Interleukin-17: Friend or foe in organ fibrosis.

Fibrosis affects all vital organs accounting for a staggering 45% of deaths worldwide and no effective therapies are currently available. Unresolved inflammation triggers downstream signaling events that lead to organ fibrosis. In recent years, proinflammatory cytokine Interleukin-17 (IL-17) has been implicated in several chronic inflammatory diseases that often culminate in organ damage followed by impaired wound healing and fibrosis. In this review, we outline the contribution of the IL-17 in mediating fibrotic diseases in various organs. A comprehensive understanding of the inflammatory events, and particularly the details of IL-17 signaling in vivo, could be beneficial in designing new therapeutic or preventive approaches to treat fibrosis. Additionally, understanding organ-specific differences in IL-17 activity could lead to targeted therapies and help spare other organs from unwanted side effects.

Loss of Twist1 in the Mesenchymal Compartment Promotes Increased Fibrosis in Experimental Lung Injury by Enhanced Expression of CXCL12.

Idiopathic pulmonary fibrosis (IPF) is a disease characterized by the accumulation of apoptosis-resistant fibroblasts in the lung. We have previously shown that high expression of the transcription factor Twist1 may explain this prosurvival phenotype in vitro. However, this observation has never been tested in vivo. We found that loss of Twist1 in COL1A2+ cells led to increased fibrosis characterized by very significant accumulation of T cells and bone marrow-derived matrix-producing cells. We found that Twist1-null cells expressed high levels of the T cell chemoattractant CXCL12. In vitro, we found that the loss of Twist1 in IPF lung fibroblasts increased expression of CXCL12 downstream of increased expression of the noncanonical NF-κB transcription factor RelB. Finally, blockade of CXCL12 with AMD3100 attenuated the exaggerated fibrosis observed in Twist1-null mice. Transcriptomic analysis of 134 IPF patients revealed that low expression of Twist1 was characterized by enrichment of T cell pathways. In conclusion, loss of Twist1 in collagen-producing cells led to increased bleomycin-induced pulmonary fibrosis, which is mediated by increased expression of CXCL12. Twist1 expression is associated with dysregulation of T cells in IPF patients. Twist1 may shape the IPF phenotype and regulate inflammation in fibrotic lung injury.

The Kallikrein-Kinin System: A Novel Mediator of IL-17-Driven Anti-Candida Immunity in the Kidney.

The incidence of life-threatening disseminated Candida albicans infections is increasing in hospitalized patients, with fatalities as high as 60%. Death from disseminated candidiasis in a significant percentage of cases is due to fungal invasion of the kidney, leading to renal failure. Treatment of candidiasis is hampered by drug toxicity, the emergence of antifungal drug resistance and lack of vaccines against fungal pathogens. IL-17 is a key mediator of defense against candidiasis. The underlying mechanisms of IL-17-mediated renal immunity have so far been assumed to occur solely through the regulation of antimicrobial mechanisms, particularly activation of neutrophils. Here, we identify an unexpected role for IL-17 in inducing the Kallikrein (Klk)-Kinin System (KKS) in C. albicans-infected kidney, and we show that the KKS provides significant renal protection in candidiasis. Microarray data indicated that Klk1 was upregulated in infected kidney in an IL-17-dependent manner. Overexpression of Klk1 or treatment with bradykinin rescued IL-17RA-/- mice from candidiasis. Therapeutic manipulation of IL-17-KKS pathways restored renal function and prolonged survival by preventing apoptosis of renal cells following C. albicans infection. Furthermore, combining a minimally effective dose of fluconazole with bradykinin markedly improved survival compared to either drug alone. These results indicate that IL-17 not only limits fungal growth in the kidney, but also prevents renal tissue damage and preserves kidney function during disseminated candidiasis through the KKS. Since drugs targeting the KKS are approved clinically, these findings offer potential avenues for the treatment of this fatal nosocomial infection.

IL-17 Receptor Signaling Negatively Regulates the Development of Tubulointerstitial Fibrosis in the Kidney.

