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1.
J Leukoc Biol ; 109(5): 877-890, 2021 05.
Article in English | MEDLINE | ID: mdl-33438263

ABSTRACT

Neutrophil-macrophage interplay is a fine-tuning mechanism that regulates the innate immune response during infection and inflammation. Cell surface receptors play an essential role in neutrophil and macrophage functions. The same receptor can provide different outcomes within diverse leukocyte subsets in different inflammatory conditions. Understanding the variety of responses mediated by one receptor is critical for the development of anti-inflammatory treatments. In this study, we evaluated the role of a leukocyte adhesive receptor, integrin αD ß2 , in the development of acute inflammation. αD ß2 is mostly expressed on macrophages and contributes to the development of chronic inflammation. In contrast, we found that αD -knockout dramatically increases mortality in the cecal ligation and puncture sepsis model and LPS-induced endotoxemia. This pathologic outcome of αD -deficient mice is associated with a reduced number of monocyte-derived macrophages and an increased number of neutrophils in their lungs. However, the tracking of adoptively transferred fluorescently labeled wild-type (WT) and αD-/- monocytes in WT mice during endotoxemia demonstrated only a moderate difference between the recruitment of these two subsets. Moreover, the rescue experiment, using i.v. injection of WT monocytes to αD -deficient mice followed by LPS challenge, showed only slightly reduced mortality. Surprisingly, the injection of WT neutrophils to the bloodstream of αD-/- mice markedly increased migration of monocyte-derived macrophage to lungs and dramatically improves survival. αD -deficient neutrophils demonstrate increased necrosis/pyroptosis. αD ß2 -mediated macrophage accumulation in the lungs promotes efferocytosis that reduced mortality. Hence, integrin αD ß2 implements a complex defense mechanism during endotoxemia, which is mediated by macrophages via a neutrophil-dependent pathway.


Subject(s)
Endotoxemia/immunology , Integrin alpha Chains/metabolism , Neutrophils/metabolism , Sepsis/immunology , Adoptive Transfer , Animals , Cecum/pathology , Cell Count , Cell Movement , Cytokines/blood , Disease Models, Animal , Endotoxemia/blood , Endotoxemia/complications , Integrin alpha Chains/deficiency , Ligation , Lipopolysaccharides , Lung/pathology , Macrophages/pathology , Male , Mice, Inbred C57BL , Monocytes/pathology , Necrosis , Neutrophils/pathology , Phagocytosis , Punctures , Pyroptosis , Sepsis/blood , Sepsis/complications , Survival Analysis , Up-Regulation
2.
Nat Commun ; 11(1): 6171, 2020 12 02.
Article in English | MEDLINE | ID: mdl-33268774

ABSTRACT

While effective in specific settings, adoptive chimeric antigen receptor (CAR) T cell therapy for cancer requires further improvement and optimization. Our previous results show that CD40L-overexpressing CAR T cells mobilize endogenous immune effectors, resulting in improved antitumor immunity. However, the cell populations required for this protective effect remain to be identified. Here we show, by analyzing Batf3-/- mice lacking the CD103+ conventional dendritic cell type 1 (cDC1) subpopulation important for antigen cross-presentation, that CD40L-overexpressing CAR T cells elicit an impaired antitumor response in the absence of cDC1s. We further find that CD40L-overexpressing CAR T cells stimulate tumor-resident CD11b-CD103- double-negative (DN) cDCs to proliferate and differentiate into cDC1s in wild-type mice. Finally, re-challenge experiments show that endogenous CD8+ T cells are required for protective antitumor memory in this setting. Our findings thus demonstrate the stimulatory effect of CD40L-overexpressing CAR T cells on innate and adaptive immune cells, and provide a rationale for using CD40L-overexpressing CAR T cells to improve immunotherapy responses.


Subject(s)
CD40 Ligand/genetics , CD8-Positive T-Lymphocytes/immunology , Dendritic Cells/immunology , Immunotherapy, Adoptive/methods , Lymphoma, B-Cell/therapy , Receptors, Chimeric Antigen/genetics , Adaptive Immunity , Animals , Antigen Presentation , Antigens, CD/genetics , Antigens, CD/immunology , Basic-Leucine Zipper Transcription Factors/deficiency , Basic-Leucine Zipper Transcription Factors/genetics , Basic-Leucine Zipper Transcription Factors/immunology , CD11b Antigen/deficiency , CD11b Antigen/genetics , CD11b Antigen/immunology , CD40 Ligand/immunology , CD8-Positive T-Lymphocytes/cytology , Dendritic Cells/cytology , Female , Gene Expression , Immunity, Innate , Immunophenotyping , Integrin alpha Chains/deficiency , Integrin alpha Chains/genetics , Integrin alpha Chains/immunology , Lymphoma, B-Cell/genetics , Lymphoma, B-Cell/immunology , Lymphoma, B-Cell/pathology , Mice , Mice, Inbred BALB C , Mice, Knockout , Neoplasm Transplantation , Receptors, Chimeric Antigen/immunology , Repressor Proteins/deficiency , Repressor Proteins/genetics , Repressor Proteins/immunology
3.
Am J Physiol Cell Physiol ; 316(6): C876-C887, 2019 06 01.
Article in English | MEDLINE | ID: mdl-30892939

ABSTRACT

Regenerative medicine treatments for severe skeletal muscle injuries are limited, resulting in persistent functional deficits. Clinical options include neglecting the wound with the expectation that fibrosis will develop or using an autologous muscle graft with minimal functional improvement. A regenerative matrix can be used, but muscle fiber development on these matrices remains a challenge in vivo. Here, we explored the fundamental mechanisms that mediate cell-substrate signaling and its effect on cell-cell communication during myoblast fusion and tube formation to improve outcomes following implantation of matrices used to stimulate muscle regeneration. We previously reported that integrin-α7 was increased on anisotropic biomaterials, suggesting a role for α7ß1 signaling in myoblast communication via connexin 43 and M-cadherin. Our results demonstrated that α7 silencing blocked expression of myogenic differentiation factor 1 (Myod), myogenin (Myog), myogenic factor 6 (Myf6), myosin heavy chain type 1 (Myh1), and transmembrane protein 8c (Tmem8c), indicating that myoblast fusion was inhibited. Expression of α5 and M-cadherin decreased but ß1 and connexin 43 increased. We examined protein production and observed reduced extracellular-signal regulated kinase 1/2 (ERK) in α7-silenced cells that correlated with upregulation of connexin 43 and M-cadherin, suggesting a compensatory pathway. These results indicate that α7 signaling plays a critical role in ex vivo fusion and implicates a relationship with connexin 43 and M-cadherin.


