Although human endogenous retroviruses (HERVs) represent a substantial proportion of the human genome and some HERVs, such as HERV-K(HML-2), are reported to be involved in neurological disorders, little is known about their biological function. We report that RNA from an HERV-K(HML-2) envelope gene region binds to and activates human Toll-like receptor (TLR) 8, as well as murine Tlr7, expressed in neurons and microglia, thereby causing neurodegeneration. HERV-K(HML-2) RNA introduced into the cerebrospinal fluid (CSF) of either C57BL/6 wild-type mice or APPPS1 mice, a mouse model for Alzheimer’s disease (AD), resulted in neurodegeneration and microglia accumulation. Tlr7-deficient mice were protected against neurodegenerative effects but were resensitized toward HERV-K(HML-2) RNA when neurons ectopically expressed murine Tlr7 or human TLR8. Transcriptome data sets of human AD brain samples revealed a distinct correlation of upregulated HERV-K(HML-2) and TLR8 RNA expression. HERV-K(HML-2) RNA was detectable more frequently in CSF from individuals with AD compared with controls. Our data establish HERV-K(HML-2) RNA as an endogenous ligand for species-specific TLRs 7/8 and imply a functional contribution of human endogenous retroviral transcripts to neurodegenerative processes, such as AD.
Paul Dembny, Andrew G. Newman, Manvendra Singh, Michael Hinz, Michal Szczepek, Christina Krüger, Robert Adalbert, Omar Dzaye, Thorsten Trimbuch, Thomas Wallach, Gunnar Kleinau, Katja Derkow, Bernhard C. Richard, Carola Schipke, Claus Scheidereit, Harald Stachelscheid, Douglas Golenbock, Oliver Peters, Michael Coleman, Frank L. Heppner, Patrick Scheerer, Victor Tarabykin, Klemens Ruprecht, Zsuzsanna Izsvák, Jens Mayer, Seija Lehnardt
Vascularized composite allotransplantation (VCA) has become a valid therapeutic option to restore form and function after devastating tissue loss. However, the need for high-dose multidrug immunosuppression to maintain allograft survival is still hampering more widespread application of VCA. In this study, we investigated the immunoregulatory potential of costimulation blockade (CoB; CTLA4-Ig and anti-CD154 mAb) combined with nonmyeoablative total body irradiation (TBI) to promote allograft survival of VCA in a fully MHC-mismatched mouse model of orthotopic hind limb transplantation. Compared with untreated controls (median survival time [MST] 8 days) and CTLA4-Ig treatment alone (MST 17 days), CoB treatment increased graft survival (MST 82 days), and the addition of nonmyeloablative TBI led to indefinite graft survival (MST > 210 days). Our analysis suggests that VCA-derived BM induced mixed chimerism in animals treated with CoB and TBI + CoB, promoting gradual deletion of alloreactive T cells as the underlying mechanism of long-term allograft survival. Acceptance of donor-matched secondary skin grafts, decreased ex vivo T cell responsiveness, and increased graft-infiltrating Tregs further indicated donor-specific tolerance induced by TBI + CoB. In summary, our data suggest that vascularized BM-containing VCAs are immunologically favorable grafts promoting chimerism induction and long-term allograft survival in the context of CoB.
Byoung Chol Oh, Georg J. Furtmüller, Madeline L. Fryer, Yinan Guo, Franka Messner, Johanna Krapf, Stefan Schneeberger, Damon S. Cooney, W.P. Andrew Lee, Giorgio Raimondi, Gerald Brandacher
The lack of sufficient functional tumor-infiltrating lymphocytes in the tumor microenvironment (TME) is one of the primary indications for the poor prognosis of patients with cancer. In this study, we developed an Erbitux-based IL-21 tumor-targeting fusion protein (Erb-IL21) to prolong the half-life and improve the antitumor efficacy of IL-21. Compared with Erb-IL2, Erb-IL21 demonstrated much lower toxicity in vivo. Mechanistically, Erb-IL21 selectively expanded functional cytotoxic T lymphocytes but not dysfunctional CD8+ T cells in the TME. We observed that the IL-21–mediated antitumor effect largely depended on the existing intratumoral CD8+ T cells, instead of newly migrated CD8+ T cells. Furthermore, Erb-IL21 overcame checkpoint blockade resistance in mice with advanced tumors. Our study reveals that Erb-IL21 can target IL-21 to tumors and maximize the antitumor potential of checkpoint blockade by expending a subset of tumor antigen–specific CD8+ T cells to achieve effective tumor control.
