BACKGROUND:Diabetic Nephropathy (DN) is a Chronic Kidney Disease (CKD), and its main pathological changes are renal tubular injury and glomerulosclerosis. Semen Ziziphi Spinosae (SZS) is the seed of Ziziphus jujuba var. spinosa (Bunge) Hu ex H.F. Chow. As a triterpene saponin, Jujuboside A (Ju A) is the main active substance isolated from SZS. This study sought to investigate the potential effect and mechanism of Jujuboside A against DN. METHODS:The anti-apoptotic effects of Ju A on renal parenchymal cells of DN were examined by in vivo and in vitro studies. Molecular docking and Molecular Dynamics (MD) simulation revealed that Ju A could bind to TNF-α and Caspase-3 via forming stable receptor-ligand complexes, respectively. Immunofluorescence (IF) staining and ELISA detection were carried out to investigate the potential mechanisms by which Ju A exerted its amelioration effect on DN. RESULTS:Our study showed that, accompanied by the restored renal function, Ju A inhibited apoptosis of renal tubules and glomeruli in vivo and in vitro. Network pharmacology revealed that 42 overlapping targets were related to Ju A and DN. Among them, IL6, IL1B, TNF, VEGFA, EGFR, ALB, IGF1, FGF2, CASP3, and ESR1 were the top 10 targets. Ju A could bind to TNF-α and Caspase-3 via forming stable receptor-ligand complexes, respectively, as demonstrated by molecular docking and MD simulation. Ju A decreased the protein levels of TNF-α and IL-1β in renal tubules and glomeruli of diabetic mice, and in HG-cultured HK-2 cells and podocytes, leading to the alleviation of inflammation. Besides, the up-regulated relative phosphorylation levels of NF-κB p65 and cleaved caspase-3 were also down-regulated by Ju A in vivo and in vitro. DISCUSSION:The research showed that Ju A had a high affinity for Caspase-3 and TNF-α, and the underlying mechanism of Ju A against DN was the inhibition of apoptosis in renal tubular epithelial cells and podocytes. These findings strengthened the evidence that Ju A could be a potential treatment strategy for DN and offered opportunities for therapeutic advances in the field. CONCLUSION:Ju A could inhibit apoptosis and alleviate inflammation of renal parenchymal cells by inactivating the TNF-α/NF-κB p65/Caspase-3 signaling pathway, exerting renal protective effect against DN.

Our research team developed AB-38b, a biphenyl diester derivative containing α,β-unsaturated carbonyl groups, inspired by investigations into the pharmacological prevention and pathophysiology of diabetic kidney disease (DKD). We have completed the synthesis and pharmacodynamic evaluation of AB-38b in preclinical DKD studies. For pharmacokinetic analysis, we established a high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method for quantifying AB-38b in rat plasma, as well as in liver and kidney tissues. Sample preparation involved protein precipitation using methanol, with tolvaptan employed as the internal standard (IS). Chromatographic separation was achieved on a Shim-pack VP-ODS C18 column (2.0 × 150 mm, 5 µm), with mobile phase A (5 mM ammonium acetate) and mobile phase B (methanol) under gradient elution at a flow rate of 0.2 mL min-1. AB-38b and the IS were detected and quantified using positive electrospray ionization in multiple reaction monitoring (MRM) mode at transitions of m/z 705.30 → 618.20 for AB-38b and m/z 449.30 → 252.20 for the IS. The method demonstrated excellent linearity, stability, accuracy, precision, recovery, and a non-significant matrix effect within the validated range. A one-compartment model with rapid absorption and distribution, quicker elimination, and clear tissue distribution was demonstrated by the mean blood concentration-time curve of AB-38b in rat plasma. The liver tissue contained high levels of AB-38b, with no evidence of tissue accumulation observed. These findings offer a reference for further research into the pharmacological mechanisms and potential therapeutic applications of AB-38b in metabolically related diseases, especially DKD.

