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Background Because of their capacity to induce antigen-specific immunosuppression, tolerogenic dendritic cells are a promising tool for treatment of autoimmune conditions, such as GN caused by autoimmunity against myeloperoxidase (MPO). Methods We sought to generate tolerogenic dendritic cells to suppress anti-MPO GN by culturing bone marrow cells with an NFkB inhibitor (BAY 11-7082) and exposing them to a pulse of MPO. After administering these MPO/BAY dendritic cells or saline to mice with established anti-MPO or anti-methylated BSA (mBSA) immunity, we assessed immune responses and GN. We also examined mechanisms of action of MPO/BAY dendritic cells. Results MPO/BAY dendritic cells decreased anti-MPO immunity and GN without inhibiting immune responses against mBSA; they also induced IL-10-producing regulatory T cells in MPO-immunized mice without affecting IL-10+ CD4+Foxp32 type 1 regulatory T cells or regulatory B cells. MPO/BAY dendritic cells did not inhibit anti-MPO immunity when CD4+Foxp3+ cells were depleted in vivo, showing that regulatory T cells are required for their effects. Coculture experiments with dendritic cells and CD4+Foxp32 or CD4+Foxp3+ cells showed that MPO/BAY dendritic cells generate Foxp3+ regulatory T cells from CD4+Foxp32 cells through several pathways, and induce IL-10+ regulatory T cells via inducible costimulator (ICOS), which was confirmed in vivo. Transfer of MPO/BAY dendritic cell-induced regulatory T cells in vivo, with or without anti-IL-10 receptor antibody, demonstrated that they suppress anti-MPO immunity and GN via IL-10. Conclusions MPO/BAY dendritic cells attenuate established anti-MPO autoimmunity and GN in an antigen-specific manner through ICOS-dependent induction of IL-10-expressing regulatory T cells. This suggests that autoantigen-loaded tolerogenic dendritic cells may represent a novel antigen-specific therapeutic option for anti-MPO GN.Copyright © 2019 by the American Society of Nephrology.

Background Because of their capacity to induce antigen-specific immunosuppression, tolerogenic dendritic cells are a promising tool for treatment of autoimmune conditions, such as GN caused by autoimmunity against myeloperoxidase (MPO). Methods We sought to generate tolerogenic dendritic cells to suppress anti-MPO GN by culturing bone marrow cells with an NFkB inhibitor (BAY 11-7082) and exposing them to a pulse of MPO. After administering these MPO/BAY dendritic cells or saline to mice with established anti-MPO or anti-methylated BSA (mBSA) immunity, we assessed immune responses and GN. We also examined mechanisms of action of MPO/BAY dendritic cells. Results MPO/BAY dendritic cells decreased anti-MPO immunity and GN without inhibiting immune responses against mBSA; they also induced IL-10-producing regulatory T cells in MPO-immunized mice without affecting IL-10+ CD4+Foxp32 type 1 regulatory T cells or regulatory B cells. MPO/BAY dendritic cells did not inhibit anti-MPO immunity when CD4+Foxp3+ cells were depleted in vivo, showing that regulatory T cells are required for their effects. Coculture experiments with dendritic cells and CD4+Foxp32 or CD4+Foxp3+ cells showed that MPO/BAY dendritic cells generate Foxp3+ regulatory T cells from CD4+Foxp32 cells through several pathways, and induce IL-10+ regulatory T cells via inducible costimulator (ICOS), which was confirmed in vivo. Transfer of MPO/BAY dendritic cell-induced regulatory T cells in vivo, with or without anti-IL-10 receptor antibody, demonstrated that they suppress anti-MPO immunity and GN via IL-10. Conclusions MPO/BAY dendritic cells attenuate established anti-MPO autoimmunity and GN in an antigen-specific manner through ICOS-dependent induction of IL-10-expressing regulatory T cells. This suggests that autoantigen-loaded tolerogenic dendritic cells may represent a novel antigen-specific therapeutic option for anti-MPO GN.Copyright © 2019 by the American Society of Nephrology.

Myeloperoxidase (MPO) is important in intracellular microbial killing by neutrophils but extracellularly causes tissue damage. Its role in adaptive immunity and T-cell2mediated 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 inMPO-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 MPOgenerated 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 DCmigration 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.Copyright © 2013 by The American Society of Hematology.

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 disodiumcromoglycate tomast cell-deficientmice had no effect on the development of MPO autoimmunity or GN. MPO-specific CD4+ effector T cell proliferationwas enhanced by co-culture with mast cells, but in the presence of disodiumcromoglycate, 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.Copyright © 2016 by the American Society of Nephrology.

Myeloperoxidase (MPO) anti-neutrophil cytoplasmic Ab (ANCA)-associated vasculitis results from autoimmunity to MPO. IL-17A plays a critical role in generating this form of autoimmune injury but its cell of origin is uncertain. We addressed the hypothesis that IL-17A-producing gd T cells are a nonredundant requisite in the development of MPO autoimmunity and glomerulonephritis (GN).We studied MPO-ANCA GN in wild type, ab, or gd T cell-deficient (C57BL/6, bTCR2/2, and dTCR2/2 respectively) mice. Both T cell populations played important roles in the generation of autoimmunity to MPO and GN. Humoral autoimmunity was dependent on intact ab T cells but was unaffected by gd T cell deletion. Following MPO immunization, activated gd T cells migrate to draining lymph nodes. Studies in dTCR2/2 and transfer of gd T cells to dTCR2/2 mice show that gd T cells facilitate the generation of anti-MPO autoimmunity and GN. dTCR2/2 mice that received IL-17A2/2 gd T cells demonstrate that the development of anti-MPO autoimmunity and GN are dependent on gd T cell IL-17A production. Finally, transfer of anti-MPO CD4+ T cell clones to naive dTCR2/2 and wild type mice with planted glomerular MPO shows that gd T cells are also necessary for recruitment of anti-MPO ab CD4+ effector T cells. This study demonstrates that IL-17A produced by gd T cells plays a critical role in the pathogenesis of MPO-ANCA GN by promoting the development of MPO-specific ab T cells.Copyright © 2017 by The American Association of Immunologists, Inc.

