RESUMO
Introduction: Spontaneous cervical artery dissection (sCAD) is a rare vasculopathy whose trigger is still unknown. We hypothesized that autoimmunity against components of the vascular wall might play a critical role in sCAD and examined anti-collagen type I antibodies in patients with sCAD, acute ischemic stroke, patients with thromboendarterectomy, and controls. Methods: Fifty-seven patients with sCAD (age 45.7 ± 10.2 years, female 18 (31.6%)) were prospectively enrolled in four German stroke centers. Blood samples were collected at baseline, at day 10 ± 3, and after 6 ± 1 months. Patients with ischemic stroke not related to CAD (n=54, age 56.7 ± 13.7 years, female 15 (27.8%)), healthy probands (n=80, age 57.4 ± 12.9 years, female 56 (70%)), and patients undergoing thromboendarterectomy of the carotid artery (n=9, age 70.7 ± 9.3 years, female 2 (22.2%)) served as controls. Anti-collagen type I antibodies were determined by enzyme-linked immunosorbent assays (ELISAs). Results: Patients with acute sCAD had higher serum levels of anti-collagen type I antibodies (33.9 ± 24.6 µg/ml) than probands (18.5 ± 11.0 µg/ml; p <0.001) but lower levels than patients with ischemic stroke not related to sCAD (47.8 ± 28.4 µg/ml; p=0.003). In patients with sCAD, serum levels of anti-collagen type I antibodies were similar in the acute, subacute, and chronic phase. Levels of anti-collagen type I antibodies significantly correlated with circulating collagen type I (rho=0.207, p=0.003). Conclusion: Anti-collagen type I antibodies seem not to represent a trigger for acute sCAD or ischemic stroke but may rather be linked to the metabolism and turnover of collagen type I.
Assuntos
Autoanticorpos , Colágeno Tipo I , AVC Isquêmico , Humanos , Feminino , Masculino , Pessoa de Meia-Idade , Colágeno Tipo I/imunologia , Colágeno Tipo I/sangue , Estudos Prospectivos , AVC Isquêmico/imunologia , AVC Isquêmico/sangue , Adulto , Idoso , Autoanticorpos/sangue , Autoanticorpos/imunologia , Dissecação da Artéria Vertebral/imunologia , Dissecação da Artéria Vertebral/sangue , Dissecação da Artéria Vertebral/cirurgiaRESUMO
CD8+ T cell dysfunction impedes antitumor immunity in solid cancers, but the underlying mechanisms are diverse and poorly understood. Extracellular matrix (ECM) composition has been linked to impaired T cell migration and enhanced tumor progression; however, impacts of individual ECM molecules on T cell function in the tumor microenvironment (TME) are only beginning to be elucidated. Upstream regulators of aberrant ECM deposition and organization in solid tumors are equally ill-defined. Therefore, we investigated how ECM composition modulates CD8+ T cell function in undifferentiated pleomorphic sarcoma (UPS), an immunologically active desmoplastic tumor. Using an autochthonous murine model of UPS and data from multiple human patient cohorts, we discovered a multifaceted mechanism wherein the transcriptional coactivator YAP1 promotes collagen VI (COLVI) deposition in the UPS TME. In turn, COLVI induces CD8+ T cell dysfunction and immune evasion by remodeling fibrillar collagen and inhibiting T cell autophagic flux. Unexpectedly, collagen I (COLI) opposed COLVI in this setting, promoting CD8+ T cell function and acting as a tumor suppressor. Thus, CD8+ T cell responses in sarcoma depend on oncogene-mediated ECM composition and remodeling.
Assuntos
Linfócitos T CD8-Positivos , Matriz Extracelular , Sarcoma , Microambiente Tumoral , Proteínas de Sinalização YAP , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/patologia , Animais , Microambiente Tumoral/imunologia , Camundongos , Proteínas de Sinalização YAP/imunologia , Proteínas de Sinalização YAP/genética , Humanos , Matriz Extracelular/imunologia , Matriz Extracelular/metabolismo , Matriz Extracelular/patologia , Sarcoma/imunologia , Sarcoma/patologia , Sarcoma/genética , Sarcoma/metabolismo , Colágeno Tipo VI/genética , Colágeno Tipo VI/imunologia , Colágeno Tipo VI/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/imunologia , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Fatores de Transcrição/imunologia , Oncogenes , Proteínas de Neoplasias/imunologia , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Colágeno Tipo I/metabolismo , Colágeno Tipo I/genética , Colágeno Tipo I/imunologiaRESUMO
Systemic sclerosis (SSc) is a chronic multisystem disorder characterized by fibrosis and autoimmunity. Interleukin (IL)-31 has been implicated in fibrosis and T helper (Th) 2 immune responses, both of which are characteristics of SSc. The exact role of IL-31 in SSc pathogenesis is unclear. Here we show the overexpression of IL-31 and IL-31 receptor A (IL-31RA) in dermal fibroblasts (DFs) from SSc patients. We elucidate the dual role of IL-31 in SSc, where IL-31 directly promotes collagen production in DFs and indirectly enhances Th2 immune responses by increasing pro-Th2 cytokine expression in DFs. Furthermore, blockade of IL-31 with anti-IL-31RA antibody significantly ameliorates fibrosis and Th2 polarization in a mouse model of SSc. Therefore, in addition to defining IL-31 as a mediator of fibrosis and Th2 immune responses in SSc, our study provides a rationale for targeting the IL-31/IL-31RA axis in the treatment of SSc.
