RESUMO
ANCA-associated vasculitis (AAV) is intricately linked with infections. Toll-like receptors (TLR) provide a potential link between infection and anti-myeloperoxidase (MPO) autoimmunity. TLR9 ligation has been shown to promote anti-MPO autoimmunity and glomerular vasculitis in murine MPO-AAV. This study investigates dendritic cell TLR9 ligation in murine experimental anti-MPO glomerulonephritis. We analyzed autoimmune responses to MPO following transfer of TLR9 stimulated, MPO pulsed dendritic cells and kidney injury following a sub-nephritogenic dose of sheep anti-mouse glomerular basement membrane globulin. TLR9 ligation enhanced dendritic cell activation upregulating CD40 and CD80 expression, promoting systemic anti-MPO autoimmunity and T cell recall responses and exacerbating kidney injury. CD40 upregulation by TLR9 was critical for the induction of nephritogenic autoimmunity. The presence of DEC205, which transports the TLR9 ligand to TLR9 located in the endosome, also promoted kidney injury. This confirms TLR9 mediated dendritic cell activation as a mechanism of anti-MPO autoimmunity in AAV and further defines the link between infection and the generation of MPO specific autoimmune inflammation.
Assuntos
Vasculite Associada a Anticorpo Anticitoplasma de Neutrófilos , Glomerulonefrite , Receptor Toll-Like 9 , Animais , Camundongos , Autoimunidade , Células Dendríticas , Glomerulonefrite/metabolismo , Peroxidase/metabolismo , Ovinos , Receptor Toll-Like 9/metabolismoRESUMO
BACKGROUND: Acute Kidney Injury (AKI) is a well recognized complication of cardiac surgery. It is associated with significant morbidity and mortality. The aims of our study are twofold; 1. To define the incidence of AKI post cardiac surgery. 2. To identify pre-morbid and operative risk factors for developing AKI and to determine if immediate post operative serum creatinine (IPOsCr) accurately predicts the development of AKI. METHODS: We prospectively studied 196 consecutive patients undergoing elective (on-pump) cardiac surgery. Baseline patient characteristics, including medical co-morbidities, proteinuria, procedural data and kidney function (serum creatinine (sCr) were collected. Internationally standardised criteria for AKI were used (sCr >1.5 times baseline, elevation in sCr >26.4 µmmol/L (0.3 mg/dl). Measurements were collected pre-operatively, within 2 h of surgical completion (IPOsCr) and daily for two days. Logistic regression was used to assess predictive factors for AKI including IPOsCr. Model discrimination was assessed using ROC AUC curves. RESULTS: Forty (20.4%) patients developed AKI postoperatively. Hypertension (OR 2.64, p = 0.02), diabetes (OR 2.25, p = 0.04), proteinuria (OR 2.48, p = 0.02) and a lower baseline eGFR (OR 0.74, p = 0.002) were associated with AKI in univariate analysis. A multivariate logistic model with preoperative and surgical factors (age, gender, eGFR, proteinuria, hypertension, diabetes and type of cardiac surgery) demonstrated moderate discrimination for AKI (ROC AUC 0.76). The addition of IPOsCr improved model discrimination for AKI (AUC 0.82, p = 0.07 versus baseline AUC) and was independently associated with AKI (OR 7.17; 95% CI 1.27-40.32; p = 0.025). CONCLUSIONS: One in 5 patients developed AKI post cardiac surgery. These patients have significantly increased morbidity and mortality. IPOsCr is significantly associated with the development of AKI, providing a cheap readily available prognostic marker.
Assuntos
Injúria Renal Aguda/sangue , Injúria Renal Aguda/mortalidade , Creatinina/sangue , Diagnóstico Precoce , Complicações Pós-Operatórias/sangue , Complicações Pós-Operatórias/mortalidade , Modelos de Riscos Proporcionais , Injúria Renal Aguda/diagnóstico , Idoso , Idoso de 80 Anos ou mais , Biomarcadores/sangue , Procedimentos Cirúrgicos Cardíacos , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Complicações Pós-Operatórias/diagnóstico , Prognóstico , Reprodutibilidade dos Testes , Medição de Risco/métodos , Fatores de Risco , Sensibilidade e Especificidade , Taxa de Sobrevida , Vitória/epidemiologiaRESUMO
Observations in experimental murine myeloperoxidase (MPO)-ANCA-associated vasculitis (AAV) show mast cells degranulate, thus enhancing injury as well as producing immunomodulatory IL-10. Here we report that, compared with biopsy specimens from control patients, renal biopsy specimens from 44 patients with acute AAV had more mast cells in the interstitium, which correlated with the severity of tubulointerstitial injury. Furthermore, most of the mast cells were degranulated and spindle-shaped in patients with acute AAV, indicating an activated phenotype. We hypothesized that the mast cell stabilizer disodium cromoglycate would attenuate mast cell degranulation without affecting IL-10 production. We induced anti-MPO GN by immunizing mice with MPO and a low dose of anti-glomerular basement membrane antibody. When administered before or after induction of MPO autoimmunity in these mice, disodium cromoglycate attenuated mast cell degranulation, development of autoimmunity, and development of GN, without diminishing IL-10 production. In contrast, administration of disodium cromoglycate to mast cell-deficient mice had no effect on the development of MPO autoimmunity or GN. MPO-specific CD4(+) effector T cell proliferation was enhanced by co-culture with mast cells, but in the presence of disodium cromoglycate, proliferation was inhibited and IL-10 production was enhanced. These results indicate that disodium cromoglycate blocks injurious mast cell degranulation specifically without affecting the immunomodulatory role of these cells. Thus as a therapeutic, disodium cromoglycate may substantially enhance the regulatory role of mast cells in MPO-AAV.
