InVivoMAb anti-mouse CD28
Product Details
The PV-1 monoclonal antibody reacts with mouse CD28, a 45 kDa costimulatory receptor and a member of the Ig superfamily. CD28 is expressed by thymocytes, most peripheral T cells, and NK cells. CD28 is a receptor for CD80 (B7-1) and CD86 (B7-2). Signaling through CD28 augments IL-2 and IL-2 receptor expression as well as cytotoxicity of CD3-activated T cells. The PV-1 antibody has been shown to stimulate the proliferation and cytokine production by activated T and NK cells.Specifications
| Isotype | Armenian Hamster IgG,Ā Īŗ |
|---|---|
| Recommended Isotype Control(s) | InVivoMAb polyclonal Armenian hamster IgG |
| Recommended Dilution Buffer | InVivoPure pH 7.0 Dilution Buffer |
| Conjugation | This product is unconjugated. Conjugation is available via our Antibody Conjugation Services. |
| Immunogen | C57BL/6 mouse T cell lymphoma EL-4 cells |
| Reported Applications | in vitro T cell stimulation/activation |
| Formulation |
PBS, pH 7.0 Contains no stabilizers or preservatives |
| Endotoxin |
<2EU/mg (<0.002EU/Ī¼g) Determined by LAL gel clotting assay |
| Purity |
>95% Determined by SDS-PAGE |
| Sterility | 0.2 Āµm filtration |
| Production | Purified from cell culture supernatant in an animal-free facility |
| Purification | Protein G |
| RRID | AB_1107628 |
| Molecular Weight | 150 kDa |
| Storage | The antibody solution should be stored at the stock concentration at 4Ā°C. Do not freeze. |
Recommended Products
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Recommended Isotype Control(s)
InVivoMAb polyclonal Armenian hamster IgG
-
Recommended Dilution Buffer
InVivoPure pH 7.0 Dilution Buffer
in vitro T cell stimulation/activation
Huang, Y., et al. (2015). "CRK proteins selectively regulate T cell migration into inflamed tissues" J Clin Invest 125(3): 1019-1032. PubMed
Effector T cell migration into inflamed sites greatly exacerbates tissue destruction and disease severity in inflammatory diseases, including graft-versus-host disease (GVHD). T cell migration into such sites depends heavily on regulated adhesion and migration, but the signaling pathways that coordinate these functions downstream of chemokine receptors are largely unknown. Using conditional knockout mice, we found that T cells lacking the adaptor proteins CRK and CRK-like (CRKL) exhibit reduced integrin-dependent adhesion, chemotaxis, and diapedesis. Moreover, these two closely related proteins exhibited substantial functional redundancy, as ectopic expression of either protein rescued defects in T cells lacking both CRK and CRKL. We determined that CRK proteins coordinate with the RAP guanine nucleotide exchange factor C3G and the adhesion docking molecule CASL to activate the integrin regulatory GTPase RAP1. CRK proteins were required for effector T cell trafficking into sites of inflammation, but not for migration to lymphoid organs. In a murine bone marrow transplantation model, the differential migration of CRK/CRKL-deficient T cells resulted in efficient graft-versus-leukemia responses with minimal GVHD. Together, the results from our studies show that CRK family proteins selectively regulate T cell adhesion and migration at effector sites and suggest that these proteins have potential as therapeutic targets for preventing GVHD.
in vitro T cell stimulation/activation
Klimatcheva, E., et al. (2015). "CXCL13 antibody for the treatment of autoimmune disorders" BMC Immunol 16: 6. PubMed
BACKGROUND: Homeostatic B Cell-Attracting chemokine 1 (BCA-1) otherwise known as CXCL13 is constitutively expressed in secondary lymphoid organs by follicular dendritic cells (FDC) and macrophages. It is the only known ligand for the CXCR5 receptor, which is expressed on mature B cells, follicular helper T cells (Tfh), Th17 cells and regulatory T (Treg) cells. Aberrant expression of CXCL13 within ectopic germinal centers has been linked to the development of autoimmune disorders (e.g. Rheumatoid Arthritis, Multiple Sclerosis, Systemic Lupus Erythematosis). We, therefore, hypothesized that antibody-mediated disruption of the CXCL13 signaling pathway would interfere with the formation of ectopic lymphoid follicles in the target organs and inhibit autoimmune disease progression. This work describes pre-clinical development of human anti-CXCL13 antibody MAb 5261 and includes therapeutic efficacy data of its mouse counterpart in murine models of autoimmunity. RESULTS: We developed a human IgG1 monoclonal antibody, MAb 5261 that specifically binds to human, rodent and primate CXCL13 with an affinity of approximately 5 nM and is capable of neutralizing the activity of CXCL13 from these various species in in vitro functional assays. For in vivo studies we have engineered a chimeric antibody to contain the same human heavy and light chain variable genes along with mouse constant regions. Treatment with this antibody led to a reduction in the number of germinal centers in mice immunized with 4-Hydroxy-3-nitrophenylacetyl hapten conjugated to Keyhole Limpet Hemocyanin (NP-KLH) and, in adoptive transfer studies, interfered with the trafficking of B cells to the B cell areas of mouse spleen. Furthermore, this mouse anti-CXCL13 antibody demonstrated efficacy in a mouse model of Rheumatoid arthritis (Collagen-Induced Arthritis (CIA)) and Th17-mediated murine model of Multiple Sclerosis (passively-induced Experimental Autoimmune Encephalomyelitis (EAE)). CONCLUSIONS: We developed a novel therapeutic antibody targeting CXCL13-mediated signaling pathway for the treatment of autoimmune disorders.
