Hans-Joachim Paust

The IL-23/Th17 Axis Contributes to Renal Injury in Experimental Glomerulonephritis

T cells infiltrate the kidney in both human and experimental glomerulonephritis, and several lines of evidence indicate that T cell-mediated tissue damage plays an important role in the immunopathogenesis of renal inflammatory diseases. However, the functions of the different T cell subsets, particularly the recently identified interleukin-17 (IL-17)-producing T cells (Th17 cells), are incompletely understood in glomerulonephritis. Here, we identified renal IL-17-producing T cells in the T cell-mediated model of nephrotoxic nephritis in mice. In vitro, IL-17 enhanced the production of the proinflammatory chemokines CCL2/MCP-1, CCL3/MIP-1alpha, and CCL20/LARC, which are implicated in the recruitment of T cells and monocytes, in mouse mesangial cells. To determine the function of Th17 cells in renal inflammation, we induced nephrotoxic nephritis in IL-23 p19(-/-) mice, which have reduced numbers of Th17 cells, and in IL-17(-/-) mice, which are deficient in the effector cytokine IL-17 itself. In comparison with nephritic wild-type mice, IL-23 p19(-/-) mice demonstrated less infiltration of Th17 cells, and both IL-23 p19(-/-) and IL-17(-/-) mice developed less severe nephritis as measured by renal function, albuminuria, and frequency of glomerular crescent formation. These results demonstrate that the IL-23/IL-17 pathway significantly contributes to renal tissue injury in experimental glomerulonephritis. Targeting the IL-23/Th17 axis may be a promising therapeutic strategy for the treatment of proliferative and crescentic glomerulonephritis.

CCR6 Recruits Regulatory T Cells and Th17 Cells to the Kidney in Glomerulonephritis

T cells recruited to the kidney contribute to tissue damage in crescentic and proliferative glomerulonephritides. Chemokines and their receptors regulate T cell trafficking, but the expression profile and functional importance of chemokine receptors for renal CD4+ T cell subsets are incompletely understood. In this study, we observed that renal FoxP3+CD4+ regulatory T cells (Tregs) and IL-17-producing CD4+ T (Th17) cells express the chemokine receptor CCR6, whereas IFNgamma-producing Th1 cells are CCR6-. Induction of experimental glomerulonephritis (nephrotoxic nephritis) in mice resulted in upregulation of the only CCR6 ligand, CCL20, followed by T cell recruitment, renal tissue injury, albuminuria, and loss of renal function. CCR6 deficiency aggravated renal injury and increased mortality (from uremia) among nephritic mice. Compared with wild-type (WT) mice, CCR6 deficiency reduced infiltration of Tregs and Th17 cells but did not affect recruitment of Th1 cells in the setting of glomerulonephritis. Adoptive transfer of WT but not CCR6-deficient Tregs attenuated morphologic and functional renal injury in nephritic mice. Furthermore, reconstitution with WT Tregs protected CCR6-/- mice from aggravated nephritis. Taken together, these data suggest that CCR6 mediates renal recruitment of both Tregs and Th17 cells and that the reduction of anti-inflammatory Tregs in the presence of a fully functional Th1 response aggravates experimental glomerulonephritis.

The Th17 immune response in renal inflammation

The discovery of interleukin (IL)-17-producing CD4(+) T (Th17) cells as a unique T-helper cell lineage has revised our understanding of T-cell-mediated tissue injury. Recent data from studies in humans and mice indicate that autoimmune diseases such as multiple sclerosis and rheumatoid arthritis, classically believed to be Th1-mediated, are predominantly driven by a Th17 immune response. IL-17 (IL-17A), IL-17F, IL-21, IL-22, and possibly also IL-9 produced by Th17 cells promote inflammation by directly causing tissue injury and enhancing secretion of pro-inflammatory cytokines and chemokines by resident cells. This results in augmented infiltration of leukocytes, in particular neutrophils, to the affected tissue where they induce organ inflammation and injury. Recent studies have highlighted the potential importance of the Th17 immune response also in renal inflammatory disease. This includes the identification and characterization of IL-17-producing T cells in nephritic kidneys of mice and humans, as well as evidence for the contribution of IL-17 and the IL-23/Th17 axis to renal tissue injury in glomerulonephritis. In this review, we will briefly summarize general characteristics of Th17 cells and discuss in detail the potential role of the Th17 immune response in human and experimental renal inflammation with a special focus on glomerulonephritis.

CXCR3 Mediates Renal Th1 and Th17 Immune Response in Murine Lupus Nephritis

Infiltration of T cells into the kidney is a typical feature of human and experimental lupus nephritis that contributes to renal tissue injury. The chemokine receptor CXCR3 is highly expressed on Th1 cells and is supposed to be crucial for their trafficking into inflamed tissues. In this study, we explored the functional role of CXCR3 using the MRL/MpJ-Faslpr (MRL/lpr) mouse model of systemic lupus erythematosus that closely resembles the human disease. CXCR3−/− mice were generated and backcrossed into the MRL/lpr background. Analysis of 20-wk-old CXCR3−/− MRL/lpr mice showed amelioration of nephritis with reduced glomerular tissue damage and decreased albuminuria and T cell recruitment. Most importantly, not only the numbers of renal IFN-γ-producing Th1 cells, but also of IL-17-producing Th17 cells were significantly reduced. Unlike in inflamed kidneys, there was no reduction in the numbers of IFN-γ- or IL-17-producing T cells in spleens, lymph nodes, or the small intestine of MRL/lpr CXCR3−/− mice. This observation suggests impaired trafficking of effector T cells to injured target organs, rather than the inability of CXCR3−/− mice to mount efficient Th1 and Th17 immune responses. These findings show a crucial role for CXCR3 in the development of experimental lupus nephritis by directing pathogenic effector T cells into the kidney. For the first time, we demonstrate a beneficial effect of CXCR3 deficiency through attenuation of both the Th1 and the newly defined Th17 immune response. Our data therefore identify the chemokine receptor CXCR3 as a promising therapeutic target in lupus nephritis.