Myeloperoxidase (MPO) is important in intracellular microbial killing by neutrophils but extracellularly causes tissue damage. Its role in adaptive immunity and T-cell2mediated 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 inMPO-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 MPOgenerated 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 DCmigration 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.Copyright © 2013 by The American Society of Hematology.

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 disodiumcromoglycate tomast cell-deficientmice had no effect on the development of MPO autoimmunity or GN. MPO-specific CD4+ effector T cell proliferationwas enhanced by co-culture with mast cells, but in the presence of disodiumcromoglycate, 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.Copyright © 2016 by the American Society of Nephrology.

Neutrophils are no longer seen as leukocytes with a sole function of being the essential first responders in the removal of pathogens at sites of infection. Being armed with numerous pro- and anti-inflammatory mediators, these phagocytes can also contribute to the development of various autoimmune diseases and can positively or negatively regulate the generation of adaptive immune responses. In this review, we will discuss how myeloperoxidase, the most abundant neutrophil granule protein, plays a key role in the various functions of neutrophils in innate and adaptive immunity.Copyright © 2016 Dragana Odobasic et al.

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, Tlr92/2 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 Rag12/2 mice were reconstituted with nonregulatory CD252 splenocytes from wild-type (WT) or Tlr92/2 mice, AKI was similarly enhanced. However, when Rag12/2 mice were reconstituted with CD4+CD25+ regulatory cells, WT CD4+CD25+ cells were more renoprotective and localized to the kidney more efficiently than Tlr92/2 CD4+CD25+ cells. In Tregdepleted Foxp3DTR mice, reconstitution with naive WT CD4+CD25+ cells resulted in less severe AKI than did reconstitution with Tlr92/2 Tregs. Tlr92/2 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 Tlr92/2 mice. In conclusion, we identified a pathway by which TLR9 promotes renal Treg accumulation in AKI.

Neutrophils are no longer seen as leukocytes with a sole function of being the essential first responders in the removal of pathogens at sites of infection. Being armed with numerous pro- and anti-inflammatory mediators, these phagocytes can also contribute to the development of various autoimmune diseases and can positively or negatively regulate the generation of adaptive immune responses. In this review, we will discuss how myeloperoxidase, the most abundant neutrophil granule protein, plays a key role in the various functions of neutrophils in innate and adaptive immunity.Copyright © 2016 Dragana Odobasic et al.

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, Tlr92/2 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 Rag12/2 mice were reconstituted with nonregulatory CD252 splenocytes from wild-type (WT) or Tlr92/2 mice, AKI was similarly enhanced. However, when Rag12/2 mice were reconstituted with CD4+CD25+ regulatory cells, WT CD4+CD25+ cells were more renoprotective and localized to the kidney more efficiently than Tlr92/2 CD4+CD25+ cells. In Tregdepleted Foxp3DTR mice, reconstitution with naive WT CD4+CD25+ cells resulted in less severe AKI than did reconstitution with Tlr92/2 Tregs. Tlr92/2 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 Tlr92/2 mice. In conclusion, we identified a pathway by which TLR9 promotes renal Treg accumulation in AKI.

Many forms of glomerulonephritis are mediated by autoimmunity. While autoantibodies are often pathogenic, cell-mediated immunity plays an important role in a number of forms of rapidly progressive glomerulonephritis. This unit describes the induction of cell-mediated autoimmune glomerular disease in mice. One disease model, experimental anti-glomerular basement membrane (GBM) disease, features autoreactivity to a well-defined component of type IV collagen found in the GBM, a3(IV)NC1. The other models the cell-mediated effector response in forms of renal vasculitis, where autoantibodies to myeloperoxidase result in systemic neutrophil activation, resulting in their localization to the glomerulus and the subsequent deposition of myeloperoxidase within glomerular capillaries. There, myeloperoxidase acts as a "planted" autoantigen and is recognized by effector autoreactive myeloperoxidase-specific T cells. These models are useful both in defining mechanisms germane to the development of autoimmunity to a3(IV)NC1 and myeloperoxidase, and in dissecting the role of cell-mediated responses in effecting glomerular injury.Copyright © 2014 by John Wiley & Sons, Inc.

The glomerulonephritides are diseases characterized by immune-mediated glomerular inflammation. Most severe and rapidly progressive forms of glomerulonephritis feature the participation of injurious leukocytes that localize to glomeruli. This unit describes classical models of rapidly progressive glomerulonephritis in mice, induced by injecting heterologous globulin (raised in sheep) that binds to the glomerular basement membrane. These models have been particularly useful in defining the participation of effector leukocytes in severe glomerular disease. In these models, injury typically occurs in two phases. In the initial, heterologous phase, injury is mediated by the globulin bound within the glomerulus acting as an antibody. The later, autologous phase of injury is mediated by the host's adaptive immunity to the heterologous globulin now functioning as a planted foreign antigen within glomeruli. As autologous phase injury is driven by immunity to sheep globulin, assessment of antigen-specific systemic immunity to sheep globulin is critical when using this model.Copyright © 2014 by John Wiley & Sons, Inc.

