Renal IL-23-Dependent Type 3 Innate Lymphoid Cells Link Crystal-induced Intrarenal Inflammasome Activation with Kidney Fibrosis.

Chronic inflammasome activation in mononuclear phagocytes (MNPs) promotes fibrosis in various tissues, including the kidney. The cellular and molecular links between the inflammasome and fibrosis are unclear. To address this question, we fed mice lacking various immunological mediators an adenine-enriched diet, which causes crystal precipitation in renal tubules, crystal-induced inflammasome activation, and renal fibrosis. We found that kidney fibrosis depended on an intrarenal inflammasome-dependent type 3 immune response driven by its signature transcription factor Rorc (retinoic acid receptor-related orphan receptor C gene), which was partially carried out by type 3 innate lymphoid cells (ILC3s). The role of ILCs in the kidney is less well known than in other organs, especially that of ILC3. In this article, we describe that depletion of ILCs or genetic deficiency for Rorc attenuated kidney inflammation and fibrosis. Among the inflammasome-derived cytokines, only IL-1ß expanded ILC3 and promoted fibrosis, whereas IL-18 caused differentiation of NKp46+ ILC3. Deficiency of the type 3 maintenance cytokine, IL-23, was more protective than IL-1ß inhibition, which may be explained by the downregulation of the IL-1R, but not of the IL-23R, by ILC3 early in the disease, allowing persistent sensing of IL-23. Mechanistically, ILC3s colocalized with renal MNPs in vivo as shown by multiepitope-ligand cartography. Cell culture experiments indicated that renal ILC3s caused renal MNPs to increase TGF-ß production that stimulated fibroblasts to produce collagen. We conclude that ILC3s link inflammasome activation with kidney inflammation and fibrosis and are regulated by IL-1ß and IL-23.

Transcriptome alterations in peripheral blood B cells of patients with multiple sclerosis receiving immune reconstitution therapy.

BACKGROUND: Multiple sclerosis (MS) is a chronic, inflammatory and neurodegenerative disease that leads to irreversible damage to the brain and spinal cord. The goal of so-called "immune reconstitution therapies" (IRTs) is to achieve long-term disease remission by eliminating a pathogenic immune repertoire through intense short-term immune cell depletion. B cells are major targets for effective immunotherapy in MS. OBJECTIVES: The aim of this study was to analyze the gene expression pattern of B cells before and during IRT (i.e., before B-cell depletion and after B-cell repopulation) to better understand the therapeutic effects and to identify biomarker candidates of the clinical response to therapy. METHODS: B cells were obtained from blood samples of patients with relapsing-remitting MS (n = 50), patients with primary progressive MS (n = 13) as well as healthy controls (n = 28). The patients with relapsing MS received either monthly infusions of natalizumab (n = 29) or a pulsed IRT with alemtuzumab (n = 15) or cladribine (n = 6). B-cell subpopulation frequencies were determined by flow cytometry, and transcriptome profiling was performed using Clariom D arrays. Differentially expressed genes (DEGs) between the patient groups and controls were examined with regard to their functions and interactions. We also tested for differences in gene expression between patients with and without relapse following alemtuzumab administration. RESULTS: Patients treated with alemtuzumab or cladribine showed on average a > 20% lower proportion of memory B cells as compared to before IRT. This was paralleled by profound transcriptome shifts, with > 6000 significant DEGs after adjustment for multiple comparisons. The top DEGs were found to regulate apoptosis, cell adhesion and RNA processing, and the most highly connected nodes in the network of encoded proteins were ESR2, PHB and RC3H1. Higher mRNA levels of BCL2, IL13RA1 and SLC38A11 were seen in patients with relapse despite IRT, though these differences did not pass the false discovery rate correction. CONCLUSIONS: We show that B cells circulating in the blood of patients with MS undergoing IRT present a distinct gene expression signature, and we delineated the associated biological processes and gene interactions. Moreover, we identified genes whose expression may be an indicator of relapse risk, but further studies are needed to verify their potential value as biomarkers.

Targeting kidney inflammation as a new therapy for primary hyperoxaluria?

