Immunology
Abstract
ADAR1 edits double-stranded RNAs (dsRNAs) by deaminating adenosines into inosines, preventing aberrant activation of innate immunity by endogenous dsRNAs, which may resemble viral structures. Several tumors exploit ADAR1 to evade immune surveillance; indeed, its deletion reduces tumor viability and reshapes infiltrating leukocytes. Here we investigated the role of ADAR1 in immune evasion mechanisms during cervical cancer (CC) progression. Patients’ biopsy samples showed higher ADAR1 expression already in premalignant lesions (squamous intraepithelial lesions [SIL]) and a substantially reduced percentage of infiltrating CD7+ innate cells in in situ and invasive carcinomas compared with normal mucosa, with CD56+ NK cells showing phenotypic alterations that may have affected their functional responses. In CC-derived cell lines (SiHa, CaSki), ADAR1 silencing reduced cell proliferation, an effect further enhanced by exogenous IFN-β administration. It also induced proinflammatory gene expression, as demonstrated by RNA-Seq analysis, and conditioned supernatants collected from these cells activated several NK cell effector functions. NK cell infiltration and activation were also confirmed in organotypic 3D tissue models of SiHa cells knocked out for ADAR1. In conclusion, ADAR1 expression increased with CC progression and was accompanied by alterations in tumor-infiltrating NK cells, but its silencing in CC-derived cell lines potentiated antitumor NK cell activities. Thus, ADAR1 inhibition may represent a therapeutic perspective for CC and possibly other malignancies.
Authors
Valentina Tassinari, Marta Kaciulis, Stefano Petrai, Helena Stabile, Angelina Pernazza, Martina Leopizzi, Valeria Di Maio, Francesca Belleudi, Danilo Ranieri, Vanessa Mancini, Innocenza Palaia, Federica Tanzi, Ludovica Lospinoso Severini, Silvia Ruggeri, Maria Emanuela Greco, Giovanni Bernardini, Alessandra Zingoni, Marco Cippitelli, Cristina Cerboni, Alessandra Soriani
Abstract
Regulatory T cells (Tregs) are known to play critical roles in tissue repair via provision of growth factors, such as amphiregulin (Areg). Areg-producing Tregs have previously been difficult to study because of an inability to isolate live Areg-producing cells. In this report, we created a reporter mouse to detect Areg expression in live cells (AregThy1.1). We employed influenza A and bleomycin models of lung damage to sort Areg-producing and non-Areg-producing Tregs for transcriptomic analyses. Single-cell RNA-Seq revealed distinct subpopulations of Tregs and allowed transcriptomic comparisons of damage-induced populations. Single-cell TCR sequencing showed that Treg clonal expansion was biased toward Areg-producing Tregs and largely occurred within damage-induced subgroups. Gene module analysis revealed functional divergence of Tregs into immunosuppression-oriented and tissue repair–oriented groups, leading to identification of candidate receptors for induction of repair activity in Tregs. We tested these using an ex vivo assay for Treg-mediated tissue repair, identifying 4-1BB agonism as a mechanism for reparative activity induction. Overall, we demonstrate that the AregThy1.1 mouse is a promising tool for investigating tissue repair activity in leukocytes.
Authors
Lucas F. Loffredo, Katherine A. Kaiser, Adam Kornberg, Samhita Rao, Kenia de los Santos-Alexis, Arnold Han, Nicholas Arpaia
Abstract
Glioblastoma (GBM) is a lethal brain tumor containing a subpopulation of GBM stem cells (GSCs) that interaction with surrounding cells, including infiltrating tumor-associated macrophages and microglia (TAMs). While GSCs and TAMs are in close proximity and likely interact to coordinate tumor growth, a limited number of mechanisms have been identified that support their communication. Here, we identified glycoprotein NMB (GPNMB) as a key factor mediating a unique bidirectional interaction between GSCs and TAMs in GBM. Specifically, GSCs educated macrophages and microglia to preferentially express GPNMB in the GBM tumor microenvironment. As a result, TAM-secreted GPNMB interacted with its receptor CD44 on GSCs to promote their glycolytic and self-renewal abilities via activating the PYK2/RSK2 signaling axis. Disrupting GPNMB-mediated GSC-TAM interplay suppressed tumor progression and self-renewal in GBM mouse models. Our study found a protumor function of GPNMB-mediated GSC-TAM bidirectional communication and supports GPNMB as a promising therapeutic target for GBM.
