Abstract Background Podocyte loss and death are pathological hallmarks of diabetic kidney disease (DKD), and PANoptosis (apoptosis, pyroptosis, and necroptosis) in podocytes is crucial to DKD progression. Regulated in development and DNA damage response 1 (REDD1) is a multifaceted regulator involved in metabolism, oxidative stress, autophagy, and cell fate. In this study, we aimed to investigate the effects and underlying mechanisms of REDD1 on podocyte PANoptosis and autophagy in DKD. Methods REDD1 knockout (KO) mice were induced to diabetes by intraperitoneal injections of streptozotocin (STZ). We assessed renal function, albuminuria, kidney pathology, and podocyte injury in diabetic mice. In vitro, mouse podocyte cells (MPCs) were transfected with REDD1 shRNA plasmid, stratifin (SFN) expression plasmid, SFN siRNA, and treated with TFEB activator 1 or GSK-872 and cultured in high glucose (HG) medium. Gene and protein expression was assessed by real-time quantitative PCR, western blotting, immunofluorescence, and immunohistochemistry. Apoptosis, cytoskeleton change, mitochondrial morphology and membrane potential were evaluated in podocytes. Results REDD1 KO improved renal function and reduced mesangial expansion, podocyte loss, and markers related to PANoptosis in podocytes in diabetic mice. In vitro, REDD1 knockdown suppressed HG-induced PANoptosis, cytoskeletal disorganization, mitochondrial damage, and mitochondrial membrane potential reduction in podocytes. In addition, REDD1 deletion restored autophagy and transcription factor EB (TFEB) expression in diabetic kidneys. Meanwhile, REDD1 knockdown alleviated autophagy dysfunction and promoted TFEB nuclear translocation in podocytes exposed to HG. Moreover, REDD1 KO inhibited podocyte SFN expression in diabetic mice. SFN knockdown or receptor interacting protein kinase 3 (RIPK3) inhibitor GSK-872 alleviated HG-induced PANoptosis and autophagy dysfunction in podocytes. Besides, overexpression of SFN reversed the effect of REDD1 knockdown on PANoptosis and autophagy in HG-treated podocytes. Conclusions REDD1 deficiency protects against podocyte injury through inhibiting PANoptosis and restoring autophagy in DKD. REDD1 is a potential therapeutic target to slow the progression of DKD.

Itch, an E3 ubiquitin ligase, is involved in various cellular functions by regulating ubiquitination and proteasomal degradation of target proteins. However, its roles are unknown in kidney fibrosis. In the present study, the decreased expression of Itch in the renal tubules of patients with chronic kidney disease (CKD) and mice induced by unilateral ureteral obstruction (UUO) or folic acid (FA) is demonstrated. Itch-/- mice exhibited more severe fibrotic lesions and inflammation after obstruction or FA treatment compared with Itch+/+ control. Itch overexpression alleviated UUO- or FA-induced kidney fibrosis and inflammation in mice. In addition, Disheveled 2 (Dvl2) deficiency reduced kidney fibrosis and inflammation in UUO mice. In vitro, Itch overexpression inhibited transforming growth factor-beta 1(TGF-β1)-induced fibrotic response in HK-2 cells. Mechanistically, Itch promoted K48-linked ubiquitination of Dvl2 at lysine 343, facilitating its proteasomal degradation, followed by suppressing GSK-3β/β-catenin signaling pathway, finally leading to the alleviation of kidney fibrosis. Collectively, our studies uncover that Itch exerts a critical role in kidney fibrosis, and it may be an attractive therapeutic target to slow the progression of fibrotic kidney disease.

Lipid accumulation and chronic inflammation are key drivers of chronic kidney disease (CKD) progression to renal fibrosis, but the underlying mechanisms remain incompletely understood. The stimulator of interferon genes (STING), a central regulator of innate immunity and inflammatory responses, has recently garnered significant attention in the context of kidney disease. In this study, we demonstrate that STING expression is markedly upregulated in fibrotic kidneys from both CKD patients and mice subjected to ischemia-reperfusion injury (IRI). Genetic ablation or pharmacological inhibition of STING alleviated renal fibrosis, inflammation, and lipid accumulation in IRI mice, effects that were closely associated with hexokinase 3 (HK3)-mediated lipid metabolism. Moreover, inhibition of STING suppressed NF-κB activation and subsequently reduced HK3 upregulation, thereby attenuating hypoxia/reoxygenation-induced fibrosis, inflammation, and lipid accumulation in primary mouse renal tubular cells (RTECs) and HK-2 cells. Mechanistically, activation of the cGAS-STING signaling pathway promotes the binding of its downstream effector NF-κB p65 to the HK3 promoter, which initiates transcriptional upregulation of HK3, leading to dysregulated lipid metabolism, enhanced inflammation, and ultimately renal fibrosis. Taken together, our findings indicate that the cGAS-STING-NF-κB-HK3 signaling axis may represent a novel therapeutic target for early-stage renal fibrosis.