The primary hyperoxalurias (PHs) are inborn errors of glyoxylate metabolism characterized by endogenous oxalate overproduction in the liver, and thus elevated urinary oxalate excretion. The urinary calcium-oxalate (CaOx) supersaturation and the continuous renal accumulation of insoluble CaOx crystals yield a progressive decline in renal function that often ends with renal failure. In PH Type 1 (AGXT mutated), the most frequent and severe condition, patients typically progress to end-stage renal disease (ESRD); in PH Type 2 (GRHPR mutated), 20% of patients develop ESRD, while only one patient with PH Type 3 (HOGA1 mutated) has been reported with ESRD so far. Patients with ESRD undergo frequent maintenance (haemo)dialysis treatment, and finally must receive a combined liver-kidney transplantation as the only curative treatment option available in PH Type 1. In experimental models using oxalate-enriched chow, CaOx crystals were bound to renal tubular cells, promoting a pro-inflammatory environment that led to fibrogenesis in the renal parenchyma by activation of a NACHT, LRR and PYD domains-containing protein 3 (NALP3)-dependent inflammasome in renal dendritic cells and macrophages. Chronic fibrogenesis progressively impaired renal function. Targeting the inflammatory response has recently been suggested as a therapeutic strategy to treat not only oxalate-induced crystalline nephropathies, but also those characterized by accumulation of cystine and urate in other organs. Herein, we summarize the pathogenesis of PH, revising the current knowledge of the CaOx-mediated inflammatory response in animal models of endogenous oxalate overproduction. Furthermore, we highlight the possibility of modifying the NLRP3-dependent inflammasome as a new and complementary therapeutic strategy to treat this severe and devastating kidney disease.

B7-H1 shapes T-cell-mediated brain endothelial cell dysfunction and regional encephalitogenicity in spontaneous CNS autoimmunity.

Molecular mechanisms that determine lesion localization or phenotype variation in multiple sclerosis are mostly unidentified. Although transmigration of activated encephalitogenic T cells across the blood-brain barrier (BBB) is a crucial step in the disease pathogenesis of CNS autoimmunity, the consequences on brain endothelial barrier integrity upon interaction with such T cells and subsequent lesion formation and distribution are largely unknown. We made use of a transgenic spontaneous mouse model of CNS autoimmunity characterized by inflammatory demyelinating lesions confined to optic nerves and spinal cord (OSE mice). Genetic ablation of a single immune-regulatory molecule in this model [i.e., B7-homolog 1 (B7-H1, PD-L1)] not only significantly increased incidence of spontaneous CNS autoimmunity and aggravated disease course, especially in the later stages of disease, but also importantly resulted in encephalitogenic T-cell infiltration and lesion formation in normally unaffected brain regions, such as the cerebrum and cerebellum. Interestingly, B7-H1 ablation on myelin oligodendrocyte glycoprotein-specific CD4+ T cells, but not on antigen-presenting cells, amplified T-cell effector functions, such as IFN-γ and granzyme B production. Therefore, these T cells were rendered more capable of eliciting cell contact-dependent brain endothelial cell dysfunction and increased barrier permeability in an in vitro model of the BBB. Our findings suggest that a single immune-regulatory molecule on T cells can be ultimately responsible for localized BBB breakdown, and thus substantial changes in lesion topography in the context of CNS autoimmunity.

Antigen-specific Helios- , Neuropilin-1- Tregs induce apoptosis of autoreactive B cells via PD-L1.

Regulatory T cells (Tregs) maintain self-tolerance and prevent autoimmunity by controlling autoreactive T cells. We recently demonstrated in vivo that Tregs can directly suppress auto-reactive B cells via programmed death ligand 1 (PD-L1) that ligated PD-1 on B cells and caused them to undergo apoptosis. Here, we asked whether this mechanism is utilized by thymus-derived natural Tregs and/or by peripheral lymphoid tissue-induced Tregs. We first demonstrated that antigen-specific PD-L1-expressing Tregs were induced in the draining lymph node of autoantigen-expressing tissue and characterized them by their lack of the transcription factor Helios and of the surface marker Neuropilin-1 (Nrp-1). Next, we established an in vitro co-culture system to study the interaction between B cells and Treg subsets under controlled conditions. We found that Nrp- Treg, but not Nrp+ Treg suppressed autoreactive B cells, whereas both were able to suppress T-helper cells. Such suppression was antigen-specific and was facilitated by PD-L1/PD-1-induced apoptosis. Furthermore, it required physical cell contact and was MHC II-restricted, providing an explanation for the antigen-specificity of peripherally-induced Tregs. These findings identify a role for peripherally induced Helios- Nrp-1- inducible Treg in controlling peripheral B-cell tolerance against tissue auto-antigens.

