commensal bacteria. IEC TLR4 activation and signaling is required for mucosal healing, leading to inflammatory factor release that recruits immune cells to limit bacteria invasion. TLR4 absence leads to mucosal damage from loss in epithelial proliferation, attenuated inflammatory response, and bacteria translocation. We hypothesize after anastomosis, an imbalance in microbiota will occur due to a decrease in TLR4 expression and will lead to changes in the immune milieu. Results We isolated fecal content and small intestinal leukocytes from murine, Roux-en-Y and end-to-end anastomoses, to identify microbiome changes and subsequent alterations in the regulatory and pro-inflammatory immune cells 3 days post-operative. TLR4.sup.+ IECs were impaired after anastomosis. Microbiome diversity was reduced, with Firmicutes, Bacteroidetes, and Saccharibacteria decreased and Proteobacteria increased. A distinct TCR[beta].sup.hi CD4.sup.+ T cells subset after anastomosis was 10-20-fold greater than in control mice. 84% were Th17 IL-17A/F.sup.+ IL-22.sup.+ and/or TNF[alpha].sup.+. iNKT cells were increased and TCR[beta].sup.hi. 75% were iNKT IL-10.sup.+ and 13% iNKTh17 IL-22.sup.+. Additionally, Treg IL-10.sup.+ and IL-22.sup.+ cells were increased. A novel dendritic cell subset was identified in anastomotic regions that was CD11b.sup.hi CD103.sup.mid and was 93% IL-10.sup.+. Conclusions This anastomotic study demonstrated a decrease in IEC TLR4 expression and microbiome diversity which then coincided with increased expansion of regulatory and pro-inflammatory immune cells and cytokines. Defining the anastomotic mucosal environment could help inform innovative therapeutics to target excessive pro-inflammatory invasion and microbiome imbalance. Keywords: Immune-microbiota crosstalk, Mucosal healing, Toll-like receptors, CD11b.sup.hi CD103.sup.mid, Author(s): Emilie E. Vomhof-DeKrey[sup.1,2], Allie Stover[sup.2] and Marc D. Basson[sup.1,2,3] Background Anastomotic failure is one of the most serious complications of intestinal surgery [24]. An anastomotic leak can lead to [...]

Sepsis survivors exhibit impaired responsiveness to antigen (Ag) challenge associated with increased mortality from infection. The contribution of follicular dendritic cells (FDCs) in the impaired humoral response in sepsis-surviving mice is investigated in this study. We demonstrated that mice subjected to sepsis from cecal ligation and puncture (CLP mice) have reduced NP- specific high-affinity class-switched Ig antibodies (Abs) compared with sham-operated control mice following immunization with the T cell-dependent Ag, NP-CGG. NP-specific germinal center (GC) B cells in CLP mice exhibited reduced TNF-[alpha] and AID mRNA expression compared with sham-operated mice. CLP mice showed a reduction in FDC clusters, a reduced binding of immune complexes on FDCs, and reduced mRNA expression of CR2, ICAM-1, VCAM-1, Fc[gamma]RIIB, TNFR1, IKK2, and LT[beta]R compared with sham-operated mice. Adoptive transfer studies showed that there was no B cell- intrinsic defect. In summary, our data suggest that the reduced Ag-specific Ab response in CLP mice is secondary to a disruption in FDC and GC B cell function., Introduction Sepsis is defined as life-threatening organ dysfunction resulting from a dysregulated host response to infection. It is a major cause of death in the United States, accounting for billions [...]