BackgroundEarly detection of mild cognitive impairment (MCI) is essential for initiating timely intervention and delaying progression to dementia. However, current diagnostic approaches often lack scalability and efficiency, limiting their utility in primary care and community settings.ObjectiveThis study developed and validated the Integrated Cognitive Screening Platform (ICSP), a self-administered, tablet-based tool designed for rapid, multidomain screening of MCI.MethodsICSP comprises five cognitive tasks involving immediate and delayed memory, attention, sensory perception, and executive function. Both accuracy scores and reaction times (RTs) were recorded and processed within five minutes. A total of 126 participants (76 with MCI, 50 cognitively normal controls) completed standard neuropsychological assessments and the ICSP battery. A multivariate logistic regression model was developed using 60% of the data as a training set and evaluated on the remaining 40% as a validation set.ResultsRTs and accuracy scores in sensory perception and executive function tasks, along with educational attainment, were identified as significant predictors of MCI. The model achieved high classification performance (AUC: 0.915; p < 0.001), with robust validation performance (AUC: 0.821; p < 0.001).ConclusionsICSP is an accurate and scalable digital screening tool capable of identifying MCI with high specificity. Its multimodal design and automated analysis make it well-suited for clinical and community-level early detection of cognitive impairments.Trial registrationThis study was registered with the Chinese Clinical Trial Registry (Registration No. ChiCTR2400082429) on March 28, 2024.

Dapagliflozin shows variable renoprotective efficacy in patients with type 2 diabetic kidney disease (T2DKD). A retrospective clinical analysis confirmed marked interindividual variability in its urinary protein-lowering effects. To investigate the mechanism, drug affinity responsive target stability (DARTS) combined with quantitative proteomics was applied for target identification. Subsequent in vivo and in vitro validation suggested that GSK3β acts as a mediator of dapagliflozin-associated nephroprotection. Dapagliflozin directly bound and partially inhibited GSK3β, and GSK3β activity influenced podocyte protection. To translate findings into clinical relevance, a prospective trial was conducted. The GSK3B rs60393216 polymorphism was associated with urinary albumin-to-creatinine ratio (UACR) reduction after dapagliflozin therapy. These findings suggest that GSK3β contributes to the renoprotective effects of dapagliflozin and that GSK3B polymorphisms may influence therapeutic responses in T2DKD.

This study investigated the effects of combined transcranial direct current stimulation (tDCS) and aerobic exercise (AE) on executive function and hematological markers in poststroke cognitive impairment (PSCI) patients. A total of 74 PSCI patients were randomly assigned to three groups: the tDCS + AE, tDCS, and AE groups. The tDCS group received 2.0 mA anodal stimulation over the left dorsolateral prefrontal cortex (DLPFC) for 30 min, five days per week for four weeks. The AE group performed moderate-intensity exercise (60–70

STUDY OBJECTIVES:Whether cerebrospinal fluid (CSF)-dynamics-related glymphatic alterations occur in middle-aged and older adults with chronic insomnia (CI) remains unknown. We therefore examined global and network-level blood oxygenation level-dependent (BOLD)-CSF coupling in this population and assessed the effects of low-frequency repetitive transcranial magnetic stimulation (LF-rTMS) during standardized hypnotic tapering. METHODS:This two-stage study included a cross-sectional comparison and a randomized, double-blind, parallel-group, sham-controlled trial. In Stage 1, 43 CI patients and 40 matched healthy controls completed sleep assessments and resting-state functional magnetic resonance imaging to quantify global and network-level BOLD-CSF coupling. In Stage 2, 26 CI patients were randomized (1:1) to receive 4 weeks of active or sham LF-rTMS during hypnotic tapering. Sleep was assessed at baseline, 2 weeks, 4 weeks, and 12 months. Neuroimaging was acquired at baseline and 4 weeks. RESULTS:CI patients showed significantly reduced global BOLD-CSF coupling, particularly in frontoparietal network (FPN) and default mode network (DMN). Global and FPN coupling correlated with sleep quality. In the randomized trial, LF-rTMS produced greater improvements in sleep at 4 weeks than sham resulted in fewer participants resuming hypnotics at 12 months. LF-rTMS increased global and DMN BOLD-CSF coupling, and these changes were associated with improvements in sleep. CONCLUSIONS:Middle-aged and older adults with CI exhibit reduced global BOLD-CSF coupling, indicating alterations in CSF dynamics that may relate to glymphatic function. LF-rTMS improved insomnia symptoms and modulated this coupling, indicating therapeutic potential for CI.Trial Registration: ChiCTR2100049455.