Subject(s)
Cadherins/metabolism , Connexin 43/metabolism , Integrin alpha Chains/deficiency , Myoblasts/metabolism , Signal Transduction/physiology , Animals , Antigens, CD/genetics , Cell Communication/physiology , Cells, Cultured , Integrin alpha Chains/genetics , Mice , Mice, Inbred C57BL
4.
PLoS One ; 13(5): e0197937, 2018.
Article in English | MEDLINE | ID: mdl-29813125

ABSTRACT

BACKGROUND: Integrin α8 (ITGA8) heterodimerizes with integrin ß1 and is highly expressed in stromal cells of the lung. Platelet-derived growth factor receptor beta (PDGFRß+) cells constitute a major population of contractile myofibroblasts in the lung following bleomycin-induced fibrosis. Integrin α8ß1 is upregulated in fibrotic foci in bleomycin-induced lung injury. However, the functional role of ITGA8 in fibrogenesis has not been characterized. In this study, we examined whether genetic deletion of ITGA8 from PDGFRß+ cells in the lung altered fibrosis. METHODS: Pdgfrb-Cre/+;Itga8flox/- or Pdgfrb-Cre/+;Itga8flox/flox (Cre+) and control mice (Cre-) were used for in vitro and in vivo studies. Primary cultures of PDGFRß+ cells were exposed to TGFß, followed by RNA isolation for qPCR. For in vivo studies, Cre+ and Cre- mice were characterized at baseline and after bleomycin-induced fibrosis. RESULTS: PDGFRß-selected cells from Cre+ animals showed higher levels of Col1a1 expression after treatment with TGFß. However, Cre- and Cre+ animals showed no significant difference in measures of acute lung injury or fibrosis following bleomycin challenge. CONCLUSION: While ITGA8 deletion in lung PDGFRß+ stromal cells showed evidence of greater Col1a1 mRNA expression after TGFß treatment in vitro, no functional difference was detected in vivo.


Subject(s)
Integrin alpha Chains/metabolism , Pulmonary Fibrosis/metabolism , Animals , Bronchoalveolar Lavage Fluid/chemistry , Disease Models, Animal , Female , Gene Deletion , Integrin alpha Chains/deficiency , Integrin alpha Chains/genetics , Mice , Pulmonary Fibrosis/genetics , Up-Regulation
5.
PLoS One ; 12(6): e0179678, 2017.
Article in English | MEDLINE | ID: mdl-28628641

ABSTRACT

BACKGROUND: Pulmonary dendritic cells drive lung responses to foreign antigens, including Saccharopolyspora rectivirgula, a causative agent of hypersensitivity pneumonitis. While the airway inflammatory mechanisms involved in hypersensitivity pneumonitis are well described, the mechanisms leading to the break in homeostasis and hypersensitivity pneumonitis onset are not well-described, and could involve CD103+ dendritic cells, which are found at baseline and during inflammatory responses in the lung. However, recent demonstration of the ability of CD103+ dendritic cells to induce inflammatory responses starkly contrasts with their classically described role as regulatory cells. These discrepancies may be attributable to the lack of current information on the importance of CD103 expression and modulation on these cells during inflammatory episodes. METHODS: To verify the importance of CD103 expression in the regulation of hypersensitivity pneumonitis, wild-type and Cd103-/- mice were exposed intranasally to S. rectivirgula and airway inflammation was quantified. Surface expression of CD103 in response to S. rectivirgula exposure was studied and cell transfers were used to determine the relative importance of CD103 expression on dendritic cells and T cells in regulating the inflammation in hypersensitivity pneumonitis. RESULTS: Cd103-/- mice developed an exacerbated inflammatory response as early as 18h following S. rectivirgula exposure. CD103 expression on dendritic cells was downregulated quickly following S. rectivirgula exposure, and cell transfers demonstrated that CD103 expression on dendritic cells specifically (and not T cells) regulates the onset and severity of this response. CONCLUSION: All in all, we demonstrate that CD103 expression by dendritic cells, but not T cells, is crucial for homeostasis maintenance and the regulation of the TH17 airway inflammatory response in hypersensitivity pneumonitis.


Subject(s)
Alveolitis, Extrinsic Allergic/pathology , Antigens, CD/metabolism , Dendritic Cells/metabolism , Integrin alpha Chains/metabolism , Alveolitis, Extrinsic Allergic/immunology , Alveolitis, Extrinsic Allergic/microbiology , Animals , Antigens, Bacterial/immunology , Antigens, CD/genetics , Bronchoalveolar Lavage Fluid/cytology , CD4-Positive T-Lymphocytes/cytology , CD4-Positive T-Lymphocytes/immunology , CD4-Positive T-Lymphocytes/metabolism , CD8-Positive T-Lymphocytes/cytology , CD8-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/metabolism , Cytokines/metabolism , Dendritic Cells/cytology , Dendritic Cells/immunology , Disease Models, Animal , Down-Regulation , Immunoglobulin G/blood , Immunoglobulin G/immunology , Integrin alpha Chains/deficiency , Integrin alpha Chains/genetics , Leukocytes/cytology , Lung/cytology , Lung/immunology , Lung/pathology , Mice , Mice, Knockout , Saccharopolyspora/metabolism , Saccharopolyspora/pathogenicity , Severity of Illness Index , Spleen/cytology , Spleen/immunology , Th17 Cells/cytology , Th17 Cells/immunology , Th17 Cells/metabolism
6.
J Immunol ; 198(12): 4855-4867, 2017 06 15.
Article in English | MEDLINE | ID: mdl-28500072