Assuntos
Vasculite Associada a Anticorpo Anticitoplasma de Neutrófilos/tratamento farmacológico , Vasculite Associada a Anticorpo Anticitoplasma de Neutrófilos/imunologia , Doenças Autoimunes/tratamento farmacológico , Doenças Autoimunes/imunologia , Cromolina Sódica/farmacologia , Cromolina Sódica/uso terapêutico , Glomerulonefrite/tratamento farmacológico , Glomerulonefrite/imunologia , Mastócitos/efeitos dos fármacos , Mastócitos/imunologia , Peroxidase/imunologia , Idoso , Animais , Feminino , Humanos , Masculino , CamundongosRESUMO
Toll-like receptor 9 (TLR9) enhances proinflammatory responses, but whether it can act in a regulatory capacity remains to be established. In experimental murine AKI induced by cisplatin, Tlr9(-/-) mice developed enhanced renal injury and exhibited fewer intrarenal regulatory T cells (Tregs) compared with genetically intact mice. A series of reconstitution and depletion studies defined a role for TLR9 in maintaining Treg-mediated homeostasis in cisplatin-induced AKI. When Rag1(-/-) mice were reconstituted with nonregulatory CD25(-) splenocytes from wild-type (WT) or Tlr9(-/-) mice, AKI was similarly enhanced. However, when Rag1(-/-) mice were reconstituted with CD4(+)CD25(+) regulatory cells, WT CD4(+)CD25(+) cells were more renoprotective and localized to the kidney more efficiently than Tlr9(-/-) CD4(+)CD25(+) cells. In Treg-depleted Foxp3(DTR) mice, reconstitution with naive WT CD4(+)CD25(+) cells resulted in less severe AKI than did reconstitution with Tlr9(-/-) Tregs. Tlr9(-/-) mice were not deficient in CD4(+)CD25(+) cells, and WT and TLR9-deficient Tregs had similar suppressive function ex vivo. However, expression of adhesion molecules important in Treg trafficking was reduced on peripheral CD4(+)CD25(+) cells from Tlr9(-/-) mice. In conclusion, we identified a pathway by which TLR9 promotes renal Treg accumulation in AKI.
Assuntos
Injúria Renal Aguda/imunologia , Injúria Renal Aguda/metabolismo , Ativação Linfocitária , Linfócitos T Reguladores/imunologia , Receptor Toll-Like 9/fisiologia , Injúria Renal Aguda/induzido quimicamente , Animais , Antígeno CD11a/metabolismo , Cisplatino , Proteínas de Homeodomínio/genética , Receptores de Hialuronatos/metabolismo , Integrina alfa4/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Linfócitos T Reguladores/metabolismo , Receptor Toll-Like 9/genética , Receptor Toll-Like 9/metabolismoRESUMO
Myeloperoxidase (MPO) is an important neutrophil lysosomal enzyme, a major autoantigen, and a potential mediator of tissue injury in MPO-ANCA-associated vasculitis (MPO-AAV) and glomerulonephritis. Here we examined MPO deposition in kidney biopsies from 47 patients with MPO-AAV. Leukocyte accumulation and fibrin deposition consistent with cell-mediated immunity was a major feature. Tubulointerstitial macrophage, CD4+ and CD8+ T-cell, and neutrophil numbers correlated with low presenting eGFR. MPO was not detected in kidneys from patients with minimal change or thin basement membrane disease, but was prominent in glomerular, periglomerular, and tubulointerstitial regions in MPO-AAV. Extracellular MPO released from leukocytes was pronounced in all MPO-AAV patients. Similar numbers of neutrophils and macrophages expressed MPO in the kidneys, but colocalization studies identified neutrophils as the major source of extracellular MPO. Extraleukocyte MPO was prominent in neutrophil extracellular traps in the majority of patients; most of which had traps in half or more glomeruli. These traps were associated with more neutrophils and more MPO within glomeruli. Glomerular MPO-containing macrophages generated extracellular trap-like structures. MPO also localized to endothelial cells and podocytes. The presence of the most active glomerular lesions (both segmental necrosis and cellular crescents) correlated with intraglomerular CD4+ cells and MPO+ macrophages. Thus, cellular and extracellular MPO may cause glomerular and interstitial injury.