All-trans retinoic acid (ATRA), the active metabolite of vitamin A, serves as the first-line therapy for acute promyelocytic leukemia (APL) in clinical. However, increasing clinical evidence indicates that ATRA treatment is frequently associated with dyslipidemia, the underlying molecular mechanisms of which remain unclear. Fenofibrate, a specific agonist of peroxisome proliferator-activated receptor alpha (PPARα), is widely used in the management of metabolic disorders, yet its potential to alleviate ATRA-induced lipid abnormalities has not been fully elucidated. In this study, we established a mouse model of ATRA-induced hyperlipidemia and hepatic steatosis to investigate the underlying mechanisms and assess the therapeutic effects of fenofibrate as a combinatorial agent. Our findings revealed that ATRA promoted the formation of RARα/RXRα heterodimers, which activated the hepatic FOXO1-APOCIII pathway, leading to hyperlipidemia and hepatic lipid accumulation in mice. Fenofibrate effectively counteracted these effects by activating PPARα, thereby competitively inhibiting RARα binding to RXRα and restoring lipid homeostasis. This study reveals a novel mechanism underlying ATRA-induced hyperlipidemia and hepatic lipid accumulation, which offers a theoretical foundation for the clinical use of fenofibrate in managing ATRA-induced hyperlipidemia and hepatic steatosis.

Renal tubular epithelial cells are among the earliest renal parenchymal cells to be injured in the context of acute kidney injury (AKI). Numerous studies have confirmed that fibrinogen-like protein 2 (FGL2) can regulate the occurrence and development of inflammation during disease progression. We found that FGL2 expression is elevated under AKI conditions. However, the role of FGL2 in AKI remains unclear. To elucidate the role of FGL2 in AKI, we employed lentiviral and adeno-associated virus for transfection in cellular and murine models. Furthermore, by leveraging datasets from the gene expression omnibus and gene set enrichment analysis databases, we identified the inflammation-related genes in AKI and predicted their interaction with FGL2. Both in vivo and in vitro studies showed that overexpression of FGL2 markedly increased complement C3a (C3a) levels and exacerbated inflammation and injury. In contrast, knockdown of FGL2 resulted in a marked decline in C3a levels, which not only conferred a substantial protective effect against hypoxia/reoxygenation-induced injury in tubular cells but also effectively alleviated kidney injury in mice. At the molecular level, FGL2 interacts with complement C3, leading to elevated C3a production. This stimulates the inflammatory response of renal tubular epithelial cells, thereby inducing tubular damage. Targeting FGL2 may hold potential prevention and treatment strategy for tubular injury in AKI.

Abstract Introduction Chronic insomnia (CI) is characterized by hyperarousal and abnormal large-scale brain dynamics, yet the energetic demands underlying transitions between brain states remain poorly understood. Network control theory provides a mechanistic framework for quantifying the minimum control energy required for the brain to maintain or transition between functional configurations. This study aimed to characterize the energy landscape of brain states in patients with CI and to examine its associations with symptom severity and neurotransmitter receptor architecture. Methods Forty-two adults with CI and forty-four matched healthy controls underwent diffusion tensor imaging, T1-weighted imaging, and resting-state functional MRI. Hidden Markov modeling identified recurrent functional states and quantified fractional occupancy and mean dwell time. Individual structural connectivity matrices served as system matrices for estimating control energy for state maintenance and transitions. Nineteen neurotransmitter receptor and transporter maps were incorporated to identify neurochemical contributors to altered energy patterns. Associations with insomnia severity and sleep quality were assessed. Results Two robust brain states were identified. State 1 was characterized by high activity in the default mode and frontoparietal control network, accompanied by reduced activation in the dorsal attention and somatomotor networks. State 2 was characterized by opposing patterns. CI participants showed significantly higher fractional occupancy (Z = 2.84, P = 0.006) and longer mean lifetime in State 1, alongside significantly reduced occupancy in State 2 (Z = 3.72, P < 0.001). The longer mean lifetime in State 1 was associated with lower habitual sleep efficiency (R2 = 0.10, P = 0.040). The transition energy from State 2 to State 1 was positively associated with sleep latency (R2 = 0.14, P = 0.024) and insomnia severity (R2 = 0.13, P = 0.028). Neurotransmitter analyses revealed strong contributions from GABAergic, dopaminergic, cholinergic, glutamatergic, serotonergic, and opioid systems to altered transition energies. Conclusion CI is characterized by a flattened brain-state energy landscape, reduced dynamical stability, and neurotransmitter-mediated abnormalities in the energy required for transitioning between functional states. These findings provide mechanistic evidence linking impaired brain-state controllability with clinical symptoms and highlight neurotransmitter systems that may shape neural dynamics in insomnia. Support (if any) This study was supported by the CYRUS TANG FUNDATION.