Assuntos
Fibroblastos/imunologia , Interleucinas/genética , Receptores de Interleucina/genética , Escleroderma Sistêmico/imunologia , Células Th2/imunologia , Adulto , Idoso , Animais , Anticorpos Monoclonais/farmacologia , Colágeno Tipo I/genética , Colágeno Tipo I/imunologia , Cadeia alfa 1 do Colágeno Tipo I , Modelos Animais de Doenças , Feminino , Fibroblastos/efeitos dos fármacos , Fibroblastos/patologia , Fibrose , Regulação da Expressão Gênica , Humanos , Interleucina-13/genética , Interleucina-13/imunologia , Interleucina-4/genética , Interleucina-4/imunologia , Interleucina-6/genética , Interleucina-6/imunologia , Interleucinas/imunologia , Masculino , Camundongos , Pessoa de Meia-Idade , Isoformas de Proteínas/genética , Isoformas de Proteínas/imunologia , Receptores de Interleucina/antagonistas & inibidores , Receptores de Interleucina/imunologia , Fator de Transcrição STAT1/genética , Fator de Transcrição STAT1/imunologia , Escleroderma Sistêmico/tratamento farmacológico , Escleroderma Sistêmico/genética , Escleroderma Sistêmico/patologia , Pele/efeitos dos fármacos , Pele/imunologia , Pele/patologia , Linfócitos T Reguladores/efeitos dos fármacos , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/patologia , Células Th2/efeitos dos fármacos , Células Th2/patologiaRESUMO
Footrot is a polymicrobial infectious disease in sheep causing severe lameness, leading to one of the industry's largest welfare problems. The complex etiology of footrot makes in situ or in vitro investigations difficult. Computational methods offer a solution to understanding the bacteria involved and how they may interact with the host, ultimately providing a way to identify targets for future hypothesis-driven investigative work. Here, we present the first combined global analysis of bacterial community transcripts together with the host immune response in healthy and diseased ovine feet during a natural polymicrobial infection state using metatranscriptomics. The intratissue and surface bacterial populations and the most abundant bacterial transcriptomes were analyzed, demonstrating that footrot-affected skin has reduced diversity and increased abundances of not only the causative bacterium Dichelobacter nodosus but also other species such as Mycoplasma fermentans and Porphyromonas asaccharolytica. Host transcriptomics reveals the suppression of biological processes related to skin barrier function, vascular functions, and immunosurveillance in unhealthy interdigital skin, supported by histological findings that type I collagen (associated with scar tissue formation) is significantly increased in footrot-affected interdigital skin compared to outwardly healthy skin. Finally, we provide some interesting indications of host and pathogen interactions associated with virulence genes and the host spliceosome, which could lead to the identification of future therapeutic targets.
Assuntos
Bactérias/imunologia , Pododermatite Necrótica dos Ovinos/imunologia , Interações Hospedeiro-Patógeno/imunologia , Imunidade/imunologia , Ovinos/imunologia , Animais , Colágeno Tipo I/imunologia , Pododermatite Necrótica dos Ovinos/microbiologia , Ovinos/microbiologia , Doenças dos Ovinos/imunologia , Doenças dos Ovinos/microbiologia , Pele/imunologia , Pele/microbiologia , Transcriptoma/imunologia , Virulência/imunologiaRESUMO
Fibrosis is a prominent feature of chronic allograft rejection, caused by an excessive production of matrix proteins, including collagen-1. Several cell types produce collagen-1, including mesenchymal fibroblasts and cells of hematopoietic origin. Here, we sought to determine whether tissue-resident donor-derived cells or allograft-infiltrating recipient-derived cells are responsible for allograft fibrosis, and whether hematopoietic cells contribute to collagen production. A fully MHC-mismatched mouse heterotopic heart transplantation model was used, with transient depletion of CD4+ T cells to prevent acute rejection. Collagen-1 was selectively knocked out in recipients or donors. In addition, collagen-1 was specifically deleted in hematopoietic cells. Tissue-resident macrophages were depleted using anti-CSF1R antibody. Allograft fibrosis and inflammation were quantified 20 days post-transplantation. Selective collagen-1 knock-out in recipients or donors showed that tissue-resident cells from donor hearts, but not infiltrating recipient-derived cells, are responsible for production of collagen-1 in allografts. Cell-type-specific knock-out experiments showed that hematopoietic tissue-resident cells in donor hearts substantially contributed to graft fibrosis. Tissue resident macrophages, however, were not responsible for collagen-production, as their deletion worsened allograft fibrosis. Donor-derived cells including those of hematopoietic origin determine allograft fibrosis, making them attractive targets for organ preconditioning to improve long-term transplantation outcomes.
Assuntos
Colágeno Tipo I/biossíntese , Rejeição de Enxerto/etiologia , Rejeição de Enxerto/metabolismo , Transplante de Coração/efeitos adversos , Doadores de Tecidos , Animais , Biomarcadores , Doença Crônica , Colágeno Tipo I/imunologia , Modelos Animais de Doenças , Suscetibilidade a Doenças , Rejeição de Enxerto/diagnóstico , Transplante de Coração/métodos , Imunofenotipagem , Camundongos , Camundongos Transgênicos , Transplante HomólogoRESUMO
Most rheumatic diseases, including rheumatoid arthritis (RA), are characterized by immune disorders that affect antibody activity. In the present study, using Dot blot and ELISA assay, we showed that patients with rheumatic disease produced significantly more antibodies against lipopolysaccharide (LPS) P. mirabilis O3 compared to healthy donors (p < 0.05), and affinity purified antibodies against LPS O3 may cross-react with collagen type I. It was demonstrated that purified of antibodies isolated from RA patients sera, reacted stronger with the collagen than healthy donors (p = 0.015), and cross-reaction was correlated with level of anti-citrullinated peptide antibodies (r = 0.7, p = 0.003). Moreover, using six different lipopolysaccharides were demonstrated the significant correlations in sera reactivity among lysine-containing lipopolysaccharides observed in patients' sera (p < 0.05). Using Attenuated Total Reflection Fourier Transform Infrared Spectroscopy (ATR-FTIR) it was shown that unique wavenumbers of sera spectra correlate with reactivity with lipopolysaccharides allowing distinguish patients from healthy blood donors. Antibodies adsorption by synthetic antigens shows that in patients' group anti-LPS O3 antibodies can be adsorbed by both amides of galacturonic acid and lysine or threonine, which suggests less specificity of antibodies binding with non-carbohydrate LPS component. The observed correlations suggest that non-carbohydrate components of LPS may be an important epitope for less specific anti-LPS antibodies, which might lead to cross-reactions and affect disease development.