Assuntos
Anticorpos Anticitoplasma de Neutrófilos , Doenças Autoimunes/enzimologia , Armadilhas Extracelulares/enzimologia , Glomerulonefrite/enzimologia , Peroxidase/metabolismo , Idoso , Anticorpos Anticitoplasma de Neutrófilos/imunologia , Doenças Autoimunes/imunologia , Doenças Autoimunes/patologia , Contagem de Linfócito CD4 , Linfócitos T CD4-Positivos , Linfócitos T CD8-Positivos , Células Dendríticas/enzimologia , Células Endoteliais/enzimologia , Líquido Extracelular/enzimologia , Feminino , Taxa de Filtração Glomerular , Glomerulonefrite/imunologia , Glomerulonefrite/patologia , Humanos , Glomérulos Renais/enzimologia , Glomérulos Renais/patologia , Macrófagos/enzimologia , Masculino , Neutrófilos/enzimologia , Podócitos/enzimologiaRESUMO
In acute kidney injury, which is a significant cause of morbidity and mortality, cytokines and leukocytes promote inflammation and injury. We examined the pathogenic role of IL-17A in cisplatin-induced acute kidney injury. Intrarenal IL-17A mRNA transcription and protein expression were increased in wild-type mice after cisplatin-induced renal injury. An important role for IL-17A in the nephrotoxicity of cisplatin was demonstrated by observing protection from cisplatin-induced functional and histological renal injury in Il17a(-/-) and Rorγt(-/-) mice, as well as in mice treated pre-emptively with anti-IL-17A antibodies. Both renal injury and renal IL-1ß and IL-17A production were attenuated in Asc(-/-) and Tlr2(-/-) mice, suggesting that cisplatin induces endogenous TLR2 ligand production and activates the ASC-dependent inflammasome complex, resulting in IL-1ß and injurious IL-17A production. Neutrophils and natural killer cells are the likely targets of these pathways, because combined depletion of these cells was strongly protective; anti-IL-17A antibodies had no additional effect in this setting. Although IL-17A can also be produced by CD4(+) and γδ T cells, IL-17A from those cells does not contribute to renal injury. Cisplatin-induced injury was unchanged in γδ T-cell-deficient mice, whereas Il17a(-/-) CD4(+) T cells induced similar injury as did wild-type CD4(+) T cells on transfer to cisplatin-injected Rag1(-/-) mice. These studies demonstrate an important role for TLR2, the ASC inflammasome, and IL-17A in innate leukocytes in cisplatin-induced renal injury.
Assuntos
Injúria Renal Aguda/patologia , Inflamassomos/metabolismo , Interleucina-17/biossíntese , Leucócitos/metabolismo , Receptor 2 Toll-Like/metabolismo , Injúria Renal Aguda/induzido quimicamente , Injúria Renal Aguda/genética , Animais , Anticorpos/farmacologia , Proteínas Reguladoras de Apoptose/metabolismo , Proteínas Adaptadoras de Sinalização CARD , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/imunologia , Quimiocinas/genética , Quimiocinas/metabolismo , Cisplatino/efeitos adversos , Regulação da Expressão Gênica/efeitos dos fármacos , Inflamassomos/efeitos dos fármacos , Interleucina-17/genética , Interleucina-17/metabolismo , Rim/metabolismo , Rim/patologia , Células Matadoras Naturais/efeitos dos fármacos , Células Matadoras Naturais/imunologia , Leucócitos/efeitos dos fármacos , Masculino , Camundongos Endogâmicos C57BL , Infiltração de Neutrófilos/efeitos dos fármacos , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismoRESUMO
Myeloperoxidase (MPO) is important in intracellular microbial killing by neutrophils but extracellularly causes tissue damage. Its role in adaptive immunity and T-cell-mediated diseases is poorly understood. Here, T-cell responses in lymph nodes (LNs) were enhanced by MPO deletion or in vivo inhibition, causing enhanced skin delayed-type hypersensitivity and antigen (Ag)-induced arthritis. Responses of adoptively transferred OT-II T cells were greater in MPO-deficient than wild-type (WT) recipients. MPO, deposited by neutrophils in LNs after Ag injection, interacted with dendritic cells (DCs) in vivo. Culture of murine or human DCs with purified MPO or neutrophil supernatant showed that enzymatically dependent MPO-mediated inhibition of DC activation occurs via MPO-generated reactive intermediates and involves DC Mac-1. Transfer of DCs cultured with WT, but not MPO-deficient, neutrophil supernatant attenuated Ag-specific immunity in vivo. MPO deficiency or in vivo inhibition increased DC activation in LNs after immunization. Studies with DQ-ovalbumin showed that MPO inhibits Ag uptake/processing by DCs. In vivo DC transfer and in vitro studies showed that MPO inhibits DC migration to LNs by reducing their expression of CCR7. Therefore, MPO, via its catalytic activity, inhibits the generation of adaptive immunity by suppressing DC activation, Ag uptake/processing, and migration to LNs to limit pathological tissue inflammation.
Assuntos
Células Dendríticas/imunologia , Inflamação/imunologia , Neutrófilos/enzimologia , Peroxidase/fisiologia , Linfócitos T/fisiologia , Imunidade Adaptativa/genética , Imunidade Adaptativa/imunologia , Animais , Apresentação de Antígeno/imunologia , Células Cultivadas , Células Dendríticas/metabolismo , Humanos , Inflamação/etiologia , Inflamação/genética , Inflamação/metabolismo , Linfonodos/imunologia , Ativação Linfocitária/imunologia , Masculino , Erros Inatos do Metabolismo/genética , Erros Inatos do Metabolismo/imunologia , Erros Inatos do Metabolismo/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neutrófilos/metabolismo , Peroxidase/genética , Peroxidase/metabolismo , Linfócitos T/metabolismoRESUMO
AIMS/HYPOTHESIS: The AGEs and the receptor for AGEs (RAGE) are known contributors to diabetic complications. RAGE also has a physiological role in innate and adaptive immunity and is expressed on immune cells. The aim of this study was to determine whether deletion of RAGE from bone-marrow-derived cells influences the pathogenesis of experimental diabetic nephropathy. METHODS: Groups (n = 8/group) of lethally irradiated 8 week old wild-type (WT) mice were reconstituted with bone marrow from WT (WT â WT) or RAGE-deficient (RG) mice (RG â WT). Diabetes was induced using multiple low doses of streptozotocin after 8 weeks of bone marrow reconstitution and mice were followed for a further 24 weeks. RESULTS: Compared with diabetic WT mice reconstituted with WT bone marrow, diabetic WT mice reconstituted with RG bone marrow had lower urinary albumin excretion and podocyte loss, more normal creatinine clearance and less tubulo-interstitial injury and fibrosis. However, glomerular collagen IV deposition, glomerulosclerosis and cortical levels of TGF-ß were not different among diabetic mouse groups. The renal tubulo-interstitium of diabetic RG â WT mice also contained fewer infiltrating CD68(+) macrophages that were activated. Diabetic RG â WT mice had lower renal cortical concentrations of CC chemokine ligand 2 (CCL2), macrophage inhibitory factor (MIF) and IL-6 than diabetic WT â WT mice. Renal cortical RAGE ligands S100 calgranulin (S100A)8/9 and AGEs, but not high mobility box protein B-1 (HMGB-1) were also decreased in diabetic RG â WT compared with diabetic WT â WT mice. In vitro, bone-marrow-derived macrophages from WT but not RG mice stimulated collagen IV production in cultured proximal tubule cells. CONCLUSIONS/INTERPRETATION: These studies suggest that RAGE expression on haemopoietically derived immune cells contributes to the functional changes seen in diabetic nephropathy by promoting macrophage infiltration and renal tubulo-interstitial damage.