in vitro T cell stimulation/activation
Bertin, S., et al. (2015). "Dual-specificity phosphatase 6 regulates CD4+ T-cell functions and restrains spontaneous colitis in IL-10-deficient mice" Mucosal Immunol 8(3): 505-515. PubMed
Mitogen-activated protein kinase (MAPK) phosphatases are dual-specificity phosphatases (DUSPs) that dephosphorylate phosphothreonine and phosphotyrosine residues within MAPKs. DUSP6 preferentially dephosphorylates extracellular signal-regulated kinases 1 and 2 (ERK1/2) rendering them inactive. Here, we study the role of DUSP6 in CD4(+) T-cell function, differentiation, and inflammatory profile in the colon. Upon T-cell receptor (TCR) stimulation, DUSP6 knockout (Dusp6(-/-)) CD4(+) T cells showed increased ERK1/2 activation, proliferation, T helper 1 differentiation, and interferon-gamma production, as well as a marked decrease in survival, interleukin- 17A (IL-17A) secretion, and regulatory T-cell function. To analyze the role of DUSP6 in vivo, we employed the Il10(-/-) model of colitis and generated Il10(-/-)/Dusp6(-/-) double-knockout mice. Il10(-/-)/Dusp6(-/-) mice suffered from accelerated and exacerbated spontaneous colitis, which was prevented by ERK1/2 inhibition. ERK1/2 inhibition also augmented regulatory T-cell differentiation in vitro and in vivo in both C57Bl/6 and Dusp6(-/-) mice. In summary, DUSP6 regulates CD4(+) T-cell activation and differentiation by inhibiting the TCR-dependent ERK1/2 activation. DUSP6 might therefore be a potential intervention target for limiting aberrant T-cell responses in T-cell-mediated diseases, such as inflammatory bowel disease.
in vitro T cell stimulation/activation
Pallandre, J. R., et al. (2015). "Novel aminotetrazole derivatives as selective STAT3 non-peptide inhibitors" Eur J Med Chem 103: 163-174. PubMed
The development of inhibitors blocking STAT3 transcriptional activity is a promising therapeutic approach against cancer and inflammatory diseases. In this context, the selectivity of inhibitors against the STAT1 transcription factor is crucial as STAT3 and STAT1 play opposite roles in the apoptosis of tumor cells and polarization of the immune response. A structure-based virtual screening followed by a luciferase-containing promoter assay on STAT3 and STAT1 signaling were used to identify a selective STAT3 inhibitor. An important role of the aminotetrazole group in modulating STAT3 and STAT1 inhibitory activities has been established. Optimization of the hit compound leads to 23. This compound inhibits growth and survival of cells with STAT3 signaling pathway while displaying a minimal effect on STAT1 signaling. Moreover, it prevents lymphocyte T polarization into Th17 and Treg without affecting their differentiation into Th1 lymphocyte.
in vitro T cell stimulation/activation
Heinemann, C., et al. (2014). "IL-27 and IL-12 oppose pro-inflammatory IL-23 in CD4+ T cells by inducing Blimp1" Nat Commun 5: 3770. PubMed
Central nervous system (CNS) autoimmunity is regulated by the balance of pro-inflammatory cytokines and IL-10. Here we identify the transcriptional regulator Blimp1 as crucial to induce IL-10 in inflammatory T helper cells. Pre-committed Th17 cells respond to IL-27 and IL-12 by upregulating Blimp1 and adopt a Tr-1-like phenotype characterized by IL-10 and IFN-gamma production. Accordingly, Blimp1-deficient effector T cells fail to produce IL-10, and deficiency in Tr-1 cell function leads to uncontrolled Th17 cell-driven CNS pathology without the need to stabilize the Th17 phenotype with IL-23. IL-23 counteracts IL-27 and IL-12-mediated effects on Tr-1-development reinforcing the pro-inflammatory phenotype of Th17 cells. Thus, the balance of IL-23 vs IL-12/IL-27 signals into CD4(+) effector T cells determines whether tissue inflammation is perpetuated or resolves.
in vitro T cell stimulation/activation
Bertin, S., et al. (2014). "The ion channel TRPV1 regulates the activation and proinflammatory properties of CD4(+) T cells" Nat Immunol 15(11): 1055-1063. PubMed
TRPV1 is a Ca(2+)-permeable channel studied mostly as a pain receptor in sensory neurons. However, its role in other cell types is poorly understood. Here we found that TRPV1 was functionally expressed in CD4(+) T cells, where it acted as a non-store-operated Ca(2+) channel and contributed to T cell antigen receptor (TCR)-induced Ca(2+) influx, TCR signaling and T cell activation. In models of T cell-mediated colitis, TRPV1 promoted colitogenic T cell responses and intestinal inflammation. Furthermore, genetic and pharmacological inhibition of TRPV1 in human CD4(+) T cells recapitulated the phenotype of mouse Trpv1(-/-) CD4(+) T cells. Our findings suggest that inhibition of TRPV1 could represent a new therapeutic strategy for restraining proinflammatory T cell responses.