Authors
Yang Liu, Lizhi Pang, Fatima Khan, Junyan Wu, Fei Zhou, Craig Horbinski, Shideng Bao, Jennifer S. Yu, Justin D. Lathia, Peiwen Chen
Abstract
The tumor microenvironment (TME) is highly heterogeneous and can dictate the success of therapeutic interventions. Identifying TMEs that are susceptible to specific therapeutic interventions paves the way for more personalized and effective treatments. In this study, using a spontaneous murine model of breast cancer, we characterize a TME that is responsive to inhibitors of the heme degradation pathway mediated by heme oxygenase (HO), resulting in CD8+ T- and NK-cell-dependent tumor control. A hallmark of this TME is a chronic type-I interferon (IFN) signal that is directly involved in orchestrating the anti-tumor immune response. Importantly, we identify that similar TMEs exist in human breast cancer which are associated with patient prognosis. Leveraging these observations, we demonstrate that combining a STING agonist, which induces type-I IFN responses, with an HO inhibitor produces a synergistic effect leading to superior tumor control. This study highlights HO activity as a potential resistance mechanism for type-I IFN responses in cancer offering a novel avenue for overcoming immune evasion in cancer therapy.
Authors
Dominika Sosnowska, Tik Shing Cheung, Jit Sarkar, James W. Opzoomer, Karen T. Feehan, Joanne E. Anstee, Chloé Amelia Woodman, Mohamed Reda Keddar, Kalum Clayton, Samira Ali, William Macmorland, Dorothy D. Yang, James Rosekilly, Cheryl E. Gillett, Francesca D. Ciccarelli, Richard Buus, James Spicer, Anita Grigoriadis, James N. Arnold
Abstract
Idiopathic pulmonary fibrosis (IPF) is a severe diffuse progressive fibrosing interstitial disease leading to respiratory failure and death in the absence of organ transplantation. Substantial evidence has confirmed the pivotal role of fibroblasts in the progression of IPF, yet effective therapeutic options are scarce. Single-cell transcriptomics profiling revealed that among the diverse fibroblast subsets, FAP1+ alveolar fibroblasts (AFs) are pivotal for the progression of IPF. On the basis of these findings, we developed FAP1-targeting chimeric antigen receptor cytotoxic effector regulatory T (CAR-cTregs) cells, which leverage the targeted killing advantage of the currently trending CAR-based immunotherapy for tumors and incorporate the immunosuppressive functions of Tregs to mitigate the inflammation caused by both the disease itself and CAR-T-cell infusion. Accordingly, CAR-cTregs were constructed to effectively eliminate FAP1+ fibroblasts in vitro. This cytotoxic effect can be abrogated by inhibitors of the granzyme-perforin pathway. In the bleomycin-induced PF model, CAR-cTregs were found to reverse fibrosis characterized by diminished recruitment of fibrocytes and improved remodeling of epithelial cells. Together, our results demonstrate that CAR-cTregs can serve as a promising therapeutic option for IPF and provide a novel strategy for treating multiple chronic inflammatory diseases by inducing both cytotoxicity and immunosuppression.
Authors
Yun-Han Jiang, Meng Zhou, Meng-Di Cheng, Sai Chen, Ying-Qiang Guo
Abstract
Hematopoietic Protein-1 (Hem1) is a component of the WASP-family verprolin-homologous protein (WAVE) actin regulatory complex, which is activated downstream of multiple immune receptors. Mutations in the NCKAP1L gene encoding HEM1 have recently been found to result in severe Primary Immunodeficiency Disease (PID), characterized by recurrent respiratory infections, hyperinflammation, autoimmunity, and high mortality. However, how loss of Hem1 results in PID is unclear. To define the importance of Hem1 specifically in T cells, we generated constitutive and T cell specific Hem1 null mice. Hem1 deficient T cells exhibited an increased shift from naïve to memory T cells, and increased ratio of immunosuppressive regulatory to effector T cells. Loss of Hem1 resulted in hallmarks of T cell exhaustion including T cell lymphopenia, decreased activation and proliferation, increased expression of PD-1 and Tim3, and increased IL-10 production. In vitro TCR stimulation of CD4 T cells resulted in increased production of Th1 (IFN), Th2 (IL-5, IL-13), Th17 (IL-17, IL-22), and Treg (IL-10) cytokines. This correlated with reduced F-actin, increased expression of CD107a, and increased granzyme release indicative of increased granule membrane fusion and exocytosis. These results suggest that Hem-1 is critical for maintaining T cell activation, homeostasis and regulated cytokine production following antigen encounter.