The cytokine-signaling inhibitor ruxolitinib causes disease flares of chronic lymphocytic leukemia (CLL). This tumor-promoting activity correlates with its ability to inhibit interleukin (IL)-10 production by CLL B cells that have been activated with IL-2 and the Toll-like receptor 7 (TLR7) agonist resiquimod (called 2S cells) in vitro. In TLR-activated normal human B cells, IL-10 production is regulated by cholesterol biosynthesis and can be inhibited by statins. The goal of this study was to determine if statins affect IL-10 production by 2S cells. Lipophilic statins decreased IL-10 production from 2S CLL cells by inhibiting activation of MYC along with secondary signaling events that amplify and maintain IL10 transcription. IL-10 production was restored when the prenylation defect imposed by statins was corrected by adding geranylgeranyl pyrophosphate. CLL cells activated by ruxolitinib in vivo were enriched with genes associated with prenylation inhibition in TLR-activated human B cells in vitro. These findings suggest IL-10 production by 2S-activated CLL cells is regulated by cholesterol biosynthesis in part through geranylgeranyl pyrophosphate production and substrate prenylation. If IL-10 production in the 2S model constitutes a surrogate test for drug responses in vivo, realization of the potential clinical benefits of statins in CLL may require coadministration of other agents.

Lysine acetylation, tightly regulated by the dynamic balance between lysine acetyltransferases (KATs) and deacetylases (KDACs), plays a critical role in cellular processes, yet its involvement in kidney diseases remains elusive due to inconsistent findings. While KDACs have been extensively studied, the role of KATs, particularly p300 and CREB-binding protein (CBP), in kidney diseases is underexplored. p300 and CBP are major KATs and key transcriptional coactivators that regulate gene expression through acetylation of histones and non-histone proteins. A-485, a potent and specific p300/CBP inhibitor, was employed to investigate their role in kidney disease. Pan-acetylation levels were evaluated in three kidney disease models: type 1 diabetes, type 2 diabetes, and unilateral ureteral obstruction (UUO). The UUO model exhibited the most pronounced upregulation of pan-acetylation in the kidney, prompting its selection for further investigation. A-485 treatment in UUO mice and cultured renal epithelial cells reduced pan-acetylation, alleviated renal fibrosis and apoptosis, and improved mitochondrial dysfunction. Mechanistically, A-485 deactivated STAT3 by inhibiting acetylation at the K685 site, which disrupted tyrosine 705 (Y705) phosphorylation, a critical step in STAT3 activation, without altering serine 727 (S727) phosphorylation or total STAT3 expression. This reveals a tandem regulatory mechanism involving acetylation and phosphorylation in STAT3 signaling during renal fibrosis. Our findings demonstrate that A-485 protects against renal fibrosis by targeting p300/CBP-mediated STAT3 acetylation, providing a strong rationale for its potential therapeutic application in kidney diseases. This study advances the understanding of molecular mechanisms underlying kidney diseases and highlights A-485 as a promising candidate for future therapeutic development.