in vitro T cell stimulation/activation
Vegran, F., et al. (2014). "The transcription factor IRF1 dictates the IL-21-dependent anticancer functions of TH9 cells" Nat Immunol 15(8): 758-766. PubMed
The TH9 subset of helper T cells was initially shown to contribute to the induction of autoimmune and allergic diseases, but subsequent evidence has suggested that these cells also exert antitumor activities. However, the molecular events that account for their effector properties are elusive. Here we found that the transcription factor IRF1 enhanced the effector function of TH9 cells and dictated their anticancer properties. Under TH9-skewing conditions, interleukin 1beta (IL-1beta) induced phosphorylation of the transcription factor STAT1 and subsequent expression of IRF1, which bound to the promoters of Il9 and Il21 and enhanced secretion of the cytokines IL-9 and IL-21 from TH9 cells. Furthermore, IL-1beta-induced TH9 cells exerted potent anticancer functions in an IRF1- and IL-21-dependent manner. Our findings thus identify IRF1 as a target for controlling the function of TH9 cells.
in vitro T cell stimulation/activation
Chen, E. J., et al. (2013). "Ezrin and moesin are required for efficient T cell adhesion and homing to lymphoid organs" PLoS One 8(2): e52368. PubMed
T cell trafficking between the blood and lymphoid organs is a complex, multistep process that requires several highly dynamic and coordinated changes in cyto-architecture. Members of the ezrin, radixin and moesin (ERM) family of actin-binding proteins have been implicated in several aspects of this process, but studies have yielded conflicting results. Using mice with a conditional deletion of ezrin in CD4+ cells and moesin-specific siRNA, we generated T cells lacking ERM proteins, and investigated the effect on specific events required for T cell trafficking. ERM-deficient T cells migrated normally in multiple in vitro and in vivo assays, and could undergo efficient diapedesis in vitro. However, these cells were impaired in their ability to adhere to the beta1 integrin ligand fibronectin, and to polarize appropriately in response to fibronectin and VCAM-1 binding. This defect was specific for beta1 integrins, as adhesion and polarization in response to ICAM-1 were normal. In vivo, ERM-deficient T cells showed defects in homing to lymphoid organs. Taken together, these results show that ERM proteins are largely dispensable for T cell chemotaxis, but are important for beta1 integrin function and homing to lymphoid organs.
in vitro T cell stimulation/activation
Berger, H., et al. (2013). "SOCS3 transactivation by PPARgamma prevents IL-17-driven cancer growth" Cancer Res 73(12): 3578-3590. PubMed
Activation of the transcription factor PPARgamma by the n-3 fatty acid docosahexaenoic acid (DHA) is implicated in controlling proinflammatory cytokine secretion, but the intracellular signaling pathways engaged by PPARgamma are incompletely characterized. Here, we identify the adapter-encoding gene SOCS3 as a critical transcriptional target of PPARgamma. SOCS3 promoter binding and gene transactivation by PPARgamma was associated with a repression in differentiation of proinflammatory T-helper (TH)17 cells. Accordingly, TH17 cells induced in vitro displayed increased SOCS3 expression and diminished capacity to produce interleukin (IL)-17 following activation of PPARgamma by DHA. Furthermore, naive CD4 T cells derived from mice fed a DHA-enriched diet displayed less capability to differentiate into TH17 cells. In two different mouse models of cancer, DHA prevented tumor outgrowth and angiogenesis in an IL-17-dependent manner. Altogether, our results uncover a novel molecular pathway by which PPARgamma-induced SOCS3 expression prevents IL-17-mediated cancer growth.
in vitro T cell stimulation/activation
Nowak, E. C., et al. (2009). "IL-9 as a mediator of Th17-driven inflammatory disease" J Exp Med 206(8): 1653-1660. PubMed
We report that like other T cells cultured in the presence of transforming growth factor (TGF) beta, Th17 cells also produce interleukin (IL) 9. Th17 cells generated in vitro with IL-6 and TGF-beta as well as purified ex vivo Th17 cells both produced IL-9. To determine if IL-9 has functional consequences in Th17-mediated inflammatory disease, we evaluated the role of IL-9 in the development and progression of experimental autoimmune encephalomyelitis, a mouse model of multiple sclerosis. The data show that IL-9 neutralization and IL-9 receptor deficiency attenuates disease, and this correlates with decreases in Th17 cells and IL-6-producing macrophages in the central nervous system, as well as mast cell numbers in the regional lymph nodes. Collectively, these data implicate IL-9 as a Th17-derived cytokine that can contribute to inflammatory disease.
Restraint of IFN-Ī³ expression through a distal silencer CNS-28 for tissue homeostasis.
In Immunity on 9 May 2023 by Cui, K., Chen, Z., et al.
PubMed
Interferon-Ī³ (IFN-Ī³) is a key cytokine in response to viral or intracellular bacterial infection in mammals. While a number of enhancers are described to promote IFN-Ī³ responses, to the best of our knowledge, no silencers for the Ifng gene have been identified. By examining H3K4me1 histone modification in naive CD4+ TĀ cells within Ifng locus, we identified a silencer (CNS-28) that restrains Ifng expression. Mechanistically, CNS-28 maintains Ifng silence by diminishing enhancer-promoter interactions within Ifng locus in a GATA3-dependent but T-bet-independent manner. Functionally, CNS-28 restrains Ifng transcription in NK cells, CD4+ cells,Ā and CD8+ TĀ cells during both innate and adaptive immune responses. Moreover, CNS-28 deficiency resulted in repressed type 2 responses due to elevated IFN-Ī³ expression, shifting Th1 and Th2 paradigm. Thus, CNS-28 activity ensures immune cell quiescence by cooperating with other regulatory cis elements within the Ifng gene locus to minimize autoimmunity. Published by Elsevier Inc.
- Immunology and Microbiology,
- Neuroscience,
- In Vitro,
- Mus musculus (House mouse)
Small intestine and colon tissue-resident memory CD8+ TĀ cells exhibit molecular heterogeneity and differential dependence on Eomes.
