Chlorogenic acid alleviates renal fibrosis by reducing lipid accumulation in diabetic kidney disease through suppressing the Notch1 and Stat3 signaling pathway.

AIMS: Abnormal renal lipid metabolism causes renal lipid deposition, which leads to the development of renal fibrosis in diabetic kidney disease (DKD). The aim of this study was to investigate the effect and mechanism of chlorogenic acid (CA) on reducing renal lipid accumulation and improving DKD renal fibrosis. METHODS: This study evaluated the effects of CA on renal fibrosis, lipid deposition and lipid metabolism by constructing in vitro and in vivo models of DKD, and detected the improvement of Notch1 and Stat3 signaling pathways. Molecular docking was used to predict the binding between CA and the extracellular domain NRR1 of Notch1 protein. RESULTS: In vitro studies have shown that CA decreased the expression of Fibronectin, α-smooth muscle actin (α-SMA), p-smad3/smad3, alleviated lipid deposition, promoted the expression of carnitine palmitoyl transferase 1 A (CPT1A), and inhibited the expression of cholesterol regulatory element binding protein 1c (SREBP1c). The expression of Notch1, Cleaved Notch1, Hes1, and p-stat3/stat3 were inhibited. These results suggested that CA might reduce intercellular lipid deposition in human kidney cells (HK2) by inhibiting Notch1 and stat3 signaling pathways, thereby improving fibrosis. Further, in vivo studies demonstrated that CA improved renal fibrosis and renal lipid deposition in DKD mice by inhibiting Notch1 and stat3 signaling pathways. Finally, molecular docking experiments showed that the binding energy of CA and NRR1 was -6.6 kcal/mol, which preliminarily predicted the possible action of CA on Notch1 extracellular domain NRR1. CONCLUSION: CA reduces renal lipid accumulation and improves DKD renal fibrosis by inhibiting Notch1 and stat3 signaling pathways.

Tonabersat suppresses priming/activation of the NOD-like receptor protein-3 (NLRP3) inflammasome and decreases renal tubular epithelial-to-macrophage crosstalk in a model of diabetic kidney disease.

BACKGROUND: Accompanied by activation of the NOD-like receptor protein 3 (NLRP3) inflammasome, aberrant connexin 43 (Cx43) hemichannel-mediated ATP release is situated upstream of inflammasome assembly and inflammation and contributes to multiple secondary complications of diabetes and associated cardiometabolic comorbidities. Evidence suggests there may be a link between Cx43 hemichannel activity and inflammation in the diabetic kidney. The consequences of blocking tubular Cx43 hemichannel-mediated ATP release in priming/activation of the NLRP3 inflammasome in a model of diabetic kidney disease (DKD) was investigated. We examined downstream markers of inflammation and the proinflammatory and chemoattractant role of the tubular secretome on macrophage recruitment and activation. METHODS: Analysis of human transcriptomic data from the Nephroseq repository correlated gene expression to renal function in DKD. Primary human renal proximal tubule epithelial cells (RPTECs) and monocyte-derived macrophages (MDMs) were cultured in high glucose and inflammatory cytokines as a model of DKD to assess Cx43 hemichannel activity, NLRP3 inflammasome activation and epithelial-to-macrophage paracrine-mediated crosstalk. Tonabersat assessed a role for Cx43 hemichannels. RESULTS: Transcriptomic analysis from renal biopsies of patients with DKD showed that increased Cx43 and NLRP3 expression correlated with declining glomerular filtration rate (GFR) and increased proteinuria. In vitro, Tonabersat blocked glucose/cytokine-dependant increases in Cx43 hemichannel-mediated ATP release and reduced expression of inflammatory markers and NLRP3 inflammasome activation in RPTECs. We observed a reciprocal relationship in which NLRP3 activity exacerbated increased Cx43 expression and hemichannel-mediated ATP release, events driven by nuclear factor kappa-B (NFκB)-mediated priming and Cx43 hemichannel opening, changes blocked by Tonabersat. Conditioned media (CM) from RPTECs treated with high glucose/cytokines increased expression of inflammatory markers in MDMs, an effect reduced when macrophages were pre-treated with Tonabersat. Co-culture using conditioned media from Tonabersat-treated RPTECs dampened macrophage inflammatory marker expression and reduced macrophage migration. CONCLUSION: Using a model of DKD, we report for the first time that high glucose and inflammatory cytokines trigger aberrant Cx43 hemichannel activity, events that instigate NLRP3-induced inflammation in RPTECs and epithelial-to-macrophage crosstalk. Recapitulating observations previously reported in diabetic retinopathy, these data suggest that Cx43 hemichannel blockers (i.e., Tonabersat) may dampen multi-system damage observed in secondary complications of diabetes.