Assuntos
Anticorpos Antibacterianos/sangue , Artrite Reumatoide/imunologia , Colágeno Tipo I/metabolismo , Lipopolissacarídeos/imunologia , Lisina/imunologia , Proteus mirabilis/imunologia , Anticorpos Antibacterianos/imunologia , Artrite Reumatoide/sangue , Artrite Reumatoide/microbiologia , Estudos de Casos e Controles , Colágeno Tipo I/imunologia , Reações Cruzadas , Epitopos/imunologia , Feminino , Humanos , Masculino , Pessoa de Meia-IdadeRESUMO
We studied the effect of KE and AED peptides on the expression of sirtuin-1, sirtuin-6, collagen I, cytokines (IL-1, TGF-ß), and transcription factor NF-κB in human skin fibroblasts during their replicative aging. Immunocytochemical analysis and confocal microscopy showed that KE peptide reduces the synthesis of factors of the inflammatory response IL-1, NF-κB, and TGF-ß and stimulates the synthesis of sirtuin-6. KE peptide normalizes the immunological function of human skin fibroblasts during their aging. AED peptide activates the synthesis of sirtuin-1, sirtuin-6, and collagen I in human skin fibroblasts during their replicative aging, which attests to its geroprotective effect.
Assuntos
Antioxidantes/farmacologia , Fibroblastos/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Fatores Imunológicos/farmacologia , Oligopeptídeos/farmacologia , Antioxidantes/síntese química , Senescência Celular/efeitos dos fármacos , Colágeno Tipo I/genética , Colágeno Tipo I/imunologia , Feminino , Fibroblastos/citologia , Fibroblastos/imunologia , Humanos , Fatores Imunológicos/síntese química , Interleucina-1/genética , Interleucina-1/imunologia , NF-kappa B/genética , NF-kappa B/imunologia , Oligopeptídeos/síntese química , Cultura Primária de Células , Transdução de Sinais , Sirtuína 1/genética , Sirtuína 1/imunologia , Sirtuínas/genética , Sirtuínas/imunologia , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta/imunologiaRESUMO
The commensal microbiota is a recognized enhancer of arterial thrombus growth. While several studies have demonstrated the prothrombotic role of the gut microbiota, the molecular mechanisms promoting arterial thrombus growth are still under debate. Here, we demonstrate that germ-free (GF) mice, which from birth lack colonization with a gut microbiota, show diminished static deposition of washed platelets to type I collagen compared with their conventionally raised (CONV-R) counterparts. Flow cytometry experiments revealed that platelets from GF mice show diminished activation of the integrin αIIbß3 (glycoprotein IIbIIIa) when activated by the platelet agonist adenosine diphosphate (ADP). Furthermore, washed platelets from Toll-like receptor-2 (Tlr2)-deficient mice likewise showed impaired static deposition to the subendothelial matrix component type I collagen compared with wild-type (WT) controls, a process that was unaffected by GPIbα-blockade but influenced by von Willebrand factor (VWF) plasma levels. Collectively, our results indicate that microbiota-triggered steady-state activation of innate immune pathways via TLR2 enhances platelet deposition to subendothelial matrix molecules. Our results link host colonization status with the ADP-triggered activation of integrin αIIbß3, a pathway promoting platelet deposition to the growing thrombus.
Assuntos
Difosfato de Adenosina/farmacologia , Plaquetas/efeitos dos fármacos , Colágeno Tipo I/genética , Complexo Glicoproteico GPIIb-IIIa de Plaquetas/genética , Trombose/microbiologia , Fator de von Willebrand/genética , Animais , Artérias/metabolismo , Artérias/patologia , Plaquetas/imunologia , Plaquetas/patologia , Adesão Celular/efeitos dos fármacos , Colágeno Tipo I/imunologia , Feminino , Microbioma Gastrointestinal/imunologia , Expressão Gênica , Vida Livre de Germes , Humanos , Imunidade Inata , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Complexo Glicoproteico GPIIb-IIIa de Plaquetas/agonistas , Complexo Glicoproteico GPIIb-IIIa de Plaquetas/imunologia , Complexo Glicoproteico GPIb-IX de Plaquetas/genética , Complexo Glicoproteico GPIb-IX de Plaquetas/imunologia , Cultura Primária de Células , Simbiose/imunologia , Trombose/genética , Trombose/imunologia , Trombose/patologia , Receptor 2 Toll-Like/deficiência , Receptor 2 Toll-Like/genética , Receptor 2 Toll-Like/imunologia , Fator de von Willebrand/imunologiaRESUMO
Collagen type I is a major constituent of animal bodies. It is found in large quantities in tendon, bone, skin, cartilage, blood vessels, bronchi, and the lung interstitium. It is also produced and accumulates in large amounts in response to certain inflammations such as lung fibrosis. Our understanding of the molecular organization of fibrillar collagen and cellular interaction motifs, such as those involved with immune-associated molecules, continues to be refined. In this study, antibodies raised against type I collagen were used to label intact D-periodic type I collagen fibrils and observed with atomic force microscopy (AFM), and X-ray diffraction (XRD) and immunolabeling positions were observed with both methods. The antibodies bind close to the C-terminal telopeptide which verifies the location and accessibility of both the major histocompatibility complex (MHC) class I (MHCI) binding domain and C-terminal telopeptide on the outside of the collagen fibril. The close proximity of the C-telopeptide and the MHC1 domain of type I collagen to fibronectin, discoidin domain receptor (DDR), and collagenase cleavage domains likely facilitate the interaction of ligands and receptors related to cellular immunity and the collagen-based Extracellular Matrix.
