Lentiviral vector-based dendritic cell vaccines induce protective T cell responses against viral infection and cancer in animal models. In this study, we tested whether preventative and therapeutic vaccination could be achieved by direct injection of antigen expressing lentiviral vector, obviating the need for ex vivo transduction of dendritic cells. Injected lentiviral vector preferentially transduced splenic dendritic cells and resulted in long-term expression. Injection of a lentiviral vector encoding an MHC class I restricted T cell epitope of LCMV and CD40L induced an antigen-specific cytolytic CD8+ T lymphocyte response that protected the mice from infection. The injection of chronically infected mice with a lentiviral vector encoding LCMV MHC class I and II T cell epitopes and a soluble PD-1 microbody rapidly cleared the virus. Vaccination by direct injection of lentiviral vector was more effective in SAMHD1 knock-out mice, suggesting that lentiviral vectors containing Vpx, a lentiviral protein that increases the efficiency of dendritic cell transduction by inducing the degradation of SAMHD1, would be an effective strategy for the treatment of chronic disease in humans.
Takuya Tada, Thomas D. Norton, Rebecca Leibowitz, Nathaniel R. Landau
B-lymphocytes have long been recognized for their critical contributions to adaptive immunity, providing defense against pathogens through cognate antigen presentation to T cells and antibody production. More recently appreciated is that B cells are also integral in securing self-tolerance; this has led to interest in their therapeutic application to down-regulate unwanted immune responses such as transplant rejection. In this study, we found that PMA and ionomycin-activated mouse B cells acquire regulatory properties following stimulation through Toll-like receptor (TLR)4/TLR9 receptors (Bregs-TLR). Bregs-TLR efficiently inhibited T cell proliferation in vitro and prevented allograft rejection. Unlike most reported Breg activities, the inhibition of alloimmune responses by Bregs-TLR relied on the expression of TGF-βand not IL-10. In vivo, Bregs-TLR interrupted donor-specific T cell expansion and induced regulatory T cells in a TGF-β dependent manner. RNA-seq analyses corroborated the involvement of TGF-β pathways in Breg-TLR function, identified potential gene pathways implicated in preventing graft rejection, and suggested new targets to foster Breg regulation.
Kang Mi Lee, Qiang Fu, Guoli Huai, Kevin Deng, Ji Lei, Lisa Kojima, Divyansh Agarwal, Peter Van Galen, Shoko Kimura, Naoki Tanimine, Laura Washburn, Heidi Yeh, Ali Naji, Charles G. Rickert, Christian LeGuern, James F. Markmann
Plasmacytoid dendritic cells (pDCs) perform dual proinflammatory and immunosuppressive roles. We recently reported the potential of pDC therapy for treatment of intractable acute liver failure. However, establishment of efficient methods to deliver pDCs to the liver is essential for future clinical therapeutic applications. The present study demonstrates a higher abundance of liver and peripheral blood pDCs in mice lacking the C-C motif chemokine receptor 9 (CCR9), a pDC gut-homing receptor, than that in wild-type (WT) mice. Adoptive pDC transfer resulted in a higher efficiency of Ccr9-/- pDC migration to the liver than that to the original target organ, the small intestine, compared with that of WT pDCs. Further, Ccr9-/- pDCs consistently migrated efficiently to the concanavalin A-induced inflamed liver, and exerted a more effective immunosuppressive effect, resulting in better protection against acute liver inflammation than that demonstrated by WT pDCs. These findings highlight the therapeutic potential of the manipulation of CCR9 axis as a novel approach to improve migration of immunosuppressive pDCs to the liver in order to exploit their beneficial effects in acute liver disease.
Yuzo Koda, Nobuhiro Nakamoto, Po-Sung Chu, Toshiaki Teratani, Akihisa Ueno, Takeru Amiya, Nobuhito Taniki, Sayako Chiba, Kentaro Miyamoto, Michiie Sakamoto, Takanori Kanai
The immune factors associated with impaired SARS-CoV-2 vaccine response in the elderly are mostly unknown. We studied >60 and <60 years old people vaccinated with SARS-CoV-2 BNT162b2 mRNA before and after the first and second dose. Aging was associated with a lower anti-RBD IgG levels and a decreased magnitude and polyfunctionality of SARS-CoV-2 specific T cell response. The dramatic decrease in thymic function in aged people with >60 years of age, which fueled alteration in T cell homeostasis, and lower CD161+ T cell levels were associated with decreased T cell response two months after vaccination. Additionally, a deficient dendritic cell (DC) homing, activation and Toll like receptor (TLR)-mediated function, along with a proinflammatory functional profile in monocytes, were observed in the >60 years old group, which was also related to lower specific T cell response after vaccination. These findings might be relevant for the improvement of the current vaccination strategies and for the development of new vaccine prototypes.
