Attenuation of renal injury by depleting cDC1 and by repurposing Flt3 inhibitor in anti-GBM disease.

Previous studies found cDC1s to be protective in early stage anti-GBM disease through Tregs, but pathogenic in late stage Adriamycin nephropathy through CD8+ T cells. Flt3 ligand is a growth factor essential for cDC1 development and Flt3 inhibitors are currently used for cancer treatment. We conducted this study to clarify the role and mechanisms of effects of cDC1s at different time points in anti-GBM disease. In addition, we aimed to utilize drug repurposing of Flt3 inhibitors to target cDC1s as a treatment of anti-GBM disease. We found that in human anti-GBM disease, the number of cDC1s increased significantly, proportionally more than cDC2s. The number of CD8+ T cells also increased significantly and their number correlated with cDC1 number. In XCR1-DTR mice, late (day 12-21) but not early (day 3-12) depletion of cDC1s attenuated kidney injury in mice with anti-GBM disease. cDC1s separated from kidneys of anti-GBM disease mice were found to have a pro-inflammatory phenotype (i.e. express high level of IL-6, IL-12 and IL-23) in late but not early stage. In the late depletion model, the number of CD8+ T cells was also reduced, but not Tregs. CD8+ T cells separated from kidneys of anti-GBM disease mice expressed high levels of cytotoxic molecules (granzyme B and perforin) and inflammatory cytokines (TNF-α and IFN-γ), and their expression reduced significantly after cDC1 depletion with diphtheria toxin. These findings were reproduced using a Flt3 inhibitor in wild type mice. Therefore, cDC1s are pathogenic in anti-GBM disease through activation of CD8+ T cells. Flt3 inhibition successfully attenuated kidney injury through depletion of cDC1s. Repurposing Flt3 inhibitors has potential as a novel therapeutic strategy for anti-GBM disease.

Regulatory innate lymphoid cells suppress innate immunity and reduce renal ischemia/reperfusion injury.

Innate lymphoid cells are a recently recognized group of immune cells with critical roles in tissue homeostasis and inflammation. Regulatory innate lymphoid cells are a newly identified subset of innate lymphoid cells, which play a suppressive role in the innate immune response, favoring the resolution of intestinal inflammation. However, the expression and role of regulatory innate lymphoid cells in kidney has not been reported. Here, we show that regulatory innate lymphoid cells are present in both human and mouse kidney, express similar surface markers and form a similar proportion of total kidney innate lymphoid cells. Regulatory innate lymphoid cells from kidney were expanded in vitro with a combination of IL-2, IL-7 and transforming growth factor-ß. These cells exhibited immunosuppressive effects on innate immune cells via secretion of IL-10 and transforming growth factor-ß. Moreover, treatment with IL-2/IL-2 antibody complexes (IL-2C) promoted expansion of regulatory innate lymphoid cells in vivo, and prevent renal ischemia/reperfusion injury in Rag-/- mice that lack adaptive immune cells including Tregs. Depletion of regulatory innate lymphoid cells with anti-CD25 antibody abolished the beneficial effects of IL-2C in the Rag-/- mice. Adoptive transfer of ex vivo expanded regulatory innate lymphoid cells improved renal function and attenuated histologic damage when given before or after induction of ischemia/reperfusion injury in association with reduction of neutrophil infiltration and induction of reparative M2 macrophages in kidney. Thus, our study shows that regulatory innate lymphoid cells suppress innate renal inflammation and ischemia/reperfusion injury.

Type 2 innate lymphoid cells are protective against hepatic ischaemia/reperfusion injury.

Background and Aims: Although type 2 innate lymphoid cells (ILC2s) were originally found to be liver-resident lymphocytes, the role and importance of ILC2 in liver injury remains poorly understood. In the current study, we sought to determine whether ILC2 is an important regulator of hepatic ischaemia/reperfusion injury (IRI). Methods: ILC2-deficient mice (ICOS-T or NSG) and genetically modified ILC2s were used to investigate the role of ILC2s in murine hepatic IRI. Interactions between ILC2s and eosinophils or macrophages were studied in coculture. The role of human ILC2s was assessed in an immunocompromised mouse model of hepatic IRI. Results: Administration of IL-33 prevented hepatic IRI in association with reduction of neutrophil infiltration and inflammatory mediators in the liver. IL-33-treated mice had elevated numbers of ILC2s, eosinophils, and regulatory T cells. Eosinophils, but not regulatory T cells, were required for IL-33-mediated hepatoprotection in IRI mice. Depletion of ILC2s substantially abolished the protective effect of IL-33 in hepatic IRI, indicating that ILC2s play critical roles in IL-33-mediated liver protection. Adoptive transfer of ex vivo-expanded ILC2s improved liver function and attenuated histologic damage in mice subjected to IRI. Mechanistic studies combining genetic and adoptive transfer approaches identified a protective role of ILC2s through promoting IL-13-dependent induction of anti-inflammatory macrophages and IL-5-dependent elevation of eosinophils in IRI. Furthermore, in vivo expansion of human ILC2s by IL-33 or transfer of ex vivo-expanded human ILC2s ameliorated hepatic IRI in an immunocompromised mouse model of hepatic IRI. Conclusions: This study provides insight into the mechanisms of ILC2-mediated liver protection that could serve as therapeutic targets to treat acute liver injury. Impact and Implications: We report that type 2 innate lymphoid cells (ILC2s) are important regulators in a mouse model of liver ischaemia/reperfusion injury (IRI). Through manipulation of macrophage and eosinophil phenotypes, ILC2s mitigate liver inflammation and injury during liver IRI. We propose that ILC2s have the potential to serve as a therapeutic tool for protecting against acute liver injury and lay the foundation for translation of ILC2 therapy to human liver disease.