Fully Automated Evaluation of Total Glomerular Number and Capillary Tuft Size in Nephritic Kidneys Using Lightsheet Microscopy.

The total number of glomeruli is a fundamental parameter of kidney function but very difficult to determine using standard methodology. Here, we counted all individual glomeruli in murine kidneys and sized the capillary tufts by combining in vivo fluorescence labeling of endothelial cells, a novel tissue-clearing technique, lightsheet microscopy, and automated registration by image analysis. Total hands-on time per organ was <1 hour, and automated counting/sizing was finished in <3 hours. We also investigated the novel use of ethyl-3-phenylprop-2-enoate (ethyl cinnamate) as a nontoxic solvent-based clearing reagent that can be handled without specific safety measures. Ethyl cinnamate rapidly cleared all tested organs, including calcified bone, but the fluorescence of proteins and immunohistochemical labels was maintained over weeks. Using ethyl cinnamate-cleared kidneys, we also quantified the average creatinine clearance rate per glomerulus. This parameter decreased in the first week of experimental nephrotoxic nephritis, whereas reduction in glomerular numbers occurred much later. Our approach delivers fundamental parameters of renal function, and because of its ease of use and speed, it is suitable for high-throughput analysis and could greatly facilitate studies of the effect of kidney diseases on whole-organ physiology.

CD103+ Kidney Dendritic Cells Protect against Crescentic GN by Maintaining IL-10-Producing Regulatory T Cells.

Kidney dendritic cells (DCs) regulate nephritogenic T cell responses. Most kidney DCs belong to the CD11b+ subset and promote crescentic GN (cGN). The function of the CD103+ subset, which represents <5% of kidney DCs, is poorly understood. We studied the role of CD103+ DCs in cGN using several lines of genetically modified mice that allowed us to reduce the number of these cells. In all lines, we detected a reduction of FoxP3+ intrarenal regulatory T cells (Tregs), which protect against cGN. Mice lacking the transcription factor Batf3 had a more profound reduction of CD103+ DCs and Tregs than did the other lines used, and showed the most profound aggravation of cGN. The conditional reduction of CD103+ DC numbers by 50% in Langerin-DTR mice halved Treg numbers, which did not suffice to significantly aggravate cGN. Mice lacking the cytokine Flt3L had fewer CD103+ DCs and Tregs than Langerin-DTR mice but exhibited milder cGN than did Batf3-/- mice presumably because proinflammatory CD11b+ DCs were somewhat depleted as well. Conversely, Flt3L supplementation increased the number of CD103+ DCs and Tregs, but also of proinflammatory CD11b+ DCs. On antibody-mediated removal of CD11b+ DCs, Flt3L supplementation ameliorated cGN. Mechanistically, CD103+ DCs caused cocultured T cells to differentiate into Tregs and produced the chemokine CCL20, which is known to attract Tregs into the kidney. Our findings show that CD103+ DCs foster intrarenal FoxP3+ Treg accumulation, thereby antagonizing proinflammatory CD11b+ DCs. Thus, increasing CD103+ DC numbers or functionality might be advantageous in cGN.

An NLRP3-specific inflammasome inhibitor attenuates crystal-induced kidney fibrosis in mice.