Assuntos
Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/terapia , Rim/metabolismo , Receptores Imunológicos/metabolismo , Animais , Diabetes Mellitus Experimental/genética , Macrófagos/metabolismo , Masculino , Camundongos , Receptor para Produtos Finais de Glicação Avançada , Receptores Imunológicos/genéticaRESUMO
BACKGROUND: A predictive histologic classification recently was proposed to determine the prognostic value of kidney biopsy in patients with antineutrophil cytoplasmic antibody-associated renal vasculitis (AAV). STUDY DESIGN: A dual-purpose retrospective observational cohort study to assess the reproducibility of the new classification and clinical variables that predict outcomes. SETTING & PARTICIPANTS: 169 consecutive patients with AAV were identified; 145 were included in the reproducibility study, and 120, in the outcomes study. PREDICTOR: Kidney biopsy specimens were classified according to the predominant glomerular lesion: focal, mixed, crescentic, and sclerotic. An assessment of tubular atrophy also was performed. OUTCOMES: The primary outcome was time to end-stage kidney disease or all-cause mortality, modeled using Cox regression analysis. MEASUREMENTS: Estimated glomerular filtration rate, requirement for renal replacement therapy. RESULTS: For the reproducibility study, the overall inter-rater reliability of the classification demonstrated variability among 3 histopathologists (intraclass correlation coefficient, 0.48; 95% CI, 0.38-0.57; κ statistic=0.46). Although agreement was high in the sclerotic group (κ=0.70), it was less consistent in other groups (κ=0.51, κ=0.47, and κ=0.23 for crescentic, focal, and mixed, respectively). For the clinical outcomes study, patients with sclerotic patterns of glomerular injury displayed the worst outcomes. Patients with focal (HR, 0.26; 95% CI, 0.12-0.58; P=0.001), crescentic (HR, 0.33; 95% CI, 0.16-0.69; P=0.003), and mixed (HR, 0.39; 95% CI, 0.18-0.81; P=0.01) patterns of injury had lower risk of the primary outcome. Tubular atrophy correlated with outcome, and advanced injury was associated with worse outcomes (HR, 5.9; 95% CI, 2.25-15.47; P<0.001). Level of kidney function at presentation strongly predicted outcome (HR per 10-mL/min/1.73m(2) increase in estimated glomerular filtration rate, 0.63; 95% CI, 0.46-0.81; P<0.001). LIMITATIONS: Data availability, given the retrospective nature of the study. CONCLUSIONS: Reproducibility of the classification was seen only in patients with sclerotic patterns of glomerular injury. Sclerotic pattern of glomerular injury, advanced chronic interstitial injury, and decreased kidney function all predicted poor outcomes.
Assuntos
Vasculite Associada a Anticorpo Anticitoplasma de Neutrófilos/diagnóstico , Vasculite Associada a Anticorpo Anticitoplasma de Neutrófilos/patologia , Progressão da Doença , Rim/patologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Vasculite Associada a Anticorpo Anticitoplasma de Neutrófilos/terapia , Criança , Estudos de Coortes , Feminino , Seguimentos , Humanos , Rim/metabolismo , Masculino , Pessoa de Meia-Idade , Valor Preditivo dos Testes , Estudos Retrospectivos , Resultado do Tratamento , Adulto JovemRESUMO
Loss of tolerance to neutrophil myeloperoxidase (MPO) underlies the development of ANCA-associated vasculitis and GN, but the mechanisms underlying this loss of tolerance are poorly understood. Here, we assessed the role of the thymus in deletion of autoreactive anti-MPO T cells and the importance of peripheral regulatory T cells in maintaining tolerance to MPO and protecting from GN. Thymic expression of MPO mRNA predominantly localized to medullary thymic epithelial cells. To assess the role of MPO in forming the T cell repertoire and the role of the autoimmune regulator Aire in thymic MPO expression, we compared the effects of immunizing Mpo(-/-) mice, Aire(-/-) mice, and control littermates with MPO. Immunized Mpo(-/-) and Aire(-/-) mice developed significantly more proinflammatory cytokine-producing anti-MPO T cells and higher ANCA titers than control mice. When we triggered GN with a subnephritogenic dose of anti-glomerular basement membrane antibody, Aire(-/-) mice had more severe renal disease than Aire(+/+) mice, consistent with a role for Aire-dependent central deletion in establishing tolerance to MPO. Furthermore, depleting peripheral regulatory T cells in wild-type mice also led to more anti-MPO T cells, higher ANCA titers, and more severe GN after immunization with MPO. Taken together, these results suggest that Aire-dependent central deletion and regulatory T cell-mediated peripheral tolerance both play major roles in establishing and maintaining tolerance to MPO, thereby protecting against the development of anti-MPO GN.