Many forms of glomerulonephritis are mediated by autoimmunity. While autoantibodies are often pathogenic, cell-mediated immunity plays an important role in a number of forms of rapidly progressive glomerulonephritis. This unit describes the induction of cell-mediated autoimmune glomerular disease in mice. One disease model, experimental anti-glomerular basement membrane (GBM) disease, features autoreactivity to a well-defined component of type IV collagen found in the GBM, a3(IV)NC1. The other models the cell-mediated effector response in forms of renal vasculitis, where autoantibodies to myeloperoxidase result in systemic neutrophil activation, resulting in their localization to the glomerulus and the subsequent deposition of myeloperoxidase within glomerular capillaries. There, myeloperoxidase acts as a "planted" autoantigen and is recognized by effector autoreactive myeloperoxidase-specific T cells. These models are useful both in defining mechanisms germane to the development of autoimmunity to a3(IV)NC1 and myeloperoxidase, and in dissecting the role of cell-mediated responses in effecting glomerular injury.Copyright © 2014 by John Wiley & Sons, Inc.

The glomerulonephritides are diseases characterized by immune-mediated glomerular inflammation. Most severe and rapidly progressive forms of glomerulonephritis feature the participation of injurious leukocytes that localize to glomeruli. This unit describes classical models of rapidly progressive glomerulonephritis in mice, induced by injecting heterologous globulin (raised in sheep) that binds to the glomerular basement membrane. These models have been particularly useful in defining the participation of effector leukocytes in severe glomerular disease. In these models, injury typically occurs in two phases. In the initial, heterologous phase, injury is mediated by the globulin bound within the glomerulus acting as an antibody. The later, autologous phase of injury is mediated by the host's adaptive immunity to the heterologous globulin now functioning as a planted foreign antigen within glomeruli. As autologous phase injury is driven by immunity to sheep globulin, assessment of antigen-specific systemic immunity to sheep globulin is critical when using this model.Copyright © 2014 by John Wiley & Sons, Inc.

Objective Myeloperoxidase (MPO) locally contributes to organ damage in various chronic inflammatory conditions by generating reactive intermediates. The contribution of MPO in the development of experimental lupus is unknown. The aim of this study was to define the role of MPO in murine lupus nephritis (LN). Methods LN was induced in C57BL/6 wild-type (WT) and MPO knockout (MPO-/-) mice by intraperitoneal injection of pristane. Autoimmunity and glomerulonephritis were assessed 20 and 40 weeks after pristane administration. Cell apoptosis, leukocyte accumulation, and cytokine levels in the peritoneal cavity of WT and MPO-/- mice were assessed 3 or 6 days after pristane injection. Results MPO-/- mice developed more severe nephritis than did WT mice 20 and 40 weeks after pristane injection, despite having reduced glomerular deposition of antibody and complement and diminished levels of markers of oxidative stress (oxidized DNA and glutathione sulfonamide). Enhancement of renal disease in MPO-deficient mice correlated with increased accumulation of CD4+ T cells and macrophages in glomeruli, which, in turn, was associated with augmented generation of CD4+ T cell responses and increased activation and migration of dendritic cells in secondary lymphoid organs. In addition, the enhanced renal injury in MPO-/- mice was associated with increased glomerular accumulation of neutrophils and deposition of neutrophil extracellular traps. MPO deficiency also increased early cell apoptosis, leukocyte accumulation, and proinflammatory cytokine expression in the peritoneum. Conclusion MPO attenuates pristane-induced LN by inhibiting early inflammatory responses in the peritoneum and limiting the generation of CD4+ T cell autoimmunity in secondary lymphoid organs.Copyright © 2015, American College of Rheumatology.

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 Rorgammat-/- mice, as well as in mice treated pre-emptively with anti-IL-17A antibodies. Both renal injury and renal IL-1beta 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-1beta 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 gammadelta T cells, IL-17A from those cells does not contribute to renal injury. Cisplatin-induced injury was unchanged in gammadelta 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. © 2014 American Society for Investigative Pathology.

Objective Myeloperoxidase (MPO) is implicated as a local mediator of tissue damage when released extracellularly in many chronic inflammatory diseases. The purpose of this study was to explore the role of endogenous MPO in experimental rheumatoid arthritis (RA). Methods K/BxN serum-transfer arthritis was induced in C57BL/6 wild-type (WT) and MPO knockout (MPO-/-) mice, and disease development was assessed. MPO activity was measured in joint tissues from mice with or without K/BxN arthritis. Collagen-induced arthritis (CIA) was induced in WT and MPO-/- mice, and disease development and immune responses were examined. MPO expression was assessed in synovial biopsy samples from patients with active RA, and the effect of MPO on synovial fibroblasts was tested in vitro. Results MPO was up-regulated in the joints of mice with K/BxN arthritis, and MPO deficiency attenuated the severity of the disease without affecting circulating cytokine levels. In CIA, MPO-/- mice had enhanced CD4+ T cell responses and reduced frequency of regulatory T cells in the lymph nodes and spleen, as well as augmented interleukin-17A and diminished interferon-gamma secretion by collagen-stimulated splenocytes, without an effect on circulating anticollagen antibody levels. Despite enhanced adaptive immunity in secondary lymphoid organs, CIA development was attenuated in MPO-/- mice. Intracellular and extracellular MPO was detected in the synovium of patients with active RA, and human MPO enhanced the proliferation and decreased the apoptosis of synovial fibroblasts in vitro. Conclusion MPO contributes to the development of arthritis despite suppressing adaptive immunity in secondary lymphoid organs. This suggests distinct effects of local MPO on arthritogenic effector responses. Copyright © 2014 by the American College of Rheumatology.