Chronic inflammation has an important role in the development and progression of most fibrotic diseases, for which no effective treatments exist. Tubulointerstitial fibrosis (TF) is characterized by irreversible deposition of fibrous tissue in chronic kidney diseases. Prolonged injurious stimuli and chronic inflammation regulate downstream events that lead to TF. In recent years, interleukin-17 (IL-17) has been strongly linked to organ fibrosis. However, the role of IL-17 receptor signaling in TF is an active area of debate. Using the unilateral ureteral obstruction (UUO) mouse model of TF, we show that IL-17 receptor A-deficient mice (Il17ra-/- ) exhibit increased TF in the obstructed kidney. Consequently, overexpression of IL-17 restored protection in mice with UUO. Reduced renal expression of matrix-degrading enzymes results in failure to degrade ECM proteins, thus contributing to the exaggerated TF phenotype in Il17ra -/- mice. We demonstrate that the antifibrotic kallikrein-kinin system (KKS) is activated in the obstructed kidney in an IL-17-dependent manner. Accordingly, Il17ra-/- mice receiving bradykinin, the major end-product of KKS activation, prevents TF development by upregulating the expression of matrix-degrading enzymes. Finally, we show that treatment with specific agonists for bradykinin receptor 1 or 2 confers renal protection against TF. Overall, our results highlight an intriguing link between IL-17 and activation of KKS in protection against TF, the common final outcome of chronic kidney conditions leading to devastating end-stage renal diseases.

Expression of RXFP1 Is Decreased in Idiopathic Pulmonary Fibrosis. Implications for Relaxin-based Therapies.

RATIONALE: Relaxin is a hormone that has been considered as a potential therapy for patients with fibrotic diseases. OBJECTIVES: To gauge the potential efficacy of relaxin-based therapies in idiopathic pulmonary fibrosis (IPF), we studied gene expression for relaxin/insulin-like family peptide receptor 1 (RXFP1) in IPF lungs and controls. METHODS: We analyzed gene expression data obtained from the Lung Tissue Research Consortium and correlated RXFP1 gene expression data with cross-sectional clinical and demographic data. We also employed ex vivo donor and IPF lung fibroblasts to test RXFP1 expression in vitro. We tested CGEN25009, a relaxin-like peptide, in lung fibroblasts and in bleomycin injury. MEASUREMENTS AND MAIN RESULTS: We found that RXFP1 is significantly decreased in IPF. In patients with IPF, the magnitude of RXFP1 gene expression correlated directly with diffusing capacity of the lung for carbon monoxide (P < 0.0001). Significantly less RXFP1 was detected in vitro in IPF fibroblasts than in donor controls. Transforming growth factor-ß decreased RXFP1 in both donor and IPF lung fibroblasts. CGEN25009 was effective at decreasing bleomycin-induced, acid-soluble collagen deposition in vivo. The relaxin-like actions of CGEN25009 were abrogated by RXFP1 silencing in vitro, and, in comparison with donor lung fibroblasts, IPF lung fibroblasts exhibited decreased sensitivity to the relaxin-like effects of CGEN25009. CONCLUSIONS: IPF is characterized by the loss of RXFP1 expression. RXFP1 expression is directly associated with pulmonary function in patients with IPF. The relaxin-like effects of CGEN25009 in vitro are dependent on expression of RXFP1. Our data suggest that patients with IPF with the highest RXFP1 expression would be predicted to be most sensitive to relaxin-based therapies.

Interleukin 17 signaling drives Type I Interferon induced proliferative crescentic glomerulonephritis in lupus-prone mice.

Crescentic glomerulonephritis (cGN) is a severe clinical manifestation in a subset of patients with Systemic lupus erythematosus. Lack of understanding of the pathogenesis of cGN act as a major constraint in treating these patients. Emerging evidence suggest a critical role of IL-17 in the pathogenesis of membranoproliferative glomerulonephritis in lupus. However, the role of IL-17 receptor (IL-17RA) signaling in cGN is unknown. Here, we developed a model of poly I:C-induced Type I Interferon (IFN-I)-dependent cGN in B6.MRL-Faslpr/J (B6.lpr) mice. B6.lpr mice deficient in IL-17RA were protected from IFN-I-dependent cGN. While systemic response was unabated, renal infiltration of alternatively activated macrophages was severely impaired in IL-17RA(-/-) mice. Finally, we show that IL-17 differentially regulates the expression of macrophage chemo-attractant genes in renal tubular epithelial cells and macrophages in association with IFN-I. These results suggest that neutralization IL-17 may confer better protection in SLE patients with high IFN-I gene signature and cGN.