Assuntos
Anticorpos Anticitoplasma de Neutrófilos/imunologia , Glomerulonefrite/imunologia , Tolerância Imunológica/imunologia , Glomérulos Renais/imunologia , Peroxidase/imunologia , Linfócitos T Reguladores/imunologia , Timo/metabolismo , Animais , Anticorpos Anticitoplasma de Neutrófilos/metabolismo , Citocinas/metabolismo , Glomerulonefrite/metabolismo , Camundongos , Peroxidase/metabolismo , RNA Mensageiro , Linfócitos T Reguladores/metabolismo , Timo/imunologiaRESUMO
Mast cells contribute to the modulation of the immune response, but their role in autoimmune renal disease is not well understood. Here, we induced autoimmunity resulting in focal necrotizing GN by immunizing wild-type or mast cell-deficient (Kit(W-sh/W-sh)) mice with myeloperoxidase. Mast cell-deficient mice exhibited more antimyeloperoxidase CD4+ T cells, enhanced dermal delayed-type hypersensitivity responses to myeloperoxidase, and more severe focal necrotizing GN. Furthermore, the lymph nodes draining the sites of immunization had fewer Tregs and reduced production of IL-10 in mice lacking mast cells. Reconstituting these mice with mast cells significantly increased the numbers of Tregs in the lymph nodes and attenuated both autoimmunity and severity of disease. After immunization with myeloperoxidase, mast cells migrated from the skin to the lymph nodes to contact Tregs. In an ex vivo assay, mast cells enhanced Treg suppression through IL-10. Reconstitution of mast cell-deficient mice with IL-10-deficient mast cells led to enhanced autoimmunity to myeloperoxidase and greater disease severity compared with reconstitution with IL-10-intact mast cells. Taken together, these studies establish a role for mast cells in mediating peripheral tolerance to myeloperoxidase, protecting them from the development of focal necrotizing GN in ANCA-associated vasculitis.
Assuntos
Vasculite Associada a Anticorpo Anticitoplasma de Neutrófilos/complicações , Glomerulonefrite/imunologia , Interleucina-10/metabolismo , Mastócitos/fisiologia , Linfócitos T Reguladores/fisiologia , Animais , Células Cultivadas , Glomerulonefrite/metabolismo , Glomerulonefrite/patologia , Humanos , Imunomodulação , Linfonodos/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Tolerância Periférica , PeroxidaseRESUMO
Progressive renal fibrosis is the final common pathway leading to renal failure irrespective of the initiating cause. Clinical studies of renal fibrosis found that prominent mast cell accumulation correlated with worse outcomes. Mast cells are pluripotent innate immune cells that synthesize and secrete profibrotic mediators. Here we use mast cell-deficient (Kit(W-sh/W-sh)) mice to define a functional pathogenic role for these cells in the development of renal fibrosis. Intrarenal collagen deposition was significantly decreased in mast cell-deficient compared to wild-type mice 7 and 14 days after unilateral ureteric obstruction. The intrarenal expression of mRNAs for transforming growth factor-ß, α-smooth muscle actin, chemokines, and renal macrophages and CD4(+) T cells were also decreased in mast cell-deficient mice. Reconstitution of the mast cell population in mast cell-deficient mice with wild-type bone marrow-derived mast cells restored the pattern and intensity of renal fibrosis to levels seen in wild-type mice following ureteric ligation. Interestingly, the mast cells were recruited, activated, and degranulated within 6 h of ureteric ligation. A mast cell stabilizer that impairs degranulation, disodium chromoglycate, significantly attenuated renal fibrosis following ureteric ligation in wild-type mice. Thus, mast cells promote renal fibrosis and their targeting may offer therapeutic potential in the treatment of renal fibrosis.