Podocyte OTUD5 alleviates diabetic kidney disease through deubiquitinating TAK1 and reducing podocyte inflammation and injury.

Recent studies have shown the crucial role of podocyte injury in the development of diabetic kidney disease (DKD). Deubiquitinating modification of proteins is widely involved in the occurrence and development of diseases. Here, we explore the role and regulating mechanism of a deubiquitinating enzyme, OTUD5, in podocyte injury and DKD. RNA-seq analysis indicates a significantly decreased expression of OTUD5 in HG/PA-stimulated podocytes. Podocyte-specific Otud5 knockout exacerbates podocyte injury and DKD in both type 1 and type 2 diabetic mice. Furthermore, AVV9-mediated OTUD5 overexpression in podocytes shows a therapeutic effect against DKD. Mass spectrometry and co-immunoprecipitation experiments reveal an inflammation-regulating protein, TAK1, as the substrate of OTUD5 in podocytes. Mechanistically, OTUD5 deubiquitinates K63-linked TAK1 at the K158 site through its active site C224, which subsequently prevents the phosphorylation of TAK1 and reduces downstream inflammatory responses in podocytes. Our findings show an OTUD5-TAK1 axis in podocyte inflammation and injury and highlight the potential of OTUD5 as a promising therapeutic target for DKD.

Roles of Mitochondrial Dysfunction in Diabetic Kidney Disease: New Perspectives from Mechanism to Therapy.

Diabetic kidney disease (DKD) is a common microvascular complication of diabetes and the main cause of end-stage renal disease around the world. Mitochondria are the main organelles responsible for producing energy in cells and are closely involved in maintaining normal organ function. Studies have found that a high-sugar environment can damage glomeruli and tubules and trigger mitochondrial dysfunction. Meanwhile, animal experiments have shown that DKD symptoms are alleviated when mitochondrial damage is targeted, suggesting that mitochondrial dysfunction is inextricably linked to the development of DKD. This article describes the mechanisms of mitochondrial dysfunction and the progression and onset of DKD. The relationship between DKD and mitochondrial dysfunction is discussed. At the same time, the progress of DKD treatment targeting mitochondrial dysfunction is summarized. We hope to provide new insights into the progress and treatment of DKD.

Nmnat1 Deficiency Causes Mitoribosome Excess in Diabetic Nephropathy Mediated by Transcriptional Repressor HIC1.

