DUSP1 overexpression attenuates renal tubular mitochondrial dysfunction by restoring Parkin-mediated mitophagy in diabetic nephropathy.

Diabetic nephropathy (DN) is the primary cause of end-stage renal disease, and renal tubular cell dysfunction contributes to the pathogenesis of many kidney diseases. Our previous study demonstrated that dual-specificity protein phosphatase 1 (DUSP1) reduced hyperglycemia-mediated mitochondrial damage; however, its role in hyperglycemia-driven dysfunction of tubular cells is still not fully understood. In this study, we found that DUSP1 is reduced in human proximal tubular epithelial (HK-2) cells under high-glucose conditions. DUSP1 overexpression in HK-2 cells partially restored autophagic flux, improved mitochondrial function, and reduced reactive oxygen species generation and cell apoptosis under high-glucose conditions. Surprisingly, overexpressing DUSP1 abolished the decrease in mitochondrial parkin expression caused by high-glucose stimulation. In addition, knockdown of parkin in HK-2 cells reversed the effects of DUSP1 overexpression on mitophagy and apoptosis under high-glucose conditions. Overall, these data indicate that DUSP1 plays a defensive role in the pathogenesis of DN by restoring parkin-mediated mitophagy, suggesting that it may be considered a prospective therapeutic strategy for the amelioration of DN.

Suppression of lncRNA Snhg1 inhibits high glucose-induced inflammation and proliferation in mouse mesangial cells.

Diabetic nephropathy (DN) is the direct cause of end-stage renal disease, and nephritic inflammation plays a role in its growth and advancement. Aberrant expression of long non-coding RNAs (lncRNAs) correlates with many diseases, including DN. In this study, we investigated whether lncRNA small nucleolar RNA host gene 1 (Snhg1) was mechanistically involved in inflammation and mesangial cell (MC) proliferation in DN. We found that Snhg1 was significantly upregulated in DN renal tissues and high glucose (HG)-treated MCs. Overexpression of Snhg1 promoted inflammatory cytokine expression in MCs and MC proliferation under low-glucose conditions; meanwhile, Snhg1 knockdown suppressed inflammatory cytokine production and MC proliferation under HG conditions. Mechanistically, Snhg1 was found to directly bind miR-27b, thereby preventing the miRNA from binding its target KDM6B mRNA. Furthermore, miR-27b overexpression recapitulated the inhibitory effects of Snhg1 knockdown, whereas restoration of Snhg1 expression attenuated the function of miR-27b in MCs under HG conditions. Taken together, these results indicate that suppression of Snhg1 inhibited HG-induced inflammation and proliferation of MCs by regulating the miR-27b/KDM6B axis.

MicroRNA-148a facilitates inflammatory dendritic cell differentiation and autoimmunity by targeting MAFB.

Monocyte-derived DCs (moDCs) have been implicated in the pathogenesis of autoimmunity, but the molecular pathways determining the differentiation potential of these cells remain unclear. Here, we report that microRNA-148a (miR-148a) serves as a critical regulator for moDC differentiation. First, miR-148a deficiency impaired the moDC development in vitro and in vivo. A mechanism study showed that MAFB, a transcription factor that hampers moDC differentiation, was a direct target of miR-148a. In addition, a promoter study identified that miR-148a could be transcriptionally induced by PU.1, which is crucial for moDC generation. miR-148a ablation eliminated the inhibition of PU.1 on MAFB. Furthermore, we found that miR-148a increased in monocytes from patients with psoriasis, and miR-148a deficiency or intradermal injection of antagomir-148a immensely alleviated the development of psoriasis-like symptoms in a psoriasis-like mouse model. Therefore, these results identify a pivotal role for the PU.1-miR-148a-MAFB circuit in moDC differentiation and suggest a potential therapeutic avenue for autoimmunity.

Identification of Renal Long Non-coding RNA RP11-2B6.2 as a Positive Regulator of Type I Interferon Signaling Pathway in Lupus Nephritis.

Objective: Lupus nephritis (LN) is one of the most serious complications of systemic lupus erythematosus (SLE). Type I interferon (IFN-I) is associated with the pathogenesis of LN. Long non-coding RNAs (lncRNAs) have been implicated in the pathogenesis of SLE, however, the roles of lncRNAs in LN are still poorly understood. Here, we identified and investigated the function of LN-associated lncRNA RP11-2B6.2 in regulating IFN-I signaling pathway. Methods: RNA sequencing was used to analyze the expression of lncRNAs in kidney biopsies from LN patients and controls. Antisense oligonucleotides and CRISPRi system or overexpression plasmids and CRISPRa system were used to perform loss or gain of function experiments. In situ hybridization, imaging flow cytometry, dual-luciferase reporter assay, and ATAC sequencing were used to study the functions of lncRNA RP11-2B6.2. RT-qPCR, ELISA, and western blotting were done to detect RNA and protein levels of specific genes. Results: Elevated lncRNA RP11-2B6.2 was observed in kidney biopsies from LN patients and positively correlated with disease activity and IFN scores. Knockdown of lncRNA RP11-2B6.2 in renal cells inhibited the expression of IFN stimulated genes (ISGs), while overexpression of lncRNA RP11-2B6.2 enhanced ISG expression. Knockdown of LncRNA RP11-2B6.2 inhibited the phosphorylation of JAK1, TYK2, and STAT1 in IFN-I pathway, while promoted the chromatin accessibility and the transcription of SOCS1. Conclusion: The expression of lncRNAs is abnormal in the kidney of LN. LncRNA RP11-2B6.2 is a novel positive regulator of IFN-I pathway through epigenetic inhibition of SOCS1, which provides a new therapeutic target to alleviate over-activated IFN-I signaling in LN.