Chemokine Receptor CXCR3 Mediates T Cell Recruitment and Tissue Injury in Nephrotoxic Nephritis in Mice

The chemokine receptor CXCR3 is highly expressed on Th1 polarized T cells and has been predicted to play an important role in T cell recruitment and immune response in a number of inflammatory and autoimmune diseases. For testing whether CXCR3 plays a role in renal inflammation, CXCR3-deficient mice were generated and nephrotoxic nephritis was induced in C57BL/6 CXCR3(-/-) and C57BL/6 wild-type mice. Induction of the nephrotoxic nephritis leads to an increased renal mRNA expression of IP-10/CXCL10 (8.6-fold), Mig/CXCL9 (2.3-fold), and I-TAC/CXCL11 (4.9-fold) during the autologous phase at days 7 and 14. This increased chemokine expression was paralleled by the renal infiltration of T cells, followed by renal tissue injury, albuminuria, and loss of renal function. Compared with wild-type mice, CXCR3-deficient mice had significantly reduced renal T cell infiltrates. Moreover, CXCR3(-/-) mice developed less severe nephritis, with significantly lower albuminuria, better renal function, and a reduced frequency of glomerular crescent formation. Nephritic wild-type and CXCR3(-/-) mice both elicited an efficient systemic nephritogenic immune response in terms of antigen-specific IgG production and IFN-gamma expression by splenocytes in response to the nephritogenic antigen. These findings indicate that the ameliorated nephritis in CXCR3-deficient mice is due to impaired renal trafficking of effector T cells rather than their inability to mount an efficient humoral or cellular immune response. The neutralization of CXCR3 might be a promising therapeutic strategy for Th1-dependent inflammatory renal disease.

Chemokines play a critical role in the cross-regulation of Th1 and Th17 immune responses in murine crescentic glomerulonephritis

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.

CCR5 Deficiency Aggravates Crescentic Glomerulonephritis in Mice

The chemokine receptor CCR5 is predominantly expressed on monocytes and Th1-polarized T cells, and plays an important role in T cell and monocyte recruitment in inflammatory diseases. To investigate the functional role of CCR5 in renal inflammation, we induced a T cell-dependent model of glomerulonephritis (nephrotoxic serum nephritis) in CCR5−/− mice. Induction of nephritis in wild-type mice resulted in up-regulation of renal mRNA expression of the three CCR5 chemokine ligands, CCL5 (15-fold), CCL3 (4.9-fold), and CCL4 (3.4-fold), in the autologous phase of the disease at day 10. The up-regulated chemokine expression was paralleled by infiltration of monocytes and T cells, followed by renal tissue injury, albuminuria, and loss of renal function. Nephritic CCR5−/− mice showed a 3- to 4-fold increased renal expression of CCL5 (61.6-fold vs controls) and CCL3 (14.1-fold vs controls), but not of CCL4, in comparison with nephritic wild-type mice, which was accompanied by augmented renal T cell and monocyte recruitment and increased lethality due to uremia. Furthermore, CCR5−/− mice showed an increased renal Th1 response, whereas their systemic humoral and cellular immune responses were unaltered. Because the CCR5 ligands CCL5 and CCL3 also act via CCR1, we investigated the effects of the pharmacological CCR1 antagonist BX471. CCR1 blockade in CCR5−/− mice significantly reduced renal chemokine expression, T cell infiltration, and glomerular crescent formation, indicating that increased renal leukocyte recruitment and consecutive tissue damage in nephritic CCR5−/− mice depended on functional CCR1. In conclusion, this study shows that CCR5 deficiency aggravates glomerulonephritis via enhanced CCL3/CCL5-CCR1-driven renal T cell recruitment.

IL-17A Production by Renal γδ T Cells Promotes Kidney Injury in Crescentic GN

The Th17 immune response appears to contribute to the pathogenesis of human and experimental crescentic GN, but the cell types that produce IL-17A in the kidney, the mechanisms involved in its induction, and the IL-17A-mediated effector functions that promote renal tissue injury are incompletely understood. Here, using a murine model of crescentic GN, we found that CD4(+) T cells, γδ T cells, and a population of CD3(+)CD4(-)CD8(-)γδT cell receptor(-)NK1.1(-) T cells all produce IL-17A in the kidney. A time course analysis identified γδ T cells as a major source of IL-17A in the early phase of disease, before the first CD4(+) Th17 cells arrived. The production of IL-17A by renal γδ T cells depended on IL-23p19 signaling and retinoic acid-related orphan receptor-γt but not on IL-1β or IL-6. In addition, depletion of dendritic cells, which produce IL-23 in the kidney, reduced IL-17A production by renal γδ T cells. Furthermore, the lack of IL-17A production in γδ T cells, as well as the absence of all γδ T cells, reduced neutrophil recruitment into the kidney and ameliorated renal injury. Taken together, these data suggest that γδ T cells produce IL-17A in the kidney, induced by IL-23, promoting neutrophil recruitment, and contributing to the immunopathogenesis of crescentic GN.

Renal IL-17 expression in human ANCA-associated glomerulonephritis

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.

CXCL5 Drives Neutrophil Recruitment in TH17-Mediated GN

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.