RESUMO
ANCA-associated vasculitis is the most frequent cause of crescentic GN. To define new molecular and/or cellular biomarkers of this disease in the kidney, we performed microarray analyses of renal biopsy samples from patients with ANCA-associated crescentic GN. Expression profiles were correlated with clinical data in a prospective study of patients with renal ANCA disease. CC chemokine ligand 18 (CCL18), acting through CC chemokine receptor 8 (CCR8) on mononuclear cells, was identified as the most upregulated chemotactic cytokine in patients with newly diagnosed ANCA-associated crescentic GN. Macrophages and myeloid dendritic cells in the kidney were detected as CCL18-producing cells. The density of CCL18(+) cells correlated with crescent formation, interstitial inflammation, and impairment of renal function. CCL18 protein levels were higher in sera of patients with renal ANCA disease compared with those in sera of patients with other forms of crescentic GN. CCL18 serum levels were higher in patients who suffered from ANCA-associated renal relapses compared with those in patients who remained in remission. Using a murine model of crescentic GN, we explored the effects of the CCL18 murine functional analog CCL8 and its receptor CCR8 on kidney function and morphology. Compared with wild-type mice, Ccr8(-/-) mice had significantly less infiltration of pathogenic mononuclear phagocytes. Furthermore, Ccr8(-/-) mice maintained renal function better and had reduced renal tissue injury. In summary, our data indicate that CCL18 drives renal inflammation through CCR8-expressing cells and could serve as a biomarker for disease activity and renal relapse in ANCA-associated crescentic GN.
Assuntos
Vasculite Associada a Anticorpo Anticitoplasma de Neutrófilos/complicações , Quimiocinas CC/sangue , Glomerulonefrite/etiologia , Glomerulonefrite/metabolismo , Idoso , Animais , Biomarcadores/sangue , Quimiocina CCL8/genética , Quimiocina CCL8/metabolismo , Quimiocinas CC/análise , Células Dendríticas/química , Feminino , Glomerulonefrite/patologia , Glomerulonefrite/fisiopatologia , Humanos , Macrófagos/química , Masculino , Camundongos , Pessoa de Meia-Idade , Estudos Prospectivos , Análise Serial de Proteínas , Receptores CCR8/genética , Receptores CCR8/metabolismo , Regulação para CimaRESUMO
Neutrophil trafficking to sites of inflammation is essential for the defense against bacterial and fungal infections, but also contributes to tissue damage in TH17-mediated autoimmunity. This process is regulated by chemokines, which often show an overlapping expression pattern and function in pathogen- and autoimmune-induced inflammatory reactions. Using a murine model of crescentic GN, we show that the pathogenic TH17/IL-17 immune response induces chemokine (C-X-C motif) ligand 5 (CXCL5) expression in kidney tubular cells, which recruits destructive neutrophils that contribute to renal tissue injury. By contrast, CXCL5 was dispensable for neutrophil recruitment and effective bacterial clearance in a murine model of acute bacterial pyelonephritis. In line with these findings, CXCL5 expression was highly upregulated in the kidneys of patients with ANCA-associated crescentic GN as opposed to patients with acute bacterial pyelonephritis. Our data therefore identify CXCL5 as a potential therapeutic target for the restriction of pathogenic neutrophil infiltration in TH17-mediated autoimmune diseases while leaving intact the neutrophil function in protective immunity against invading pathogens.
Assuntos
Quimiocina CXCL5/metabolismo , Glomerulonefrite/patologia , Neutrófilos/metabolismo , Células Th17/citologia , Animais , Quimiocina CXCL1/metabolismo , Quimiocinas/metabolismo , Modelos Animais de Doenças , Células Epiteliais/citologia , Feminino , Glomerulonefrite/metabolismo , Glomerulonefrite/microbiologia , Inflamação , Interleucina-17/metabolismo , Rim/metabolismo , Túbulos Renais/metabolismo , Masculino , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Infiltração de Neutrófilos/imunologia , Regulação para CimaRESUMO
Immature renal dendritic cells (DCs) are protective early in murine crescentic GN, but the mechanisms underlying this protection are unknown. Here, depletion of DCs reduced the recruitment of invariant natural killer T (iNKT) cells, which attenuate GN, into the kidney in the early stage of experimental crescentic GN. More than 90% of renal iNKT cells expressed the chemokine receptor CXCR6, and renal DCs produced high amounts of the cognate ligand CXCL16 early after induction of nephritis, suggesting that renal DC-derived CXCL16 might attract protective CXCR6(+) iNKT cells. Consistent with this finding, CXCR6-deficient mice exhibited less iNKT cell recruitment and developed nephritis that was more severe, similar to the aggravated nephritis observed in mice depleted of immature DCs. Finally, adoptive transfer of CXCR6-competent NKT cells ameliorated nephritis. Taken together, these results suggest an immunoprotective mechanism involving immature DCs, CXCL16, CXCR6, and regulatory iNKT cells, which might stimulate the development of new therapeutic strategies for GN.