Emerging roles of the Th17/IL-17-axis in glomerulonephritis.

Different T cell subsets have been implicated in the pathogenesis of glomerulonephritis. Several lines of evidence indicate the recently identified interleukin-17 (IL-17)-producing T cells (Th17 cells) to be involved in the renal inflammatory cascade associated with glomerulonephritis. In this review we outline different forms of glomerulonephritis and the contribution of the Th17/IL-17-axis in mediating the downstream effects and pathology associated with the disease. Learning more about the Th17/IL-17-axis can help to develop promising therapeutic strategy for the treatment of various forms of glomerulonephritis.

Complement component C5a permits the coexistence of pathogenic Th17 cells and type I IFN in lupus.

Systemic lupus erythematosus (SLE) is a type I IFN (IFN-I)-driven autoimmune disorder with exaggerated B and Th cell responses. Th17 cells, a recently identified Th cell subset, have been strongly implicated in the pathogenesis of SLE. Because IFN-I suppresses the generation and expansion of Th17 cells in an IL-27-dependent manner, it is unclear how pathogenic Th17 cells are generated in SLE in the presence of an environment characterized by high IFN-I levels. In this study, we showed that activation of c5aR on murine macrophages blocked IFN-I-mediated IL-27 production, thus permitting the development of Th17 cells. C5aR activation on IFN-I-responsive macrophages inhibits IRF-1-mediated transactivation of IL-27 gene expression via the PI3K/Akt pathway. Consistently, C5aR-deficient mice exhibited increased IL-27 expression and fewer Th17 cells and consequently developed reduced lupus nephritis in comparison with wild-type mice. In support of these findings in mice, we found that C5a inhibited IFN-I-induced IL-27 production from macrophages of lupus subjects. Moreover, the level of serum C5a correlated with Th17 frequency in peripheral blood. Collectively, these data indicate an essential role for C5a in the generation of pathogenic Th17 responses in SLE. Thus, therapeutic strategies to block C5aR activation may be beneficial for controlling pathogenic Th17-mediated inflammation in SLE.

Clinical translation of anti-inflammatory effects of Prevotella histicola in Th1, Th2, and Th17 inflammation.

Introduction: EDP1815 is a non-colonizing pharmaceutical preparation of a single stain of Prevotella histicola isolated from the duodenum of a human donor. We report here preclinical and clinical studies showing that the action of EDP1815, an orally delivered and gut restricted single strain of commensal bacteria can regulate inflammatory responses throughout the body. Methods: Supported by evidence for anti-inflammatory activity in three preclinical mouse models of Th1-, TH2-, and Th17-mediated inflammation, EDP1815 was tested clinically in three Phase 1b studies in patients with psoriasis, patients with atopic dermatitis, and healthy volunteers in a KLH skin challenge model. Results: Preclinically, EDP1815 was efficacious in all three mouse models of inflammation, showing reduction in skin inflammation as well as related tissue cytokines. In the Phase 1b studies, EDP1815 was found to be well tolerated by participants, with a safety profile comparable to placebo, including no severe or consistent side-effects reported, and no evidence of immunosuppression with no opportunistic infection occurring in these studies. In psoriasis patients, signs of clinical efficacy were seen after 4 weeks of treatment, which continued beyond the treatment period in the higher-dose cohort. In atopic dermatitis patients, improvements were seen throughout the key physician-and patient-reported outcomes. In a healthy-volunteer study of a KLH-induced skin inflammatory response, consistent anti-inflammatory effects were seen in two cohorts through imaging-based measures of skin inflammation. Discussion: This is the first report demonstrating clinical effects from targeting peripheral inflammation with a non-colonizing gut-restricted single strain of commensal bacteria, providing proof of concept for a new class of medicines. These clinical effects occur without systemic exposure of EDP1815 or modification of the resident gut microbiota, and with placebo-like safety and tolerability. The breadth of these clinical effects of EDP1815, combined with its excellent safety and tolerability profile and oral administration, suggests the potential for a new type of effective, safe, oral, and accessible anti-inflammatory medicine to treat the wide range of diseases driven by inflammation.Clinical Trial Registration: EudraCT # 2018-002807-32; EudraCT # 2018-002807-32; NL8676; https://clinicaltrials.gov/ct2/show/NCT03733353; http://www.trialregister.nl.