In Immunity on 10 January 2023 by Lin, Y. H., Duong, H. G., et al.
PubMed
Tissue-resident memory CD8+ T (TRM) cells are a subset of memory TĀ cells that play a critical role in limiting early pathogen spread and controlling infection. TRM cells exhibit differences across tissues, but their potential heterogeneity among distinct anatomic compartments within the small intestine and colon has not been well recognized. Here, by analyzing TRM cells from the lamina propria and epithelial compartments of the small intestine and colon, we showed that intestinal TRM cells exhibited distinctive patterns of cytokine and granzyme expression along with substantial transcriptional, epigenetic, and functional heterogeneity. The T-box transcription factor Eomes, which represses TRM cell formation in some tissues, exhibited unexpected context-specific regulatory roles in supporting the maintenance of established TRM cells in the small intestine, but not in the colon. Taken together, these data provide previously unappreciated insights into the heterogeneity and differential requirements for the formation vs. maintenance of intestinal TRM cells. Copyright Ā© 2022 Elsevier Inc. All rights reserved.
- Genetics,
- Immunology and Microbiology
Forced expression of the non-coding RNA miR-17ā¼92 restores activation and function in CD28-deficient CD4+ TĀ cells.
In IScience on 18 November 2022 by Dƶlz, M., Hasiuk, M., et al.
PubMed
CD28 provides the prototypical costimulatory signal required for productive T-cell activation. Known molecular consequences of CD28 costimulation are mostly based on studies of protein signaling molecules. The microRNA cluster miR-17ā¼92 is induced by TĀ cell receptor stimulation and further enhanced by combined CD28 costimulation. We demonstrate that transgenic miR-17ā¼92 cell-intrinsically largely overcomes defects caused by CD28 deficiency. Combining genetics, transcriptomics, bioinformatics, and biochemical miRNA:mRNA interaction maps we empirically validate miR-17ā¼92 target genes that include several negative regulators of TĀ cell activation. CD28-deficient TĀ cells exhibit derepressed miR-17ā¼92 target genes during activation. CRISPR/Cas9-mediated ablation of the miR-17ā¼92 targets Pten and Nrbp1 in naive CD28-/- CD4+ TĀ cells differentially increases proliferation and expression of the activation markers CD25 and CD44, respectively. Thus, we propose that miR-17ā¼92 constitutes a central mediator for TĀ cell activation, integrating signals by the TCR and CD28 costimulation by dampening multiple brakes that prevent TĀ cell activation. Ā© 2022 The Author(s).
- Mus musculus (House mouse),
- Biochemistry and Molecular biology,
- Cancer Research,
- Immunology and Microbiology
Tumor cells dictate anti-tumor immune responses by altering pyruvate utilization and succinate signaling in CD8+ TĀ cells.
In Cell Metabolism on 2 August 2022 by Elia, I., Rowe, J. H., et al.
PubMed
The tumor microenvironment (TME) is a unique metabolic niche that can inhibit TĀ cell metabolism and cytotoxicity. To dissect the metabolic interplay between tumors and TĀ cells, we establish an inĀ vitro system that recapitulates the metabolic niche of the TME and allows us to define cell-specific metabolism. We identify tumor-derived lactate as an inhibitor of CD8+ TĀ cell cytotoxicity, revealing an unexpected metabolic shunt in the TCA cycle. Metabolically fit cytotoxic TĀ cells shunt succinate out of the TCA cycle to promote autocrine signaling via the succinate receptor (SUCNR1). Cytotoxic TĀ cells are reliant on pyruvate carboxylase (PC) to replenish TCA cycle intermediates. By contrast, lactate reduces PC-mediated anaplerosis. The inhibition of pyruvate dehydrogenase (PDH) is sufficient to restore PC activity, succinate secretion, and the activation of SUCNR1. These studies identify PDH as a potential drug target to allow CD8+ TĀ cells to retain cytotoxicity and overcome a lactate-enriched TME. Copyright Ā© 2022 Elsevier Inc. All rights reserved.
- Biochemistry and Molecular biology,
- Immunology and Microbiology,
- Stem Cells and Developmental Biology,
- Mus musculus (House mouse)
Localization of a TORC1-eIF4F translation complex during CD8+ TĀ cell activation drives divergent cell fate.
In Molecular Cell on 7 July 2022 by Liedmann, S., Liu, X., et al.
PubMed
Activated CD8+ T lymphocytes differentiate into heterogeneous subsets. Using super-resolution imaging, we found that prior to the first division, dynein-dependent vesicular transport polarized active TORC1 toward the microtubule-organizing center (MTOC) at the proximal pole. This active TORC1 was physically associated with active eIF4F, required for the translation of c-myc mRNA. As a consequence, c-myc-translating polysomes polarized toward the cellular pole proximal to the immune synapse, resulting in localized c-myc translation. Upon division, the TORC1-eIF4A complex preferentially sorted to the proximal daughter cell, facilitating asymmetric c-Myc synthesis. Transient disruption of eIF4A activity at first division skewed long-term cell fate trajectories to memory-like function. Using a genetic barcoding approach, we found that first-division sister cells often displayed differences in transcriptional profiles that largely correlated with c-Myc and TORC1 target genes. Our findings provide mechanistic insights as to how distinct TĀ cell fate trajectories can be established during the first division. Copyright Ā© 2022 The Author(s). Published by Elsevier Inc. All rights reserved.