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 Rorgammat-/- mice, as well as in mice treated pre-emptively with anti-IL-17A antibodies. Both renal injury and renal IL-1beta 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-1beta 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 gammadelta T cells, IL-17A from those cells does not contribute to renal injury. Cisplatin-induced injury was unchanged in gammadelta 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. © 2014 American Society for Investigative Pathology.

Objective Myeloperoxidase (MPO) is implicated as a local mediator of tissue damage when released extracellularly in many chronic inflammatory diseases. The purpose of this study was to explore the role of endogenous MPO in experimental rheumatoid arthritis (RA). Methods K/BxN serum-transfer arthritis was induced in C57BL/6 wild-type (WT) and MPO knockout (MPO-/-) mice, and disease development was assessed. MPO activity was measured in joint tissues from mice with or without K/BxN arthritis. Collagen-induced arthritis (CIA) was induced in WT and MPO-/- mice, and disease development and immune responses were examined. MPO expression was assessed in synovial biopsy samples from patients with active RA, and the effect of MPO on synovial fibroblasts was tested in vitro. Results MPO was up-regulated in the joints of mice with K/BxN arthritis, and MPO deficiency attenuated the severity of the disease without affecting circulating cytokine levels. In CIA, MPO-/- mice had enhanced CD4+ T cell responses and reduced frequency of regulatory T cells in the lymph nodes and spleen, as well as augmented interleukin-17A and diminished interferon-gamma secretion by collagen-stimulated splenocytes, without an effect on circulating anticollagen antibody levels. Despite enhanced adaptive immunity in secondary lymphoid organs, CIA development was attenuated in MPO-/- mice. Intracellular and extracellular MPO was detected in the synovium of patients with active RA, and human MPO enhanced the proliferation and decreased the apoptosis of synovial fibroblasts in vitro. Conclusion MPO contributes to the development of arthritis despite suppressing adaptive immunity in secondary lymphoid organs. This suggests distinct effects of local MPO on arthritogenic effector responses. Copyright © 2014 by the American College of Rheumatology.

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.

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.

Myeloperoxidase (MPO) is an enzyme that is found in neutrophils and monocytes/macrophages. Intracellularly, it plays a major role in microbial killing, but extracellularly, it may cause host tissue damage. The role of endogenous MPO was studied during neuhophil-mediated (heterologous) and T helper 1 (Th1)/macrophage-mediated (autologous) phases of crescentic glomerulonephritis. Glomerulonephritis was induced in C57BL/6 wild-type (WT) and MPO-deficient (MPO-/-) mice by intravenous injection of sheep anti-mouse glomerular basement membrane globulin. MPO activity was increased in kidneys of WT mice during both the heterologous and autologous phases of glomerulonephritis. During the heterologous phase of glomerulonephritis, proteinuria was decreased, whereas glomerular neutrophil accumulation and F-selectin expression were enhanced in MPO-/- mice. In the autologous, crescentic phase of glomerulonephritis, MPO-/- mice had increased accumulation of CD4+ cells and macrophages in glomeruli compared with WT mice. However, no difference in renal injury (crescent formation, proteinuria, and serum creatinine levels) was observed. Neutrophils and macrophages from MPO-/- mice exhibited reduced production of reactive oxygen species. Assessment of systemic immunity to sheep globulin showed that MPO-/- mice had increased splenic CD4+ cell proliferation, cytokine production, and dermal delayed-type hypersensitivity, as well as enhanced levels of circulating IgG, IgG1, and IgG3. MPO-/- mice also had an augmented Th1:Th2 ratio compared with WT mice (IFN-gamma:IL-4 and IgG3:IgG1 ratios). These results suggest that endogenous MPO locally contributes to glomerular damage during neutrophil-mediated glomerulonephritis, whereas it attenuates initiation of the adaptive immune response inducing crescentic, autologous-phase glomerulonephritis by suppressing T cell proliferation, cytokine production, and Th1:Th2 ratio. Copyright © 2007 by the American Society of Nephrology.

The inducible co-stimulatory molecule (ICOS)/ICOS ligand (ICOSL) co-stimulatory pathway is critical in T cell activation, differentiation, and effector function. Its role was investigated in a model of Th1-driven crescentic glomerulonephritis (GN). GN was induced by sensitizing mice to sheep globulin (day 0) and challenging them with sheep anti-mouse glomerular basement membrane antibody (Ab; day 10). Disease and immune responses were assessed on day 20. For testing the role of ICOSL in the induction of GN, control or anti-ICOSL mAb were administered from days 0 to 8. For examining the role of ICOSL in the effector phase of GN, treatment lasted from days 10 to 18. Blockade of ICOSL during the induction of GN increased glomerular accumulation of CD4+ T cells and macrophages and augmented renal injury. These results correlated with attenuated splenocyte production of protective Th2 cytokines IL-4 and IL-10 and decreased apoptosis of splenic CD4+ T cells. ICOSL was upregulated within glomeruli of mice with GN. Inhibition of ICOSL during the effector phase of GN enhanced glomerular T cell and macrophage accumulation and augmented disease, without affecting the systemic immune response (cytokine production, T cell apoptosis/proliferation, Ab levels). Increased presence of leukocytes in glomeruli of mice that received anti-ICOSL mAb was associated with enhanced cellular proliferation and upregulation of P-selectin and intercellular adhesion molecule-1 within glomeruli. These studies demonstrate that ICOSL is protective during the induction of GN by augmenting Th2 responses and CD4+ T cell apoptosis. They also show that ICOSL is upregulated in nephritic glomeruli, where it locally reduces accumulation of T cells and macrophages and attenuates renal injury. Copyright © 2006 by the American Society of Nephrology.