Assuntos
Colágeno Tipo I/metabolismo , Colágeno Tipo I/ultraestrutura , Receptores Imunológicos/imunologia , Animais , Sítios de Ligação , Colágeno Tipo I/química , Colágeno Tipo I/imunologia , Receptor com Domínio Discoidina 1/metabolismo , Módulo de Elasticidade , Análise de Fourier , Ouro/química , Imunoglobulinas/imunologia , Microscopia de Força Atômica , Peptídeos/metabolismo , Ratos Wistar , Espalhamento a Baixo Ângulo , Difração de Raios XRESUMO
AIMS: To date, the ROS-generating capacities of macrophages in different activation states have not been thoroughly compared. This study is aimed at determining the nature and levels of ROS generated following stimulation with common activators of M1 and M2 macrophages and investigating the potential for this to impact fibrosis. RESULTS: Human primary and THP-1 macrophages were treated with IFN-γ+LPS or IL-4-activating stimuli, and mRNA expression of established M1 (CXCL11, CCR7, IL-1ß) and M2 (MRC-1, CCL18, CCL22) markers was used to confirm activation. Superoxide generation was assessed by L-012-enhanced chemiluminescence and was increased in both M(IFN-γ+LPS) and M(IL-4) macrophages, as compared to unpolarised macrophages (MΦ). This signal was attenuated with NOX2 siRNA. Increased expression of the p47phox and p67phox subunits of the NOX2 oxidase complex was evident in M(IFN-γ+LPS) and M(IL-4) macrophages, respectively. Amplex Red and DCF fluorescence assays detected increased hydrogen peroxide generation following stimulation with IL-4, but not IFN-γ+LPS. Coculture with human aortic adventitial fibroblasts revealed that M(IL-4), but not M(IFN-γ+LPS), enhanced fibroblast collagen 1 protein expression. Macrophage pretreatment with the hydrogen peroxide scavenger, PEG-catalase, attenuated this effect. CONCLUSION: We show that superoxide generation is not only enhanced with stimuli associated with M1 macrophage activation but also with the M2 stimulus IL-4. Macrophages activated with IL-4 also exhibited enhanced hydrogen peroxide generation which in turn increased aortic fibroblast collagen production. Thus, M2 macrophage-derived ROS is identified as a potentially important contributor to aortic fibrosis.
Assuntos
Fibroblastos/efeitos dos fármacos , Interferon gama/farmacologia , Interleucina-4/farmacologia , Lipopolissacarídeos/farmacologia , Macrófagos/efeitos dos fármacos , Transdução de Sinais/genética , Catalase/farmacologia , Quimiocina CCL22/genética , Quimiocina CCL22/imunologia , Quimiocina CXCL11/genética , Quimiocina CXCL11/imunologia , Quimiocinas CC/genética , Quimiocinas CC/imunologia , Técnicas de Cocultura , Colágeno Tipo I/genética , Colágeno Tipo I/imunologia , Fibroblastos/citologia , Fibroblastos/imunologia , Regulação da Expressão Gênica , Humanos , Peróxido de Hidrogênio/metabolismo , Interferon gama/antagonistas & inibidores , Interleucina-1beta/genética , Interleucina-1beta/imunologia , Interleucina-4/antagonistas & inibidores , Lipopolissacarídeos/antagonistas & inibidores , Ativação de Macrófagos/efeitos dos fármacos , Macrófagos/citologia , Macrófagos/imunologia , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/imunologia , NADPH Oxidase 2/genética , NADPH Oxidase 2/imunologia , Oxirredução/efeitos dos fármacos , Polietilenoglicóis/farmacologia , Cultura Primária de Células , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Receptores CCR7/genética , Receptores CCR7/imunologia , Receptores Imunológicos/genética , Receptores Imunológicos/imunologia , Superóxidos/metabolismo , Células THP-1RESUMO
Interferon (IFN)-γ is mainly secreted by CD4+ T helper 1 (Th1), natural killer (NK) and NKT cells after skin injury. Although IFN-γ is well known regarding its inhibitory effects on collagen synthesis by fibroblasts in vitro, information is limited regarding its role in wound healing in vivo. In the present study, we analyzed how the defect of IFN-γ affects wound healing. Full-thickness wounds were created on the backs of wild type (WT) C57BL/6 and IFN-γ-deficient (KO) mice. We analyzed the percent wound closure, wound breaking strength, accumulation of leukocytes, and expression levels of COL1A1, COL3A1, and matrix metalloproteinases (MMPs). IFN-γKO mice exhibited significant attenuation in wound closure on Day 10 and wound breaking strength on Day 14 after wound creation, characteristics that are associated with prolonged neutrophil accumulation. Expression levels of COL1A1 and COL3A1 mRNA were lower in IFN-γKO than in WT mice, whereas expression levels of MMP-2 (gelatinase) mRNA were significantly greater in IFN-γKO than in WT mice. Moreover, under neutropenic conditions created with anti-Gr-1 monoclonal antibodies, wound closure in IFN-γKO mice was recovered through low MMP-2 expression levels. These results suggest that IFN-γ may be involved in the proliferation and maturation stages of wound healing through the regulation of neutrophilic inflammatory responses.