- Mus musculus (House mouse),
- Immunology and Microbiology
STAT1 signaling protects self-reactive T cells from control by innate cells during neuroinflammation.
In JCI Insight on 22 June 2022 by Arbelaez, C. A., Palle, P., et al.
PubMed
The transcription factor STAT1 plays a critical role in modulating the differentiation of CD4+ T cells producing IL-17 and GM-CSF, which promote the development of experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis (MS). The protective role of STAT1 in MS and EAE has been largely attributed to its ability to limit pathogenic Th cells and promote Tregs. Using mice with selective deletion of STAT1 in T cells (STAT1CD4-Cre), we identified a potentially novel mechanism by which STAT1 regulates neuroinflammation independently of Foxp3+ Tregs. STAT1-deficient effector T cells became the target of NK cell-mediated killing, limiting their capacity to induce EAE. STAT1-deficient T cells promoted their own killing by producing more IL-2 that, in return, activated NK cells. Elimination of NK cells restored EAE susceptibility in STAT1CD4-Cre mice. Therefore, our study suggests that the STAT1 pathway can be manipulated to limit autoreactive T cells during autoimmunity directed against the CNS.
- FC/FACS,
- Mus musculus (House mouse)
Ursodeoxycholic acid reduces antitumor immunosuppression by inducing CHIP-mediated TGF-Ī² degradation.
In Nature Communications on 14 June 2022 by Shen, Y., Lu, C., et al.
PubMed
TGF-Ī² is essential for inducing systemic tumor immunosuppression; thus, blocking TGF-Ī² can greatly enhance antitumor immunity. However, there are still no effective TGF-Ī² inhibitors in clinical use. Here, we show that the clinically approved compound ursodeoxycholic acid (UDCA), by degrading TGF-Ī², enhances antitumor immunity through restraining Treg cell differentiation and activation in tumor-bearing mice. Furthermore, UDCA synergizes with anti-PD-1 to enhance antitumor immunity and tumor-specific immune memory in tumor-bearing mice. UDCA phosphorylates TGF-Ī² at T282 site via TGR5-cAMP-PKA axis, causing increased binding of TGF-Ī² to carboxyl terminus of Hsc70-interacting protein (CHIP). Then, CHIP ubiquitinates TGF-Ī² at the K315 site, initiating p62-dependent autophagic sorting and subsequent degradation of TGF-Ī². Notably, results of retrospective analysis shows that combination therapy with anti-PD-1 or anti-PD-L1 and UDCA has better efficacy in tumor patients than anti-PD-1 or anti-PD-L1 alone. Thus, our results show a mechanism for TGF-Ī² regulation and implicate UDCA as a potential TGF-Ī² inhibitor to enhance antitumor immunity. Ā© 2022. The Author(s).
- Genetics,
- Immunology and Microbiology,
- Mus musculus (House mouse)
The RNA-binding protein IMP2 drives a stromal-Th17 cell circuit in autoimmune neuroinflammation.
In JCI Insight on 8 February 2022 by Bechara, R., Amatya, N., et al.
PubMed
Stromal cells are emerging as key drivers of autoimmunity, partially because they produce inflammatory chemokines that orchestrate inflammation. Chemokine expression is regulated transcriptionally but also through posttranscriptional mechanisms, the specific drivers of which are still incompletely defined. CCL2 (MCP1) is a multifunctional chemokine that drives myeloid cell recruitment. During experimental autoimmune encephalomyelitis (EAE), an IL-17-driven model of multiple sclerosis, CCL2 produced by lymph node (LN) stromal cells was essential for immunopathology. Here, we showed that Ccl2 mRNA upregulation in human stromal fibroblasts in response to IL-17 required the RNA-binding protein IGF-2 mRNA-binding protein 2 (IGF2BP2, IMP2), which is expressed almost exclusively in nonhematopoietic cells. IMP2 binds directly to CCL2 mRNA, markedly extending its transcript half-life, and is thus required for efficient CCL2 secretion. Consistent with this, Imp2-/- mice showed reduced CCL2 production in LNs during EAE, causing impairments in monocyte recruitment and Th17 cell polarization. Imp2-/- mice were fully protected from CNS inflammation. Moreover, deletion of IMP2 after EAE onset was sufficient to mitigate disease severity. These data showed that posttranscriptional control of Ccl2 in stromal cells by IMP2 was required to permit IL-17-driven progression of EAE pathogenesis.
- In Vitro,
- Mus musculus (House mouse),
- Immunology and Microbiology,
- Neuroscience
Astrocyte-Derived Pleiotrophin Mitigates Late-Stage Autoimmune CNS Inflammation.
In Frontiers in Immunology on 21 January 2022 by Linnerbauer, M., LƶĆlein, L., et al.
PubMed
Astrocytes are the most abundant glial cells in the central nervous system (CNS) with the capacity to sense and react to injury and inflammatory events. While it has been widely documented that astrocytes can exert tissue-degenerative functions, less is known about their protective and disease-limiting roles. Here, we report the upregulation of pleiotrophin (PTN) by mouse and human astrocytes in multiple sclerosis (MS) and its preclinical model experimental autoimmune encephalomyelitis (EAE). Using CRISPR-Cas9-based genetic perturbation systems, we demonstrate in vivo that astrocyte-derived PTN is critical for the recovery phase of EAE and limits chronic CNS inflammation. PTN reduces pro-inflammatory signaling in astrocytes and microglia and promotes neuronal survival following inflammatory challenge. Finally, we show that intranasal administration of PTN during the late phase of EAE successfully reduces disease severity, making it a potential therapeutic candidate for the treatment of progressive MS, for which existing therapies are limited. Copyright Ā© 2022 Linnerbauer, LƶĆlein, Farrenkopf, Vandrey, Tsaktanis, Naumann and Rothhammer.