Thioredoxin-interacting protein (TXNIP) is an important regulatory protein for thioredoxin (TRX) that elicits the generation of reactive oxygen species (ROS) by inhibiting the redox function of TRX. Abundant evidence suggests that TXNIP is involved in the fibrotic process of diabetic kidney disease (DKD). However, the potential mechanism of TXNIP in DKD is not yet well understood. In this study, we found that TXNIP knockout suppressed renal fibrosis and activation of mammalian target of rapamycin complex 1 (mTORC1) and restored transcription factor EB (TFEB) and autophagy activation in diabetic kidneys. Simultaneously, TXNIP interference inhibited epithelial-to-mesenchymal transformation (EMT), collagen I and fibronectin expression, and mTORC1 activation, increased TFEB nuclear translocation, and promoted autophagy restoration in HK-2 cells exposed to high glucose (HG). Rapamycin, an inhibitor of mTORC1, increased TFEB nuclear translocation and autophagy in HK-2 cells under HG conditions. Moreover, the TFEB activators, curcumin analog C1 and trehalose, effectively restored HG-induced autophagy, and abrogated HG-induced EMT and collagen I and fibronectin expression in HK-2 cells. Taken together, these findings suggest that TXNIP deficiency ameliorates renal fibrosis by regulating mTORC1/TFEB-mediated autophagy in diabetic kidney diseases.

Kidney fibrosis is considered to be the ultimate aggregation pathway of chronic kidney disease (CKD), but its underlying mechanism remains elusive. Protein kinase C-delta (PKC-δ) plays critical roles in the control of growth, differentiation, and apoptosis. In this study, we found that PKC-δ was highly upregulated in human biopsy samples and mouse kidneys with fibrosis. Rottlerin, a PKC-δ inhibitor, alleviated unilateral ureteral ligation (UUO)-induced kidney fibrosis, inflammation, VDAC1 expression, and cGAS-STING signaling pathway activation. Adeno-associated virus 9 (AAV9)-mediated VDAC1 silencing or VBIT-12, a VDAC1 inhibitor, attenuated renal injury, inflammation, and activation of cGAS-STING signaling pathway in UUO mouse model. Genetic and pharmacologic inhibition of STING relieved renal fibrosis and inflammation in UUO mice. In vitro, hypoxia resulted in PKC-δ phosphorylation, VDAC1 oligomerization, and activation of cGAS-STING signaling pathway in HK-2 cells. Inhibition of PKC-δ, VDAC1 or STING alleviated hypoxia-induced fibrotic and inflammatory responses in HK-2 cells, respectively. Mechanistically, PKC-δ activation induced mitochondrial membrane VDAC1 oligomerization via direct binding VDAC1, followed by the mitochondrial DNA (mtDNA) release into the cytoplasm, and subsequent activated cGAS-STING signaling pathway, which contributed to the inflammation leading to fibrosis. In conclusion, this study has indicated for the first time that PKC-δ is an important regulator in kidney fibrosis by promoting cGAS-STING signaling pathway which mediated by VDAC1. PKC-δ may be useful for treating renal fibrosis and subsequent CKD.

Renal fibrosis is the most common pathway in progressive kidney diseases. The unilateral ureteral obstruction (UUO) model is used to induce progressive renal fibrosis. We evaluated the effects of irisin on renal interstitial fibrosis in UUO mice. The GSE121190, GSE36496, GSE42303, and GSE96101 datasets were downloaded from the Gene Expression Omnibus (GEO) database. In total, 656 differentially expressed genes (DEGs) were identified in normal and UUO mouse renal samples. Periostin and matrix metalloproteinase-2 (MMP-2) were selected to evaluate the effect of irisin on renal fibrosis in UUO mice. In UUO mice, irisin ameliorated renal function, decreased the expression of periostin and MMP-2, and attenuated epithelial-mesenchymal transition and extracellular matrix deposition in renal tissues. In HK-2 cells, irisin treatment markedly attenuated TGF-β1-induced expression of periostin and MMP-2. Irisin treatment also inhibited TGF-β1-induced epithelial-mesenchymal transition, extracellular matrix formation, and inflammatory responses. These protective effects of irisin were abolished by the overexpression of periostin and MMP-2. In summary, irisin treatment can improve UUO-induced renal interstitial fibrosis through the TGF-β1/periostin/MMP-2 signaling pathway, suggesting that irisin may be used for the treatment of renal interstitial fibrosis.

