STING1 deficiency ameliorates immune-mediated crescentic glomerulonephritis in mice.

Rapidly progressive/crescentic glomerulonephritis (RPGN/CGN) involves the formation of glomerular crescents by maladaptive differentiation of parietal epithelial cells that leads to rapid loss of renal function. The molecular mechanisms of crescent formation are poorly understood. Therefore, new insights into molecular mechanisms could identify alternative therapeutic targets for RPGN/CGN. Analysis of kidney biopsies from patients with RPGN revealed increased interstitial, glomerular, and tubular expression of STING1, an accessory protein of the c-GAS-dependent DNA-sensing pathway, which was also observed in murine nephrotoxic nephritis induced by an anti-GBM antibody. STING1 was expressed by key cell types involved in RPGN and crescent formation such as glomerular parietal epithelial cells, and tubular cells as well as by inflammation accessory cells. In functional in vivo studies, Sting1-/- mice with nephrotoxic nephritis had lower kidney cytokine expression, milder kidney infiltration by innate and adaptive immune cells, and decreased disease severity. Pharmacological STING1 inhibition mirrored these findings. Direct STING1 agonism in parietal and tubular cells activated the NF-κB-dependent cytokine response and the interferon-induced genes (ISGs) program. These responses were also triggered in a STING1-dependent manner by the pro-inflammatory cytokine TWEAK. These results identify STING1 activation as a pathological mechanism in RPGN/CGN and TWEAK as an activator of STING1. Pharmacological strategies targeting STING1, or upstream regulators may therefore be potential alternatives to treat RPGN. © 2023 The Pathological Society of Great Britain and Ireland.

Iodotyrosines Are Biomarkers for Preclinical Stages of Iodine-Deficient Hypothyroidism in Dehal1-Knockout Mice.

Background: Iodine is required for the synthesis of thyroid hormone (TH), but its natural availability is limited. Dehalogenase1 (Dehal1) recycles iodine from mono- and diiodotyrosines (MIT, DIT) to sustain TH synthesis when iodine supplies are scarce, but its role in the dynamics of storage and conservation of iodine is unknown. Methods: Dehal1-knockout (Dehal1KO) mice were generated by gene trapping. The timing of expression and distribution was investigated by X-Gal staining and immunofluorescence using recombinant Dehal1-beta-galactosidase protein produced in fetuses and adult mice. Adult Dehal1KO and wild-type (Wt) animals were fed normal and iodine-deficient diets for 1 month, and plasma, urine, and tissues were isolated for analyses. TH status was monitored, including thyroxine, triiodothyronine, MIT, DIT, and urinary iodine concentration (UIC) using a novel liquid chromatography with tandem mass spectrometry method and the Sandell-Kolthoff (S-K) technique throughout the experimental period. Results: Dehal1 is highly expressed in the thyroid and is also present in the kidneys, liver, and, unexpectedly, the choroid plexus. In vivo transcription of Dehal1 was induced by iodine deficiency only in the thyroid tissue. Under normal iodine intake, Dehal1KO mice were euthyroid, but they showed negative iodine balance due to a continuous loss of iodotyrosines in the urine. Counterintuitively, the UIC of Dehal1KO mice is twofold higher than that of Wt mice, indicating that S-K measures both inorganic and organic iodine. Under iodine restriction, Dehal1KO mice rapidly develop profound hypothyroidism, while Wt mice remain euthyroid, suggesting reduced retention of iodine in the thyroids of Dehal1KO mice. Urinary and plasma iodotyrosines were continually elevated throughout the life cycles of Dehal1KO mice, including the neonatal period, when pups were still euthyroid. Conclusions: Plasma and urine iodotyrosine elevation occurs in Dehal1-deficient mice throughout life. Therefore, measurement of iodotyrosines predicts an eventual iodine shortage and development of hypothyroidism in the preclinical phase. The prompt establishment of hypothyroidism upon the start of iodine restriction suggests that Dehal1KO mice have low iodine reserves in their thyroid glands, pointing to defective capacity for iodine storage.

Crosstalk between TBK1/IKKε and the type I interferon pathway contributes to tubulointerstitial inflammation and kidney tubular injury.

The type I interferon (TI-IFN) pathway regulates innate immunity, inflammation, and apoptosis during infection. However, the contribution of the TI-IFN pathway or upstream signaling pathways to tubular injury in kidney disease is poorly understood. Upon observing evidence of activation of upstream regulators of the TI-IFN pathway in a transcriptomics analysis of murine kidney tubulointerstitial injury, we have now addressed the impact of the TI-IFN and upstream signaling pathways on kidney tubulointerstitial injury. In cultured tubular cells and kidney tissue, IFNα/ß binding to IFNAR activated the TI-IFN pathway and recruited antiviral interferon-stimulated genes (ISG) and NF-κB-associated proinflammatory responses. TWEAK and lipopolysaccharide (LPS) signaled through TBK1/IKKε and IRF3 to activate both ISGs and NF-κB. In addition, TWEAK recruited TLR4 to stimulate TBK1/IKKε-dependent ISG and inflammatory responses. Dual pharmacological inhibition of TBK1/IKKε with amlexanox decreased TWEAK- or LPS-induced ISG and cytokine responses, as well as cell death induced by a complex inflammatory milieu that included TWEAK. TBK1 or IRF3 siRNA prevented the TWEAK-induced ISG and inflammatory gene expression while IKKε siRNA did not. In vivo, kidney IFNAR and IFNß were increased in murine LPS and folic acid nephrotoxicity while IFNAR was increased in human kidney biopsies with tubulointerstitial damage. Inhibition of TBK1/IKKε with amlexanox or IFNAR neutralization decreased TI-IFN pathway activation and protected from kidney injury induced by folic acid or LPS. In conclusion, TI-IFNs, TWEAK, and LPS engage interrelated proinflammatory and antiviral responses in tubular cells. Moreover, inhibition of TBK1/IKKε with amlexanox, and IFNAR targeting, may protect from tubulointerstitial kidney injury.

CCL20 Expression by Tumor-Associated Macrophages Predicts Progression of Human Primary Cutaneous Melanoma.

The chemokine axis CCR6/CCL20 is involved in cancer progression in a variety of tumors. Here, we show that CCR6 is expressed by melanoma cells. The CCR6 ligand, CCL20, induces migration and proliferation in vitro, and enhances tumor growth and metastasis in vivo Confocal analysis of melanoma tissues showed that CCR6 is expressed by tumor cells, whereas CCL20 is preferentially expressed by nontumoral cells in the stroma of certain tumors. Stromal CCL20, but not tumoral CCR6, predicted poor survival in a cohort of 40 primary melanoma patients. Tumor-associated macrophages (TAM), independently of their M1/M2 polarization profile, were identified as the main source of CCL20 in primary melanomas that developed metastasis. In addition to CCL20, TAMs expressed TNF and VEGF-A protumoral cytokines, suggesting that melanoma progression is supported by macrophages with a differential activation state. Our data highlight the synergistic interaction between melanoma tumor cells and prometastatic macrophages through a CCR6/CCL20 paracrine loop. Stromal levels of CCL20 in primary melanomas may be a clinically useful marker for assessing patient risk, making treatment decisions, and planning or analyzing clinical trials. Cancer Immunol Res; 6(3); 267-75. ©2018 AACR.