Regulation of Peripheral Inflammation by a Non-Viable, Non-Colonizing Strain of Commensal Bacteria.

The gastrointestinal tract represents one of the largest body surfaces that is exposed to the outside world. It is the only mucosal surface that is required to simultaneously recognize and defend against pathogens, while allowing nutrients containing foreign antigens to be tolerated and absorbed. It differentiates between these foreign substances through a complex system of pattern recognition receptors expressed on the surface of the intestinal epithelial cells as well as the underlying immune cells. These immune cells actively sample and evaluate microbes and other particles that pass through the lumen of the gut. This local sensing system is part of a broader distributed signaling system that is connected to the rest of the body through the enteric nervous system, the immune system, and the metabolic system. While local tissue homeostasis is maintained by commensal bacteria that colonize the gut, colonization itself may not be required for the activation of distributed signaling networks that can result in modulation of peripheral inflammation. Herein, we describe the ability of a gut-restricted strain of commensal bacteria to drive systemic anti-inflammatory effects in a manner that does not rely upon its ability to colonize the gastrointestinal tract or alter the mucosal microbiome. Orally administered EDP1867, a gamma-irradiated strain of Veillonella parvula, rapidly transits through the murine gut without colonization or alteration of the background microbiome flora. In murine models of inflammatory disease including delayed-type hypersensitivity (DTH), atopic dermatitis, psoriasis, and experimental autoimmune encephalomyelitis (EAE), treatment with EDP1867 resulted in significant reduction in inflammation and immunopathology. Ex vivo cytokine analyses revealed that EDP1867 treatment diminished production of pro-inflammatory cytokines involved in inflammatory cascades. Furthermore, blockade of lymphocyte migration to the gut-associated lymphoid tissues impaired the ability of EDP1867 to resolve peripheral inflammation, supporting the hypothesis that circulating immune cells are responsible for promulgating the signals from the gut to peripheral tissues. Finally, we show that adoptively transferred T cells from EDP1867-treated mice inhibit inflammation induced in recipient mice. These results demonstrate that an orally-delivered, non-viable strain of commensal bacteria can mediate potent anti-inflammatory effects in peripheral tissues through transient occupancy of the gastrointestinal tract, and support the development of non-living bacterial strains for therapeutic applications.

Uremia Coupled with Mucosal Damage Predisposes Mice with Kidney Disease to Systemic Infection by Commensal Candida albicans.

Infections are the second major cause of mortality in patients with kidney disease and accompanying uremia. Both vascular access and non-access-related infections contribute equally to the infection-related deaths in patients with kidney disease. Dialysis is the most common cause of systemic infection by Candida albicans in these patients. C albicans also reside in the gastrointestinal tract as a commensal fungus. However, the contribution of gut-derived C albicans in non-access-related infections in kidney disease is unknown. Using a mouse model of kidney disease, we demonstrate that uremic animals showed increased gut barrier permeability, impaired mucosal defense, and dysbiosis. The disturbance in gut homeostasis is sufficient to drive the translocation of microbiota and intestinal pathogen Citrobacter rodentium to extraintestinal sites but not C albicans Interestingly, a majority of uremic animals showed fungal translocation only when the gut barrier integrity is disrupted. Our data demonstrate that uremia coupled with gut mucosal damage may aid in the translocation of C. albicans and cause systemic infection in kidney disease. Because most of the individuals with kidney disease suffer from some form of gut mucosal damage, these results have important implications in the risk stratification and control of non-access-related opportunistic fungal infections in these patients.

RTEC-intrinsic IL-17-driven inflammatory circuit amplifies antibody-induced glomerulonephritis and is constrained by Regnase-1.