Background: Diabetic kidney disease (DKD) is characterized by renal fibrosis, and the pathogenesis of renal fibrosis is still not definitely confirmed. MiR-204-5p plays an important role in the regulation of fibrosis, autophagy and oxidative stress. In this study, we aimed to investigate the role of miR-204-5p on renal damage in diabetic kidneys and the underlying mechanisms involved. Methods: In vivo, AAV-Ksp-miR-204-5p mimics were injected into mice via tail vein. In vitro, high glucose-induced HK-2 cells were treated with miR-204-5p inhibitor, miR-204-5p mimics, ATG5 siRNA, tertiary butyl hydroquinone (TBHQ), ML385, or 3-Methyladenine (3-MA). FISH and qRT-PCR were used to detect miR-204-5p expression. The expressions of protein and mRNA were detected by Western blotting, immunofluorescence, immunohistochemistry and qRT-PCR. The concentration of fibronectin in HK-2 cells culture medium was detected by ELISA. Results: The expression of miR-204-5p in diabetic kidneys was significantly inhibited than that in control group. Delivering miR-204-5p mimics increased miR-204-5p expression, improved renal function, inhibited renal fibrosis and oxidative stress, and restored autophagy in db/db mice. In vitro, the expression of miR-204-5p was inhibited by HG treatment in HK-2 cells. MiR-204-5p mimics effectively increased miR-204-5p expression and reduced fibronectin and collagen I expression, restored autophagy dysfunction, and increased Nrf2 expression, whereas these alterations were abrogated by Nrf2 inhibitor ML385, autophagy inhibitor 3-methyladenine (3-MA, 5 mM) treatment or ATG5 siRNA transfection in HG-induced HK-2 cells. In addition, miR-204-5p inhibitor significantly inhibited miR-204-5p expression and aggravated HG-induced fibronectin and collagen I expression, autophagy dysfunction, and decreased Nrf2 expression, while these alterations were abolished by Nrf2 activator TBHQ. Furthermore, the binding of miR-204-5p with Keap1 was confirmed by luciferase reporter assay and miR-204-5p negatively regulated Keap1 expression, resulting in the activation of Nrf2 pathway. Conclusion: MicroRNA-204-5p protects against the progression of diabetic renal fibrosis by restoring autophagy via regulating Keap1/Nrf2 pathway.

Nuclear factor kappa B (NFκB) is a pathogenic factor in chronic lymphocytic leukemia (CLL) that is not addressed specifically by current therapies. NFκB is activated by inflammatory factors that stimulate toll-like receptors (TLRs) and receptors for interleukin-1 (IL-1) family members. IL-1 is considered a master regulator of inflammation, and IL-1 receptor signaling is inhibited by the IL-1 receptor antagonist anakinra. These considerations suggested that anakinra might have a role in the treatment of CLL. Consistent with this idea, anakinra inhibited spontaneous and TLR7-mediated activation of the canonical NFκB pathway in CLL cells in vitro. However, CLL cells exhibited only weak signaling responses to IL-1 itself, and anakinra was found to inhibit NFκB along with oxidative stress in an IL-1 receptor-independent manner. Anakinra was then administered with minimal toxicity to 11 previously untreated CLL patients in a phase I dose-escalation trial (NCT04691765). A stereotyped clinical response was observed in all patients. Anakinra lowered blood lymphocytes and lymph node sizes within the first month that were associated with downregulation of NFκB and oxidative stress in the leukemia cells. However, inhibition of NFκB was accompanied by upregulation of type 1 interferon (IFN) signaling, c-MYC-regulated genes and proteins, and loss of the initial clinical response. Anakinra increased IFN signaling and survival of CLL cells in vitro that were, respectively, phenocopied by mitochondrial antioxidants and reversed by IFN receptor blocking antibodies. These observations suggest that anakinra has activity in CLL and may be a useful adjunct for conventional therapies as long as compensatory IFN signaling is blocked at the same time.

Background:The primary pathophysiology of diabetic kidney disease (DKD) is tubulointerstitial fibrosis (TIF), and an essential contributing element is excessive extracellular matrix deposition. Irisin is a polypeptide formed by splitting fibronectin type III domain containing 5 (FNDC5), which participates in a number of physiological and pathological processes.Methods:The purpose of this article is to examine irisin's function in DKD and analyze both its in vitro and in vivo effects. The Gene Expression Omnibus (GEO) database was used to download GSE30122, GSE104954, and GSE99325. Analysis of renal tubule samples from nondiabetic and diabetic mice identified 94 differentially expressed genes (DEGs). The transforming growth factor beta receptor 2 (TGFBR2), irisin, and TGF-β1 were utilized as DEGs to examine the impact of irisin on TIF in diabetic kidney tissue, according to the datasets retrieved from the GEO database and Nephroseq database. Additionally, the therapeutic impact of irisin was also examined using Western blot, RT-qPCR, immunofluorescence, immunohistochemistry, and kits for detecting mouse biochemical indices.Results:In vitro, the findings demonstrated that irisin not only down-regulated the expression of Smad4 and β-catenin but also reduced the expression of proteins linked to fibrosis, the epithelial-mesenchymal transition (EMT), and mitochondrial dysfunction in HK-2 cells maintained in high glucose (HG) environment. In vivo, overexpressed FNDC5 plasmid was injected into diabetic mice to enhance its expression. Our studies found that overexpressed FNDC5 plasmid not only reversed the biochemical parameters and renal morphological characteristics of diabetic mice but also alleviated EMT and TIF by inhibiting Smad4/β-catenin signaling pathway.Conclusion:The above experimental results revealed that irisin could reduce TIF in diabetic mice via regulating the Smad4/β-catenin pathway.