Intrarenal crystal formation activates the Nlrp3 inflammasome in myeloid cells and triggers a profound inflammatory response. Here, we studied whether a specific inhibitor of the Nlrp3 inflammasome, CP-456,773, can prevent kidney fibrosis in a murine model of crystal nephropathy induced by diets rich in oxalate or adenine. Inflammasome activation in renal dendritic cells and the resulting interleukin (IL)-1ß and IL-18 production were markedly reduced by CP-456,773 treatment both ex vivo and in vivo. We directly visualized intrarenal inflammasome activation and its inhibition by CP-456,773 in vivo by adoptive transfer of bone marrow cells transduced with interleukin-1ß-Gaussia luciferase, a proteolytic luciferase-based reporter for inflammasome activation, into irradiated mice. CP-456,773 treatment strongly attenuated kidney fibrosis when given early in the genesis of crystal nephropathy, but was unable to reverse established crystal-induced fibrosis. The urinary IL-18 concentration appeared to be a useful noninvasive biomarker for renal inflammasome activation. Finally, NLRP3 inhibition did not compromise adaptive immune responses as previously reported for the global inhibition of IL-1 signaling. Thus, early NLRP3 inhibition by CP-456,773 may be an effective treatment for crystal nephropathy. Use of iGLuc transfected cells introduces a novel imaging technique for inflammasome activation in mice.

Expression of type I interferon by splenic macrophages suppresses adaptive immunity during sepsis.

Early during Gram-negative sepsis, excessive release of pro-inflammatory cytokines can cause septic shock that is often followed by a state of immune paralysis characterized by the failure to mount adaptive immunity towards secondary microbial infections. Especially, the early mechanisms responsible for such immune hypo-responsiveness are unclear. Here, we show that TLR4 is the key immune sensing receptor to initiate paralysis of T-cell immunity after bacterial sepsis. Downstream of TLR4, signalling through TRIF but not MyD88 impaired the development of specific T-cell immunity against secondary infections. We identified type I interferon (IFN) released from splenic macrophages as the critical factor causing T-cell immune paralysis. Early during sepsis, type I IFN acted selectively on dendritic cells (DCs) by impairing antigen presentation and secretion of pro-inflammatory cytokines. Our results reveal a novel immune regulatory role for type I IFN in the initiation of septic immune paralysis, which is distinct from its well-known immune stimulatory effects. Moreover, we identify potential molecular targets for therapeutic intervention to overcome impairment of T-cell immunity after sepsis.

Liver inflammation abrogates immunological tolerance induced by Kupffer cells.

UNLABELLED: The liver is essential for inducing immunological tolerance toward harmless antigens to maintain immune system homeostasis. However, the precise cellular mechanisms of tolerance induction against particle-bound antigens, the role of the local hepatic microenvironment, and implications for therapeutic targets in immune-mediated diseases are currently unclear. In order to elucidate cellular mechanisms of tolerance induction in healthy and injured liver, we developed a novel in vivo system combining the systemic delivery of low-dose peptide antigens coupled to inert particles, immunological readouts, and sophisticated intravital multiphoton microscopy-based imaging of liver in mice. We show that liver resident macrophages, Kupffer cells (KCs), but not hepatic monocyte-derived macrophages or dendritic cells (DCs), are the central cellular scavenger for circulating particle-associated antigens in homeostasis. KC-associated antigen presentation induces CD4 T-cell arrest, expansion of naturally occurring Foxp3(+) CD25(+) interleukin-10-producing antigen-specific regulatory T cells (Tregs) and tolerogenic immunity. Particle-associated tolerance induction in the liver protected mice from kidney inflammation in T-cell-mediated glomerulonephritis, indicating therapeutic potential of targeting KC for immune-mediated extrahepatic disorders. Liver inflammation in two independent experimental models of chronic liver injury and fibrosis abrogated tolerance induction and led to an immunogenic reprogramming of antigen-specific CD4 T cells. In injured liver, infiltrating monocyte-derived macrophages largely augment the hepatic phagocyte compartment, resulting in antigen redistribution between myeloid cell populations and, simultaneously, KCs lose signature markers of their tolerogenic phenotype. CONCLUSIONS: Hepatic induction of tissue-protective immunological tolerance against particulate antigens is dependent on KCs as well as on a noninflamed liver microenvironment, thereby providing mechanistic explanations for the clinical observation of immune dysfunction and tolerance break in patients with advanced liver diseases.

Regulatory T cells use programmed death 1 ligands to directly suppress autoreactive B cells in vivo.