Assuntos
Regulação Enzimológica da Expressão Gênica/imunologia , Interferon gama/deficiência , Metaloproteinase 2 da Matriz/imunologia , Neutrófilos/imunologia , Cicatrização/imunologia , Animais , Colágeno Tipo I/genética , Colágeno Tipo I/imunologia , Cadeia alfa 1 do Colágeno Tipo I , Colágeno Tipo III/genética , Colágeno Tipo III/imunologia , Ativação Enzimática/genética , Ativação Enzimática/imunologia , Inflamação/genética , Inflamação/imunologia , Inflamação/patologia , Interferon gama/imunologia , Metaloproteinase 2 da Matriz/genética , Camundongos , Camundongos Knockout , Neutrófilos/patologia , Cicatrização/genéticaRESUMO
Proliferative vitreoretinopathy (PVR) is a visionthreatening disease. It is also a common complication resulting from surgery to correct rhegmatogenous retinal detachment. Proliferation and migration of retinal pigment epithelial (RPE) cells and the secretion of extracellular matrix molecules play an important role in the formation of the preretinal membrane in PVR patients. Furthermore, upregulated expression of inflammatory cytokines within the vitreous or subretinal fluid of patients experiencing vitreoretinal disorders may aggravate the inflammatory response and be involved in the development of PVR. PVR is triggered by many inflammatory cytokines and growth factors. Macrophage migration inhibitory factor (MIF), an inflammatory cytokine, is upregulated in the vitreous in PVR patients. However, there is little known concerning the connection between MIF and the expression of inflammatory cytokines, interleukin6 (IL6) and monocyte chemotactic1 (MCP1), and the fibrogenic gene, collagen I, in human RPE cells. Cell proliferation, migration, and expression of IL6, MCP1 and collagen I were assessed using an MTT assay, a Transwell assay, realtime PCR analysis and ELISA kits. Westernblot analysis was used to detect phosphorylation of p38 mitogen activated protein kinase (MAPK) and extracellular signalregulated kinase (ERK) signaling pathways. The data revealed that MIF promoted the proliferation, migration and expression of IL6, MCP1 and collagen I, and phosphorylation of p38 and ERK signaling pathways in RPE cells in vitro. These findings suggest that MIF plays a proinflammatory and profibrotic role in the development of PVR.
Assuntos
Quimiocina CCL2/imunologia , Colágeno Tipo I/imunologia , Interleucina-6/imunologia , Oxirredutases Intramoleculares/imunologia , Fatores Inibidores da Migração de Macrófagos/imunologia , Epitélio Pigmentado da Retina/imunologia , Movimento Celular , Proliferação de Células , Quimiocina CCL2/genética , Colágeno Tipo I/genética , Humanos , Interleucina-6/genética , Epitélio Pigmentado da Retina/citologia , Epitélio Pigmentado da Retina/metabolismo , Epitélio Pigmentado da Retina/patologia , Regulação para Cima , Vitreorretinopatia Proliferativa/genética , Vitreorretinopatia Proliferativa/imunologia , Vitreorretinopatia Proliferativa/patologiaRESUMO
Primary Sjögren's syndrome (pSS) is a chronic inflammatory, autoimmune and systemic disorder commonly associated with dry eyes and a dry mouth. Recently, the hypothetical link between epithelial-mesenchymal transition (EMT)-dependent salivary gland (SG) fibrosis and chronic inflammatory conditions has been suggested. In this study, we present data demonstrating a negative correlation of the epithelial marker E-cadherin expression and a positive correlation of mesenchymal vimentin and collagen type I expression with increasing degrees of tissue inflammation in pSS SG specimens. In addition, as it is not clear whether dysregulated cytokines in pSS, interleukin (IL)-17 and IL-22 may also contribute to the EMT-dependent fibrosis process, the effect of IL-17 and IL-22 treatment on EMT-dependent SG fibrosis was evaluated in primary human salivary gland epithelial cells (SGEC) isolated from healthy subjects. Here we present data demonstrating that IL-17 and IL-22 can induce SGEC to undergo a morphological and phenotypical transition to a mesenchymal phenotype. In support of this, vimentin and collagen type I were up-regulated while a decreased expression of E-cadherin occurs after interleukin treatment, and co-operation between IL-17 and Il-22 was required to induce the EMT.