Background:The study aimed to identify core biomarkers related to diagnosis and immune microenvironment regulation and explore the immune molecular mechanism of diabetic nephropathy (DN) through bioinformatics analysis. Methods:GSE30529, GSE99325, and GSE104954 were merged with removing batch effects, and different expression genes (DEGs) were screened at a criterion |log2FC| >0.5 and adjusted P <0.05. KEGG, GO, and GSEA analyses were performed. Hub genes were screened by conducting PPI networks and calculating node genes using five algorithms with CytoHubba, followed by LASSO and ROC analysis to accurately identify diagnostic biomarkers. In addition, two different GEO datasets, GSE175759 and GSE47184, and an experiment cohort with 30 controls and 40 DN patients detected by IHC, were used to validate the biomarkers. Moreover, ssGSEA was performed to analyze the immune microenvironment in DN. Wilcoxon test and LASSO regression were used to determine the core immune signatures. The correlation between biomarkers and crucial immune signatures was calculated by Spearman analysis. Finally, cMap was used to explore potential drugs treating renal tubule injury in DN patients. Results:A total of 509 DEGs, including 338 upregulated and 171 downregulated genes, were screened out. "chemokine signaling pathway" and "cell adhesion molecules" were enriched in both GSEA and KEGG analysis. CCR2, CX3CR1, and SELP, especially for the combination model of the three genes, were identified as core biomarkers with high diagnostic capabilities with striking AUC, sensitivity, and specificity in both merged and validated datasets and IHC validation. Immune infiltration analysis showed a notable infiltration advantage for APC co-stimulation, CD8+ T cells, checkpoint, cytolytic activity, macrophages, MHC class I, and parainflammation in the DN group. In addition, the correlation analysis showed that CCR2, CX3CR1, and SELP were strongly and positively correlated with checkpoint, cytolytic activity, macrophages, MHC class I, and parainflammation in the DN group. Finally, dilazep was screened out as an underlying compound for DN analyzed by CMap. Conclusions:CCR2, CX3CR1, and SELP are underlying diagnostic biomarkers for DN, especially in their combination. APC co-stimulation, CD8+ T cells, checkpoint, cytolytic activity, macrophages, MHC class I, and parainflammation may participate in the occurrence and development of DN. At last, dilazep may be a promising drug for treating DN.

Background: Diabetic nephropathy is one of the main complications of diabetes, inflammation and fibrosis play an important role in its progress. NAD (P) H: quinone oxidoreductase 1 (NQO1) protects cells from oxidative stress and toxic quinone damage. In present study, we aimed to investigate the protective effects and underlying mechanisms of NQO1 on diabetes-induced renal inflammation and fibrosis. Methods: In vivo, adeno-associated virus serotype 9 was used to infect the kidneys of type 2 diabetes model db/db mice to overexpress NQO1. In vitro, human renal tubular epithelial cells (HK-2) transfected with NQO1 pcDNA were cultured in high glucose. The gene and protein expression were assessed by quantitative real-time PCR, western blot, immunofluorescence, and immunohistochemical staining. Mitochondrial reactive oxygen species was detected by MitoSox red. Result: Our study revealed that the expression of NQO1 was markedly down-regulated, Toll-like receptor 4 (TLR4) and TGF-β1 upregulated in vivo and in vitro under diabetic conditions. Overexpression of NQO1 suppressed pro-inflammatory cytokines secretion (IL-6, TNF-α, MCP-1), extracellular matrix (ECM) accumulation (collagen Ⅳ, Fibronectin) and epithelial-mesenchymal transition (EMT) (α-SMA, E-cadherin) in db/db mice kidney and high glucose cultured human renal tubular cells (HK-2). Furthermore, NQO1 overexpression ameliorated high glucose-induced TLR4/NF-κB and TGF-β/Smad pathway activation. Mechanistic studies demonstrated that TLR4 inhibitor (TAK-242) suppressed TLR4/NF-κB signaling pathway, pro-inflammatory cytokines secretion, EMT and ECM-related protein expression in HG-exposed HK-2 cells. In addition, we found that antioxidants NAC and tempol increased the expression of NQO1, decreased the expression of TLR4, TGF-β1, Nox1, Nox4 and ROS production in HK-2 cells cultured with high glucose. Conclusions: These above data suggest that NQO1 alleviates diabetes-induced renal inflammation and fibrosis by regulating TLR4/NF-κB and TGF-β/Smad signaling pathways.