Authors
Alexandra Christodoulou, Nutthakarn Suwankitwat, Jacob T. Tietsort, Ryan Z. Culbert, Julia Y. Tsai, Fatima A. Tarbal, Chengsong Zhu, Brian M. Iritani
Abstract
MICB is a ligand for NKG2D. We have shown NK cells are central to lung transplant acute lung injury (ALI) via NKG2D activation, and increased MICB in bronchoalveolar lavage predicts ALI severity. Separately, we found a MICB polymorphism (MICBG406A) is associated with decreased ALI risk. We hypothesized this polymorphism would protect against severe SARS-CoV-2 respiratory disease. We analyzed 1,036 patients hospitalized with SARS-CoV-2 infection from the IMPACC cohort. Associations between MICBG406A and outcomes were determined by linear regression or Cox Proportional Hazards models. We also measured immune profiles of peripheral blood, upper and lower airway. We identified 560 major allele homozygous patients, and 426 and 50 with one or two copies of the variant allele. MICBG406A conferred reduced odds of severe COVID-19 (OR = 0.73, CI = 0.58–0.93, P = 0.04). MICBG406A homozygous participants demonstrated 34% reduced cumulative odds for mechanical ventilation or death (CI = 0.51–0.85, P = 0.005) and 43% reduced risk for mortality (CI = 0.35–0.77, P = 0.001). Patients with MICBG406A variant alleles had reduced soluble inflammatory mediators and differential regulation of multiple immune pathways. These findings demonstrate a novel association between increasing MICBG406A variant allele copies and reduced COVID-19 severity, independent of SARS-CoV-2 viral burden and humoral immunity, suggesting the NKG2D-ligand pathway as an intervention target.
Authors
Harry Pickering, Narges Alipanah-Lechner, Ernie Chen, Dylan Duchen, Holden T. Maecker, Seunghee Kim-Schulze, Ruth R. Montgomery, Chris Cotsapas, Hanno Steen, Florian Krammer, Charles R. Langelier, Ofer Levy, Lindsey R. Baden, Esther Melamed, Lauren I.R. Ehrlich, Grace A. McComsey, Rafick P. Sekaly, Charles B. Cairns, Elias K. Haddad, Albert C. Shaw, David A. Hafler, David B. Corry, Farrah Kheradmand, Mark A. Atkinson, Scott C. Brakenridge, Nelson I. Agudelo Higuita, Jordan P. Metcalf, Catherine L. Hough, William B. Messer, Bali Pulendran, Kari C. Nadeau, Mark M. Davis, Ana Fernandez-Sesma, Viviana Simon, Monica Kraft, Christian Bime, David J. Erle, Joanna Schaenman, Al Ozonoff, Bjoern Peters, Steven H. Kleinstein, Alison D. Augustine, Joann Diray-Arce, Patrice M. Becker, Nadine Rouphael, Matthew C. Altman, Steven E. Bosinger, Walter L. Eckalbar, IMPACC Network, Carolyn S. Calfee, Oscar A. Aguilar, Elaine F. Reed, John R. Greenland, Daniel R. Calabrese
Abstract
Pulmonary fibrosis (PF) is a life-threatening disease that requires effective and well-tolerated therapeutic modalities. Previously, the distinct pathogenic roles of cannabinoid receptor 1 (CB1R) and inducible nitric oxide synthase (iNOS) in the lungs and their joint therapeutic targeting were highlighted in PF. However, the cell-specific role of CB1R in PF has not been explored. Here, we demonstrate that CB1R in alveolar macrophages (AMs) mediates the release of anandamide into the alveoli, which promotes PF by inducing profibrotic macrophages that are accessible to locally delivered antifibrotic therapy. A multitargeted therapy may improve therapeutic efficacy in PF. Pulmonary delivery of 0.5 mg/kg/day MRI-1867 (zevaquenabant), a peripherally acting hybrid CB1R/iNOS inhibitor, is as effective as systemic delivery of 10 mg/kg/day, and also matches the efficacy of nintedanib in mitigating bleomycin-induced PF. A systems pharmacology approach reveals that zevaquenabant and nintedanib treatments reverse pathologic changes in both distinct and shared PF-related pathways, which are conserved in human and mouse. Moreover, zevaquenabant treatment also attenuated fibrosis and profibrotic mediators in human precision-cut lung slices. These findings establish CB1R-expressing AMs as a therapeutic target and support local delivery of dual CB1R/iNOS inhibitor zevaquenabant by inhalation as an effective, well-tolerated, and safer strategy for PF.