- Cancer Research,
- Immunology and Microbiology,
- Mus musculus (House mouse)
Divergent fates of antigen-specific CD8+ TĀ cell clones in mice with acute leukemia.
In Cell Reports on 9 November 2021 by Chen, X., MacNabb, B. W., et al.
PubMed
The existence of a dysfunctional CD8+ TĀ cell state in cancer is well established. However, the degree to which CD8+ TĀ cell fates are influenced by the context in which they encounter cognate tumor antigen is less clear. We previously demonstrated that CD8+ TĀ cells reactive to a model leukemia antigen were deleted by antigen cross-presenting type 1 conventional dendritic cells (cDC1s). Here, through a study of TĀ cell receptor (TCR) transgenic CD8+ TĀ cells (TCRTg101) reactive to a native C1498 leukemia cell antigen, we uncover a different mode of TĀ cell tolerance in which TCRTg101 undergo progressive expansion and differentiation into an exhausted state. Antigen encounter by TCRTg101 requires leukemia cell major histocompatibility complex (MHC)-I expression and is independent of DCs, implying that leukemia cells directly mediate the exhausted TCRTg101 phenotype. Collectively, our data reveal that leukemia antigens are presented to CD8+ TĀ cells via discrete pathways, leading to distinct tolerant states. Copyright Ā© 2021 The Author(s). Published by Elsevier Inc. All rights reserved.
- Cell Culture,
- Mus musculus (House mouse),
- Immunology and Microbiology
Urolithin A ameliorates experimental autoimmune encephalomyelitis by targeting aryl hydrocarbon receptor.
In EBioMedicine on 1 February 2021 by Shen, P. X., Li, X., et al.
PubMed
Urolithin A (URA) is an intestinal microbiota metabolic product from ellagitannin-containing foods with multiple biological activities. However, its role in autoimmune diseases is largely unknown. Here, for first time, we demonstrate the therapeutic effect of URA in an experimental autoimmune encephalomyelitis (EAE) animal model. Therapeutic effect was evaluated via an active and passive EAE animal model in vivo. The function of URA on bone marrow-derived dendritic cells (BM-DCs), T cells, and microglia were tested in vitro. Oral URA (25 mg/kg/d) suppressed disease progression at prevention, induction, and effector phases of preclinical EAE. Histological evaluation showed that significantly fewer inflammatory cells, decreased demyelination, lower numbers of M1-type microglia and activated DCs, as well as reduced infiltrating Th1/Th17 cells were present in the central nervous system (CNS) of the URA-treated group. URA treatment at 25 Ī¼M inhibited the activation of BM-DCs in vitro, restrained Th17 cell differentiation in T cell polarization conditions, and in a DC-CD4+ T cell co-culture system. Moreover, we confirmed URA inhibited pathogenicity of Th17 cells in adoptive EAE. Mechanism of URA action was directly targeting Aryl Hydrocarbon Receptor (AhR) and modulating the signaling pathways. Collectively, our study offers new evidence that URA, as a human microbial metabolite, is valuable to use as a prospective therapeutic candidate for autoimmune diseases. Copyright Ā© 2021 The Authors. Published by Elsevier B.V. All rights reserved.
- Mus musculus (House mouse),
- Biochemistry and Molecular biology
An IL-27-Driven Transcriptional Network Identifies Regulators of IL-10 Expression across T Helper Cell Subsets.
In Cell Reports on 24 November 2020 by Zhang, H., Madi, A., et al.
PubMed
Interleukin-27 (IL-27) is an immunoregulatory cytokine that suppresses inflammation through multiple mechanisms, including induction of IL-10, but the transcriptional network mediating its diverse functions remains unclear. Combining temporal RNA profiling with computational algorithms, we predict 79 transcription factors induced by IL-27 in TĀ cells. We validate 11 known and discover 5 positive (Cebpb, Fosl2, Tbx21, Hlx, and Atf3) and 2 negative (Irf9 and Irf8) Il10 regulators, generating an experimentally refined regulatory network for Il10. We report two central regulators, Prdm1 and Maf, that cooperatively drive the expression of signature genes induced by IL-27 in type 1 regulatory TĀ cells, mediate IL-10 expression in all T helper cells, and determine the regulatory phenotype of colonic Foxp3+ regulatory TĀ cells. Prdm1/Maf double-knockout mice develop spontaneous colitis, phenocopying ll10-deficient mice. Our work provides insights into IL-27-driven transcriptional networks and identifies two shared Il10 regulators that orchestrate immunoregulatory programs across T helper cell subsets. Copyright Ā© 2020 The Authors. Published by Elsevier Inc. All rights reserved.
- Mus musculus (House mouse),
- Immunology and Microbiology
Mouse T cell priming is enhanced by maturation-dependent stiffening of the dendritic cell cortex.
In eLife on 27 July 2020 by Blumenthal, D., Chandra, V., et al.