Assuntos
Degranulação Celular , Nefropatias/etiologia , Rim/imunologia , Mastócitos/imunologia , Obstrução Ureteral/complicações , Actinas/genética , Actinas/metabolismo , Animais , Linfócitos T CD4-Positivos/imunologia , Degranulação Celular/efeitos dos fármacos , Células Cultivadas , Quimiocinas/genética , Quimiocinas/metabolismo , Colágeno/metabolismo , Cromolina Sódica/farmacologia , Modelos Animais de Doenças , Progressão da Doença , Fibrose , Regulação da Expressão Gênica , Hidronefrose/etiologia , Hidronefrose/imunologia , Hidronefrose/patologia , Rim/efeitos dos fármacos , Rim/patologia , Nefropatias/genética , Nefropatias/imunologia , Nefropatias/patologia , Nefropatias/prevenção & controle , Macrófagos/imunologia , Masculino , Mastócitos/efeitos dos fármacos , Mastócitos/transplante , Metaloproteinase 12 da Matriz/genética , Metaloproteinase 12 da Matriz/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Mutação , Fenótipo , Proteínas Proto-Oncogênicas c-kit/genética , Fatores de Tempo , Fator de Crescimento Transformador beta/genética , Fator de Crescimento Transformador beta/metabolismo , Obstrução Ureteral/genética , Obstrução Ureteral/imunologia , Obstrução Ureteral/patologiaRESUMO
Despite its initially defined role as a T-helper type 1 cell (Th1)-inducing cytokine, interleukin-27 (IL-27) has complex roles in vivo. The role of IL-27 receptor (IL-27R) was defined in experimental crescentic glomerulonephritis induced by a foreign antigen, sheep globulin, which is planted in glomeruli. This lesion is dependent on a Th1 effector cellular response. Twenty-one days after the administration of sheep anti-mouse glomerular basement membrane antibody, wild-type mice developed histologic and functional inflammatory renal injury. Injury was attenuated in the absence of IL-27R α chain (IL-27Rα), the unique component of the IL-27R complex. In contrast to the attenuated renal injury on day 21, Il27ra(-/-) mice exhibited enhanced systemic immune responses, including Th1 responses, with increased IL-2-dependent interferon-γ (IFN-γ) production. However, earlier in the development of the nephritogenic immune response, IFN-γ production was decreased, with reduced early immune responses translating into attenuated renal injury. Having demonstrated decreased early Th1 systemic immune responses, followed by enhanced nephritogenic Th1 immune responses, renal injury was studied at later time points. On days 28 and 35 after injection of the nephritogenic antigen, renal injury was enhanced in Il27ra(-/-) mice compared with wild-type mice in an at least partially IFN-γ-dependent manner. In Th1-dependent autoinflammatory lesions, IL-27Rα has a biphasic role in vivo, initially pathogenic, but ultimately playing a protective role by regulating immune responses and attenuating disease.
Assuntos
Glomerulonefrite/imunologia , Glomerulonefrite/patologia , Receptores de Citocinas/metabolismo , Células Th1/imunologia , Animais , Especificidade de Anticorpos/imunologia , Antígenos/imunologia , Glomerulonefrite/genética , Imunidade/imunologia , Mediadores da Inflamação/metabolismo , Interferon gama/biossíntese , Interleucina-2/biossíntese , Interleucinas/genética , Interleucinas/metabolismo , Rim/imunologia , Rim/patologia , Camundongos , Camundongos Endogâmicos C57BL , Testes de Neutralização , Receptores de Citocinas/genética , Receptores de Interleucina , Ovinos , Fatores de Tempo , Fatores de Transcrição/metabolismoRESUMO
T helper (Th)17 cells might contribute to immune-mediated renal injury. Thus, we sought to define the time course of IL-17A-induced kidney damage and examined the relation between Th17 and Th1 cells in a model of crescentic anti-glomerular basement membrane glomerulonephritis. Renal injury and immune responses were assessed in wild-type and in IL-17A-deficient mice on days 6, 14, and 21 of disease development. On day 6, when mild glomerulonephritis developed, IL-17A-deficient mice were protected from renal injury. On day 14, when more severe disease developed, protection from renal injury due to IL-17A deficiency was less evident. On day 21, when crescentic glomerulonephritis was fully established, disease was enhanced in IL-17A(-/-) mice, with increased glomerular T-cell accumulation and fibrin deposition, and augmented Th1 responses. Mice lacking the Th17-promoting cytokine, IL-23 (p19), also developed more severe disease than wild-type animals on day 21. In contrast, mice deficient in the key Th1-promoting cytokine, IL-12 (p35), had decreased Th1 and increased Th17 responses and developed less severe crescentic glomerulonephritis than wild-type animals. These studies show that IL-17A contributes to early glomerular injury, but it attenuates established crescentic glomerulonephritis by suppressing Th1 responses. They provide further evidence that Th1 cells mediate crescentic injury in this model and that Th1 and Th17 cells counterregulate each other during disease development.
Assuntos
Injúria Renal Aguda/imunologia , Doença Antimembrana Basal Glomerular/imunologia , Interleucina-17/deficiência , Células Th1/fisiologia , Animais , Moléculas de Adesão Celular/metabolismo , Sobrevivência Celular , Fibrina/metabolismo , Imunidade Celular/imunologia , Imunoglobulinas , Interferon gama/metabolismo , Interleucina-12/deficiência , Interleucina-17/metabolismo , Interleucina-23/deficiência , Interleucina-23/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BLRESUMO
OBJECTIVE: Autoantibodies constitute the hallmark of antineutrophil cytoplasmic antibody-associated vasculitis (AAV); however, CD4+ T cells play an essential role in the development of autoimmunity. Infection is associated with vasculitis, with Toll-like receptors (TLRs) a potential link between infection and autoimmunity. This study was undertaken to investigate the role of TLR ligation on cellular and humoral autoimmunity and glomerular injury in experimental myeloperoxidase (MPO)-induced AAV. METHODS: We analyzed autoimmune responses in wild-type mice immunized with MPO alone or coimmunized with MPO and a TLR-2 or TLR-9 ligand. The major vascular injury found in human disease, glomerulonephritis with focal necrosis, was triggered by administering a subnephritogenic dose of nephrotoxic serum. RESULTS: MPO alone induced low-titer antineutrophil cytoplasmic antibodies (ANCAs) without delayed-type hypersensitivity or CD4 cytokine responses. However, when MPO was given with either TLR ligand, cellular and humoral autoimmunity was enhanced, but with distinctly different CD4 subsets and IgG ANCA isotypes. TLR-2 ligand induced Th17 autoimmunity, with retinoic acid receptor-related orphan nuclear receptor γt-dependent interleukin-17A (IL-17A) production. TLR-9 ligand promoted Th1 autoimmunity, with enhanced production of interferon-γ (IFNγ) and Th1-associated IgG subclasses. Glomerular vasculitis developed only after the administration of nephrotoxic serum in mice immunized with either TLR ligand and MPO. Glomerulonephritis directed by MPO and TLR-2 ligation was attenuated when IL-17A was neutralized, while glomerulonephritis induced by MPO and TLR-9 ligation was attenuated when IFNγ was neutralized. CONCLUSION: Our findings indicate a pathogenic role of TLRs in initiating autoimmune AAV. TLR-2 induces Th17 CD4 cells while TLR-9 can also direct vasculitis, by directing Th1 autoimmunity.