The mechanisms by which regulatory T cells (T(regs)) suppress autoantibody production are unclear. Here we have addressed this question using transgenic mice expressing model antigens in the kidney. We report that T(regs) were essential and sufficient to suppress autoreactive B cells in an antigen-specific manner and to prevent them from producing autoantibodies. Most of this suppression was mediated through the inhibitory cell-surface-molecule programmed death-1 (PD-1). Suppression required PD-1 expression on autoreactive B cells and expression of the two PD-1 ligands on T(regs). PD-1 ligation inhibited activation of autoreactive B cells, suppressed their proliferation, and induced their apoptosis. Intermediate PD-1(+) cells, such as T helper cells, were dispensable for suppression. These findings demonstrate in vivo that T(regs) use PD-1 ligands to directly suppress autoreactive B cells, and they identify a previously undescribed peripheral B-cell tolerance mechanism against tissue autoantigens.

Batf3-dependent dendritic cells in the renal lymph node induce tolerance against circulating antigens.

Although the spleen is a major site where immune tolerance to circulating innocuous antigens occurs, the kidney also contributes. Circulating antigens smaller than albumin are constitutively filtered and concentrated in the kidney and reach the renal lymph node by lymphatic drainage, where resident dendritic cells (DCs) capture them and induce tolerance of specific cytotoxic T cells through unknown mechanisms. Here, we found that the coinhibitory cell surface receptor programmed death 1 (PD-1) on cytotoxic T cells mediates to their tolerance. Renal lymph node DCs of the CD8(+) XCR1(+) subset, which depend on the transcription factor Batf3, expressed the PD-1 cognate ligand PD-L1. Batf3-dependent DCs in the renal lymph node presented antigen that had been concentrated in the kidney and used PD-L1 to induce apoptosis of cytotoxic T cells. In contrast, T cell tolerance in the spleen was independent of PD-1, PD-L1, and Batf3. In summary, these results clarify how the kidney/renal lymph node system tolerizes the immune system against circulating innocuous antigens.

Differential gene expression in B cells and T helper cells following high-dose glucocorticoid therapy for multiple sclerosis relapse.

BACKGROUND: Despite remarkable advances in the therapy of multiple sclerosis (MS), patients with MS may still experience relapses. High-dose short-term methylprednisolone (MP) remains the standard treatment in the acute management of MS relapses due to its potent anti-inflammatory and immunosuppressive properties. However, there is a lack of studies on the cell type-specific transcriptome changes that are induced by this synthetic glucocorticoid (GC). Moreover, it is not well understood why some patients do not benefit adequately from MP therapy. METHODS: We collected peripheral blood from MS patients in relapse immediately before and after ∼3-5 days of therapy with MP at 4 study centers. CD19+ B cells and CD4+ T cells were then isolated for profiling the transcriptome with high-density arrays. The patients' improvement of neurological symptoms was evaluated after ∼2 weeks by the treating physicians. We finally analyzed the data to identify genes that were differentially expressed in response to the therapy and whose expression differed between clinical responders and non-responders. RESULTS: After MP treatment, a total of 33 genes in B cells and 55 genes in T helper cells were significantly up- or downregulated. The gene lists overlap in 10 genes and contain genes that have already been described as GC-responsive genes in the literature on other cell types and diseases. Their differential expression points to a rapid and coordinated modulation of multiple signaling pathways that influence transcription. Genes that were previously suggested as potential prognostic biomarkers of the clinical response to MP therapy could not be confirmed in our data. However, a greater increase in the expression of genes encoding proteins with antimicrobial activity was detected in CD4+ T cells from non-responders compared to responders. CONCLUSION: Our study delved into the cell type-specific effects of MP at the transcriptional level. The data suggest a therapy-induced ectopic expression of some genes (e.g., AZU1, ELANE and MPO), especially in non-responders. The biological consequences of this remain to be explored in greater depth. A better understanding of the molecular mechanisms underlying clinical recovery from relapses in patients with MS will help to optimize future treatment decisions.

Endogenous foxp3(+) T-regulatory cells suppress anti-glomerular basement membrane nephritis.