Joana Vitallé, Alberto Pérez-Gómez, Francisco José Ostos, Carmen Gasca-Capote, Maria Reyes Jiménez-Leon, Sara Bachiller, Inmaculada Rivas-Jeremías, Maria del Mar Silva-Sánchez, Anabel M. Ruiz-Mateos, María Ángeles Martín-Sánchez, Luis Fernando López-Cortes, Mohammed Rafii El Idrissi Benhnia, Ezequiel Ruiz-Mateos
Allogeneic hematopoietic stem cell transplantation (allo-HSCT) efficacy is complicated by graft-versus-host disease (GVHD), a leading cause of morbidity and mortality after transplant. Despite GVHD prophylaxis, 30-70% of patients develop GVHD resulting in susceptibility to infections, relapse and secondary malignancies. Regulatory T-cells (Tregs) have shown efficacy in preventing GVHD, but variably suppressive at high doses. To enhance in vivo suppressor function, murine Treg were transduced to express an anti-human CD19 chimeric antigen receptor (hCAR19) and infused into lethally irradiated hCD19 transgenic recipients for allo-HSCT. As compared to recipients receiving controlled transduced Tregs, those receiving hCAR19 Tregs had a significant decrease in acute GVHD lethality. GVHD amelioration was accomplished with not only maintenance but potentiation of the graft-versus tumor (GVT) response, as recipient hCD19 B-cells and murine hCD19TBL12luc lymphoma cells were both cleared by allogeneic hCAR19 Tregs. Mechanistically, hCAR19 Tregs killed syngeneic hCD19+ but not hCD19- murine TBL12luc cells in vitro in a perforin-dependent, granzyme B-independent manner. Importantly, cyclophosphamide treated hCD19 transgenic mice given hCAR19 cytotoxic T-lymphocytes without allo-HSCT experienced rapid lethality due to systemic toxicity, whereas hCAR19 Tregs avoided this severe complication. In conclusion, CAR19 Tregs are a novel and effective strategy to suppress GVHD without loss of GVT responses.
Sara Bolivar-Wagers, Michael L. Loschi, Sujeong Jin, Govindarajan Thangavelu, Jemma H. Larson, Cameron S. McDonald-Hyman, Ethan A. Aguilar, Asim Saha, Brent H. Koehn, Mehrdad Hefazi, Mark J. Osborn, Michael C. Jensen, John E. Wagner, Christopher A. Pennell, Bruce R. Blazar.
Lysosomal acid lipase (LAL) is a key enzyme in the metabolic pathway of neutral lipids. In the blood of LAL deficient (lal-/-) mice, increased CD11c+ cells were accompanied by up-regulated PD-L1 expression. Single cell RNA sequencing of lal-/- CD11c+ cells identified two distinctive clusters with a major metabolic shift towards glucose utilization and reactive oxygen species (ROS) over-production. Pharmacologically blocking pyruvate dehydrogenase in glycolysis not only reduced CD11c+ cells and their PD-L1 expression, but also reversed their capabilities of T cell suppression and tumor growth stimulation. Colony-stimulating factor 1 receptor (CSF1R) plays an essential role in controlling lal-/- CD11c+ cell homeostasis and function and PD-L1 expression. Inhibition of LAL activity by pharmacological inhibitor increased CD11c, PD-L1 and CSF1R levels in both normal murine myeloid cells and human blood cells. Tumor-bearing mice and human non-small-cell lung cancer (NSCLC) patients also showed CD11c+ cell expansion with PD-L1 and CSF1R up-regulation and immunosuppression. There were positive correlations among CD11c, PD-L1 and CSF1R expression and negative correlations with LAL expression in lung cancer and melanoma patients using the TCGA database and patient samples. Therefore, CD11c+ cells switched their functions to immune suppression and tumor growth stimulation through CSF1R/PD-L1 upregulation and metabolic reprogramming.
Ting Zhao, Sheng Liu, Xinchun Ding, Erica M. Johnson, Nasser H. Hanna, Kanhaiya Singh, Chandan K. Sen, Jun Wan, Hong Du, Cong Yan
Pneumocystis is the most common fungal pulmonary infection in children under 5. In children with primary immunodeficiency, Pneumocystis often presents at 3-6 months that coincides with the nadir of maternal IgG and where IgM is the dominant immunoglobulin isotype. Since B cells are the dominant antigen-presenting cells for Pneumocystis, we hypothesized the presence of fungal specific IgMs in human and mice and that these IgM specificities would predict T cell antigens. We detected fungal specific IgMs in human and mouse serum and utilized immunoprecipitation to determine if any antigens were similar across donors. We then assessed T cell responses to these antigens. We found anti-Pneumocystis IgM in wild-type mice as well as Aicda-/- mice and in human cord blood. Immunoprecipitation of Pneumocystis murina with human cord blood identified shared antigens among these donors. Using class II MHC binding prediction, we designed peptides with these antigens and identified robust peptide specific lung T cell responses after P. murina infection. After mice were immunized with two of the antigens, adoptive transfer of vaccine elicited CD4+ T cells showed effector activity suggesting that these antigens contain protective Pneumocystis epitopes. These data support the notion that germline encoded IgM B-cell receptors are critical in antigen presentation and T cell priming in early Pneumocystis infection.