IntroductionChronic lymphocytic leukemia (CLL) is characterized by an aberrant cytokine network that can support tumor growth by triggering janus kinase (JAK)/STAT pathways. Targeting cytokine-signaling should then be a rational therapeutic strategy but the JAK inhibitor ruxolitinib failed to control and seemingly accelerated the disease in clinical trials.MethodsThe effect of ruxolitinib on primary human CLL cells was studied in vitro and in vivo.ResultsRuxolitinib increased phosphorylation of IRAK4, an important toll-like receptor (TLR)- signaling intermediate, in circulating CLL cells in vitro. It also enhanced p38 and NFKB1 phosphorylation while lowering STAT3 phosphorylation in CLL cells activated with TLR-7/8 agonists and IL-2. Among the cytokines made by activated CLL cells, high levels of IL-10 contributed strongly to STAT3 phosphorylation and inhibited TLR7 activity. Ruxolitinib limited TLR-mediated IL10 transcription and markedly reduced IL-10 production in vitro. It also decreased blood levels of IL-10 while increasing TNFα along with phospho-p38 expression and gene sets associated with TLR-activation in CLL cells in vivo. The bruton's tyrosine kinase inhibitor ibrutinib decreased IL-10 production in vitro but, in contrast to ruxolitinib, blocked initial IL10 transcription induced by TLR-signaling in vitro, decreased TNFα production, and deactivates CLL cells in vivo.DiscussionThese findings suggest the possible benefits of inhibiting growth factors with JAK inhibitors in CLL are outweighed by negative effects on potential tumor suppressors such as IL-10 that allow unrestrained activation of NFκB by drivers such as TLRs. Specific inhibition of growth-promoting cytokines with blocking antibodies or infusing suppressive cytokines like IL-10 might be better strategies to manipulate cytokines in CLL.

To observe the role of transforming growth factor beta-activated kinase 1 (TAK1)/p38 MAPK/TGF-β1 signal pathway plays in oxidative stress and apoptosis in human renal tubular epithelial cells (HK-2) under high glucose induction. HK-2 cells were cultured in high glucose medium with and without TAK1 inhibitor 5Z-7-oxozeaenol. TUNEL and flow cytometry were used to detect cell apoptosis. The protein expression of TAK1, TGF-β1, Bax and Bcl-2 was detected by immunofluorescence. Meanwhile, flow cytometry was used to detect the production of reactive oxygen species (ROS), and MitoSOX staining was performed to detect the production of mitochondrial ROS. Moreover, real-time quantitative PCR and Western blotting was used to measure the expression of TAK1, TGF-β1, NOX1, NOX4 and HO-1, Bax, Bcl-2, p38MAPK, p-p38MAPK and TGF-β1. Results showed that high glucose up-regulated the protein expression of p-TAK1, p-p38 MAPK and TGF-β1, which induced the aggravation of oxidative stress by promoting the production of ROS, thus promote the apoptosis in HK-2 cells. However, addition of 5z -7-oxozeaenol in HK-2 cells reversed all the above functions induced by high glucose. Another experimental result also showed that SB203580, a p38MAPK inhibitor can down-regulated TGF-β1 expression and reduce ROS production, thus alleviate cell apoptosis in TAK1 overexpression group. In summary, high glucose intervention could activate TAK1 and promote apoptosis in HK-2 cells. Inhibition of TAK1 expression could block p38 MAPK/TGF-β1 signaling pathway and reduce ROS production and oxidative stress, which may be one of the signal pathways of TAK1 to reduce apoptosis of HK-2 cells induced by high glucose.Abbreviations: DN, Diabetic nephropathy; TAK1, transforming growth factor β-activated kinase-1; TGF-β, transforming growth factor-β; NG, normal glucose; HG, high glucose; p38 MAPK, p38 mitogen-activated protein kinase; ROS, reactive oxygen species.