Assuntos
Células Epiteliais/imunologia , Transição Epitelial-Mesenquimal/imunologia , Interleucina-17/imunologia , Interleucinas/imunologia , Glândulas Salivares/imunologia , Síndrome de Sjogren/imunologia , Idoso , Antígenos CD/imunologia , Caderinas/imunologia , Colágeno Tipo I/imunologia , Células Epiteliais/patologia , Feminino , Humanos , Inflamação/imunologia , Inflamação/patologia , Masculino , Pessoa de Meia-Idade , Glândulas Salivares/patologia , Síndrome de Sjogren/patologia , Regulação para Cima/imunologia , Vimentina/imunologia , Interleucina 22RESUMO
Conventional bone decalcification is a time-consuming process and is therefore unsuitable for clinical applications and time-limited research projects. Consequently, we compared the effect of four different decalcification solutions applied at three different temperatures, and assessed the rate of decalcification and the implications on tissue morphology and antigenicity of mouse and rat tibiae. Bones were decalcified with 10% ethylenediaminetetraacetic acid (EDTA), 10% formic acid, 5% hydrochloric acid, and 5% nitric acid at 4C, 25C, and 37C. Decalcification in both species was fastest in nitric acid at 37C and slowest in EDTA at 4C. Histological and immunohistochemical staining confirmed that the conventional protocols of EDTA at 4C and 25C remain the best option regarding the quality of tissue preservation. Whereas formic acid at 4C is a good alternative saving about 90% of the decalcification time, hydrochloric and nitric acids should be avoided particularly in case of rat tibia. By contrast, due to their smaller size, mouse tibiae had shorter decalcification times and tolerated higher temperatures and exposure to acids much better. In conclusion, this study demonstrated that depending on the specific research question and sample size, alternative decalcification methods could be used to decrease the time of decalcification while maintaining histological accuracy.
Assuntos
Técnica de Descalcificação/métodos , Tíbia/citologia , Tíbia/imunologia , Animais , Colágeno Tipo I/imunologia , Masculino , Camundongos , Ratos , Fator de Transcrição Sp7/imunologia , Fator de von Willebrand/imunologiaRESUMO
Dietary products may protect against inflammatory bowel disease (IBD) through mechanisms such as forming gut microbiota structures and providing substrates for microbial metabolism. Recently, many studies have been conducted on diets that potentially alleviate or suppress IBD development. To assess the efficacy of dietary oils in treating IBD, we examined the protective effects of olive oil, coconut oil, corn oil, and cottonseed oil in a dextran sodium sulfate (DSS)-induced colitis mouse model. Treatment with cottonseed oil or corn oil ameliorated the severity of DSS-induced colitis, alleviating weight loss and preventing the shortening of the intestine. Moreover, cottonseed oil or corn oil treatment significantly reduced the expression of inflammatory cytokines such as tumor necrosis factor-α, interleukin (IL)-1ß, IL-6, and IL-17, as well as the expression of oxidative stress markers, including 8-hydroxyguanosine and nitrotyrosine in colon sections, compared with vehicle treatment. Cottonseed oil treatment inhibited intestinal fibrosis by reducing the expression of α-smooth muscle actin and type I collagen, compared with vehicle treatment in mice with DSS-induced colitis. Cottonseed oil protects against intestinal inflammation and the development of intestinal fibrosis by reducing inflammatory cytokines and oxidative stress markers, and may therefore be useful as a dietary product with therapeutic benefits for IBD.
Assuntos
Colite/prevenção & controle , Óleo de Sementes de Algodão/administração & dosagem , Substâncias Protetoras/administração & dosagem , Actinas/genética , Actinas/imunologia , Animais , Colite/induzido quimicamente , Colite/genética , Colite/imunologia , Colágeno Tipo I/genética , Colágeno Tipo I/imunologia , Sulfato de Dextrana/efeitos adversos , Modelos Animais de Doenças , Humanos , Interleucina-1beta/genética , Interleucina-1beta/imunologia , Interleucina-6/genética , Interleucina-6/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Sulfatos/efeitos adversos , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologiaRESUMO
Lung transplant is a definitive treatment for several end-stage lung diseases. However, the high incidence of allograft rejection limits the overall survival following lung transplantation. Traditionally, alloimmunity directed against human leukocyte antigens (HLA) has been implicated in transplant rejection. Recently, the clinical impact of non-HLA lung-restricted antibodies (LRA) has been recognized and extensive research has demonstrated that they may play a dominant role in the development of lung allograft rejection. The immunogenic lung-restricted antigens that have been identified include amongst others, collagen type I, collagen type V, and k-alpha 1 tubulin. Pre-existing antibodies against these lung-restricted antigens are prevalent in patients undergoing lung transplantation and have emerged as one of the predominant risk factors for primary graft dysfunction which limits short-term survival following lung transplantation. Additionally, LRA have been shown to predispose to chronic lung allograft rejection, the predominant cause of poor long-term survival. This review will discuss ongoing research into the mechanisms of development of LRA as well as the pathogenesis of associated lung allograft injury.
Assuntos
Autoanticorpos/metabolismo , Bronquiolite Obliterante/imunologia , Rejeição de Enxerto/imunologia , Transplante de Pulmão , Pulmão/imunologia , Animais , Autoantígenos/imunologia , Autoimunidade , Colágeno Tipo I/imunologia , Colágeno Tipo V/imunologia , Humanos , Especificidade de Órgãos , Tubulina (Proteína)/imunologiaRESUMO
PURPOSE: For patients with end stage lung disease, lung transplantation (LT) remains the only definitive treatment option. Long term survival post LT is limited by acute and chronic allograft dysfunction. Antibodies to lung self-antigens Kα1Tubulin and collagen V (autoantibodies) have been implicated in adverse outcomes post LT. The aim of our study was to determine the prevalence of autoantibodies in pre- and post-transplant sera, evaluate the impact on post-transplant outcomes. METHODS: In a prospective observational cohort analysis, 44 patients were enrolled who received LT between 09/01/2014 and 10/31/2015. Pre- and post-transplant sera were analyzed using enzyme-linked immunosorbent assay (ELISA) for the presence of antibodies to collagen I, collagen V, and K-alpha 1 tubulin. The outcome variables are presence of primary graft dysfunction (PGD), cumulative acute cellular rejection (ACR), treatment with pulse steroids for clinical rejection, association with DSA, and onset of Bronchiolitis Obliterans Syndrome (BOS). RESULTS: In our cohort, 33 patients (75%) tested positive for the presence of autoantibodies. Pre-transplant autoantibodies were present in 23 patients (70%). Only a small percentage (26%) cleared these antibodies with standard immunosuppression. Some developed de novo post-transplant (nâ¯=â¯10). PGD was observed in 34% of our cohort, however the presence of autoantibodies did not correlate with increase in the incidence or severity of PGD. The prevalence of donor specific antibodies (DSA) in the entire cohort was 73%, with an increased prevalence of DSA noted in the autoantibody positive group (78.7% vs. 54.5%) than in the autoantibody negative group. BOS was observed in 20% of the cohort, with a median time to onset of 291â¯days' post-transplant. Patients with pre-transplant autoantibodies had a statistically significant decrease in BOS-free survival (pâ¯=â¯0.029 by log-rank test). CONCLUSIONS: In our cohort, we observed a high prevalence of autoantibodies and DSA in lung transplant recipients. Pre-transplant autoantibodies were associated with de novo development of DSA along with a decrease in BOS-free survival. Limitations to our study include the small sample size and single center enrollment, along with limited time for follow-up.