Assuntos
Quimiocina CXCL6/metabolismo , Células Dendríticas/fisiologia , Glomerulonefrite/imunologia , Leucócitos Mononucleares/fisiologia , Receptores CXCR/metabolismo , Animais , Quimiocina CXCL16 , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Receptores CXCR6 , OvinosRESUMO
Interleukin-17A (IL-17) promotes inflammatory renal tissue damage in mouse models of crescentic glomerulonephritis, including murine experimental autoimmune anti-myeloperoxidase glomerulonephritis, which most likely depends on IL-17-producing Th17 cells. In human anti-neutrophil cytoplasmic antibody (ANCA)-associated glomerulonephritis, however, the cellular sources of IL-17 remain to be elucidated. Therefore, we analyzed human kidney biopsies of active necrotizing and crescentic ANCA-associated glomerulonephritis by immunohistochemistry using an IL-17-specific antibody and by immunofluorescent colocalization with cell type markers. We detected numerous IL-17-expressing (IL-17(+)) cells in the glomeruli and in the tubulointerstitium. Unexpectedly, most of these IL-17(+) cells were polymorphonuclear neutrophilic granulocytes, while IL-17(+) T cells and IL-17(+) mast cells were present at significantly lower frequencies. IL-17 was not detected in other infiltrating or resident kidney cells. In those patients who had not received immunosuppressive treatment before biopsy, serum creatinine levels were positively correlated with tubulointerstitial IL-17(+) neutrophils as well as IL-17(+) T cells. Furthermore, we could demonstrate that purified human blood neutrophils expressed IL-17 protein and released it upon stimulation in vitro. In conclusion, these results support a pathogenic role for IL-17 in human ANCA-associated glomerulonephritis. Our data suggest that in the acute stage of the disease neutrophils may act as an important immediate-early innate source of IL-17 and may thereby initiate and promote ongoing renal inflammation. IL-17 may thus be a target for treating acute ANCA-associated glomerulonephritis.
Assuntos
Anticorpos Anticitoplasma de Neutrófilos/metabolismo , Glomerulonefrite/metabolismo , Interleucina-17/metabolismo , Rim/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Anticorpos Anticitoplasma de Neutrófilos/imunologia , Feminino , Glomerulonefrite/imunologia , Humanos , Rim/imunologia , Masculino , Mastócitos/imunologia , Mastócitos/metabolismo , Pessoa de Meia-Idade , Neutrófilos/imunologia , Neutrófilos/metabolismoRESUMO
Th1 and Th17 subtype effector CD4(+) T cells are thought to play a critical role in the pathogenesis of human and experimental crescentic glomerulonephritis. The time course, mechanism, and functions of Th1 and Th17 cell recruitment, and their potential interaction in glomerulonephritis, however, remain to be elucidated. We performed interventional studies using IL-17- and IFN-γ-gene-deficient mice, as well as neutralizing antibodies that demonstrated the importance of the Th17-mediated immune response during the early phase of the disease. At a later stage, we found that Th1 cells were critical mediators of renal tissue injury. Early recruitment of IL-17-producing Th17 cells triggered expression of the chemokine CXCL9 in the kidney that drove the infiltration of Th1 cells bearing its receptor CXCR3. At a later stage, Th1 cell-derived IFN-γ was found to inhibit local chemokine CCL20 expression, acting through its receptor CCR6 on Th17 cells, thereby limiting the renal Th17 immune response. Thus, our findings provide mechanistic evidence for a cytokine-chemokine-driven feedback loop that orchestrates the observed differential Th1 and Th17 cell infiltration into the inflamed kidney. This contributes to the observed time-dependent function of these two major pathogenic effector CD4(+) T cell subsets in crescentic glomerulonephritis.