Glomerular podocytes injury represents a critical pathological hallmark of diabetic kidney disease (DKD), in which lipotoxicity plays a central pathogenic role. Our previous investigations in type 2 diabetes mellitus (T2DM) have demonstrated that Jujuboside A (Ju A), a triterpene saponin isolated from Semen Ziziphi Spinosae (SZS), exerted dual therapeutic effects in T2DM by ameliorating hepatic steatosis and renal dysfunction. However, the role of podocytes lipid metabolism in Ju A-mediated protection against DKD remain undefined prior to the present study. In this work, we reported that Ju A significantly attenuated glomerular podocytes injury and lipotoxicity in DKD, while concurrently improving renal function and preserving glomerular morphology. Mechanistically, Yin Yang 1 (YY1)-mediated alleviation of lipotoxicity contributed to the protective effect of Ju A against glomerular podocytes injury, primarily by promoting intracellular cholesterol transport and efflux. In conclusion, our findings demonstrated that Ju A mitigated lipid overload in glomerular podocytes by modulating cholesterol homeostasis via YY1, which not only intercepted the pathological progression of DKD but also provided a potential therapeutic target (YY1) and candidate agent (Ju A) for DKD intervention.

INTRODUCTION:Liver fibrosis is a common pathological process in chronic liver disease, reflecting the advanced stage of the disease. Liver endothelial cells (ECs), especially liver sinusoidal endothelial cells (LSECs), are recognized as critical modulators of liver homeostasis and play essential roles in the recruitment and function of liver immune cells. In this study, we aimed to explore the mechanism of hepatic EC injury and the potential regulatory pathways of intercellular communication in liver fibrosis. METHODS:In this study, C57BL/6 male mice were treated with CCl4 for 6 weeks to establish a liver fibrosis model. Masson staining and immunohistochemistry were performed to assess the extent of liver fibrosis. Hepatic endothelial injury was detected by using scanning electron microscopy (SEM) and PCR technology. Single-cell RNA sequencing (scRNA-seq) was performed to analyze phenotypic changes in nonparenchymal cells and dissect intercellular crosstalk. RESULTS:A total of 24,534 cells were clustered into 10 main cell subsets. The LSEC fenestrae and surface receptor expression were reduced, and the expression of Cd34 was upregulated. Liver ECs exhibited dense cellular crosstalk with immune cells (macrophages, T and B cells). The analysis of intercellular signaling pathways revealed that immune cells targeted liver ECs through the Ptprc-Mrc1 and Sell-Podxl signaling pathways to maintain cellular interactions during liver fibrosis. CONCLUSION:We revealed apparent damage and capillarization of liver ECs and demonstrated the cell-cell communications among liver immune cells and ECs during the development of liver fibrosis. The Ptprc-Mrc1 and Sell-Podxl signaling pathways exerted prominent roles in liver immune cell-EC interactions.