Most humans with microscopic polyarteritis and anti-myeloperoxidase (anti-MPO), anti-neutrophil cytoplasmic antibodies (ANCA) develop "pauci-immune" crescentic glomerulonephritis. For dissection of the roles of ANCA and cell-mediated effectors in microscopic polyarteritis, experimental autoimmune anti-MPO glomerulonephritis was induced by immunizing C57BL/6 mice with human MPO. Autoimmunity to mouse MPO (ANCA and CD4+ cell reactivity) was induced. Challenge with anti-glomerular basement membrane globulin resulted in accumulation of neutrophils, CD4+ cells and macrophages, and significant numbers of crescentic glomeruli compared with similarly challenged control-immunized mice. MPO-deficient (Mpo-/-) mice immunized with MPO developed similar immune responses to MPO but failed to recruit effector cells to glomeruli or develop significant crescent formation, suggesting that MPO is acting as a planted glomerular autoantigen. Effector CD4+ cell depletion in this model attenuated crescentic glomerulonephritis and effector cell influx without altering ANCA titers. However, B cell-deficient mice, with no ANCA, still developed severe crescentic glomerulonephritis with accumulation of effector cells. Intravital microscopy studies demonstrated that passive transfer of sera from MPO-immunized Mpo -/- mice to LPS-primed mice rapidly induced glomerular neutrophil accumulation and release of MPO. These studies provide in vivo evidence in a relevant vascular bed for both humoral and cellular anti-MPO responses as key inducers of injury. ANCA induces glomerular neutrophil infiltration and MPO deposition. Subsequently, anti-MPO CD4+ cells recognize MPO as a planted glomerular antigen and act with macrophages to amplify severe glomerular injury. Copyright © 2006 by the American Society of Nephrology.

T-bet is a transcription factor that is essential for T helper (Th)1 lineage commitment and optimal IFN-gamma production by CD4+ T cells. We examined the role of T-bet in the development of experimental crescentic glomerulonephritis, which is induced by Th1-predominant, delayed-type hypersensitivity-like responses directed against a nephritogenic antigen. Anti-glomerular basement membrane (GBM) glomerulonephritis was induced in T-bet-/- and wild-type C57BL/6 mice. Compared with wild-type controls, renal injury was attenuated in T-bet-/- mice with glomerulonephritis, evidenced by less proteinuria, glomerular crescents, and tubulointerstitial inflammation. Accumulation of glomerular CD4+ T cells and macrophages was decreased, and was associated with reduced intrarenal expression of the potent Th1 chemoattractants CCL5/RANTES and CXCL9/Mig. Supporting the pro-inflammatory nature of T-bet signaling, assessment of systemic immunity confirmed that T-bet-/- mice had a reduction in Th1 immunity. The kinetic profile of T-bet mRNA in wild-type mice supported the hypothesis that T-bet deficiency attenuates renal injury in part by shifting the Th1/Th2 balance away from a Th1 phenotype. Expression of renal and splenic IL-17A, characteristically expressed by the Th17 subset of effector T cells, which have been implicated in the pathogenesis of autoimmune disease, was increased in T-bet-/- mice. We conclude that T-bet directs Th1 responses that induce renal injury in experimental crescentic glomerulonephritis. Copyright © 2008 by the American Society of Nephrology.

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. Method(s): 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. Result(s): MPO alone induced low-titer antineutrophil cytoplasmic antibodies (ANCAs) without delayedtype 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 kappat-dependent interleukin-17A (IL-17A) production. TLR-9 ligand promoted Th1 autoimmunity, with enhanced production of interferon-kappa (IFNkappa) 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 IFNkappa was neutralized. Conclusion(s): 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. © 2011, American College of Rheumatology.

Myeloperoxidase (MPO) is an enzyme that is found in neutrophils and monocytes/macrophages. Intracellularly, it plays a major role in microbial killing, but extracellularly, it may cause host tissue damage. The role of endogenous MPO was studied during neuhophil-mediated (heterologous) and T helper 1 (Th1)/macrophage-mediated (autologous) phases of crescentic glomerulonephritis. Glomerulonephritis was induced in C57BL/6 wild-type (WT) and MPO-deficient (MPO-/-) mice by intravenous injection of sheep anti-mouse glomerular basement membrane globulin. MPO activity was increased in kidneys of WT mice during both the heterologous and autologous phases of glomerulonephritis. During the heterologous phase of glomerulonephritis, proteinuria was decreased, whereas glomerular neutrophil accumulation and F-selectin expression were enhanced in MPO-/- mice. In the autologous, crescentic phase of glomerulonephritis, MPO-/- mice had increased accumulation of CD4+ cells and macrophages in glomeruli compared with WT mice. However, no difference in renal injury (crescent formation, proteinuria, and serum creatinine levels) was observed. Neutrophils and macrophages from MPO-/- mice exhibited reduced production of reactive oxygen species. Assessment of systemic immunity to sheep globulin showed that MPO-/- mice had increased splenic CD4+ cell proliferation, cytokine production, and dermal delayed-type hypersensitivity, as well as enhanced levels of circulating IgG, IgG1, and IgG3. MPO-/- mice also had an augmented Th1:Th2 ratio compared with WT mice (IFN-gamma:IL-4 and IgG3:IgG1 ratios). These results suggest that endogenous MPO locally contributes to glomerular damage during neutrophil-mediated glomerulonephritis, whereas it attenuates initiation of the adaptive immune response inducing crescentic, autologous-phase glomerulonephritis by suppressing T cell proliferation, cytokine production, and Th1:Th2 ratio. Copyright © 2007 by the American Society of Nephrology.