Assuntos
Bronquiolite Obliterante/epidemiologia , Colágeno Tipo V/metabolismo , Rejeição de Enxerto/imunologia , Transplante de Pulmão , Pulmão/metabolismo , Tubulina (Proteína)/metabolismo , Doença Aguda , Adulto , Idoso , Autoanticorpos/sangue , Estudos de Coortes , Colágeno Tipo I/imunologia , Colágeno Tipo I/metabolismo , Colágeno Tipo V/imunologia , Feminino , Rejeição de Enxerto/epidemiologia , Antígenos HLA/imunologia , Humanos , Imunidade Humoral , Isoanticorpos/sangue , Pulmão/imunologia , Masculino , Pessoa de Meia-Idade , Prevalência , Estudos Prospectivos , Tubulina (Proteína)/imunologia , Estados Unidos/epidemiologiaRESUMO
The immunomodulatory effect of IL-10 as an immunosuppressive and anti-inflammatory cytokine is well known. Taking advantage of our established mouse model of autoimmune cholangitis using 2-octynoic acid conjugated ovalbumin (2-OA-OVA) induction, we compared liver pathology, immune cell populations and antimitochondrial antibodies between IL-10 knockout and wild type mice immunized with 2-OA-OVA. At 10 weeks post immunization, portal inflammation and fibrosis were more severe in 2-OA-OVA immunized IL-10 knockout mice than in wild type mice. This was accompanied by significant higher levels of collagen I and III expression, T, NK and NKT subsets in liver and IgG anti-mitochondrial autoantibodies (AMAs) compared to 2-OA-OVA immunized wild type mice, suggesting that endogenous IL-10 is necessary for the maintenance of immune tolerance in primary biliary cholangitis (PBC). Further, we investigated whether administration of exogenous IL-10 could prevent PBC by administration of IL-10 expressing recombinant adeno-associated virus (AAV-IL-10) either 3 days before or 3 weeks after the establishment of liver pathology. Interestingly, administration of AAV-IL-10 resulted in increased liver inflammation and fibrosis, accompanied by increases in IFN-γ in liver CD4+ T cell, granzyme B, FasL, and CD107a in liver CD8+ T and NKT cells, and granzyme B and FasL in liver NK cells of AAV-IL-10 administered mice compared with control mice. Furthermore, administration of AAV-IL-10 significantly increased levels of proinflammatory cytokines and chemokines (IFN-γ, TNF-α, CXCL9 and CXCL10) and collagen I and III production in naïve mice, together with increase in immune cell infiltration and collagen deposition in the liver, suggesting a role of IL-10 in fibrosis. In conclusion, our data demonstrate that endogenous IL-10 is critical in the maintenance of immune tolerance but exogenous administration of IL-10 exacerbates liver inflammation and fibrosis. Furthermore, the distinctive presence of inflammatory immune cell populations and collagen expression in AAV-IL-10 treated naïve mice cautions against the clinical use of exogenous IL-10 in patients with autoimmune cholangitis.