Type I IFN production is essential for innate control of acute viral infection; however, prolonged high-level IFN production is associated with chronic immune activation in HIV-infected individuals. Although plasmacytoid DCs (pDCs) are a primary source of IFN, the mechanisms that regulate IFN levels following the acute phase are unknown. We hypothesized that HIV-specific Ab responses regulate late IFN production. We evaluated the mechanism through which HIV-activated pDCs produce IFN as well as how both monoclonal HIV-specific Abs and Abs produced in natural HIV infection modulated normal pDC sensing of HIV. We found that HIV-induced IFN production required TLR7 signaling, receptor-mediated entry, fusion, and viral uncoating, but not endocytosis or HIV life cycle stages after uncoating. Abs directed against the HIV envelope that do not interfere with CD4 binding markedly enhanced the IFN response, irrespective of their ability to neutralize [CD4.sup.+] T cell infection. Ab-mediated enhancement of IFN production required Fc [gamma] receptor engagement, bypassed fusion, and initiated signaling through both TLR7 and TLR9, which was not utilized in the absence of Ab. Polyclonal Abs isolated from HIV-infected subjects also enhanced pDC production of IFN in response to HIV. Our data provide an explanation for high levels of IFN production and immune activation in chronic HIV infection., Introduction Type I IFN production is essential in innate control of acute viral infection, but can enhance pathogenesis during chronic infections such as HIV, SIV, and lymphocytic choriomeningitis virus infection [...]

Human TRIM5[alpha] restricts HIV-1 infection of Langerhans cells through Langerin-dependent autophagy pathway. C-type lectin receptor-mediated anti-HIV activity Teunis Geijtenbeek and colleagues find a role for human E3-ubiquitin ligase tri-partite-containing motif 5[alpha] (TRIM5[alpha]) in restricting HIV-1 infection in Langerhans cells, a subset of dendritic cells present at the mucosal barrier. They show that capture of HIV-1 by the C-type lectin receptor Langerin serves to route the virus to a TRIM5[alpha]- and Langerin-dependent autophagy pathway. This mechanism of TRIM5[alpha]-mediated restriction differs from the proteasome-dependent mechanism by which rhesus TRIM5[alpha] is thought to restrict HIV-1, and seems to be a Langerhans-cell-specific restriction mechanism operating at the mucosal barrier. Langerhans cells are important in the defence against HIV-1 infection during sexual transmission, and this work highlights the potential of interventions involving C-type lectin receptors and autophagy-targeting strategies to promote cell-mediated resistance to HIV-1. The most prevalent route of HIV-1 infection is across mucosal tissues after sexual contact. Langerhans cells (LCs) belong to the subset of dendritic cells (DCs) that line the mucosal epithelia of vagina and foreskin and have the ability to sense and induce immunity to invading pathogens.sup.1. Anatomical and functional characteristics make LCs one of the primary targets of HIV-1 infection.sup.2. Notably, LCs form a protective barrier against HIV-1 infection and transmission.sup.3,4,5. LCs restrict HIV-1 infection through the capture of HIV-1 by the C-type lectin receptor Langerin and subsequent internalization into Birbeck granules.sup.5. However, the underlying molecular mechanism of HIV-1 restriction in LCs remains unknown. Here we show that human E3-ubiquitin ligase tri-partite-containing motif 5[alpha] (TRIM5[alpha]) potently restricts HIV-1 infection of LCs but not of subepithelial DC-SIGN.sup.+ DCs. HIV-1 restriction by TRIM5[alpha] was thus far considered to be reserved to non-human primate TRIM5[alpha] orthologues.sup.6,7,8,9, but our data strongly suggest that human TRIM5[alpha] is a cell-specific restriction factor dependent on C-type lectin receptor function. Our findings highlight the importance of HIV-1 binding to Langerin for the routeing of HIV-1 into the human TRIM5[alpha]-mediated restriction pathway. TRIM5[alpha] mediates the assembly of an autophagy-activating scaffold to Langerin, which targets HIV-1 for autophagic degradation and prevents infection of LCs. By contrast, HIV-1 binding to DC-SIGN.sup.+ DCs leads to disassociation of TRIM5[alpha] from DC-SIGN, which abrogates TRIM5[alpha] restriction. Thus, our data strongly suggest that restriction by human TRIM5[alpha] is controlled by C-type-lectin-receptor-dependent uptake of HIV-1, dictating protection or infection of human DC subsets. Therapeutic interventions that incorporate C-type lectin receptors and autophagy-targeting strategies could thus provide cell-mediated resistance to HIV-1 in humans., Author(s): Carla M. S. Ribeiro [sup.1] , Ramin Sarrami-Forooshani [sup.1] , Laurentia C. Setiawan [sup.1] , Esther M. Zijlstra-Willems [sup.1] , John L. van Hamme [sup.1] , Wikky Tigchelaar [sup.2] [...]