Sisi Deng, Zhichen Sun, Jian Qiao, Yong Liang, Longchao Liu, Chunbo Dong, Aijun Shen, Yang Wang, Hong Tang, Yang-Xin Fu, Hua Peng
Immune suppressive donor regulatory T cells (Tregs) can prevent graft-versus-host disease (GVHD) or solid organ allograft rejection. We previously demonstrated inhibiting STAT3 phosphorylation (pSTAT3) augments FOXP3 expression, stabilizing induced Tregs (iTregs). Here we report human pSTAT3-inhibited iTregs prevent human skin graft rejection and xenogeneic GVHD yet spare donor anti-leukemia immunity. pSTAT3-inhibited iTregs express increased levels of skin-homing CLA antigen, immune suppressive GARP and PD-1, and IL-9 that supports tolerizing mast cells. Further, pSTAT3-inhibited iTregs significantly reduce alloreactive conventional T cells, Th1, and Th17 cells implicated in GVHD and tissue rejection, and impair infiltration by pathogenic Th2 cells. Mechanistically, pSTAT3 inhibition of iTregs provokes a shift in metabolism from oxidative phosphorylation (OxPhos) to glycolysis and reduced electron transport chain activity. Strikingly, co-treatment with coenzyme Q10 (coQ10) restores OxPhos in pSTAT3-inhibited iTregs and augments their suppressive potency. These findings support the rationale for clinically testing the safety and efficacy of metabolically tuned, human pSTAT3-inhibited iTregs to control alloreactive T cells.
Kelly Walton, Mario R. Fernandez, Elizabeth M. Sagatys, Jordan Reff, Jongphil Kim, Marie Catherine Lee, John Kiluk, Jane Yuet Ching Hui, David McKenna, Meghan Hupp, Colleen Forster, Michael A. Linden, Nicholas J. Lawrence, Harshani R. Lawrence, Joseph Pidala, Steven Z. Pavletic, Bruce R. Blazar, Said M. Sebti, John L. Cleveland, Claudio Anasetti, Brian C. Betts
Chronic beryllium disease (CBD) is a metal hypersensitivity/autoimmune disease in which damage-associated molecular patterns (DAMPs) promote a break in T cell tolerance and expansion of Be2+/self-peptide reactive CD4+ T cells. In this study, we investigated the mechanism of cell death induced by beryllium particles (Be) in alveolar macrophages (AMΦs) and its impact on DAMP release. We found that phagocytosis of Be led to AM cell death independently of caspase, RIP1K, RIP3K or ROS activity. Prior to cell death, Be-exposed AMΦs secreted TNFalpha that boosted intracellular stores of IL-1alpha followed by caspase 8-dependent fragmentation of DNA. IL-1alpha and nucleosomal DNA were subsequently released from AMΦs upon loss of plasma membrane integrity. In contrast, necrotic AMs released only unfragmented DNA and necroptotic AMΦs released only IL-1alpha. In mice exposed to Be, TNFalpha promoted release of both DAMPs and was required for the mobilization of immunogenic DCs, expansion of Be-reactive CD4+ T cells and pulmonary inflammation in a mouse model of CBD. Thus, early autocrine effects of particle-induced TNFalpha on AMs led to a break in peripheral tolerance. This novel mechanism may underlie the known relationship between fine particle inhalation, TNFalpha and loss of peripheral tolerance in T cell-mediated autoimmune disease and hypersensitivities.