Nicotinamide adenine dinucleotide (NAD) is involved in renal physiology and is synthesized by nicotinamide mononucleotide adenylyltransferase (NMNAT). NMNAT exists as three isoforms, namely, NMNAT1, NMNAT2, and NMNAT3, encoded by Nmnat1, Nmnat2, and Nmnat3, respectively. In diabetic nephropathy (DN), NAD levels decrease, aggravating renal fibrosis. Conversely, sodium-glucose cotransporter-2 inhibitors increase NAD levels, mitigating renal fibrosis. In this regard, renal NAD synthesis has recently gained attention. However, the renal role of Nmnat in DN remains uncertain. Therefore, we investigated the role of Nmnat by establishing genetically engineered mice. Among the three isoforms, NMNAT1 levels were markedly reduced in the proximal tubules (PTs) of db/db mice. We examined the phenotypic changes in PT-specific Nmnat1 conditional knockout (CKO) mice. In CKO mice, Nmnat1 expression in PTs was downregulated when the tubules exhibited albuminuria, peritubular type IV collagen deposition, and mitochondrial ribosome (mitoribosome) excess. In CKO mice, Nmnat1 deficiency-induced mitoribosome excess hindered mitoribosomal translation of mitochondrial inner membrane-associated oxidative phosphorylation complex I (CI), CIII, CIV, and CV proteins and mitoribosomal dysfunction. Furthermore, the expression of hypermethylated in cancer 1, a transcription repressor, was downregulated in CKO mice, causing mitoribosome excess. Nmnat1 overexpression preserved mitoribosomal function, suggesting its protective role in DN.

LncRNA KIFAP3-5:1 inhibits epithelial-mesenchymal transition of renal tubular cell through PRRX1 in diabetic nephropathy.

Long noncoding RNAs play an important role in several pathogenic processes in diabetic nephropathy, but the relationship with epithelial-mesenchymal transition in DN is unclear. Herein, we found that KIFAP3-5:1 expression was significantly down-regulated in DN plasma samples, db/db mouse kidney tissues and high glucose treated renal tubular epithelial cells compared to normal healthy samples and untreated cells. Overexpression of KIFAP3-5:1 improved renal fibrosis in db/db mice and rescued epithelial-mesenchymal transition of high glucose cultured renal tubular epithelial cells. The silence of KIFAP3-5:1 will exacerbate the progression of EMT. Mechanistically, KIFAP3-5:1 was confirmed to directly target to the -488 to -609 element of the PRRX1 promoter and negatively modulate PRRX1 mRNA and protein expressions. Furthermore, rescue assays demonstrated that the knockdown of PRRX1 counteracted the KIFAP3-5:1 low expression-mediated effects on EMT in hRPTECs cultured under high glucose. The plasma KIFAP3-5:1 of DN patients is highly correlated with the severity of renal dysfunction and plays an important role in the prediction model of DN diseases. These findings suggested that KIFAP3-5:1 plays a critical role in regulation of renal EMT and fibrosis through suppress PRRX1, and highlight the clinical potential of KIFAP3-5:1 to assist in the diagnosis of diabetic nephropathy.

High glucose-induced downregulation of PTEN-Long is sufficient for proximal tubular cell injury in diabetic kidney disease.

During the progression of diabetic kidney disease, proximal tubular epithelial cells respond to high glucose to induce hypertrophy and matrix expansion leading to renal fibrosis. Recently, a non-canonical PTEN has been shown to be translated from an upstream initiation codon CUG (leucine) to produce a longer protein called PTEN-Long (PTEN-L). Interestingly, the extended sequence present in PTEN-L contains cell secretion/penetration signal. Role of this non-canonical PTEN-L in diabetic renal tubular injury is not known. We show that high glucose decreases expression of PTEN-L. As a mechanism of its function, we find that reduced PTEN-L activates Akt-2, which phosphorylates and inactivate tuberin and PRAS40, resulting in activation of mTORC1 in tubular cells. Antibacterial agent acriflavine and antiviral agent ATA regulate translation from CUG codon. Acriflavine and ATA, respectively, decreased and increased expression of PTEN-L to altering Akt-2 and mTORC1 activation in the absence of change in expression of canonical PTEN. Consequently, acriflavine and ATA modulated high glucose-induced tubular cell hypertrophy and lamininγ1 expression. Importantly, expression of PTEN-L inhibited high glucose-stimulated Akt/mTORC1 activity to abrogate these processes. Since PTEN-L contains secretion/penetration signals, addition of conditioned medium containing PTEN-L blocked Akt-2/mTORC1 activity. Notably, in renal cortex of diabetic mice, we found reduced PTEN-L concomitant with Akt-2/mTORC1 activation, leading to renal hypertrophy and lamininγ1 expression. These results present first evidence for involvement of PTEN-L in diabetic kidney disease.