ABSTRACT

Macrophage accumulation is a critical step during development of chronic inflammation, initiating progression of many devastating diseases. Leukocyte-specific integrin αDß2 (CD11d/CD18) is dramatically upregulated on macrophages at inflammatory sites. Previously we found that CD11d overexpression on cell surfaces inhibits in vitro cell migration due to excessive adhesion. In this study, we have investigated how inflammation-mediated CD11d upregulation contributes to macrophage retention at inflammatory sites during atherogenesis. Atherosclerosis was evaluated in CD11d-/-/ApoE-/- mice after 16 wk on a Western diet. CD11d deficiency led to a marked reduction in lipid deposition in aortas and isolated macrophages. Macrophage numbers in aortic sinuses of CD11d-/- mice were reduced without affecting their apoptosis and proliferation. Adoptive transfer of fluorescently labeled wild-type and CD11d-/- monocytes into ApoE-/- mice demonstrated similar recruitment from circulation, but reduced accumulation of CD11d-/- macrophages within the aortas. Furthermore, CD11d expression was significantly upregulated on macrophages in atherosclerotic lesions and M1 macrophages in vitro. Interestingly, expression of the related ligand-sharing integrin CD11b was not altered. This difference defines their distinct roles in the regulation of macrophage migration. CD11d-deficient M1 macrophages demonstrated improved migration in a three-dimensional fibrin matrix and during resolution of peritoneal inflammation, whereas migration of CD11b-/- M1 macrophages was not affected. These results prove the contribution of high densities of CD11d to macrophage arrest during atherogenesis. Because high expression of CD11d was detected in several inflammation-dependent diseases, we suggest that CD11d/CD18 upregulation on proinflammatory macrophages may represent a common mechanism for macrophage retention at inflammatory sites, thereby promoting chronic inflammation and disease development.


Subject(s)
Atherosclerosis/immunology , Blood Vessels/pathology , CD11 Antigens/genetics , CD18 Antigens/genetics , Integrin alpha Chains/genetics , Macrophages/immunology , Animals , Aorta/immunology , Aorta/pathology , Apolipoproteins E/deficiency , Atherosclerosis/etiology , Atherosclerosis/pathology , Blood Vessels/immunology , CD11 Antigens/immunology , CD18 Antigens/immunology , Diet, Western , Humans , Inflammation/pathology , Inflammation Mediators/metabolism , Integrin alpha Chains/deficiency , Integrin alpha Chains/immunology , Macrophages/metabolism , Macrophages/pathology , Mice , Mice, Knockout , Peritonitis/immunology , Peritonitis/pathology , Transcriptional Activation , Up-Regulation
7.
Am J Physiol Heart Circ Physiol ; 311(1): H96-H106, 2016 07 01.
Article in English | MEDLINE | ID: mdl-27199132

ABSTRACT

Excessive cardiac interstitial fibrosis impairs normal cardiac function. We have shown that the α11ß1 (α11) integrin mediates fibrotic responses to glycated collagen in rat myocardium by a pathway involving transforming growth factor-ß. Little is known of the role of the α11 integrin in the developing mammalian heart. Therefore, we examined the impact of deletion of the α11 integrin in wild-type mice and in mice treated with streptozotocin (STZ) to elucidate the role of the α11 integrin in normal cardiac homeostasis and in the pathogenesis of diabetes-related fibrosis. As anticipated, cardiac fibrosis was reduced in α11 integrin knockout mice (α11(-/-); C57BL/6 background) treated with STZ compared with STZ-treated wild-type mice (P < 0.05). Unexpectedly, diastolic function was impaired in both vehicle and STZ-treated α11(-/-) mice, as shown by the decreased minimum rate of pressure change and prolonged time constant of relaxation in association with increased end-diastolic pressure (all P < 0.05 compared with wild-type mice). Accordingly, we examined the phenotype of untreated α11(-/-) mice, which demonstrated a reduced cardiomyocyte cross-sectional cell area and myofibril thickness (all P < 0.05 compared with wild-type mice) and impaired myofibril arrangement. Immunostaining for desmin and connexin 43 showed abnormal intermediate filament organization at intercalated disks and impaired gap-junction development. Overall, deletion of the α11 integrin attenuates cardiac fibrosis in the mammalian mouse heart and reduces ECM formation as a result of diabetes. Furthermore, α11 integrin deletion impairs cardiac function and alters cardiomyocyte morphology. These findings shed further light on the poorly understood interaction between the fibroblast-cardiomyocyte and the ECM.


Subject(s)
Diabetic Cardiomyopathies/metabolism , Extracellular Matrix/metabolism , Fibroblasts/metabolism , Integrin alpha Chains/metabolism , Myocytes, Cardiac/metabolism , Animals , Cell Size , Connexin 43/metabolism , Desmin/metabolism , Diabetes Mellitus, Experimental/genetics , Diabetes Mellitus, Experimental/metabolism , Diabetic Cardiomyopathies/genetics , Diabetic Cardiomyopathies/pathology , Diabetic Cardiomyopathies/physiopathology , Female , Fibroblasts/pathology , Fibrosis , Genotype , Integrin alpha Chains/deficiency , Integrin alpha Chains/genetics , Male , Mice, Inbred C57BL , Mice, Knockout , Myocytes, Cardiac/pathology , Myofibrils/metabolism , Myofibrils/pathology , Phenotype , Signal Transduction , Streptozocin , Stroke Volume , Ventricular Dysfunction, Left/genetics , Ventricular Dysfunction, Left/metabolism , Ventricular Dysfunction, Left/physiopathology , Ventricular Function, Left , Ventricular Pressure , Ventricular Remodeling
8.
Virulence ; 7(7): 826-35, 2016 10 02.
Article in English | MEDLINE | ID: mdl-27191829

ABSTRACT

Dendritic cells (DCs) are critical for defense against a variety of pathogens and the formation of adaptive immune responses. The transcription factor Batf3 is critical for the development of CD103(+)CD11b(-) DCs, which promote IL-12-dependent protective immunity during viral and parasitic infections, dampen Th2 immunity during helminthic infection, and exert detrimental effects during bacterial infection. Whether CD103(+) DCs modulate immunity during systemic or mucosal fungal disease remains unknown. Herein, we report that Batf3 is critical for accumulation of CD103(+) DCs in the kidney and tongue at steady state, for their expansion during systemic and oropharyngeal candidiasis, and for tissue-specific production of IL-12 in kidney but not tongue during systemic and oropharyngeal candidiasis, respectively. Importantly, deficiency of CD103(+) DCs does not impair survival or fungal clearance during systemic or oropharyngeal candidiasis, indicating that Batf3-dependent CD103(+) DC accumulation mediates pathogen- and tissue-specific immune effects.