Morgan K. Collins, Abigail M. Shotland, Morgan F. Wade, Shaikh M. Atif, Denay J.K. Richards, Manolo Torres-Llompart, Douglas G. Mack, Allison K. Martin, Andrew P. Fontenot, Amy S. McKee
While blockade of PD-1/PD-L1 immune checkpoint revolutionized cancer treatment, how it works on tumor-infiltrating CD8+ T cells recognizing the same antigen at various differentiation stages remains elusive. Here, we found that the chemokine receptor CX3CR1 identified three distinct differentiation states of intratumoral CD8+ T-cell subsets. Adoptively transferred antigen-specific CX3CR1neg CD8+ T cells generated phenotypically and functionally distinct CX3CR1int and CX3CR1hi subsets in the periphery. Notably, expression of co-inhibitory receptors and Tcf1 inversely correlated with the degree of T-cell differentiation defined by CX3CR1. Despite significantly lower expression of co-inhibitory receptors and potent cytolytic activity, in vivo depletion of the CX3CR1hi subset did not alter the antitumor efficacy of adoptively transferred CD8+ T cells. Furthermore, differentiated CX3CR1int and CX3CR1hi subsets were impaired in their ability to undergo proliferation upon re-stimulation, and had no impact on established tumors upon second adoptive transfer compared with the CX3CR1neg subset that remained effective. Accordingly, anti-PD-L1 therapy preferentially rescued proliferation and cytokine production of the CX3CR1neg subset, and significantly enhanced antitumor efficacy of adoptively transferred CD8+ T cells. These findings provide a better understanding of the phenotypic and functional heterogeneity of tumor-infiltrating CD8+ T cells, and can be exploited to develop more effective immunotherapy.
Takayoshi Yamauchi, Toshifumi Hoki, Takaaki Oba, Hidehito Saito, Kristopher Attwood, Michael S. Sabel, Alfred E. Chang, Kunle Odunsi, Fumito Ito
Background: Prehospital plasma improves survival in severely injured trauma patients at risk for hemorrhagic shock and transported by air ambulance. We hypothesized that prehospital plasma would be associated with a reduction in immune imbalance and endothelial damage. Methods: We collected blood samples from 405 trauma patients enrolled in the Prehospital Air MedicalPlasma (PAMPer) trial upon hospital admission (0 hours) and 24 hours post admission across 6 U.S. sites(9 level-one trauma centers) with air medical transport services. We assayed samples for 21 inflammatory mediators and 7 markers of endothelial damage. We performed hierarchical clustering analysis (HCA) on principal components of these biomarkers of the immune response and endothelial injury. Regression analysis was used to control for known differences across study arms near the time of randomization and to assess any association with prehospital plasma administration. Results: HCA based on inflammatory mediator and endothelial damage marker concentrations distinguished two patient clusters, each with different injury patterns and outcomes. Patients in cluster A had greater injury severity and incidence of blunt trauma, traumatic brain injury, and mortality. Cluster A patients that received prehospital plasma as compared to standard care fluid resuscitation showed improved 30-day survival. Prehospital plasma did not improve survival in cluster B patients. In an adjusted analysis of themost seriously injured patients (ISS>30), plasma was associated with a an increase in circulating levels of adiponectin, IL-1β, IL-17A, IL-23, and IL-17E upon admission. One day following admission, prehospital plasmas was associated with a reduction in syndecan-1, TM, VEGF, IL-6, IP-10, MCP-1, and TNF-α, and an increase in IL-33, IL-21, IL-23, and IL-17E. Conclusion: This is the first human study to suggest that prehospital plasma may ameliorate the endotheliopathy of trauma and modulate an imbalance between pro-inflammatory (e.g. IL-6, TNF-α, and MCP-1) and protective (e.g. IL-33 and IL-17E) mediators. These effects of early plasma administration may contribute to improved survival in severely injured patients. Trial Registration: ClinicalTrials.gov NCT01818427 Funding: National Institutes of Health T32; U.S. Army Medical Research and Materiel Command W81XWH-12-2-0023; National Institutes of Health R35; National Institutes of Health 1R35GM119526-01; the Office of the Assistant Secretary of Defense for Health Affairs, through the Defense Medical Research and Development Program W81XWH-18-2-0051 and W81XWH-15-PRORP-OCRCA. Opinions, interpretations, conclusions and recommendations are those of the authors and not necessarily endorsed by the Department of Defense.