Kristin Noell, Guixiang Dai, Dora Pungan, Anna Ebacher, Janet E. McCombs, Samuel J. Landry, Jay K. Kolls
Regulatory CD4+Foxp3+ T cells (Treg) restrain inflammation and immunity. However, the mechanisms underlying Treg suppressor function in inflamed non-lymphoid tissues remain largely unexplored. Here, we restricted immune responses to non-lymphoid tissues and used intravital microscopy to visualize Treg suppression of rejection by effector T cells (Teff) within inflamed allogeneic islet transplants. Despite their elevated motility, Treg preferentially contact antigen-presenting cells (APCs) over Teff. Interestingly, Treg specifically target APCs that are extensively and simultaneously contacted by Teff. In turn, Treg decrease MHC-II expression on APCs and hinder Teff function. Lastly, we demonstrate that Treg suppressor function within inflamed allografts requires ecto-nucleotidase CD73 activity, which generates the anti-inflammatory adenosine. Consequently, CD73-/- Treg exhibit reduced contacts with APCs within inflamed allografts compared to wt Treg, but not in spleen. Overall, our findings demonstrate that Treg suppress immunity within inflamed grafts through CD73 activity and suggest that Treg-APC direct contacts are central to this process.
Hehua Dai, Andressa Pena, Lynne Bauer, Amanda Williams, Simon c. Watkins, Geoffrey Camirand
Accumulation of activated natural killer (NK) cells in tissues during Ebola virus infection contributes to Ebola virus disease (EVD) pathogenesis. Yet, immunization with Ebola virus-like particles (VLPs) comprising glycoprotein (GP) and matrix protein VP40 provides rapid, NK cell-mediated protection against Ebola challenge. We used Ebola VLPs as the viral surrogates to elucidate the molecular mechanism by which Ebola virus triggers heightened NK cell activity. Incubation of human peripheral blood mononuclear cells (PBMCs) with Ebola VLPs or VP40 protein led to increased expression of IFN-γ, TNF-α, granzyme B, and perforin by CD3-CD56+ NK cells, along with concomitant increase in degranulation and cytotoxic activity of these cells. Optimal activation required accessory cells like CD14+ myeloid and CD14- cells and triggered increased secretion of numerous inflammatory cytokines. VP40-induced IFN-γ and TNF-α secretion by NK cells was dependent on IL-12 and IL-18 and suppressed by IL-10. In contrast, their increased degranulation was dependent on IL-12 with little influence of IL-18 or IL-10. These results demonstrate that Ebola VP40 stimulates NK cell functions in an IL-12 and IL-18 dependent manner that involves CD14+ and CD14- accessory cells. These novel findings may help in designing improved intervention strategies required to control viral transmission during Ebola outbreaks.
Hung Le, Paul Spearman, Stephen N. Waggoner, Karnail Singh
Although published studies have demonstrated that interferon epsilon (IFNε) has a crucial role in regulating protective immunity in the mouse female reproductive tract (mFRT), expression and regulation of IFNε in the human female reproductive tract (hFRT) have not been characterised. To characterise human IFNε, we obtained hFRT samples from a well- characterized cohort of women, enabling us to comprehensively assess ex vivo IFNε expression in the hFRT at various stages of the menstrual cycle. We found that among the various types of IFNs, IFNε is uniquely selectively and constitutively expressed in the hFRT epithelium. It has distinct expression patterns in the surface and glandular epithelia of the upper hFRT compared with basal layers of the stratified squamous epithelia of the lower hFRT. There is cyclical variation of IFNε expression in the endometrial epithelium of the upper hFRT and not in the distal FRT, consistent with selective endometrial expression of the progesterone receptor and regulation of the IFNE promoter by progesterone. Since we show IFNε stimulates important protective IFN-regulated genes (IRGs) in FRT epithelium, this characterisation is a key element in understanding the mechanisms of hormonal control of mucosal immunity.
Nollaig M. Bourke, Sharon L. Achilles, Stephanie U-Shane Huang, Helen E. Cumming, San S. Lim, Irene Papageorgiou, Linden J. Gearing, Ross Chapman, Suruchi Thakore, Niamh E. Mangan, Sam Mesiano, Paul J. Hertzog
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