Subject(s)
Antigens, CD/analysis , Basic-Leucine Zipper Transcription Factors/immunology , Candidiasis, Invasive/immunology , Candidiasis, Oral/immunology , Dendritic Cells/immunology , Integrin alpha Chains/analysis , Repressor Proteins/immunology , Animals , Basic-Leucine Zipper Transcription Factors/genetics , Candidiasis, Invasive/microbiology , Candidiasis, Oral/microbiology , Cell Proliferation , Dendritic Cells/physiology , Immunity, Innate , Integrin alpha Chains/deficiency , Interleukin-12/biosynthesis , Interleukin-12/immunology , Kidney/immunology , Mice , Mice, Inbred C57BL , Repressor Proteins/genetics , Tongue/immunology
9.
J Immunol ; 196(9): 3665-76, 2016 05 01.
Article in English | MEDLINE | ID: mdl-27009492

ABSTRACT

Resting central Tregs (cTregs) and activated effector Tregs (eTregs) are required for self-tolerance, but the heterogeneity and relationships within and between phenotypically distinct subsets of cTregs and eTregs are poorly understood. By extensive immune profiling and deep sequencing of TCR-ß V regions, two subsets of cTregs, based on expression of Ly-6C, and three subsets of eTregs, based on distinctive expression of CD62L, CD69, and CD103, were identified. Ly-6C(+) cTregs exhibited lower basal activation, expressed on average lower affinity TCRs, and less efficiently developed into eTregs when compared with Ly-6C(-) cTregs. The dominant TCR Vßs of Ly-6C(+) cTregs were shared by eTregs at a low frequency. A single TCR clonotype was also identified that was largely restricted to Ly-6C(+) cTregs, even under conditions that promoted the development of eTregs. Collectively, these findings indicate that some Ly-6C(+) cTregs may persist as a lymphoid-specific subset, with minimal potential to develop into highly activated eTregs, whereas other cTregs readily develop into eTregs. In contrast, subsets of CD62L(lo) eTregs showed higher clonal expansion and were more highly interrelated than cTreg subsets based on their TCR-ß repertoires, but exhibited varied immune profiles. The CD62L(lo) CD69(-) CD103(-) eTreg subset displayed properties of a transitional intermediate between cTregs and more activated eTreg subsets. Thus, eTreg subsets appear to exhibit substantial flexibility, most likely in response to environmental cues, to adopt defined immune profiles that are expected to optimize suppression of autoreactive T cells.


Subject(s)
Genes, T-Cell Receptor beta , Self Tolerance , T-Lymphocyte Subsets/immunology , T-Lymphocytes, Regulatory/immunology , Animals , Antigens, CD/genetics , Antigens, Differentiation, T-Lymphocyte/genetics , Antigens, Ly/genetics , High-Throughput Nucleotide Sequencing , Integrin alpha Chains/deficiency , Integrin alpha Chains/genetics , L-Selectin/genetics , Lectins, C-Type/deficiency , Lectins, C-Type/genetics , Mice , T-Lymphocyte Subsets/physiology
10.
Gastroenterology ; 148(7): 1417-26, 2015 Jun.
Article in English | MEDLINE | ID: mdl-25747597

ABSTRACT

BACKGROUND & AIMS: Intraepithelial lymphocytes that express the γδ T-cell receptor (γδ IELs) limit pathogen translocation across the intestinal epithelium by unknown mechanisms. We investigated whether γδ IEL migration and interaction with epithelial cells promote mucosal barrier maintenance during enteric infection. METHODS: Salmonella typhimurium or Toxoplasma gondii were administered to knockout (KO) mice lacking either the T cell receptor δ chain (Tcrd) or CD103, or control TcrdEGFP C57BL/6 reporter mice. Intravital microscopy was used to visualize migration of green fluorescent protein (GFP)-tagged γδ T cells within the small intestinal mucosa of mice infected with DsRed-labeled S typhimurium. Mixed bone marrow chimeras were generated to assess the effects of γδ IEL migration on early pathogen invasion and chronic systemic infection. RESULTS: Morphometric analyses of intravital video microscopy data showed that γδ IELs rapidly localized to and remained near epithelial cells in direct contact with bacteria. Within 1 hour, greater numbers of T gondii or S typhimurium were present within mucosae of mice with migration-defective occludin KO γδ T cells, compared with controls. Pathogen invasion in Tcrd KO mice was quantitatively similar to that in mice with occludin-deficient γδ T cells, whereas invasion in CD103 KO mice, which have increased migration of γδ T cells into the lateral intercellular space, was reduced by 63%. Consistent with a role of γδ T-cell migration in early host defense, systemic salmonellosis developed more rapidly and with greater severity in mice with occludin-deficient γδ IELs, relative to those with wild-type or CD103 KO γδ IELs. CONCLUSIONS: In mice, intraepithelial migration to epithelial cells in contact with pathogens is essential to γδ IEL surveillance and immediate host defense. γδ IEL occludin is required for early surveillance that limits systemic disease.