Antibody-mediated glomerulonephritis (AGN) is a clinical manifestation of many autoimmune kidney diseases for which few effective treatments exist. Chronic inflammatory circuits in renal glomerular and tubular cells lead to tissue damage in AGN. These cells are targeted by the cytokine IL-17, which has recently been shown to be a central driver of the pathogenesis of AGN. However, surprisingly little is known about the regulation of pathogenic IL-17 signaling in the kidney. Here, using a well-characterized mouse model of AGN, we show that IL-17 signaling in renal tubular epithelial cells (RTECs) is necessary for AGN development. We also show that Regnase-1, an RNA binding protein with endoribonuclease activity, is a negative regulator of IL-17 signaling in RTECs. Accordingly, mice with a selective Regnase-1 deficiency in RTECs exhibited exacerbated kidney dysfunction in AGN. Mechanistically, Regnase-1 inhibits IL-17-driven expression of the transcription factor IκBξ and, consequently, its downstream gene targets, including Il6 and Lcn2. Moreover, deletion of Regnase-1 in human RTECs reduced inflammatory gene expression in a IκBξ-dependent manner. Overall, these data identify an IL-17-driven inflammatory circuit in RTECs during AGN that is constrained by Regnase-1.

Restoring glucose uptake rescues neutrophil dysfunction and protects against systemic fungal infection in mouse models of kidney disease.

Disseminated candidiasis caused by the fungus Candida albicans is a major clinical problem in individuals with kidney disease and accompanying uremia; disseminated candidiasis fatality is twice as common in patients with uremia as those with normal kidney function. Many antifungal drugs are nephrotoxic, making treatment of these patients particularly challenging. The underlying basis for this impaired capacity to control infections in uremic individuals is poorly understood. Here, we show in multiple models that uremic mice exhibit an increased susceptibility to systemic fungal infection. Uremia inhibits Glut1-mediated uptake of glucose in neutrophils by causing aberrant activation of GSK3ß, resulting in reduced ROS generation and hence impaired killing of C. albicans in mice. Consequently, pharmacological inhibition of GSK3ß restored glucose uptake and rescued ROS production and candidacidal function of neutrophils in uremic mice. Similarly, neutrophils isolated from patients with kidney disease and undergoing hemodialysis showed similar defect in the fungal killing activity, a phenotype rescued in the presence of a GSK3ß inhibitor. These findings reveal a mechanism of neutrophil dysfunction during uremia and suggest a potentially translatable therapeutic avenue for treatment of disseminated candidiasis.

Unexpected kidney-restricted role for IL-17 receptor signaling in defense against systemic Candida albicans infection.

Kidney injury is a frequent outcome in patients with disseminated Candida albicans fungal infections. IL-17 receptor (IL-17R) signaling is critical for renal protection against disseminated candidiasis, but the identity and function of IL-17-responsive cells in mediating renal defense remains an active area of debate. Using BM chimeras, we found that IL-17R signaling is required only in nonhematopoietic cells for immunity to systemic C. albicans infection. Since renal tubular epithelial cells (RTEC) are highly responsive to IL-17 in vitro, we hypothesized that RTEC might be the dominant target of IL-17 activity in the infected kidney. We generated mice with a conditional deletion of IL-17 receptor A (Il17ra) in RTEC (Il17raΔRTEC). Strikingly, Il17raΔRTEC mice showed enhanced kidney damage and early mortality following systemic infection, very similar to Il17ra-/- animals. Increased susceptibility to candidiasis in Il17raΔRTEC mice was associated with diminished activation of the renal protective Kallikrein-kinin system (KKS), resulting in reduced apoptosis of kidney-resident cells during hyphal invasion. Moreover, protection was restored by treatment with bradykinin, the major end-product of KKS activation, which was mediated dominantly via bradykinin receptor b1. These data show that IL-17R signaling in RTEC is necessary and likely sufficient for IL-17-mediated renal defense against fatal systemic C. albicans infection.

Oral epithelial cells orchestrate innate type 17 responses to Candida albicans through the virulence factor candidalysin.