Mesangial cell proliferation is an early pathological indicator of diabetic nephropathy (DN). Growing evidence highlights the pivotal role of paired-related homeobox 1 (Prrx1), a key regulator of cellular proliferation and tissue differentiation, in various disease pathogenesis. Notably, Prrx1 is highly expressed in mesangial cells under DN conditions. Both in vitro and in vivo studies have demonstrated that Prrx1 overexpression promotes mesangial cell proliferation and contributes to renal fibrosis in db/m mice. Conversely, Prrx1 knockdown markedly suppresses hyperglycemia-induced mesangial cell proliferation and mitigates renal fibrosis in db/db mice. Mechanistically, Prrx1 directly interacts with the Yes-associated protein 1 (YAP) promoter, leading to the upregulation of YAP expression. This upregulation promotes mesangial cell proliferation and exacerbates renal fibrosis. These findings emphasize the crucial role of Prrx1 upregulation in high glucose-induced mesangial cell proliferation, ultimately leading to renal fibrosis in DN. Therefore, targeting Prrx1 to downregulate its expression presents a promising therapeutic strategy for treating renal fibrosis associated with DN.

Pragmatics plays a crucial role in effectively conveying messages across various social communication contexts. This aspect is frequently highlighted in the challenges experienced by children diagnosed with autism spectrum disorder (ASD). Notably, there remains a paucity of research investigating how the structural connectome (SC) predicts pragmatic language abilities within this population. Using diffusion tensor imaging (DTI) and deterministic tractography, we constructed the whole-brain white matter structural network (WMSN) in a cohort comprising 92 children with ASD and 52 typically developing (TD) preschoolers, matched for age and gender. We employed network-based statistic (NBS)-Predict, a novel methodology that integrates machine learning (ML) with NBS, to identify dysconnected subnetworks associated with ASD, and then to predict pragmatic language abilities based on the SC derived from the whole-brain WMSN in the ASD group. Initially, NBS-Predict identified a subnetwork characterized by 42 reduced connections across 37 brain regions (p = 0.01), achieving a highest classification accuracy of 79.4

Obeticholic acid (OCA) was approved for the treatment of primary biliary cholangitis (PBC) patients. However, it can cause severe drug-induced liver injury (DILI), which may put PBC patients at risk of acute-on-chronic liver failure (ACLF) and even death. Farnesoid X receptor (FXR) is considered as the target of OCA for cholestasis, but there is still a lack of research on whether hepatic and ileal FXR have different effects after OCA treatment. The aim of this study was to investigate the mechanism of OCA aggravating liver fibrosis in cholestasis. The results showed that 40 mg/kg OCA elevated serum AST, ALT, ALP and γ-GT levels in bile duct ligation (BDL) mice. Besides, severe fibrosis and necrosis were observed in the OCA-treated BDL mice, which was related to hepatic apoptosis pathway activation. Both hepatic and ileal FXR signaling could be significantly activated by OCA. However, ileum-specific knockout of Fxr aggravated OCA-induced liver injury in BDL mice. On the contrary, hepatic-specific knockout of Fxr structurally and functionally ameliorated liver pathological processes in the OCA-treated BDL mice, which was due to the blockade of hepatic FXR-induced apoptosis. In conclusion, the mechanism of OCA aggravating liver fibrosis in cholestasis was based on the activation of hepatic FXR-induced apoptosis. It was also indicated ileal FXR might be a safer pharmacological target for bile acids regulation.