Subject(s)
Bacterial Translocation , Chemotaxis, Leukocyte , Epithelial Cells/immunology , Intestinal Mucosa/immunology , Lymphocytes/immunology , Receptors, Antigen, T-Cell, gamma-delta/metabolism , Salmonella Infections, Animal/immunology , Salmonella typhimurium/pathogenicity , Toxoplasmosis, Animal/immunology , Animals , Antigens, CD/genetics , Bone Marrow Transplantation , Disease Models, Animal , Epithelial Cells/metabolism , Epithelial Cells/microbiology , Epithelial Cells/parasitology , Host-Pathogen Interactions , Immunity, Innate , Integrin alpha Chains/deficiency , Integrin alpha Chains/genetics , Intestinal Mucosa/metabolism , Intestinal Mucosa/microbiology , Intestinal Mucosa/parasitology , Lymphocytes/metabolism , Lymphocytes/microbiology , Lymphocytes/parasitology , Mice, Inbred C57BL , Mice, Knockout , Occludin/deficiency , Occludin/drug effects , Permeability , Receptors, Antigen, T-Cell, gamma-delta/deficiency , Receptors, Antigen, T-Cell, gamma-delta/drug effects , Salmonella Infections, Animal/genetics , Salmonella Infections, Animal/metabolism , Salmonella Infections, Animal/microbiology , Salmonella typhimurium/immunology , Time Factors , Toxoplasmosis, Animal/genetics , Toxoplasmosis, Animal/parasitology , Transplantation Chimera , Virulence
11.
Nat Immunol ; 16(4): 406-14, 2015 Apr.
Article in English | MEDLINE | ID: mdl-25706747

ABSTRACT

We report that oral infection with Yersinia pseudotuberculosis results in the development of two distinct populations of pathogen-specific CD8(+) tissue-resident memory T cells (TRM cells) in the lamina propria. CD103(-) T cells did not require transforming growth factor-ß (TGF-ß) signaling but were true resident memory cells. Unlike CD103(+)CD8(+) T cells, which were TGF-ß dependent and were scattered in the tissue, CD103(-)CD8(+) T cells clustered with CD4(+) T cells and CX3CR1(+) macrophages and/or dendritic cells around areas of bacterial infection. CXCR3-dependent recruitment of cells to inflamed areas was critical for development of the CD103(-) population and pathogen clearance. Our studies have identified the 'preferential' development of CD103(-) TRM cells in inflammatory microenvironments within the lamina propria and suggest that this subset has a critical role in controlling infection.


Subject(s)
Antigens, CD/immunology , CD8-Positive T-Lymphocytes/immunology , Integrin alpha Chains/immunology , Intestinal Mucosa/immunology , Yersinia pseudotuberculosis Infections/immunology , Animals , Antigens, CD/genetics , CD4-Positive T-Lymphocytes/immunology , CD4-Positive T-Lymphocytes/microbiology , CD4-Positive T-Lymphocytes/pathology , CD8-Positive T-Lymphocytes/microbiology , CD8-Positive T-Lymphocytes/pathology , Cell Movement , Cellular Microenvironment , Dendritic Cells/immunology , Dendritic Cells/microbiology , Dendritic Cells/pathology , Gene Expression Regulation , Immunologic Memory , Immunophenotyping , Integrin alpha Chains/deficiency , Integrin alpha Chains/genetics , Intestinal Mucosa/microbiology , Intestinal Mucosa/pathology , Macrophages/immunology , Macrophages/microbiology , Macrophages/pathology , Mice , Mice, Inbred C57BL , Mice, Knockout , Receptors, CXCR3/genetics , Receptors, CXCR3/immunology , Signal Transduction , Transforming Growth Factor beta/genetics , Transforming Growth Factor beta/immunology , Yersinia pseudotuberculosis/immunology , Yersinia pseudotuberculosis Infections/genetics , Yersinia pseudotuberculosis Infections/microbiology , Yersinia pseudotuberculosis Infections/pathology
12.
Proc Natl Acad Sci U S A ; 111(9): E856-65, 2014 Mar 04.
Article in English | MEDLINE | ID: mdl-24550510

ABSTRACT

Intestinal CD103(-) dendritic cells (DCs) are pathogenic for colitis. Unveiling molecular mechanisms that render these cells proinflammatory is important for the design of specific immunotherapies. In this report, we demonstrated that mesenteric lymph node CD103(-) DCs express, among other proinflammatory cytokines, high levels of osteopontin (Opn) during experimental colitis. Opn expression by CD103(-) DCs was crucial for their immune profile and pathogenicity, including induction of T helper (Th) 1 and Th17 cell responses. Adoptive transfer of Opn-deficient CD103(-) DCs resulted in attenuated colitis in comparison to transfer of WT CD103(-) DCs, whereas transgenic CD103(-) DCs that overexpress Opn were highly pathogenic in vivo. Neutralization of secreted Opn expressed exclusively by CD103(-) DCs restrained disease severity. Also, Opn deficiency resulted in milder disease, whereas systemic neutralization of secreted Opn was therapeutic. We determined a specific domain of the Opn protein responsible for its CD103(-) DC-mediated proinflammatory effect. We demonstrated that disrupting the interaction of this Opn domain with integrin α9, overexpressed on colitic CD103(-) DCs, suppressed the inflammatory potential of these cells in vitro and in vivo. These results add unique insight into the biology of CD103(-) DCs and their function during inflammatory bowel disease.


Subject(s)
Colitis/immunology , Dendritic Cells/metabolism , Osteopontin/metabolism , Adoptive Transfer , Animals , Antibodies, Neutralizing/immunology , Antigens, CD , Colitis/physiopathology , DNA Primers/genetics , Flow Cytometry , Integrin alpha Chains/deficiency , Integrins/metabolism , Lymph Nodes/metabolism , Male , Mice , Mice, Inbred C57BL , Osteopontin/genetics , Protein Structure, Tertiary , Reverse Transcriptase Polymerase Chain Reaction , Trinitrobenzenesulfonic Acid
13.
J Immunol ; 190(8): 4305-14, 2013 Apr 15.
Article in English | MEDLINE | ID: mdl-23509351

ABSTRACT

Integrins are adhesion molecules critical for the recruitment of leukocytes from blood into peripheral tissues. However, whether integrins are also involved in leukocyte exit from peripheral tissues via afferent lymphatics to the draining lymph node remains poorly understood. In this article, we show that adhesion by the collagen IV-binding integrin α1ß1 unexpectedly inhibited macrophage exit from inflamed skin. We monitored macrophages exiting mouse footpads using a newly developed in situ pulse labeling technique. Blockade of α1ß1 integrin or genetic deletion (Itga1(-/-)) increased macrophage exit efficiency. Chemotaxis assays through collagen IV showed more efficient migration of Itga1(-/-) macrophages relative to wild type. Given that macrophages are key orchestrators of inflammation, α1ß1 integrin adhesion may represent a mechanism for regulating inflammatory responses by controlling macrophage exit or persistence in inflamed tissues.