Sestrin2 is identified as a stress-induced protein and could functionate in many aspects. In our study, we investigated the latent impact of Sestrin2 on podocyte injury and its molecular mechanism in vivo and in vitro in diabetic kidney disease (DKD). Sestrin2 was low-expressed in renal biopsies from individuals with DKD, the glomeruli from diabetic mice, and mouse podocytes exposed to high glucose (HG). Sestrin2 overexpression ameliorated HG-induced phenotypic alterations, apoptosis, and oxidative stress in conditionally immortalized mouse podocytes and modulated the activity of Thrombospondin-1 (TSP-1)/transforming growth factor (TGF-β1)/Smad3 pathway in podocytes. Moreover, TSP-1 inhibitor LSKL or TGF-β blocker Pirfenidone arrested podocyte injury induced by HG. Streptozotocin (STZ) was employed to render equivalent diabetes in B6-TgN (CMV-Sestrin2) (TgN) and wild-type (WT) control mice. Sestrin2 alleviated increased levels of 24‐h urinary protein, blood urea nitrogen, serum creatinine and triglyceride, and urine 8-OHdG in diabetic mice. Podocyte phenotypic alterations, increased expression of apoptosis-associated proteins and podocyte loss were observed in WT but not in diabetic TgN mice, as well as oxidative stress. Additionally, TSP-1/TGF-β1/Smad3 signaling pathway was also suppressed in glomeruli of diabetic TgN mice. Thus, Sestrin2 mitigates podocyte injury in DKD via orchestrating TSP-1/TGF-β1/Smad3 pathway, underlining Sestrin2 as a promising therapeutic target for DKD.

Lipid accumulation and progressive necroinflammation play pivotal roles in the development of diabetic nephropathy. C1q tumour necrosis factor-related protein-3 (CTRP3) is an adipokine with pleiotropic functions in cell proliferation, glucose and lipid metabolism, and inflammation. However, the mechanism and involvement of CTRP3 in lipid metabolism and the necroinflammation of renal tubular cells remain unclear. Here, we report that CTRP3 expression decreased in a time- and concentration-dependent manner in high glucose-stimulated HK-2 cells. We noted that the overexpression of CTRP3 or recombinant CTRP3 (rCTRP3) treatment prevented high glucose-induced lipid accumulation by inhibiting the expression of sterol regulatory element-binding protein-1 and increasing the expression of peroxisome proliferator-activated receptor-α and ATP-binding cassette A1. Moreover, the nucleotide-binding oligomerisation domain-like receptor protein 3-mediated inflammatory response and mixed lineage kinase domain-like protein-dependent necroinflammation were inhibited by CTRP3 overexpression or rCTRP3 treatment in HK-2 cells cultured in high glucose. Furthermore, lipotoxicity-induced by palmitic acid was found to be involved in necroinflammation in HK-2 cells, and CTRP3 displayed the same protective effect. CTRP3 also activated the adenosine monophosphate-activated protein kinase (AMPK) pathway, whereas adenine 9-β-D-arabinofuranoside, an AMPK inhibitor, replicated the protective effects of CTRP3. Besides, using kidney biopsies from patients with diabetes, we found that decreased CTRP3 expression was accompanied by increased lipid deposition, as well as the structural and functional injury of renal tubular cells. Our findings demonstrate that CTRP3 affects lipid metabolism and necroinflammation in renal tubular cells via the AMPK signalling pathway. Thus, CTRP3 may be a potential therapeutic target in diabetic renal injury.