The inducible co-stimulatory molecule (ICOS)/ICOS ligand (ICOSL) co-stimulatory pathway is critical in T cell activation, differentiation, and effector function. Its role was investigated in a model of Th1-driven crescentic glomerulonephritis (GN). GN was induced by sensitizing mice to sheep globulin (day 0) and challenging them with sheep anti-mouse glomerular basement membrane antibody (Ab; day 10). Disease and immune responses were assessed on day 20. For testing the role of ICOSL in the induction of GN, control or anti-ICOSL mAb were administered from days 0 to 8. For examining the role of ICOSL in the effector phase of GN, treatment lasted from days 10 to 18. Blockade of ICOSL during the induction of GN increased glomerular accumulation of CD4+ T cells and macrophages and augmented renal injury. These results correlated with attenuated splenocyte production of protective Th2 cytokines IL-4 and IL-10 and decreased apoptosis of splenic CD4+ T cells. ICOSL was upregulated within glomeruli of mice with GN. Inhibition of ICOSL during the effector phase of GN enhanced glomerular T cell and macrophage accumulation and augmented disease, without affecting the systemic immune response (cytokine production, T cell apoptosis/proliferation, Ab levels). Increased presence of leukocytes in glomeruli of mice that received anti-ICOSL mAb was associated with enhanced cellular proliferation and upregulation of P-selectin and intercellular adhesion molecule-1 within glomeruli. These studies demonstrate that ICOSL is protective during the induction of GN by augmenting Th2 responses and CD4+ T cell apoptosis. They also show that ICOSL is upregulated in nephritic glomeruli, where it locally reduces accumulation of T cells and macrophages and attenuates renal injury. Copyright © 2006 by the American Society of Nephrology.

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 (KitW-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 cellsmigrated fromthe 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. Copyright © 2012 by the American Society of Nephrology.

Most humans with microscopic polyarteritis and anti-myeloperoxidase (anti-MPO), anti-neutrophil cytoplasmic antibodies (ANCA) develop "pauci-immune" crescentic glomerulonephritis. For dissection of the roles of ANCA and cell-mediated effectors in microscopic polyarteritis, experimental autoimmune anti-MPO glomerulonephritis was induced by immunizing C57BL/6 mice with human MPO. Autoimmunity to mouse MPO (ANCA and CD4+ cell reactivity) was induced. Challenge with anti-glomerular basement membrane globulin resulted in accumulation of neutrophils, CD4+ cells and macrophages, and significant numbers of crescentic glomeruli compared with similarly challenged control-immunized mice. MPO-deficient (Mpo-/-) mice immunized with MPO developed similar immune responses to MPO but failed to recruit effector cells to glomeruli or develop significant crescent formation, suggesting that MPO is acting as a planted glomerular autoantigen. Effector CD4+ cell depletion in this model attenuated crescentic glomerulonephritis and effector cell influx without altering ANCA titers. However, B cell-deficient mice, with no ANCA, still developed severe crescentic glomerulonephritis with accumulation of effector cells. Intravital microscopy studies demonstrated that passive transfer of sera from MPO-immunized Mpo -/- mice to LPS-primed mice rapidly induced glomerular neutrophil accumulation and release of MPO. These studies provide in vivo evidence in a relevant vascular bed for both humoral and cellular anti-MPO responses as key inducers of injury. ANCA induces glomerular neutrophil infiltration and MPO deposition. Subsequently, anti-MPO CD4+ cells recognize MPO as a planted glomerular antigen and act with macrophages to amplify severe glomerular injury. Copyright © 2006 by the American Society of Nephrology.

T-bet is a transcription factor that is essential for T helper (Th)1 lineage commitment and optimal IFN-gamma production by CD4+ T cells. We examined the role of T-bet in the development of experimental crescentic glomerulonephritis, which is induced by Th1-predominant, delayed-type hypersensitivity-like responses directed against a nephritogenic antigen. Anti-glomerular basement membrane (GBM) glomerulonephritis was induced in T-bet-/- and wild-type C57BL/6 mice. Compared with wild-type controls, renal injury was attenuated in T-bet-/- mice with glomerulonephritis, evidenced by less proteinuria, glomerular crescents, and tubulointerstitial inflammation. Accumulation of glomerular CD4+ T cells and macrophages was decreased, and was associated with reduced intrarenal expression of the potent Th1 chemoattractants CCL5/RANTES and CXCL9/Mig. Supporting the pro-inflammatory nature of T-bet signaling, assessment of systemic immunity confirmed that T-bet-/- mice had a reduction in Th1 immunity. The kinetic profile of T-bet mRNA in wild-type mice supported the hypothesis that T-bet deficiency attenuates renal injury in part by shifting the Th1/Th2 balance away from a Th1 phenotype. Expression of renal and splenic IL-17A, characteristically expressed by the Th17 subset of effector T cells, which have been implicated in the pathogenesis of autoimmune disease, was increased in T-bet-/- mice. We conclude that T-bet directs Th1 responses that induce renal injury in experimental crescentic glomerulonephritis. Copyright © 2008 by the American Society of Nephrology.

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. Method(s): 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. Result(s): MPO alone induced low-titer antineutrophil cytoplasmic antibodies (ANCAs) without delayedtype 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 kappat-dependent interleukin-17A (IL-17A) production. TLR-9 ligand promoted Th1 autoimmunity, with enhanced production of interferon-kappa (IFNkappa) 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 IFNkappa was neutralized. Conclusion(s): 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. © 2011, American College of Rheumatology.