Subject(s)
Cell Migration Inhibition/immunology , Inflammation Mediators/physiology , Integrin alpha1beta1/physiology , Macrophages, Peritoneal/immunology , Macrophages, Peritoneal/pathology , Adaptor Proteins, Signal Transducing , Animals , Antigens, CD/biosynthesis , Antigens, CD/genetics , Cell Adhesion/genetics , Cell Adhesion/immunology , Cell Migration Inhibition/genetics , Foot , Inflammation/immunology , Inflammation/metabolism , Inflammation/pathology , Inflammation Mediators/metabolism , Integrin alpha Chains/biosynthesis , Integrin alpha Chains/deficiency , Integrin alpha Chains/genetics , Integrin alpha1beta1/biosynthesis , Integrin alpha1beta1/deficiency , Lymph Nodes/immunology , Lymph Nodes/metabolism , Lymph Nodes/pathology , Macrophages, Peritoneal/metabolism , Mice , Mice, Inbred C57BL , Mice, Knockout , Monocytes/immunology , Monocytes/metabolism , Monocytes/pathology , Nerve Tissue Proteins/biosynthesis , Nerve Tissue Proteins/deficiency , Nerve Tissue Proteins/physiology
14.
Mucosal Immunol ; 5(6): 646-57, 2012 Nov.
Article in English | MEDLINE | ID: mdl-22643849

ABSTRACT

Human immunodeficiency virus (HIV) and Simian immunodeficiency virus (SIV) disease progression is associated with multifocal damage to the gastrointestinal tract epithelial barrier that correlates with microbial translocation and persistent pathological immune activation, but the underlying mechanisms remain unclear. Investigating alterations in mucosal immunity during SIV infection, we found that damage to the colonic epithelial barrier was associated with loss of multiple lineages of interleukin (IL)-17-producing lymphocytes, cells that microarray analysis showed expressed genes important for enterocyte homeostasis, including IL-22. IL-22-producing lymphocytes were also lost after SIV infection. Potentially explaining coordinate loss of these distinct populations, we also observed loss of CD103+ dendritic cells (DCs) after SIV infection, which associated with the loss of IL-17- and IL-22-producing lymphocytes. CD103+ DCs expressed genes associated with promotion of IL-17/IL-22+ cells, and coculture of CD103+ DCs and naïve T cells led to increased IL17A and RORc expression in differentiating T cells. These results reveal complex interactions between mucosal immune cell subsets providing potential mechanistic insights into mechanisms of mucosal immune dysregulation during HIV/SIV infection, and offer hints for development of novel therapeutic strategies to address this aspect of AIDS virus pathogenesis.


Subject(s)
Colon/immunology , Dendritic Cells/immunology , Enterocytes/immunology , Immunity, Mucosal , Simian Acquired Immunodeficiency Syndrome/immunology , Simian Immunodeficiency Virus/physiology , Th17 Cells/immunology , Animals , Antigens, CD/immunology , Cell Differentiation , Cell Lineage , Coculture Techniques , Colon/pathology , Colon/virology , Dendritic Cells/pathology , Dendritic Cells/virology , Enterocytes/pathology , Enterocytes/virology , Gene Expression Regulation , Integrin alpha Chains/deficiency , Integrin alpha Chains/immunology , Interleukin-17/deficiency , Interleukin-17/genetics , Interleukin-17/immunology , Interleukins/deficiency , Interleukins/genetics , Interleukins/immunology , Macaca mulatta , Nuclear Receptor Subfamily 1, Group F, Member 3/genetics , Nuclear Receptor Subfamily 1, Group F, Member 3/immunology , Simian Acquired Immunodeficiency Syndrome/pathology , Simian Acquired Immunodeficiency Syndrome/virology , Th17 Cells/pathology , Th17 Cells/virology , Interleukin-22
15.
J Immunol ; 188(10): 4921-30, 2012 May 15.
Article in English | MEDLINE | ID: mdl-22508930

ABSTRACT

Dendritic cells (DC) play important roles in both tolerance and immunity to ß cells in type 1 diabetes. How and why DC can have diverse and opposing functions in islets remains elusive. To answer these questions, islet DC subsets and their specialized functions were characterized. Under both homeostatic and inflammatory conditions, there were two main tissue-resident DC subsets in islets, defined as CD11b(lo/-)CD103(+)CX3CR1(-) (CD103(+) DC), the majority of which were derived from fms-like tyrosine kinase 3-dependent pre-DC, and CD11b(+)CD103(-)CX3CR1(+) (CD11b(+) DC), the majority of which were derived from monocytes. CD103(+) DC were the major migratory DC and cross-presented islet-derived Ag in the pancreatic draining lymph node, although this DC subset displayed limited phagocytic activity. CD11b(+) DC were numerically the predominant subset (60-80%) but poorly migrated to the draining lymph node. Although CD11b(+) DC had greater phagocytic activity, they poorly presented Ag to T cells. CD11b(+) DC increased in numbers and percentage during T cell-mediated insulitis, suggesting that this subset might be involved in the pathogenesis of diabetes. These data elucidate the phenotype and function of homeostatic and inflammatory islet DC, suggesting differential roles in islet immunity.