Assuntos
Comunicação Celular , Quimiocinas/metabolismo , Glomerulonefrite/imunologia , Rim/imunologia , Células Th1/imunologia , Células Th17/imunologia , Transferência Adotiva , Animais , Anticorpos Neutralizantes , Células Cultivadas , Quimiocina CCL20/metabolismo , Quimiocina CXCL9/metabolismo , Quimiocinas/genética , Quimiotaxia de Leucócito , Modelos Animais de Doenças , Retroalimentação Fisiológica , Regulação da Expressão Gênica , Glomerulonefrite/genética , Glomerulonefrite/patologia , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Imunoglobulina G , Interferon gama/deficiência , Interferon gama/genética , Interleucina-17/deficiência , Interleucina-17/genética , Rim/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores CCR6/deficiência , Receptores CCR6/genética , Receptores CXCR3/deficiência , Receptores CXCR3/genética , Ovinos , Transdução de Sinais , Baço/imunologia , Fatores de TempoRESUMO
Estrogen-induced synaptic plasticity was frequently shown by an increase of spines at apical dendrites of CA1 pyramidal neurons after systemic application of estradiol to ovariectomized rats. Surprisingly, exogenous application of estradiol to hippocampal cultures had no effect on spines and on spine synapses, although quantitative immunohistochemistry revealed an upregulation of spinophilin and of synaptophysin, in these cultures. The role of synaptophysin as a presynaptic marker and of spinophilin as a postsynaptic marker, appears questionable from these discrepancies. In contrast, synaptopodin, a marker protein of "mature" mushroom-shaped spines, was downregulated after treatment of hippocampal cultures with estradiol. Synaptopodin is strongly associated to the spine apparatus, a spine-specific cell organelle, which is present in 80% of all mushroom-shaped spines. Consistently, we found a reduction in the number of spines, containing a spine apparatus in response to estradiol, suggesting that the presence of a spine apparatus in many but not all spines is very likely a result of their dynamic character. In summary, synaptic proteins appear to be regulated by estradiol, independent of its function on spine and spine synapse formation.
Assuntos
Estradiol/farmacologia , Estradiol/fisiologia , Hipocampo/efeitos dos fármacos , Sinapses/metabolismo , Animais , Espinhas Dendríticas/metabolismo , Espinhas Dendríticas/ultraestrutura , Regulação para Baixo/efeitos dos fármacos , Hipocampo/metabolismo , Camundongos , Proteínas dos Microfilamentos/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Neurônios/metabolismo , Receptores de Estrogênio/efeitos dos fármacos , Receptores de Estrogênio/metabolismo , Sinapses/ultraestrutura , Sinaptofisina/metabolismo , Regulação para Cima/efeitos dos fármacosRESUMO
Estrogen has been suggested to be pro-epileptic by reducing GABA synthesis, resulting in increased spine density and a decreased threshold for seizures in the hippocampus, which, once they occur, are characterized by a dramatic spine loss in the affected brain areas. As considerable amounts of estradiol are synthesized in the hippocampus, in this study we focused on aromatase, the rate-limiting enzyme in estrogen synthesis in order to examine the role of locally synthesized estrogens in epilepsy. To this end, we first examined the effects of letrozole, a potent aromatase inhibitor, on GABA metabolism in single interneurons of hippocampal dispersion cultures. Letrozole downregulated estradiol release into the medium, as well as glutamate decarboxylase (GAD) expression and GABA synthesis, and decreased the number of GAD positive cells in the cultures. Next, we counted spine synapses and measured estradiol release of hippocampal slice cultures, in which GABA(A) receptors had been blocked by bicuculline, in order to mimic epileptic activity. Treatment of slice cultures with bicuculline resulted in a dramatic decrease in the number of spine synapses and in a significant suppression of estrogen synthesis. The decrease in synapse number in response to bicuculline was restored by combined application of estradiol and bicuculline. Surprisingly, estradiol alone had no effect on either spine synapse number or on GAD expression and GABA synthesis. "Rescue" of synapse number in "epileptic slices" by estradiol and maintenance of GABA metabolism by hippocampus-derived estradiol points to a neuroprotective role of aromatase in epilepsy. Re-filling of estradiol stores after their depletion due to overexcitation may therefore add to therapeutical strategies in epilepsy.