Experimental crescentic glomerulonephritis (GN) is characterized by T helper 1 (Th1) directed nephritogenic immune responses and cell-mediated glomerular injury. IL-12p40, the common cytokine chain for both IL-12 and IL-23, is important in the generation and potentially the maintenance of Th1 responses, whereas IL-18 is a co-factor for Th1 responses that may have systemic and local proinflammatory effects. For testing the hypothesis that both endogenous IL-12p40 and endogenous IL-18 play pathogenetic roles in crescentic GN, accelerated anti-glomerular basement membrane GN was induced in mice genetically deficient in IL-12p40 (IL-12p40-/-), IL-18 (IL-18 -/-), or both IL-12p40 and IL-18 (IL-12p4-/-IL-18 -/-). Compared with wild-type C57BL/6 mice, IL-12p40-/- mice failed to make a nephritogenic Th1 response and developed markedly reduced crescent formation and renal leukocytic infiltration, despite renal production of chemoattractants and adhesion molecules. IL-18-/- mice developed an intact antigen-specific systemic Th1 response, a similar degree of crescent formation, but fewer glomeruli affected by other severe histologic changes and fewer leukocytes in glomeruli and interstitium. IL-18 was expressed within diseased kidneys. Local production of TNF, IL-1beta, IFN-gamma, CCL3 (MIP-1alpha), and CCL4 (MIP-1beta) was reduced in IL-18-/- mice, demonstrating a local proinflammatory role for IL-18. Combined deletion of IL-12p40 and IL-18 did not result in synergistic effects. Consistent with the hypothesis that inflammation leads to fibrosis, all three groups of deficient mice expressed lower levels of intrarenal TGF-beta1 and/or alpha1(I) procollagen mRNA. These studies demonstrate that in severe experimental crescentic GN, IL-12p40 is the key Th1-defining cytokine chain, whereas IL-18 has local proinflammatory roles. Copyright © 2005 by the American Society of Nephrology.

The participation of renal expression of CD80 and CD86 in the immunopathogenesis of crescentic Th1-mediated antiglomerular basement membrane (anti-GBM) glomerulonephritis (GN) has not been assessed. Immunohistochemical staining demonstrated prominent upregulation of both molecules in glomeruli of mice with anti-GBM GN, suggesting a potential role for the local expression of CD80 and CD86 in nephritogenic effector T cell responses. For testing this hypothesis, control or inhibitory anti-CD80 and/or anti-CD86 mAb were administered to mice during the effector phase of the disease but after the establishment of a systemic immune response. Anti-CD80 or anti-CD86 mAb treatment had no effect on the development of GN or infiltration of leukocytes into glomeruli; however, administration of anti-CD80/86 mAb attenuated glomerular accumulation of CD4+ T cells and macrophages, crescent formation, and proteinuria, correlating with reduced antigen-specific skin delayed-type hypersensitivity. Attenuated glomerular infiltration of leukocytes in mice that were treated with anti-CD80/86 mAb was associated with decreased intraglomerular expression of adhesion molecules P-selectin and intercellular adhesion molecule-1, as well as attenuated renal mRNA levels of proinflammatory cytokines IFN-gamma and migration inhibitory factor, without reducing chemokine and chemokine receptor expression in the kidney or intraglomerular apoptosis and proliferation. The systemic Th1/Th2 balance (assessed by splenocyte production of IFN-gamma and IL-4 and circulating levels of IgG1 and IgG2a) was not affected by the inhibition of CD80 and CD86. These studies show that CD80 and CD86 are expressed in glomeruli of mice with crescentic anti-GBM GN, in which they play a critical role in facilitating accumulation of Th1 effectors and macrophages, thus exacerbating renal injury. Copyright © 2005 by the American Society of Nephrology.

Experimental crescentic glomerulonephritis (GN) is characterized by T helper 1 (Th1) directed nephritogenic immune responses and cell-mediated glomerular injury. IL-12p40, the common cytokine chain for both IL-12 and IL-23, is important in the generation and potentially the maintenance of Th1 responses, whereas IL-18 is a co-factor for Th1 responses that may have systemic and local proinflammatory effects. For testing the hypothesis that both endogenous IL-12p40 and endogenous IL-18 play pathogenetic roles in crescentic GN, accelerated anti-glomerular basement membrane GN was induced in mice genetically deficient in IL-12p40 (IL-12p40-/-), IL-18 (IL-18 -/-), or both IL-12p40 and IL-18 (IL-12p4-/-IL-18 -/-). Compared with wild-type C57BL/6 mice, IL-12p40-/- mice failed to make a nephritogenic Th1 response and developed markedly reduced crescent formation and renal leukocytic infiltration, despite renal production of chemoattractants and adhesion molecules. IL-18-/- mice developed an intact antigen-specific systemic Th1 response, a similar degree of crescent formation, but fewer glomeruli affected by other severe histologic changes and fewer leukocytes in glomeruli and interstitium. IL-18 was expressed within diseased kidneys. Local production of TNF, IL-1beta, IFN-gamma, CCL3 (MIP-1alpha), and CCL4 (MIP-1beta) was reduced in IL-18-/- mice, demonstrating a local proinflammatory role for IL-18. Combined deletion of IL-12p40 and IL-18 did not result in synergistic effects. Consistent with the hypothesis that inflammation leads to fibrosis, all three groups of deficient mice expressed lower levels of intrarenal TGF-beta1 and/or alpha1(I) procollagen mRNA. These studies demonstrate that in severe experimental crescentic GN, IL-12p40 is the key Th1-defining cytokine chain, whereas IL-18 has local proinflammatory roles. Copyright © 2005 by the American Society of Nephrology.