Subject(s)
Antigen Presentation/immunology , Dendritic Cells/immunology , Dendritic Cells/metabolism , Islets of Langerhans/immunology , Islets of Langerhans/metabolism , Animals , Antigen Presentation/genetics , Antigens, CD/physiology , Cell Movement/genetics , Cell Movement/immunology , Cells, Cultured , Coculture Techniques , Dendritic Cells/pathology , Inflammation/genetics , Inflammation/immunology , Inflammation/pathology , Integrin alpha Chains/deficiency , Integrin alpha Chains/physiology , Islets of Langerhans/pathology , Male , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Mice, Knockout , Mice, Mutant Strains , Mice, Transgenic , Phagocytes/immunology , Phagocytes/pathology
16.
J Exp Med ; 209(1): 139-55, 2012 Jan 16.
Article in English | MEDLINE | ID: mdl-22231304

ABSTRACT

Dendritic cells (DCs) and macrophages (MPs) are important for immunological homeostasis in the colon. We found that F4/80(hi)CX3CR1(hi) (CD11b(+)CD103(-)) cells account for 80% of mouse colonic lamina propria MHC-II(hi) cells. Both CD11c(+) and CD11c(-) cells within this population were identified as MPs based on multiple criteria, including an MP transcriptome revealed by microarray analysis. These MPs constitutively released high levels of IL-10 at least partially in response to the microbiota via an MyD88-independent mechanism. In contrast, cells expressing low to intermediate levels of F4/80 and CX3CR1 were identified as DCs based on phenotypic and functional analysis and comprise three separate CD11c(hi) cell populations: CD103(+)CX3CR1(-)CD11b(-) DCs, CD103(+)CX3CR1(-)CD11b(+) DCs, and CD103(-)CX3CR1(int)CD11b(+) DCs. In noninflammatory conditions, Ly6C(hi) monocytes (MOs) differentiated primarily into CD11c(+) but not CD11c(-) MPs. In contrast, during colitis, Ly6C(hi) MOs massively invaded the colon and differentiated into proinflammatory CD103(-)CX3CR1(int)CD11b(+) DCs, which produced high levels of IL-12, IL-23, iNOS, and TNF. These findings demonstrate the dual capacity of Ly6C(hi) blood MOs to differentiate into either regulatory MPs or inflammatory DCs in the colon and that the balance of these immunologically antagonistic cell types is dictated by microenvironmental conditions.


Subject(s)
Cell Differentiation/immunology , Colon/immunology , Dendritic Cells/immunology , Inflammation/immunology , Macrophages/immunology , Monocytes/cytology , Adoptive Transfer , Animals , Antigens, CD/genetics , Antigens, Ly/analysis , Colon/metabolism , Cytokines/biosynthesis , Cytokines/immunology , Dendritic Cells/cytology , Inflammation/metabolism , Inflammation Mediators/immunology , Inflammation Mediators/metabolism , Integrin alpha Chains/deficiency , Integrin alpha Chains/genetics , Interferon-gamma/biosynthesis , Interleukin-10/biosynthesis , Interleukin-10/immunology , Lipopolysaccharides/immunology , Macrophages/cytology , Mice , Mice, Inbred C57BL , Mice, Knockout , Mucous Membrane/immunology , Mucous Membrane/metabolism , T-Lymphocytes/immunology , T-Lymphocytes/metabolism , Transposases/genetics
17.
PLoS One ; 7(12): e52519, 2012.
Article in English | MEDLINE | ID: mdl-23300694

ABSTRACT

In all metazoa, the response of cells to molecular stimuli from their environment represents a fundamental principle of regulatory processes controlling cell growth and differentiation. Among the membrane-linked receptors mediating extracellular communication processes are integrin receptors. Besides managing adhesion to the extracellular matrix or to other cells, they arrange information flow into the cells by activating intracellular signaling pathways often acting synergistically through cooperation with growth factor receptors. Although a wealth of information exists on integrins in different model organisms, there is a big gap of knowledge for platyhelminths. Here we report on the in silico detection and reconstruction of α and ß integrins from free-living and parasitic platyhelminths, which according to structural and phylogenetic analyses form specific clades separate from each other and from further metazoan integrins. As representative orthologs of parasitic platyhelminths we have cloned one beta-integrin (Smß-Int1) and four alpha-integrins (Smα-Int1 - Smα-Int4) from Schistosoma mansoni; they were characterized by molecular and biochemical analyses. Evidence is provided that Smß-Int1 interacts and co-localizes in the reproductive organs with known schistosome cellular tyrosine kinases (CTKs), of which the Syk kinase SmTK4 appeared to be the strongest interaction partner as shown by yeast two-hybrid analyses and coimmunoprecipitation experiments. By a novel RNAi approach with adult schistosomes in vitro we demonstrate for the first time multinucleated oocytes in treated females, indicating a decisive role Smß-Int1 during oogenesis as phenotypically analyzed by confocal laser scanning microscopy (CLSM). Our findings provide a first comprehensive overview about platyhelminth integrins, of which the parasite group exhibits unique features allowing a clear distinction from the free-living groups. Furthermore, we shed first lights on the functions of integrins in a trematode model parasite, revealing the complexity of molecular processes involved in its reproductive biology, which may be representative for other platyhelminths.


Subject(s)
Integrin alpha Chains/metabolism , Integrin beta Chains/metabolism , Schistosoma mansoni/metabolism , Animals , Cloning, Molecular , Female , Integrin alpha Chains/deficiency , Integrin alpha Chains/genetics , Integrin beta Chains/genetics , Male , Oogenesis , Ovary/metabolism , Ovary/physiology , Protein-Tyrosine Kinases/genetics , Protein-Tyrosine Kinases/metabolism , RNA, Small Interfering/genetics , Schistosoma mansoni/genetics , Schistosoma mansoni/physiology , Sequence Analysis , Species Specificity , Testis/metabolism , Testis/physiology , Transcription, Genetic
18.
PLoS One ; 6(7): e21968, 2011.
Article in English | MEDLINE | ID: mdl-21779359