Diabetic kidney disease (DKD) is the most-common cause of chronic renal failure and end-stage renal disease (ERSD) in diabetes mellitus. Autophagy impairment is implicated in the progression of DKD. REDD1 is a stress-regulated protein whose levels are increased in several neurodegenerative diseases. The effect of REDD1 on autophagy has been investigated; however, it has not been investigated whether and how REDD1 regulates autophagy in DKD. Here, we identified that REDD1 was up-regulated both in the renal samples of patients with diabetic nephropathy and diabetic mice from GEO database. Next, defective autophagy was observed in kidneys of db/db mice and in high glucose (HG) - treated HK-2 cells, which were restored by REDD1 deficiency. Consequently, we indicated that REDD1 was attributed to the activation of Nucleotide leukin-rich polypeptide 3 (NLRP3) inflammasome and Notch1 signaling pathway both in db/db mice and HG-treated HK-2 cells. Then, further assessment showed that NLRP3 inhibition protected against defective autophagy, and block Notch1 signaling pathway under diabetic conditions using streptozotocin (STZ)-induced diabetic NLRP3 knockout (NKO) mice and MCC950-treated db/db mice. Meanwhile, we detected that both NLRP3 and Notch1 deficiency promoted autophagy. Importantly, IL-1β aggravated the above-mentioned defective autophagy, and promoted Notch1 expression in HK-2 cells under HG conditions. Taken together, the results of our study demonstrated that REDD1 governed autophagy with the participation of NLRP3-mediated Notch1 signaling. Regulation of REDD1 may be a potential therapeutic approach for DKD.

BACKGROUND AND AIM:Diabetic kidney disease (DKD) is the most-common cause of chronic renal failure and end-stage renal disease (ERSD) in diabetes mellitus (DM) patients. Renal inflammation and glomerular or interstitial fibrosis are mainly associated with the progression of DKD. Carbohydrate response element binding protein (ChREBP) is activated and transcribed in a glucose dependent manner. This study is aimed at exploring the role and underlying mechanisms of ChREBP in DKD.METHODS:ChREBP knockout mice, obtained by CRISPR Cas9 gene editing technology, were used to study the effects of ChREBP on inflammation and fibrosis in diabetic kidney of mice. Human renal tubular epithelial (HK-2) cells were cultured in a medium containing normal or high glucose levels. Additionally, the role of ChREBP in high glucose (HG)-induced NLRP3 inflammasome activation was assessed.RESULTS:We identified that renal inflammation, renal extracellular matrix deposition, and renal fibrosis were restored by ChREBP deficiency in diabetic mouse kidney. Consequently, ChREBP deficiency decreased the activation of nucleotide leukin-rich polypeptide 3 (NLRP3) inflammasome, which later restrained hyperglycemia-induced renal fibrosis. Importantly, NLRP3 inflammasome aggravated the above-mentioned renal fibrosis via TGF-β1 expression and the signaling pathways of Smad2/3 and the p38 MAPK. Additionally, ChREBP deficiency inhibited NLRP3 inflammasome activation both in HG-induced HK-2 cells and diabetic mouse kidney.CONCLUSION:Our findings establish a critical role of ChREBP in engaging inflammation and renal fibrosis by regulating NLRP3 inflammasome activation in DKD.

Type I IFN is made by cells in response to stress. Cancer cells exist in a state of stress, but their IFN response is complex and not completely understood. This study investigated the role of autocrine IFN in human chronic lymphocytic leukemia (CLL) cells. CLL cells were found to make low amounts of IFN via TANK-binding kinase 1 pathways, but p-STAT1 and -STAT2 proteins along with IFN-stimulated genes that reflect IFN activation were variably downregulated in cultured CLL cells by the neutralizing IFNAR1 Ab anifrolumab. Patients with CLL were segregated into two groups based on the response of their leukemia cells to anifrolumab. Samples associated with more aggressive clinical behavior indicated by unmutated IGHV genes along with high CD38 and p-Bruton’s tyrosine kinase expression exhibited responses to low amounts of IFN that were blocked by anifrolumab. Samples with more indolent behavior were unaffected by anifrolumab. Hypersensitivity to IFN was associated with higher expression of IFNAR1, MX1, STAT1, and STAT2 proteins and lower activity of negative regulatory tyrosine phosphatases. Autocrine IFN protected responsive CLL cells from stressful tissue culture environments and therapeutic drugs such as ibrutinib and venetoclax in vitro, in part by upregulating Mcl-1 expression. These findings suggest hypersensitivity to IFN may promote aggressive clinical behavior. Specific blockade of IFN signaling may improve outcomes for patients with CLL with higher-risk disease.