The participation of renal expression of CD80 and CD86 in the immunopathogenesis of crescentic Th1-mediated antiglomerular basement membrane (anti-GBM) glomerulonephritis (GN) has not been assessed. Immunohistochemical staining demonstrated prominent upregulation of both molecules in glomeruli of mice with anti-GBM GN, suggesting a potential role for the local expression of CD80 and CD86 in nephritogenic effector T cell responses. For testing this hypothesis, control or inhibitory anti-CD80 and/or anti-CD86 mAb were administered to mice during the effector phase of the disease but after the establishment of a systemic immune response. Anti-CD80 or anti-CD86 mAb treatment had no effect on the development of GN or infiltration of leukocytes into glomeruli; however, administration of anti-CD80/86 mAb attenuated glomerular accumulation of CD4+ T cells and macrophages, crescent formation, and proteinuria, correlating with reduced antigen-specific skin delayed-type hypersensitivity. Attenuated glomerular infiltration of leukocytes in mice that were treated with anti-CD80/86 mAb was associated with decreased intraglomerular expression of adhesion molecules P-selectin and intercellular adhesion molecule-1, as well as attenuated renal mRNA levels of proinflammatory cytokines IFN-gamma and migration inhibitory factor, without reducing chemokine and chemokine receptor expression in the kidney or intraglomerular apoptosis and proliferation. The systemic Th1/Th2 balance (assessed by splenocyte production of IFN-gamma and IL-4 and circulating levels of IgG1 and IgG2a) was not affected by the inhibition of CD80 and CD86. These studies show that CD80 and CD86 are expressed in glomeruli of mice with crescentic anti-GBM GN, in which they play a critical role in facilitating accumulation of Th1 effectors and macrophages, thus exacerbating renal injury. Copyright © 2005 by the American Society of Nephrology.

BACKGROUND: CD80 and CD86 costimulatory molecules have been shown to affect the induction of Th1-mediated crescentic antiglomerular basement membrane (GBM) antibody-initiated glomerulonephritis (GN). The aim of the current studies was to define the mechanisms by which CD80 and CD86 regulate the development of this disease. METHOD(S): Anti-GBM GN was induced in CD80-/-, CD86-/-, and CD80/86-/- mice, as well as in C57BL/6 controls. Renal injury and immune responses were assessed after 21 days. To examine whether costimulation by OX40-ligand compensates for the absence of CD80 and CD86 in inducing GN, OX40-ligand was blocked in wild-type and CD80/86-/- mice. RESULT(S): Crescentic GN and glomerular accumulation of CD4+ T cells and macrophages were attenuated in CD80-/- mice, correlating with significantly enhanced apoptosis and decreased proliferation of spleen CD4+ T cells. GN was exacerbated in CD86-/- mice, which was associated with attenuated IL-4 and enhanced IFN-gamma levels. In contrast, CD80/86-/- mice developed crescentic GN similar to that in controls. Inhibition of OX40-ligand exacerbated GN in wild-type mice by enhancing IFN-gamma production, and attenuated disease in CD80/86-/- mice by reducing glomerular CD4+ T-cell and macrophage accumulation. CONCLUSION(S): CD80 is pathogenic in crescentic GN by enhancing survival and proliferation of CD4+ T cells, whereas CD86 is protective by enhancing Th2 and attenuating Th1 responses. Furthermore, in the presence of CD80 and CD86, OX40-ligand attenuates, whereas in their absence it enhances GN, suggesting that, in the absence of CD80 and CD86, the OX40/OX40-ligand pathway is an alternative costimulatory pathway in inducing crescentic GN.

BACKGROUND: CD80 and CD86 costimulatory molecules have been shown to affect the induction of Th1-mediated crescentic antiglomerular basement membrane (GBM) antibody-initiated glomerulonephritis (GN). The aim of the current studies was to define the mechanisms by which CD80 and CD86 regulate the development of this disease. METHOD(S): Anti-GBM GN was induced in CD80-/-, CD86-/-, and CD80/86-/- mice, as well as in C57BL/6 controls. Renal injury and immune responses were assessed after 21 days. To examine whether costimulation by OX40-ligand compensates for the absence of CD80 and CD86 in inducing GN, OX40-ligand was blocked in wild-type and CD80/86-/- mice. RESULT(S): Crescentic GN and glomerular accumulation of CD4+ T cells and macrophages were attenuated in CD80-/- mice, correlating with significantly enhanced apoptosis and decreased proliferation of spleen CD4+ T cells. GN was exacerbated in CD86-/- mice, which was associated with attenuated IL-4 and enhanced IFN-gamma levels. In contrast, CD80/86-/- mice developed crescentic GN similar to that in controls. Inhibition of OX40-ligand exacerbated GN in wild-type mice by enhancing IFN-gamma production, and attenuated disease in CD80/86-/- mice by reducing glomerular CD4+ T-cell and macrophage accumulation. CONCLUSION(S): CD80 is pathogenic in crescentic GN by enhancing survival and proliferation of CD4+ T cells, whereas CD86 is protective by enhancing Th2 and attenuating Th1 responses. Furthermore, in the presence of CD80 and CD86, OX40-ligand attenuates, whereas in their absence it enhances GN, suggesting that, in the absence of CD80 and CD86, the OX40/OX40-ligand pathway is an alternative costimulatory pathway in inducing crescentic GN.