Identification of LncRNA Linc00513 Containing Lupus-Associated Genetic Variants as a Novel Regulator of Interferon Signaling Pathway.

Systemic lupus erythematosus (SLE) is a complex autoimmune disease characterized by augmented type I interferon signaling. High-throughput technologies have identified plenty of SLE susceptibility single-nucleotide polymorphisms (SNPs) yet the exact roles of most of them are still unknown. Functional studies are principally focused on SNPs in the coding regions, with limited attention paid to the SNPs in non-coding regions. Long non-coding RNAs (lncRNAs) are important players in shaping the immune response and show relationship to autoimmune diseases. In order to reveal the role of SNPs located near SLE related lncRNAs, we performed a transcriptome profiling of SLE patients and identified linc00513 as a significantly over expressed lncRNA containing functional SLE susceptibility loci in the promoter region. The risk-associated G allele of rs205764 and A allele of rs547311 enhanced linc00513 promoter activity and related to increased expression of linc00513 in SLE. We also identified linc00513 to be a novel positive regulator of type I interferon pathway by promoting the phosphorylation of STAT1 and STAT2. Elevated linc00513 expression positively correlated with IFN score in SLE patients. Linc00513 expression was higher in active disease patients than those inactive ones. In conclusion, our data identify two functional promoter variants of linc00513 that contribute to increased level of linc00513 and confer susceptibility on SLE. The study provides new insights into the genetics of SLE and extends the role of lncRNAs in the pathogenesis of SLE.

Association of Abnormal Elevations in IFIT3 With Overactive Cyclic GMP-AMP Synthase/Stimulator of Interferon Genes Signaling in Human Systemic Lupus Erythematosus Monocytes.

OBJECTIVE: Increasing evidence indicates that the cyclic GMP-AMP synthase/stimulator of interferon genes (cGAS/STING) signaling pathway has a critical pathogenic role in systemic lupus erythematosus (SLE). Expression levels of the interferon (IFN)-inducible gene IFIT3 are elevated in SLE patients. However, it is still not clear how IFIT3 contributes to the pathogenesis of SLE. This study was undertaken to investigate the activation of the cGAS/STING signaling pathway in human SLE monocytes, and to determine how elevated expression of IFIT3 could contribute to overactive cGAS/STING signaling in patients with SLE. METHODS: Monocytes from SLE patients or healthy controls were examined for activity of the cGAS/STING signaling pathway and expression levels of IFIT3. Correlations between cGAS/STING signaling activity and SLE clinical features were analyzed. Gain- or loss-of-function experiments were used to determine the role of IFIT3 in cGAS/STING signaling. Coimmunoprecipitation assays were used to identify the interaction between IFIT3 and other proteins. RESULTS: The cGAS/STING signaling pathway was found to have enhanced activity in monocytes from SLE patients compared to healthy controls, as indicated by the higher expression of IFNß downstream. Levels of IFIT3 were significantly elevated in human SLE monocytes, and this was positively correlated with the activity of the cGAS/STING signaling pathway. In vitro, the expression of VACV70-induced IFNß was reduced by knockdown of IFIT3, whereas overexpression of IFIT3 produced an opposite effect. Finally, IFIT3 was found to interact with both STING and TANK-binding kinase 1. CONCLUSION: These findings suggest that IFIT3 is one of the genes that contributes to the overactive cGAS/STING signaling pathway in human SLE monocytes. IFIT3 may therefore serve as a novel therapeutic target for blocking the production of type I IFN and other proinflammatory cytokines by the cGAS/STING signaling pathway in patients with SLE.

T-bet+CD11c+ B cells are critical for antichromatin immunoglobulin G production in the development of lupus.

BACKGROUND: A hallmark of systemic lupus erythematosus is high titers of circulating autoantibodies. Recently, a novel CD11c+ B-cell subset has been identified that is critical for the development of autoimmunity. However, the role of CD11c+ B cells in the development of lupus is unclear. Chronic graft-versus-host disease (cGVHD) is a lupus-like syndrome with high autoantibody production. The purpose of this study was to explore the role of CD11c+ B cells in the pathogenesis of lupus in cGVHD mice. METHODS: cGVHD was induced by an intraperitoneal injection of 5 × 107 Bm12 splenocytes into B6 mice. Flow cytometry was used to analyze mice splenocytes and human samples. Magnetic beads were used to isolate mice B cells. Gene expression was determined by real-time quantitative polymerase chain reaction (RT-qPCR). Enzyme-linked immunosorbent assay (ELISA) was used to detect antibodies in serum and supernatants. RESULTS: The percentage and absolute number of CD11c+ B cells was increased in cGVHD-induced lupus, with elevated levels of antichromatin immunoglobulin (Ig)G and IgG2a in sera. CD11c+ plasma cells from cGVHD mice produced large amounts of antichromatin IgG2a upon stimulation. Depletion of CD11c+ B cells reduced antichromatin IgG and IgG2a production. T-bet was upregulated in CD11c+ B cells. Knockout of T-bet in B cells alleviated cGVHD-induced lupus. Importantly, the percentage of T-bet+CD11c+ B cells increased in lupus patients and positively correlated with serum antichromatin levels. CONCLUSION: T-bet+CD11c+ B cells promoted high antichromatin IgG production in the lupus-like disease model cGVHD. In lupus patients, the percentage of T-bet+CD11c+ B cells was elevated and positively correlated with antichromatin antibodies. The findings provide potential therapeutic insight into lupus disease treatment.