PubMed
T cell activation by dendritic cells (DCs) involves forces exerted by the T cell actin cytoskeleton, which are opposed by the cortical cytoskeleton of the interacting antigen-presenting cell. During an immune response, DCs undergo a maturation process that optimizes their ability to efficiently prime naĆÆve T cells. Using atomic force microscopy, we find that during maturation, DC cortical stiffness increases via a process that involves actin polymerization. Using stimulatory hydrogels and DCs expressing mutant cytoskeletal proteins, we find that increasing stiffness lowers the agonist dose needed for T cell activation. CD4+ T cells exhibit much more profound stiffness dependency than CD8+ T cells. Finally, stiffness responses are most robust when T cells are stimulated with pMHC rather than anti-CD3Īµ, consistent with a mechanosensing mechanism involving receptor deformation. Taken together, our data reveal that maturation-associated cytoskeletal changes alter the biophysical properties of DCs, providing mechanical cues that costimulate T cell activation. Ā© 2020, Blumenthal et al.
- In Vitro,
- Mus musculus (House mouse)
Mouse CD163 deficiency strongly enhances experimental collagen-induced arthritis.
In Scientific Reports on 24 July 2020 by Svendsen, P., Etzerodt, A., et al.
PubMed
The scavenger receptor CD163 is highly expressed in macrophages in sites of chronic inflammation where it has a not yet defined role. Here we have investigated development of collagen-induced arthritis (CIA) and collagen antibody-induced arthritis (CAIA) in CD163-deficient C57BL/6 mice. Compared to wild-type mice, the CIA in CD163-deficient mice had a several-fold higher arthritis score with early onset, prolonged disease and strongly enhanced progression. Further, the serum anti-collagen antibody isotypes as well as the cytokine profiles and T cell markers in the inflamed joints revealed that CD163-deficient mice after 52Ā days had a predominant Th2 response in opposition to a predominant Th1 response in CD163+/+ mice. Less difference in disease severity between the CD163+/+ and CD163-/- mice was seen in the CAIA model that to a large extent induces arthritis independently of T-cell response and endogenous Th1/Th2 balance. In conclusion, the present set of data points on a novel strong anti-inflammatory role of CD163.
- In Vitro,
- Mus musculus (House mouse),
- Immunology and Microbiology
T Cell-Intrinsic IRF5 Regulates T Cell Signaling, Migration, and Differentiation and Promotes Intestinal Inflammation.
In Cell Reports on 30 June 2020 by Yan, J., Pandey, S. P., et al.
PubMed
IRF5 polymorphisms are associated with multiple immune-mediated diseases, including ulcerative colitis. IRF5 contributions are attributed to its role in myeloid lineages. How TĀ cell-intrinsic IRF5 contributes to inflammatory outcomes is not well understood. We identify a previously undefined key role for TĀ cell-intrinsic IRF5. In mice, IRF5 in CD4+ TĀ cells promotes Th1- and Th17-associated cytokines and decreases Th2-associated cytokines. IRF5 is required for the optimal assembly of the TCR-initiated signaling complex and downstream signaling at early times, and at later times binds to promoters of Th1- and Th17-associated transcription factors and cytokines. IRF5 also regulates chemokine receptor-initiated signaling and, in turn, TĀ cell migration. InĀ vivo, IRF5 in CD4+ TĀ cells enhances the severity of experimental colitis. Importantly, human CD4+ TĀ cells from high IRF5-expressing disease-risk genetic carriers demonstrate increased chemokine-induced migration and Th1/Th17 cytokines and reduced Th2-associated and anti-inflammatory cytokines. These data demonstrate key roles for TĀ cell-intrinsic IRF5 in inflammatory outcomes.Copyright Ā© 2020 The Author(s). Published by Elsevier Inc. All rights reserved.
- FC/FACS,
- Cancer Research,
- Immunology and Microbiology,
- Mus musculus (House mouse)
CTL Attenuation Regulated by PS1 in Cancer-Associated Fibroblast.
In Frontiers in Immunology on 27 June 2020 by Zhang, H., Jiang, R., et al.
PubMed
Objective: Cancer-associated fibroblasts (CAFs) were associated with tumor progression in the tumor microenvironment (TME). However, their immunosuppressive roles in protecting cancer cells from the attack by cytotoxic T lymphocytes (CTLs) are not fully clear. In this study, we investigated whether and how CAFs regulate tumor-infiltrating lymphocytes as well as their role in tumor immunosuppression. Methods: Eighty-three cases of ovarian cancer and 10 controls were analyzed for CAFs and CD8+ tumor-infiltrating lymphocytes by gene array and immunohistochemistry. We evaluated presenilin 1 (PS1) expression in CAFs, CTL penetration, tumor burden, dendritic cell function, and migration of tumor-infiltrating lymphocytes and their function in vivo and in vitro after silencing PS1. In addition, the pathway via which PS1 affects the TME was also evaluated. Results: PS1 was highly expressed in CAFs, and its silencing significantly promoted CD8+ CTL proliferation and penetration in multiple ovarian models (p 0.05), resulting in tumor regression and growth inhibition. Interleukin (IL)-1Ī² was identified as a major immune inhibitor in the TME, and it was significantly decreased after PS1 silencing (p 0.05), which was regulated by the WNT/Ī²-catenin pathway. It was also showed that high expression of IL-1Ī² in CAFs inhibits CTL penetration significantly (p 0.05). Conclusion: Highly expressed PS1 in CAFs plays a crucial role in regulating tumor-infiltrating lymphocyte populations in the TME via the WNT/Ī²-catenin pathway. Targeting PS1 may retrieve functional CTLs in the TME and improve the efficacy of current immunotherapies. Copyright Ā© 2020 Zhang, Jiang, Zhou, Wang, Xu, Zhang, Gu, Fu, Shen, Zhang, Feng, Zhang, Chen and Shen.