Metabolic dysfunction-associated steatotic liver disease (MASLD) is the most prevalent chronic liver disease worldwide. Fatty-acid metabolism disorders, especially long-chain fatty acids (LCFA) accumulation, is the main pathological feature of high fat diet-induced MASLD. Fenofibrate is mainly used for the treatment of hyperlipidemia and metabolic disorders in clinical settings. In recent years, its therapeutic effect on MASLD has also been reported, but the mechanism is still unclear. Here, we aimed to investigate the effect and mechanism of fenofibrate on hepatic steatosis via fatty-acid metabolism regulation. It was found that fenofibrate strongly reduced hepatic LCFA accumulation, especially decreased the content of erucic acid (EA). In AML-12 cells treated with EA, fenofibrate improved hepatic lipid accumulation by accelerating EA metabolism. In vivo and in vitro experiments have proven that peroxidase enoyl-CoA hydratase and 3-hydroxyacyl CoA dehydrogenase is the key enzyme of fenofibrate in promoting LCFA metabolism. This study confirmed that fenofibrate upregulated peroxisome enzyme enoyl-CoA hydratase and 3-hydroxyacyl CoA dehydrogenase expression to promote LCFA oxidation, which provided a novel strategy for the treatment of high-fat diet-induced steatotic liver disease in clinical settings. SIGNIFICANCE STATEMENT: We found that long-chain fatty acid overload was a characteristic of high-fat diet-induced fatty liver, and fenofibrate ameliorated high-fat diet-induced fatty liver by upregulating enoyl-CoA hydratase and 3-hydroxyacyl CoA dehydrogenase to promote the oxidation of long-chain fatty acids, especially erucic acid. This study may contribute to the use of fenofibrate in the treatment of fatty liver disease.

BACKGROUND:Disruptions in synaptic plasticity and alterations in effective connectivity (EC) involving the hippocampus and amygdala are hallmarks of early Alzheimer's disease (AD). However, the interplay between these neurophysiological changes and their relationships with cognitive functions in subjective cognitive decline (SCD) and mild cognitive impairment (MCI) remains poorly understood. METHODS:Transcranial magnetic stimulation (TMS) and resting-state functional magnetic resonance imaging (rs-fMRI) were used to assess long-term potentiation (LTP)-like plasticity and EC involving the amygdala and hippocampus in 34 individuals with SCD, 27 with MCI, and 35 healthy controls (HC). Between-group differences in cognitive performance, EC alterations, and LTP-like plasticity were examined and their relationships were assessed via correlation and mediation analyses. RESULTS:Both SCD and MCI groups exhibited disrupted EC between the amygdala/hippocampus and the inferior occipital gyrus (IOG), inferior parietal lobule (IPL), medial frontal lobe (MFL), and precuneus. Also, both LTP-5min and LTP-10min were significantly reduced in MCI group compared to SCD and HC groups. Importantly, EC from the left hippocampus to the IPL and from the IPL, MFL, and precuneus to the hippocampus was correlated with memory and executive functions. Moreover, precuneus-to-hippocampus EC was positively correlated with LTP-10min and mediated the relationship between LTP-like plasticity and cognitive performance. CONCLUSIONS:This study provides novel evidence that precuneus-to-hippocampus EC mediates the link between synaptic plasticity and cognitive function in SCD and MCI, suggesting the precuneus-hippocampus pathway as a promising target for early diagnosis and intervention.

Pyroptosis, a lytic inflammatory cell death process, plays a critical role in diabetic nephropathy (DN) progression. Our study identifies Hematopoietic Prostaglandin D Synthase (HPGDS) as a key regulatory factor in this process. Analysis demonstrated that HPGDS expression is positively correlated with pyroptosis-related inflammatory injury in DN patients and db/db mice. In diabetic mice and high glucose-stimulated HK-2 cells, HPGDS drives DN advancement through GSDMD-mediated pyroptosis, leading to renal interstitial inflammation. HPGDS overexpression increased pyroptosis in HK-2 cells and aggravated renal interstitial inflammation. Whereas, HPGDS knockdown experiments yielded opposite results. Mechanistically, we confirmed that HPGDS triggers NLRP3/Caspase1 activation, promoting pyroptosis mediated renal tubular injury and interstitial inflammation. These findings elucidate HPGDS-mediated pyroptosis as a therapeutic strategy for DN.