Authors
Abhishek Basu, Muhammad Arif, Kaelin M. Wolf, Madeline Behee, Natalie L. Johnson, Lenny Pommerolle, Ricardo H. Pineda, John Sembrat, Charles N. Zawatsky, Szabolcs Dvorácskó, Nathan J. Coffey, Joshua K. Park, Seray B. Karagoz, Grzegorz Godlewski, Tony Jourdan, Judith Harvey-White, Melanie Königshoff, Malliga R. Iyer, Resat Cinar
Abstract
The loss of integrity of the blood retina barrier (BRB) is a key pathological hallmark of vision-threatening complications in diabetic retinopathy (DR). Although DR is considered a microvascular disease, mounting evidence from mouse models and patients show that inflammation is closely connected with microvasculopathy. Inflammatory responses during retinal pathophysiology are often orchestrated by microglia, resident innate immune cells of the retina. However, the precise role of microglia activity during DR pathogenesis remains elusive. Here, we used an anti PDGFRβ antibody and inducible endothelial cell-specific PDGFB-KO during postnatal development of retinal vasculature to reproduce key features of DR pathology in mice. In addition, we applied a minocycline therapy to modulate retinal inflammation. Postnatal depletion of pericytes or loss of PDGFB in retinal vessels altered BRB integrity, triggered secretion of angiogenic and inflammatory factors with concomitant microglia reactivity, which was sustained in mature retinas. Microglia reactivity was accompanied by upregulation of disease-associated genes. Notably, minocycline attenuated the cycle of inflammatory responses in young and mature retinas, thereby preserving retinal vascular and structural integrity in mice. Together, our findings suggest that immunomodulation of microglia-driven inflammatory responses preserves retinal vasculature and maintains BRB integrity in two different mouse models of human DR.
Authors
Urbanus Muthai Kinuthia, Christoph Möhle, Ralf H. Adams, Thomas Langmann
Abstract
BACKGROUND. Thymic involution with age leads to reduced T cell output and impaired adaptive immunity. However, the extent to which thymic activity persists later in life and how this contributes to immunological ageing remains unclear. This study aimed to assess the presence and function of thymic tissue in older adults and identify factors influencing residual thymopoiesis. METHODS. Patients aged ≥ 50 undergoing cardiothoracic surgery were recruited. Thymic structures within mediastinal adipose tissue were evaluated using histology, immunofluorescence, flow cytometry, TCR sequencing, and RNA sequencing. Recent thymic emigrants (RTEs) were quantified in peripheral blood and correlated with transcriptomic, epigenetic, and TCR repertoire data. Primary outcomes included thymic tissue identification, RTE frequency, and immune correlates. RESULTS. Functional thymic tissue was identified in mediastinal adipose tissue of older individuals. The frequency of CD31+CD4+ T cells (RTEs) positively correlated with the presence of thymic tissue. Thymic output showed substantial heterogeneity and was influenced by sex and smoking history. Thymic activity was associated with increased TCR repertoire diversity, improved immune protection to infections, and reduced epigenetic ageing. Detailed profiling uncovered functional and phenotypic heterogeneity within naïve CD4+ T cell subsets shaped by thymic activity. CONCLUSION. This study demonstrates that thymic function can persist into later life and is modulated by factors such as sex and smoking. These findings suggest that thymic activity during ageing is heterogenous and influenced by more than chronological age alone, with potential implications for immune competence in older adults.
Authors
Balraj Sandhar, Vishal Vyas, Daniel Harding, Roberta Ragazzini, Paola Bonfanti, Federica M. Marelli-Berg, Christopher G. Bell, Benny M. Chain, M. Paula Longhi
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