Assuntos
Vasculite Associada a Anticorpo Anticitoplasma de Neutrófilos/imunologia , Autoimunidade/fisiologia , Peroxidase/efeitos adversos , Células Th1/imunologia , Células Th17/imunologia , Receptor 2 Toll-Like/metabolismo , Receptor Toll-Like 9/metabolismo , Animais , Vasculite Associada a Anticorpo Anticitoplasma de Neutrófilos/metabolismo , Vasculite Associada a Anticorpo Anticitoplasma de Neutrófilos/patologia , Anticorpos Monoclonais/uso terapêutico , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD4-Positivos/patologia , Modelos Animais de Doenças , Glomerulonefrite/imunologia , Glomerulonefrite/metabolismo , Glomerulonefrite/prevenção & controle , Imunidade Humoral/fisiologia , Interferon gama/metabolismo , Rim/imunologia , Rim/metabolismo , Rim/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Células Th1/metabolismo , Células Th1/patologia , Células Th17/metabolismoRESUMO
Although Th17 responses may contribute to the pathogenesis of glomerulonephritis, whether the key transcription factor in Th17 cell development, RORγt, also promotes glomerulonephritis is unknown. Here, we induced crescentic glomerulonephritis in wild-type and RORγt-deficient (RORγt(-/-)) mice. RORγt(-/-) mice were protected from disease, with reduced histologic and functional injury and decreased leukocyte infiltration. Because RORγt(-/-) mice lack lymph nodes, which may influence the development of nephritis, we performed cell-transfer studies. We reconstituted Rag1(-/-) mice, which lack adaptive immunity but otherwise have normal architecture of the lymphatic system, with splenocytes from naïve wild-type or RORγt(-/-) mice. Mice receiving wild-type splenocytes exhibited high mortality from renal failure after induction of nephritis whereas mice receiving RORγt(-/-) cells were protected. To determine the effect of RORγt deficiency specifically in T helper cells, we isolated naïve CD4(+) T cells from wild-type and RORγt(-/-) mice and transferred them into Rag1(-/-) animals. Recipients of wild-type CD4(+) T cells developed severe glomerulonephritis whereas recipients of RORγt(-/-) cells developed less severe disease. To exclude effects of altered regulatory T cell (Treg) development caused by RORγt deficiency, we transferred naïve CD4(+) T cells depleted of Tregs into Rag1(-/-) mice. Recipients of wild-type, Treg-depleted, CD4(+) T cells developed severe glomerulonephritis whereas recipients of RORγt(-/-), Treg-depleted CD4(+) T cells did not. Taken together, this study demonstrates that RORγt promotes the development of crescentic glomerulonephritis by directing nephritogenic Th17 responses.
Assuntos
Glomerulonefrite/patologia , Glomerulonefrite/fisiopatologia , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/fisiologia , Células Th17/fisiologia , Imunidade Adaptativa/fisiologia , Transferência Adotiva , Animais , Linfócitos T CD4-Positivos/patologia , Linfócitos T CD4-Positivos/transplante , Modelos Animais de Doenças , Feminino , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/genética , Linfócitos T Reguladores/patologia , Células Th1/imunologia , Células Th1/patologia , Células Th1/fisiologia , Células Th17/imunologia , Células Th17/patologiaRESUMO
Leukocyte recruitment contributes to acute kidney injury (AKI), but the mechanisms by which leukocytes promote injury are not completely understood. The degranulation of mast cells releases inflammatory molecules, including TNF, but whether these cells participate in the pathogenesis of AKI is unknown. Here, we induced AKI with cisplatin in mast cell-deficient and wild-type mice. Compared with wild-type mice, deficiency of mast cells attenuated renal injury, reduced serum levels of TNF, and reduced recruitment of leukocytes to the inflamed kidney. Mast cell-deficient mice also exhibited significantly lower intrarenal expression of leukocyte chemoattractants. Mast cell-deficient mice reconstituted with mast cells from wild-type mice exhibited similar cisplastin-induced renal damage and serum levels of TNF as wild-type mice. In contrast, mast cell-deficient mice reconstituted with mast cells from TNF-deficient mice continued to demonstrate significant attenuation of cisplatin-induced renal injury. Furthermore, the mast-cell stabilizer sodium chromoglycate also significantly abrogated renal injury in this model of AKI. Taken together, these results suggest that mast cells mediate AKI through the production of TNF.