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Autoanticorpos/sangue , Doenças Autoimunes/genética , Tolerância Imunológica , Interleucina-10/imunologia , Cirrose Hepática Biliar/imunologia , Animais , Doenças Autoimunes/sangue , Doenças Autoimunes/patologia , Quimiocina CXCL10/genética , Quimiocina CXCL10/imunologia , Colágeno Tipo I/genética , Colágeno Tipo I/imunologia , Colágeno Tipo III/genética , Colágeno Tipo III/imunologia , Dependovirus/genética , Dependovirus/imunologia , Modelos Animais de Doenças , Proteína Ligante Fas/genética , Proteína Ligante Fas/imunologia , Feminino , Regulação da Expressão Gênica , Vetores Genéticos/administração & dosagem , Vetores Genéticos/química , Vetores Genéticos/imunologia , Granzimas/genética , Granzimas/imunologia , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Interferon gama/genética , Interferon gama/imunologia , Interleucina-10/administração & dosagem , Interleucina-10/deficiência , Interleucina-10/genética , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/patologia , Fígado/imunologia , Fígado/patologia , Cirrose Hepática Biliar/sangue , Cirrose Hepática Biliar/genética , Cirrose Hepática Biliar/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Transdução de Sinais , Linfócitos T/imunologia , Linfócitos T/patologia , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologiaRESUMO
Periodontitis, an inflammatory disease of the gingival tissue, triggered by microbial-derived elements, such as lipopolysaccharide (LPS), collapses the periodontal tissues and resorbs the alveolar bone. This study evaluated the inhibitory effects of standardized Boesenbergia pandurata extract (BPE) and panduratin A (PAN) on periodontitis-induced inflammation and alveolar bone loss. Sprague-Dawley rats with LPS-induced periodontitis were orally administered BPE (50 and 200 mg/kg/day) and PAN (20 mg/kg/day) for 8 days. Histological analysis revealed that BPE- and PAN-administered groups showed decreased cell infiltration and alveolar bone resorption. Furthermore, the BPE and PAN significantly alleviated the mRNA and protein expression levels of nuclear factor kappa B (NF-κB), interleukin-1ß, matrix metalloproteinase (MMP)-2, and MMP-8. BPE and PAN also inhibited the expression of nuclear factor of activated T cells, cytoplasmic 1, c-Fos, and ostoclastogenesis-related enzymes, including cathepsin K and tartrate-resistant acid phosphatase (ALP). BPE and PAN not only upregulated the osteoblastogenesis-associated markers, such as collagen type I (COL1A1) and ALP, but also increased the ratio of osteoprotegerin to receptor activator of NF-κB ligand. Collectively, BPE and PAN efficiently prevent destruction of periodontal tissues and stimulating the loss of alveolar bone tissues, strongly indicative of their potential as natural antiperiodontitis agents.
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Perda do Osso Alveolar/tratamento farmacológico , Chalconas/administração & dosagem , Doenças Periodontais/tratamento farmacológico , Extratos Vegetais/administração & dosagem , Zingiberaceae/química , Perda do Osso Alveolar/induzido quimicamente , Perda do Osso Alveolar/imunologia , Animais , Chalconas/química , Colágeno Tipo I/genética , Colágeno Tipo I/imunologia , Humanos , Interleucina-1beta/genética , Interleucina-1beta/imunologia , Lipopolissacarídeos/efeitos adversos , Masculino , Metaloproteinase 2 da Matriz/genética , Metaloproteinase 2 da Matriz/imunologia , NF-kappa B/genética , NF-kappa B/imunologia , Osteoprotegerina/genética , Osteoprotegerina/imunologia , Doenças Periodontais/induzido quimicamente , Doenças Periodontais/imunologia , Extratos Vegetais/química , Ratos , Ratos Sprague-DawleyRESUMO
BACKGROUND: In systemic sclerosis (SSc), autoantibodies provide the most accurate tool to predict the disease subset and pattern of organ involvement. Scleroderma autoantibodies target nucleic acids or DNA/RNA-binding proteins, thus SSc immune complexes (ICs) can embed nucleic acids. Our working hypothesis envisaged that ICs containing scleroderma-specific autoantibodies might elicit proinflammatory and profibrotic effects in skin fibroblasts. METHODS: Fibroblasts were isolated from skin biopsies obtained from healthy subjects and patients with diffuse cutaneous SSc (dcSSc). ICs were purified by polyethylene-glycol precipitation from sera of SSc patients bearing different autoantibodies. ICs from patients with systemic lupus erythematosus (SLE) and primary anti-phospholipid syndrome (PAPS) and from normal healthy subjects (NHS) were used as controls. After incubation with ICs, fibroblasts were evaluated for ICAM-1 expression, interleukin (IL)-6, IL-8, monocyte chemoattractant protein (MCP)-1, matrix metalloproteinase (MMP)-2, tumor growth factor (TGF)-ß1 and Pro-CollagenIα1 secretion, collagen (col)Iα1, mmp-1, toll-like receptor (tlr)2, tlr3, tlr4, tlr7, tlr8, tlr9, interferon (ifn)-α, ifn-ß and endothelin-1 mRNA, and NFκB, p38MAPK and SAPK-JNK activation rate. Experiments were also performed after pretreatment with DNase I/RNase and NFκB/p38MAPK inhibitors. RESULTS: The antigenic reactivity for each SSc-IC mirrored the corresponding serum autoantibody specificity, while no positivity was observed in NHS-ICs or sera. SSc-ICs but not NHS-ICs increased ICAM-1 expression, stimulated IL-6, IL-8, MMP-2, MCP-1, TGF-ß1 and Pro-CollagenIα1 secretion, upregulated et-1, ifn-α, ifn-ß, tlr2, tlr3 and tlr4, and activated NFκB, p38MAPK and SAPK-JNK. tlr9 was significantly upregulated by ARA-ICs, mmp-1 was significantly induced by ACA-ICs whereas colIα1 was not modulated by any SSc-ICs. SLE-ICs and PAPS-ICs significantly upregulated MMP-2 and activated NFκB, p38MAPK and SAPK-JNK. SLE-ICs and PAPS-ICs did not affect colIα1, mmp-1 and Pro-CollagenIα1. DNase I and RNase treatment significantly reduced the upregulation of study mediators induced by SSc-ICs. Pretreatment with NFκB/p38MAPK inhibitors suggested that response to anti-Th/To-ICs was preferentially mediated by p38MAPK whereas ATA-ICs, ACA-ICs and ARA-ICs engaged both mediators. In dcSSc fibroblasts, stimulation with SSc-ICs and NHS-ICs upregulated IL-6 and IL-8. CONCLUSIONS: These data provide the first demonstration of the proinflammatory and profibrotic effects of SSc-ICs on fibroblasts, suggesting the potential pathogenicity of SSc autoantibodies. These effects might be mediated by Toll-like receptors via the interaction with nucleic acid fragments embedded in SSc-ICs.