Foxp3(+) T-regulatory cells (Tregs) may suppress pathogenic inflammation; however, although transferred Tregs lessen glomerulonephritis in mice, the role of endogenous foxp3(+) cells is not known. To study this, we characterized endogenous foxp3(+) cells in accelerated anti-glomerular basement membrane (GBM) nephritis by using foxp3(GFP) reporter mice to track their responses in early and established disease. Further, diphtheria toxin was used to ablate foxp3(+) Tregs in foxp3(DTR) mice after establishing an immune response. In this model, mice were immunized with sheep globulin in adjuvant, and sheep anti-mouse GBM globulin was injected after 4 days to initiate progressive histological and functional injury. Intrarenal leukocytic infiltrates were increased by day 3 but intrarenal foxp3(+) Tregs, present in interstitial and periglomerular areas, were only increased at day 7. Ablation of foxp3(+) Tregs after injection of anti-GBM globulin increased renal injury and systemic T-cell responses, including increased interferon-γ and interleukin-17A (IL-17A) production, but no change in antibody titers. Compared with foxp3(+) Tregs isolated from naive mice, those from immunized mice produced more IL-10 and more effectively regulated CD4(+)foxp3(-) responder T cells. Thus, endogenous foxp3(+) Tregs infiltrate the kidney in glomerulonephritis, and deleting foxp3(+) cells after the induction of immune responses upregulated T-cell reactions and enhanced disease. Hence, endogenous foxp3(+) cells have increased suppressive capacity after immune stimuli.

New Aspects of Kidney Fibrosis-From Mechanisms of Injury to Modulation of Disease.

Organ fibrogenesis is characterized by a common pathophysiological final pathway independent of the underlying progressive disease of the respective organ. This makes it particularly suitable as a therapeutic target. The Transregional Collaborative Research Center "Organ Fibrosis: From Mechanisms of Injury to Modulation of Disease" (referred to as SFB/TRR57) was hosted from 2009 to 2021 by the Medical Faculties of RWTH Aachen University and the University of Bonn. This consortium had the ultimate goal of discovering new common but also different fibrosis pathways in the liver and kidneys. It finally successfully identified new mechanisms and established novel therapeutic approaches to interfere with hepatic and renal fibrosis. This review covers the consortium's key kidney-related findings, where three overarching questions were addressed: (i) What are new relevant mechanisms and signaling pathways triggering renal fibrosis? (ii) What are new immunological mechanisms, cells and molecules that contribute to renal fibrosis?, and finally (iii) How can renal fibrosis be modulated?

CD25+ T(reg) specifically suppress auto-Ab generation against pancreatic tissue autoantigens.

To study B-cell tolerance against non-lymphoid tissue autoantigens, we generated transgenic rat insulin promoter (RIP)-OVA/hen egg lysozyme (HEL) mice expressing the model antigens, OVA and HEL, in pancreatic islets. Their vaccination with OVA or HEL induced far less auto-Ab titers compared with non-transgenic controls. Depletion of CD25(+) cells during immunization completely restored auto-Ab production, but did not affect antibodies against a foreign control antigen. Depletion at later time-points was not effective. OVA-specific CD25(+) FoxP3(+) T(reg) were more frequent in the autoantigen-draining pancreatic LN than in other secondary lymphatics of RIP-OVA/HEL mice. Consistently, B cells were suppressed in that LN and also in the spleen, which is known to concentrate circulating antigen, such as the antigens used for vaccination. Suppression involved preventing expansion of autoreactive B cells in response to autoantigen, reducing antibody production per B-cell and isotype changes. These findings demonstrate that CD25(+) T(reg) suppress auto-Ab production against non-lymphoid tissue antigens in an antigen-specific manner.

Cutting edge: CD25+ regulatory T cells prevent expansion and induce apoptosis of B cells specific for tissue autoantigens.

To study the role of CD25(+) regulatory T cells (T(regs)) in peripheral B cell tolerance, we generated transgenic rat insulin promoter RIP-OVA/HEL mice expressing the model Ags OVA and HEL in pancreatic islet beta cells (where RIP is rat insulin promoter and HEL is hen egg lysozyme). Adoptively transferred transgenic OVA-specific CD4(+) and CD8(+) T cells proliferated only in the autoantigen-draining pancreatic lymph node (PLN), demonstrating pancreas-specific Ag expression. Transferred HEL-specific transgenic B cells (IgHEL cells) disappeared within 3 wk from transgenic but not from nontransgenic mice immunized with autoantigen. Depletion of CD25(+) FoxP3(+) cells completely restored IgHEL cell numbers. T(reg) exerted an analogous suppressive effect on endogenous HEL-specific autoreactive B cells. T(regs) acted by inhibiting the proliferation of IgHEL cells in the spleen and PLN and by systemic induction of their apoptosis. Furthermore, they reduced BCR and MHC II surface expression on IgHEL cells in the PLN. These findings demonstrate that autoreactive B cells specific for a nonlymphoid tissue autoantigen are controlled by T(regs).