Over 30% of patients with type 2 diabetes develop diabetic kidney disease (DKD), which has emerged as a major contributor to end stage renal disease. Renal fibrosis represents the final pathological outcome of most chronic kidney disease, particularly DKD. This study demonstrates elevated levels of Galectin-3 (Gal3), a lectin associated with inflammatory and fibrotic conditions, in the plasma and kidneys of DKD mice. Positive correlations between Gal3 expression and renal fibrosis are observed in both DKD patients and mice. Macrophage-derived Gal3 is found to promote Transforming growth factor beta 1 (TGFβ1) signaling activation and renal fibrogenesis. Genetic ablation of Gal3 globally or specifically in macrophages, as well as pharmacological inhibition of Gal3, significantly attenuated kidney fibrosis in diabetic mice. Mechanistically, macrophage-derived Gal3 interacted with TGFβ receptor2 (TGFBR2) and Pro-TGFβ1, preventing TGFBR2 proteasomal degradation in fibroblasts and increasing TGFβ1 levels in the diabetic kidney. These events enhances TGFβ1 signaling activation and ultimately facilitated kidney fibrosis. The findings of this study suggest Gal3 as a potential therapeutic target for renal fibrosis and DKD.

The role of genetic susceptibility in early warning and precise treatment of diabetic kidney disease (DKD) requires further investigation. A case-control study was conducted to evaluate the predictive effect of GSK3B genetic polymorphisms on the susceptibility to DKD, with the aim of providing a theoretical basis and laboratory rationale for the prediction of the risk of developing DKD in patients with type 2 diabetes mellitus (T2DM). The GSK3B genotyping was performed by SNaPshot method based on Genotype-Tissue Expression database and thousand genomes database to screen tag SNPs. The polymorphisms of GSK3B tag SNPs were statistically analyzed for their effects on DKD susceptibility and clinical indicators. Urinary exosomes from DKD patients were extracted, protein expression levels of GSK3β were detected by ELISA kits, and kinase activity of GSK3β was quantified by kinase activity spectrometry to evaluate the correlation between the gene polymorphisms of GSK3B and the expression levels and activities of GSK3β. A machine learning model was constructed for assessing the efficacy of GSK3B polymorphisms in predicting the risk of developing DKD in patients with T2DM. A total of 800 subjects who met the inclusion and exclusion criteria were included in the case-control study, including 200 healthy control subjects, 300 patients with T2DM and 300 patients with DKD. Genetic analysis identified five tag SNPs (rs60393216, rs3732361, rs2199503, rs1488766, and rs59669360) associated with the susceptibility to DKD. The protein level and activity of GSK3β were significantly elevated in DKD patients. On the other hand, the expression levels and kinase activity of GSK3β in exosomes differed significantly between patients with different genotypes of the GSK3B, suggesting that the effect of GSK3B gene polymorphisms on GSK3β expression and activity may be an important mechanism leading to individual differences in susceptibility to DKD. XG Boost algorithm model identified rs60393216 and rs1488766 as important biomarkers for clinical early warning of DKD.

Mesangial cells (MCs) are the most active intrinsic cells in the glomerulus. MCs excessively proliferate at the early stage of diabetic kidney disease (DKD), eventually causing glomerular sclerosis and even renal failure; inhibiting glomerular MC proliferation in early DKD is a promising prevention and treatment strategy for early DKD. Our previous study shows that Yin Yang 1 (YY1), a zinc finger protein, is a novel regulator of DKD-induced renal fibrosis. In this study we investigated the role of YY1 in glomerular MC proliferation in DKD in vivo and in vitro. We first showed that YY1 expression levels were significantly increased in the glomerular MCs of DKD patients and db/db mice and in high glucose (HG)-treated SV40-MES13 cells. By using YY1 expression/knockdown plasmids, we confirmed that YY1 contributed to glomerular MC proliferation in vitro. We demonstrated that YY1 upregulated hypoxia-inducible factor-1 alpha (HIF-1α) expression and activity in HG-treated SV40-MES13 cells, leading to overproduction of mROS. Moreover, mROS contributed to positive feedback regulation of YY1/HIF-1α signaling, and the YY1/HIF-1α/mROS positive feedback loop exacerbated glomerular MC proliferation in HG-treated SV40-MES13 cells. In addition, renal-specific YY1 overexpression promoted glomerular MC proliferation in normal mice, whereas renal-specific YY1 knockdown mitigated MC proliferation in early diabetic mice by inactivating HIF-1α/ROS signaling. In conclusion, the YY1/HIF-1α/mROS positive feedback loop might be an attractive therapeutic target for overcoming glomerulosclerosis in early DKD.