- Mus musculus (House mouse),
- Biochemistry and Molecular biology,
- Cell Biology
CD28 Regulates Metabolic Fitness for Long-Lived Plasma Cell Survival.
In Cell Reports on 23 June 2020 by Utley, A., Chavel, C., et al.
PubMed
Durable humoral immunity against epidemic infectious disease requires the survival of long-lived plasma cells (LLPCs). LLPC longevity is dependent on metabolic programs distinct from short-lived plasma cells (SLPCs); however, the mechanistic basis for this difference is unclear. We have previously shown that CD28, the prototypic TĀ cell costimulatory receptor, is expressed on both LLPCs and SLPCs but is essential only for LLPC survival. Here we show that CD28 transduces pro-survival signaling specifically in LLPCs through differential SLP76 expression. CD28 signaling in LLPCs increased glucose uptake, mitochondrial mass/respiration, and reactive oxygen species (ROS) production. Unexpectedly, CD28-mediated regulation of mitochondrial respiration, NF-ĪŗB activation, and survival was ROS dependent. IRF4, a target of NF-ĪŗB, was upregulated by CD28 activation in LLPCs and decreased IRF4 levels correlated with decreased glucose uptake, mitochondrial mass, ROS, and CD28-mediated survival. Altogether, these data demonstrate that CD28 signaling induces a ROS-dependent metabolic program required for LLPC survival. Copyright Ā© 2020 The Author(s). Published by Elsevier Inc. All rights reserved.
- Mus musculus (House mouse),
- Immunology and Microbiology
Inhibition of IRF4 in dendritic cells by PRR-independent and -dependent signals inhibit Th2 and promote Th17 responses.
In eLife on 4 February 2020 by Lee, J., Zhang, J., et al.
PubMed
Cyclic AMP (cAMP) is involved in many biological processes but little is known regarding its role in shaping immunity. Here we show that cAMP-PKA-CREB signaling (a pattern recognition receptor [PRR]-independent mechanism) regulates conventional type-2 Dendritic Cells (cDC2s) in mice and reprograms their Th17-inducing properties via repression of IRF4 and KLF4, transcription factors essential for cDC2-mediated Th2 induction. In mice, genetic loss of IRF4 phenocopies the effects of cAMP on Th17 induction and restoration of IRF4 prevents the cAMP effect. Moreover, curdlan, a PRR-dependent microbial product, activates CREB and represses IRF4 and KLF4, resulting in a pro-Th17 phenotype of cDC2s. These in vitro and in vivo results define a novel signaling pathway by which cDC2s display plasticity and provide a new molecular basis for the classification of novel cDC2 and cDC17 subsets. The findings also reveal that repressing IRF4 and KLF4 pathway can be harnessed for immuno-regulation. Ā© 2020, Lee et al.
- In Vitro,
- Mus musculus (House mouse),
- Biochemistry and Molecular biology,
- Cell Biology,
- Immunology and Microbiology
The Transcription Factor Bhlhe40 Programs Mitochondrial Regulation of Resident CD8+ T Cell Fitness and Functionality.
In Immunity on 17 September 2019 by Li, C., Zhu, B., et al.
PubMed
Tissue-resident memory CD8+ T (Trm) cells share core residency gene programs with tumor-infiltrating lymphocytes (TILs). However, the transcriptional, metabolic, and epigenetic regulation of Trm cell and TIL development and function is largely undefined. Here, we found that the transcription factor Bhlhe40 was specifically required for Trm cell and TIL development and polyfunctionality. Local PD-1 signaling inhibited TIL Bhlhe40 expression, and Bhlhe40 was critical for TIL reinvigoration following anti-PD-L1 blockade. Mechanistically, Bhlhe40 sustained Trm cell and TIL mitochondrial fitness and a functional epigenetic state. Building on these findings, we identified an epigenetic and metabolic regimen that promoted Trm cell and TIL gene signatures associated with tissue residency and polyfunctionality. This regimen empowered the anti-tumor activity of CD8+ TĀ cells and possessed therapeutic potential even at an advanced tumor stage in mouse models. Our results provide mechanistic insights into the local regulation of Trm cell and TIL function. Copyright Ā© 2019 Elsevier Inc. All rights reserved.
- Mus musculus (House mouse),
- Immunology and Microbiology,
- Neuroscience
CD8+ T Cell Priming in Established Chronic Viral Infection Preferentially Directs Differentiation of Memory-like Cells for Sustained Immunity.
In Immunity on 16 October 2018 by Snell, L. M., Macleod, B. L., et al.
PubMed
CD8+ TĀ cell exhaustion impedes control of chronic viral infection; yet how new TĀ cell responses are mounted during chronic infection is unclear. Unlike TĀ cells primed at the onset of infection that rapidly differentiate into effectors and exhaust, we demonstrate that virus-specific CD8+ TĀ cells primed after establishment of chronic LCMV infection preferentially generate memory-like transcription factor TCF1+ cells that were transcriptionally and proteomically distinct, less exhausted, and more responsive to immunotherapy. Mechanistically, adaptations of antigen-presenting cells and diminished TĀ cell signaling intensity promoted differentiation of the memory-like subset at the expense of rapid effector cell differentiation, which was now highly dependent on IL-21-mediated CD4+ TĀ cell help for its functional generation. Chronic viral infection similarly redirected de novo differentiation of tumor-specific CD8+ TĀ cells, ultimately preventing cancer control. Thus, targeting these TĀ cell stimulatory pathways could enable strategies to control chronic infection, tumors, and enhance immunotherapeutic efficacy. Copyright Ā© 2018 Elsevier Inc. All rights reserved.