Danielle S. Gruen, Joshua B. Brown, Francis X. Guyette, Yoram Vodovotz, Par I. Johansson, Jakob Stensballe, Derek A. Barclay, Jinling Yin, Brian J. Daley, Richard S. Miller, Brian G. Harbrecht, Jeffrey A. Claridge, Herb A. Phelan, Matthew D. Neal, Brian Zuckerbraun, Timothy R. Billiar, Jason L. Sperry
Septic cardiomyopathy is a life-threatening organ dysfunction caused by sepsis. Ribonuclease 1 (RNase 1) belongs to a group of host-defense peptides that specifically cleave extracellular RNA (eRNA). The activity of RNase1 is inhibited by ribonuclease-inhibitor 1 (RNH1). The role of RNase 1 in septic cardiomyopathy and associated cardiac apoptosis, however, is completely unknown. Here, we showed that sepsis resulted in a significant increase in RNH1 and eRNA serum levels compared to those of healthy subjects (p < 0.05). Treatment with RNase 1 resulted in a significant decrease of apoptosis, induced by the intrinsic pathway, and TNF expression in murine cardiomyocytes exposed to either necrotic cardiomyocytes or serum of septic patients for 16 h (p < 0.05). Furthermore, treatment of septic mice with RNase 1 resulted in a reduction in cardiac apoptosis, TNF expression and septic cardiomyopathy (p < 0.05). These data demonstrate that eRNA plays a crucial role in the pathophysiology of the organ (cardiac) dysfunction in sepsis and RNase and RNH1 may be new therapeutic targets/strategies to reduce the cardiac injury and dysfunction caused by sepsis.
Elisabeth Zechendorf, Caroline E O'Riordan, Lara Stiehler, Natalie Wischmeyer, Fausto Chiazza, Debora Collotta, Bernd Denecke, Sabrina Ernst, Gerhard Müller-Newen, Sina M. Coldewey, Bianka Wissuwa, Massimo Collino, Tim-Philipp Simon, Tobias Schuerholz, Christian Stoppe, Gernot Marx, Christoph Thiemermann, Lukas Martin
Acinetobacter baumannii is an extremely versatile multidrug resistant pathogen with a very high mortality rate therefore, it has become crucial to understand the host response during its infection. Given the importance of mice for modelling infection and their role in pre-clinical drug development equal emphasis should be placed on the utilization of both sexes. Through our studies using a murine model of acute pneumonia with A. baumannii, we observed that female mice were more susceptible to infection. Likewise, treatment of male mice with estradiol increased their susceptibility to infection. Analysis of the airway compartment revealed enhanced inflammation and reduced neutrophil and alveolar macrophage numbers compared to male mice. Depletion of either neutrophils or alveolar macrophages was important for bacterial clearance however, depletion of alveolar macrophages further exacerbated female susceptibility due to severe alterations in metabolic homoeostasis. Our data highlights the importance of utilizing both sexes when assessing host immune pathways.
Silvia Pires, Adeline C. Peignier, Jeremy Seto, Davida S. Smyth, Dane Parker
Acute Graft-Versus-Host Disease (aGVHD) is a T cell mediated immunological disorder and the leading cause of non-relapse mortality in patients who receive allogeneic hematopoietic cell transplants. Based on recent observations that PRMT5 and arginine methylation is upregulated in activated memory T cells, we hypothesized that PRMT5 is involved in the pathogenesis of aGVHD. Here, we show that PRMT5 expression and enzymatic activity is upregulated in activated T cells in vitro and in T cells from mice developing aGVHD after allogeneic transplant. PRMT5 expression is also upregulated in T cells of patients who developed aGVHD after allogeneic hematopoietic cell transplant compared to those who did not develop aGVHD.PRMT5 inhibition using a selective small-molecule inhibitor (C220) significantly reduces mouse and human allogeneic T cell proliferation and inflammatory IFN-γ and IL-17 cytokine production. Administration of PRMT5 small-molecule inhibitors significantly improves survival, reducing disease incidence and clinical severity in mouse models of aGVHD without adversely affecting engraftment. Importantly, we show that PRMT5 inhibition retains the beneficial graft versus leukemia (GVL) effect by maintaining cytotoxic CD8 T cell responses. Mechanistically, we show that PRMT5 inhibition potently reduces STAT-1 phosphorylation as well as transcription of pro-inflammatory genes including Interferon Stimulated Genes (ISG) and IL-17. Additionally, PRMT5 inhibition deregulates cell-cycle in activated T cells and disrupts signaling by impacting ERK1/2 phosphorylation. Thus, we have identified PRMT5 as a regulator of T cell responses and as a therapeutic target in aGVHD.
Katiri Snyder, Nina C. Zitzer, Yandi Gao, Hannah K. Choe, Natalie E. Sell, Lotus Neidemire-Colley, Anora Ignaci, Charuta Kale, Raymond D. Devine, Maria G. Abad, Maciej Pietrzak, Min Wang, Hong Lin, Yang W. Zhang, Gregory K. Behbehani, Jane E. Jackman, Ramiro Garzon, Kris Vaddi, Robert A. Baiocchi, Parvathi Ranganathan
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