Role of immune cells in crystal-induced kidney fibrosis.

Chronic kidney diseases can lead to kidney fibrosis, which can be considered a futile attempt of tissue healing to replaces functional kidney tissue with connective tissue, basically forming a scar. Chronic inflammation is a frequent cause of kidney fibrosis. Classical as well as recently discovered immune cell subsets and their molecular mediators have been intensively investigated for their contribution to kidney fibrosis and their potential as therapeutic targets. Here we review the current knowledge about the role of immune cells in crystal-induced renal fibrosis.

Recombinant HLA-G as Tolerogenic Immunomodulant in Experimental Small Bowel Transplantation.

The non-classical MHC I paralogue HLA-G is expressed by cytotrophoblast cells and implicated with fetomaternal tolerance by downregulating the maternal adaptive and innate immune response against the fetus. HLA-G expression correlates with favorable graft outcome in humans and recently promising immunosuppressive effects of therapeutic HLA-G in experimental transplantation (skin allograft acceptance) were shown. Consequently, we examined this novel therapeutic approach in solid organ transplantation. In this study, therapeutic recombinant HLA-G5 was evaluated for the first time in a solid organ model of acute rejection (ACR) after orthotopic intestinal transplantation (ITX). Allogenic ITX was performed in rats (Brown Norway to Lewis) with and without HLA-G treatment. It was found that HLA-G treatment significantly reduced histologically proven ACR at both an early and late postoperative timepoint (POD 4/7), concomitant to a functionally preserved graft contractility at POD 7. Interestingly, graft infiltration by myeloperoxidase+ cells was significantly reduced at POD7 by HLA-G treatment. Moreover, HLA-G treatment showed an effect on the allogenic T-cell immune response as assessed by flow cytometry: The influx of recipient-derived CD8+ T-cells into the graft mesenteric lymphnodes at POD7 was significantly reduced while CD4+ populations were not affected. As a potential mechanism of action, an induction of T-reg populations in the mesenteric lymphnodes was postulated, but flow cytometric analysis of classical CD4+/CD25+/FoxP3+Treg-cells showed no significant alteration by HLA-G treatment. The novel therapeutic approach using recombinant HLA-G5 reported herein demonstrates a significant immunosuppressive effect in this model of allogenic experimental intestinal transplantation. This effect may be mediated via inhibition of recipient-derived CD8+ T-cell populations either directly or by induction of non-classical Treg populations.

Differential induction of Ly6G and Ly6C positive myeloid derived suppressor cells in chronic kidney and liver inflammation and fibrosis.

CD11b+Gr1+ myeloid derived suppressor cells (MDSC) are known to be very potent suppressors of T cell immunity and can be further stratified into granulocytic MDSC and monocytic MDSC in mice based on expression of Ly6G or Ly6C, respectively. Here, using these markers and functional assays, we aimed to identify whether MDSC are induced during chronic inflammation leading to fibrosis in both kidney and liver and whether additional markers could more specifically identify these MDSC subsets. In an adenine-induced model of kidney inflammation/fibrosis suppressive Ly6Gpos MDSC were induced. The suppressive function within the Ly6G+ MDSC population was exclusively present in IFNγRß expressing cells. In contrast, in chronic inflammation in the liver induced by bile duct ligation, suppressive capacity was exclusively present in the Ly6Cpos MDSC subset. Gene expression analyses confirmed the differential origins and regulation of those MDSC subsets. Additionally, depletion of MDSC in either kidney or liver fibrosis enhanced fibrosis markers, indicating a protective role for MDSC in organ fibrosis. Thus, our data demonstrate that during liver inflammation and kidney fibrosis MDSC with similar function arise bearing a distinct marker profile and arising from different cell populations.