Assuntos
Injúria Renal Aguda/etiologia , Mastócitos/fisiologia , Fator de Necrose Tumoral alfa/biossíntese , Injúria Renal Aguda/induzido quimicamente , Animais , Cisplatino/administração & dosagem , CamundongosRESUMO
Glomerular disease can be triggered or exacerbated by microbes that activate the immune system by Toll-like receptor (TLR) ligation. TLR9 activation promotes host defenses through the enhancement of innate and adaptive immune responses that facilitate the recruitment of leukocytes to areas of inflammation. We defined the role of TLR9 in experimental crescentic glomerulonephritis. Wild-type mice administered a TLR9 ligand and sheep anti-mouse glomerular basement membrane antibody developed histological injury with impaired renal function, which was attenuated in TLR9 knockout mice. Consistent with enhanced renal injury, wild-type mice exhibited enhanced T helper 1 and T helper 17 cellular immune responses. Kidney mRNA expression of inflammatory cytokines and chemokines as well as leukocyte recruitment were increased in wild-type mice. The use of bone marrow chimeric mice demonstrated that while both bone marrow and tissue cell TLR9 are required for maximal injury, bone marrow TLR9 is more important. Administration of a TLR9 inhibitor before sheep anti-mouse glomerular basement membrane globulin in wild-type mice attenuated cellular nephritogenic immunity that resulted in decreased renal injury. Administration of the inhibitor 7 days after disease initiation decreased glomerular leukocyte recruitment as well as renal injury. These results define the role of TLR9 in experimental crescentic glomerulonephritis and identify therapeutic potential for TLR9 inhibitors in attenuating renal injury, decreasing cellular nephritogenic immunity early in disease, and decreasing kidney effector responses later.
Assuntos
Glomerulonefrite/imunologia , Leucócitos/imunologia , Receptor Toll-Like 9/imunologia , Animais , Quimiocinas/genética , Quimiocinas/imunologia , Citocinas/genética , Citocinas/imunologia , Modelos Animais de Doenças , Glomerulonefrite/patologia , Glomerulonefrite/fisiopatologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Subpopulações de Linfócitos T/imunologia , Linfócitos T/imunologiaRESUMO
Antimyeloperoxidase antibodies can cause crescentic glomerulonephritis and pulmonary hemorrhage. Toll-like receptors (TLRs) respond to infectious agents activating host defenses, whereas infections potentially initiate disease and provoke relapses. Neutrophils were found to be key effector cells of injury in experimental models, as disease does not occur in their absence and injury is enhanced by lipopolysaccharide (LPS). In this study, highly purified LPS (a pure TLR4 ligand) acted with antimyeloperoxidase antibodies to synergistically increase kidney and lung neutrophil recruitment and functional injury; effects abrogated in TLR4-deficient mice. Increased kidney TLR4 expression after stimulation predominantly occurred in glomerular endothelial cells. Enhanced glomerular neutrophil recruitment correlated with increased kidney mRNA expression of CXCL1 and CXCL2 (homologs of human CXCL8), whereas their preemptive neutralization decreased neutrophil recruitment. Disease induction in bone marrow chimeric mice showed that TLR4 in both bone marrow and renal parenchymal cells is required for maximal neutrophil recruitment and glomerular injury. Further studies in human glomerular cell lines stimulated with LPS found that glomerular endothelial cells were the prominent sources of CXCL8. Thus, our results define a role for TLR4 expression in bone marrow-derived and glomerular endothelial cells in neutrophil recruitment and subsequent functional and histological renal injury in experimental antimyeloperoxidase glomerulonephritis.
Assuntos
Anticorpos Anti-Idiotípicos/efeitos adversos , Glomerulonefrite/induzido quimicamente , Glomerulonefrite/metabolismo , Rim/metabolismo , Leucócitos/metabolismo , Peroxidase/imunologia , Receptor 4 Toll-Like/metabolismo , Animais , Anticorpos Anti-Idiotípicos/imunologia , Anticorpos Anti-Idiotípicos/farmacologia , Anticorpos Anticitoplasma de Neutrófilos/efeitos adversos , Anticorpos Anticitoplasma de Neutrófilos/farmacologia , Linhagem Celular , Quimiocina CXCL1/metabolismo , Quimiocina CXCL2/metabolismo , Modelos Animais de Doenças , Glomerulonefrite/patologia , Humanos , Interleucina-8/metabolismo , Rim/efeitos dos fármacos , Rim/patologia , Glomérulos Renais/metabolismo , Glomérulos Renais/patologia , Leucócitos/efeitos dos fármacos , Leucócitos/patologia , Lipopolissacarídeos/efeitos adversos , Lipopolissacarídeos/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos KnockoutRESUMO
Th1 effector CD4+ cells contribute to the pathogenesis of proliferative and crescentic glomerulonephritis, but whether effector Th17 cells also contribute is unknown. We compared the involvement of Th1 and Th17 cells in a mouse model of antigen-specific glomerulonephritis in which effector CD4+ cells are the only components of adaptive immunity that induce injury. We planted the antigen ovalbumin on the glomerular basement membrane of Rag1(-/-) mice using an ovalbumin-conjugated non-nephritogenic IgG1 monoclonal antibody against alpha3(IV) collagen. Subsequent injection of either Th1- or Th17-polarized ovalbumin-specific CD4+ effector cells induced proliferative glomerulonephritis. Mice injected with Th1 cells developed progressive albuminuria over 21 d, histologic injury including 5.5 +/- 0.9% crescent formation/segmental necrosis, elevated urinary nitrate, and increased renal NOS2, CCL2, and CCL5 mRNA. Mice injected with Th17 cells developed albuminuria by 3 d; compared with Th1-injected mice, their glomeruli contained more neutrophils and greater expression of renal CXCL1 mRNA. In conclusion, Th1 and Th17 effector cells can induce glomerular injury. Understanding how these two subsets mediate proliferative forms of glomerulonephritis may lead to targeted therapies.