ABSTRACT

BACKGROUND: Graft-versus-host disease (GVHD) remains the main barrier to broader application of allogeneic hematopoietic stem cell transplantation (alloSCT) as a curative therapy for host malignancy. GVHD is mediated by allogeneic T cells directed against histocompatibility antigens expressed by host tissues. Based on previous studies, we postulated that the integrin CD103 is required for CD8-mediated GVHD, but not for graft-versus-tumor effects (GVT). METHODOLOGY/PRINCIPAL FINDINGS: We herein provide evidence in support of this hypothesis. To circumvent the potentially confounding influence of donor CD4 T cells, we developed an alloSCT model in which GVHD mortality is mediated by purified CD8 T cells. In this model, host-reactive CD8 T cells receive CD4 T cell help at the time of initial activation but not in the effector phase in which mature CD8 T effectors migrate into host tissues. We show that donor CD8 T cells from wild-type BALB/c mice primed to host alloantigens induce GVHD pathology and eliminate tumors of host origin in the absence of host CD4 T cells. Importantly, CD103 deficiency dramatically attenuated GVHD mortality, but had no detectable impact on the capacity to eliminate a tumor line of host origin. We provide evidence that CD103 is required for accumulation of donor CD8 T cells in the host intestinal epithelium but not in the tumor or host lymphoid compartments. Consistent with these data, CD103 was preferentially expressed by CD8 T cells infiltrating the host intestinal epithelium but not by those infiltrating the tumor, lamina propria, or lymphoid compartments. We further demonstrate that CD103 expression is not required for classic CD8 effector activities including cytokine production and cytotoxicity. CONCLUSIONS/SIGNIFICANCE: These data indicate that CD103 deficiency inhibits GVHD pathology while sparing anti-tumor effects mediated by CD8 T cells, identifying CD103 blockade as an improved strategy for GVHD prophylaxis.


Subject(s)
CD8-Positive T-Lymphocytes/immunology , Graft vs Host Disease/immunology , Graft vs Host Disease/prevention & control , Graft vs Tumor Effect/immunology , Integrin alpha Chains/deficiency , Animals , Antigens, CD , Flow Cytometry , Isoantigens/immunology , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL
19.
J Cell Sci ; 124(Pt 13): 2287-97, 2011 Jul 01.
Article in English | MEDLINE | ID: mdl-21652631

ABSTRACT

Merosin-deficient congenital muscular dystrophy 1A (MDC1A) is a devastating neuromuscular disease that results in children being confined to a wheelchair, requiring ventilator assistance to breathe and premature death. MDC1A is caused by mutations in the LAMA2 gene, which results in the partial or complete loss of laminin-211 and laminin-221, the major laminin isoforms found in the basal lamina of skeletal muscle. MDC1A patients exhibit reduced α7ß1 integrin; however, it is unclear how the secondary loss of α7ß1 integrin contributes to MDC1A disease progression. To investigate whether restoring α7 integrin expression can alleviate the myopathic phenotype observed in MDC1A, we produced transgenic mice that overexpressed the α7 integrin in the skeletal muscle of the dy(W⁻/⁻) mouse model of MDC1A. Enhanced expression of the α7 integrin restored sarcolemmal localization of the α7ß1 integrin to laminin-α2-deficient myofibers, changed the composition of the muscle extracellular matrix, reduced muscle pathology, maintained muscle strength and function and improved the life expectancy of dy(W⁻/⁻) mice. Taken together, these results indicate that enhanced expression of α7 integrin prevents muscle disease progression through augmentation and/or stabilization of the existing extracellular matrix in laminin-α2-deficient mice, and strategies that increase α7 integrin in muscle might provide an innovative approach for the treatment of MDC1A.


Subject(s)
Antigens, CD/biosynthesis , Integrin alpha Chains/biosynthesis , Laminin/metabolism , Muscular Dystrophy, Animal/metabolism , Animals , Disease Progression , Extracellular Matrix/metabolism , Fluorescent Antibody Technique , Integrin alpha Chains/deficiency , Laminin/deficiency , Laminin/genetics , Mice , Mice, Transgenic , Muscle Strength , Muscle, Skeletal/pathology , Muscular Dystrophies/genetics , Muscular Dystrophies/metabolism , Muscular Dystrophy, Animal/pathology , Muscular Dystrophy, Animal/physiopathology , Polymerase Chain Reaction
20.
PLoS One ; 6(5): e19580, 2011 May 05.
Article in English | MEDLINE | ID: mdl-21573179

ABSTRACT

BACKGROUND: In the intestine, the integrin CD103 is expressed on a subset of T regulatory (T(reg)) cells and a population of dendritic cells (DCs) that produce retinoic acid and promote immune homeostasis. However, the role of CD103 during intestinal helminth infection has not been tested. METHODOLOGY/PRINCIPAL FINDINGS: We demonstrate that CD103 is dispensable for the development of protective immunity to the helminth parasite Trichuris muris. While we observed an increase in the frequency of CD103(+) DCs in the lamina propria (LP) following acute high-dose infection with Trichuris, lack of CD103 had no effect on the frequency of CD11c(+) DCs in the LP or mesenteric lymph nodes (mLN). CD103-deficient (CD103(-/-)) mice develop a slightly increased and earlier T cell response but resolve infection with similar kinetics to control mice. Similarly, low-dose chronic infection of CD103(-/-) mice with Trichuris resulted in no significant difference in immunity or parasite burden. Absence of CD103 also had no effect on the frequency of CD4(+)CD25(+)Foxp3(+) T(reg) cells in the mLN or LP. CONCLUSIONS/SIGNIFICANCE: These results suggest that CD103 is dispensable for intestinal immunity during helminth infection. Furthermore, lack of CD103 had no effect on DC or T(reg) recruitment or retention within the large intestine.


Subject(s)
Antigens, CD/immunology , Immunity/immunology , Integrin alpha Chains/immunology , Intestines/immunology , Intestines/parasitology , Parasites/immunology , Trichuris/immunology , Animals , Cell Movement , Chronic Disease , Dendritic Cells/cytology , Dendritic Cells/immunology , Integrin alpha Chains/deficiency , Intestines/pathology , Mice , Mice, Inbred C57BL , T-Lymphocytes, Regulatory/cytology , T-Lymphocytes, Regulatory/immunology , Trichuriasis/immunology , Trichuriasis/parasitology
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