Conventional Type 1 Dendritic Cells (cDC1) in Human Kidney Diseases: Clinico-Pathological Correlations.

Background: cDC1 is a subset of conventional DCs, whose most recognized function is cross-presentation to CD8+ T cells. We conducted this study to investigate the number and location of cDC1s in various human kidney diseases as well as their correlation with clinico-pathological features and CD8+ T cells. Methods: We analyzed 135 kidney biopsies samples. Kidney diseases included: acute tubular necrosis (ATN), acute interstitial nephritis (AIN), proliferative glomerulonephritis (GN) (IgA nephropathy, lupus nephritis, pauci-immune GN, anti-GBM disease), non-proliferative GN (minimal change disease, membranous nephropathy) and diabetic nephropathy. Indirect immunofluorescence staining was used to quantify cDC1s, CD1c+ DCs, and CD8+ T cells. Results: cDC1s were rarely present in normal kidneys. Their number increased significantly in ATN and proliferative GN, proportionally much more than CD1c+ DCs. cDC1s were mainly found in the interstitium, except in lupus nephritis, pauci-immune GN and anti-GBM disease, where they were prominent in glomeruli and peri-glomerular regions. The number of cDC1s correlated with disease severity in ATN, number of crescents in pauci-immune GN, interstitial fibrosis in IgA nephropathy and lupus nephritis, as well as prognosis in IgA nephropathy. The number of CD8+ T cells also increased significantly in these conditions and cDC1 number correlated with CD8+ T cell number in lupus nephritis and pauci-immune GN, with many of them closely co-localized. Conclusions: cDC1 number correlated with various clinic-pathological features and prognosis reflecting a possible role in these conditions. Their association with CD8+ T cells suggests a combined mechanism in keeping with the results in animal models.

The potential role of Ets-1 and miR-326 in CD19+B cells in the pathogenesis of patients with systemic lupus erythematosus.

OBJECTIVES: The aim of this study was to investigate the B cell-associated transcription factors, Ets-1 and microRNA, miR-326 in systemic lupus erythematosus (SLE) patients, and their correlation with the pathogenesis of SLE. METHOD: A total of 44 SLE patients and 20 healthy controls were enrolled in this research, all patients fulfilled the American College of Rheumatology classification criteria for SLE. The mRNA expression of Ets-1 and miR-326 in CD19+B cells from SLE patients were examined by qRT-PCR. The percentages of CD19+CD138+plasma cells were analyzed by Flow cytometry. RESULTS: We found decreased expression of Ets-1 mRNA in SLE patients compared with the healthy controls ([0.228 (0.145, 0.507)] vs [0.583 (0.452, 0.763)], p = 0.001),while increased expression of miR-326 mRNA in CD19+B cells SLE patients compared with the healthy controls([1.092 (0.457, 2.855)] vs [0.685 (0.274, 0.819)], p = 0.008). The percentage of CD19+CD138+plasma cells in SLE patients was higher than that of healthy controls (0.55 ± 0.21% vs 0.36 ± 0.21%, p = 0.002). Moreover, a negative correlation between expression of Ets-1 mRNA and miR-326 mRNA in CD19+B cells was detected (r = - 0.334, p = 0.027). A significant association between the occurrences of CD19+CD138+plasma cells and the levels of Ets-1 mRNA and miR-326 mRNA was observed (r = - 0.417, p = 0.005 and r = 0.482, p = 0.001, respectively). CONCLUSIONS: Our results suggest that miR-326 might promote B cells differentiation by targeting Ets-1, a negative regulator of B cells differentiation and therefore participate in the pathogenesis of SLE.