Candida albicans is a dimorphic commensal fungus that causes severe oral infections in immunodeficient patients. Invasion of C. albicans hyphae into oral epithelium is an essential virulence trait. Interleukin-17 (IL-17) signaling is required for both innate and adaptive immunity to C. albicans During the innate response, IL-17 is produced by γδ T cells and a poorly understood population of innate-acting CD4+ αß T cell receptor (TCRαß)+ cells, but only the TCRαß+ cells expand during acute infection. Confirming the innate nature of these cells, the TCR was not detectably activated during the primary response, as evidenced by Nur77eGFP mice that report antigen-specific signaling through the TCR. Rather, the expansion of innate TCRαß+ cells was driven by both intrinsic and extrinsic IL-1R signaling. Unexpectedly, there was no requirement for CCR6/CCL20-dependent recruitment or prototypical fungal pattern recognition receptors. However, C. albicans mutants that cannot switch from yeast to hyphae showed impaired TCRαß+ cell proliferation and Il17a expression. This prompted us to assess the role of candidalysin, a hyphal-associated peptide that damages oral epithelial cells and triggers production of inflammatory cytokines including IL-1. Candidalysin-deficient strains failed to up-regulate Il17a or drive the proliferation of innate TCRαß+ cells. Moreover, candidalysin signaled synergistically with IL-17, which further augmented the expression of IL-1α/ß and other cytokines. Thus, IL-17 and C. albicans, via secreted candidalysin, amplify inflammation in a self-reinforcing feed-forward loop. These findings challenge the paradigm that hyphal formation per se is required for the oral innate response and demonstrate that establishment of IL-1- and IL-17-dependent innate immunity is induced by tissue-damaging hyphae.

Type I interferon and T helper 17 cells co-exist and co-regulate disease pathogenesis in lupus patients.

AIM: T-helper 17 cells (Th17) and type I interferon (IFN-I) play a critical role in the pathogenesis of systemic lupus erythematosus (SLE). Previous studies have suggested that IFN-I suppresses Th17 development under autoimmune settings. Therefore, the main objective of this study was to define the association between IFN-I and Th17 pathways in SLE. METHODS: Peripheral blood samples and disease activity measures were collected from 31 patients fulfilling the American College of Rheumatology revised criteria for SLE. Serum was evaluated for IFN-α bioactivity and interleukin (IL)-6 levels by cell-based bioluminescence assay and enzyme-linked immunosorbent assay, respectively. The frequency of Th17 cells in peripheral blood was determined by intracellular cytokine staining for IL-17. RESULTS: IFN-α bioactivity in the serum of lupus subjects (mean ± SD: 6.510 ± 3.686) was significantly higher (P = 0.001) compared to healthy controls (2.9 ± 1.061). Additionally, 58.1% and 41.9% of SLE subjects displayed high and low IFN-α bioactivity, respectively. We observed a significant increase (P = 0.04) in the percentage of Th17 cells in lupus subjects with high IFN-α bioactivity (1.9 ± 1.0) compared to lupus subjects with low IFN-α bioactivity (1.2 ± 0.9). Lupus subjects with high IFN-α bioactivity and Th17 cells had significantly higher disease activity (P = 0.04) and serum IL-6 levels (P = 0.01) compared to patients with low IFN-α activity and low Th17 cells. CONCLUSION: Type I interferon and Th17 pathways co-exist and co-regulate the pathogenic processes in SLE. Additionally, these studies clearly identify IL-6 as a common link between IFN-I and Th17 pathways in SLE pathogenesis.

An essential role of interleukin-17 receptor signaling in the development of autoimmune glomerulonephritis.

In recent years, proinflammatory cytokines in the nephritic kidney appear to contribute to the pathogenesis of AGN. The complex inflammatory cytokine network that drives renal pathology is poorly understood. IL-17, the signature cytokine of Th17 cells, which promotes autoimmune pathology in a variety of settings, is beginning to be identified in acute and chronic kidney diseases as well. However, the role of IL-17-mediated renal damage in the nephritic kidney has not been elucidated. Here, with the use of a murine model of experimental AGN, we showed that IL-17RA signaling is critical for the development of renal pathology. Despite normal systemic autoantibody response and glomerular immune-complex deposition, IL-17RA(-/-) mice exhibit a diminished influx of inflammatory cells and kidney-specific expression of IL-17 target genes correlating with disease resistance in AGN. IL-17 enhanced the production of proinflammatory cytokines and chemokines from tECs. Finally, we were able to show that neutralization of IL-17A ameliorated renal pathology in WT mice following AGN. These results clearly demonstrated that IL-17RA signaling significantly contributes to renal tissue injury in experimental AGN and suggest that blocking IL-17RA may be a promising therapeutic strategy for the treatment of proliferative and crescentic glomerulonephritis.