Ginsenoside Rg3 induces mesangial cells proliferation and attenuates apoptosis by miR-216a-5p/MAPK pathway in diabetic kidney disease.

BACKGROUND: Ginsenoside Rg3 is an active saponin isolated from ginseng, which can reduce renal inflammation. However, the role and mechanism of Rg3 in diabetic kidney disease (DKD) are far from being studied. METHODS: The effects of Rg3 and miR-216a-5p on the proliferation, apoptosis, and MAPK pathway in high glucose (HG)-induced SV40 MES 13 were monitored by CCK-8, TUNEL staining, and western blot. RESULTS: Rg3 treatment could accelerate proliferation and suppress apoptosis in HG-induced SV40 MES. Moreover, miR-216a-5p inhibition also could alleviate renal injury, prevent apoptosis, and activate the MAPK pathway in kidney tissues of diabetic model mice. CONCLUSION: Rg3 could attenuate DKD progression by downregulating miR-216a-5p, suggesting Rg3 and miR-216a-5p might be the potential drug and molecular targets for DKD therapy.

MiR-33a Overexpression Exacerbates Diabetic Nephropathy Through Sirt6-dependent Notch Signaling.

INTRODUCTION: Diabetic nephropathy (DN) belongs to the major cause of end-stage kidney disease. We probed the functions of a microRNA miR-33a in inducing podocytes injury during childhood  DN (CDN). METHODS: Kidney samples were collected from 20 children with DN. Matrix deposition and glomerular basement membranes thickness were examined by periodic acid-Schiff staining. Immunofluorescence staining was performed to assess kidney function-related proteins. MicroRNA (MiR)-33a mimic together with miR-33a inhibitor was transfected into podocytes for determining the roles of miR-33a. Glomerular podocyte apoptosis was determined by terminal deoxynucleotidyl transferase (TdT) dUTP Nick-End Labeling (TUNEL) staining along with flow cytometry. RESULTS: Down-regulation of Nephrin and Podocin and increased podocyte apoptosis rate were observed in the glomerulus of CDN as well as podocytes treated with high glucose. MiR-33a was up regulated in the glomeruli and glucose-treated podocytes. Injury in podocytes was aggravated with miR-33a elevation but alleviated with miR-33a inhibition. Moreover, the expression of Sirtuin 6 (Sirt6) was decreased while the levels of notch receptor 1 (Notch1) and notch receptor 4 (Notch4) were elevated in the glomerulus and glucose-treated podocytes. Decreased level of Sirt6 upon glucose treatment was abrogated by miR-33a inhibition, and the podocytes injury induced by glucose exposure was relieved by Sirt6 via Notch signaling. CONCLUSION: These findings indicated that miR-33a promoted podocyte injury via targeting Sirt6-dependent Notch signaling in CDN, which might provide a novel sight for CDN treatment. DOI: 10.52547/ijkd.7904.

Screening of potential drugs for the treatment of diabetic kidney disease using single-cell transcriptome sequencing and connectivity map data.

OBJECTIVE: To explore the feasibility of screening potential drugs for the treatment of diabetic kidney disease (DKD) using a single-cell transcriptome sequencing dataset and Connectivity Map (CMap) database screening. METHODS: A DKD single-nucleus transcriptome sequencing dataset was analyzed using Seurat 4.0 to obtain specific podocyte subclusters and differentially expressed genes (DEGs) related to DKD. These DEGs were subsequently subjected to a search against the CMap database to screen for drug candidates. Cell and animal experiments were conducted to evaluate the efficacy of the top 3 drug candidates. RESULTS: Initially, we analyzed the DKD single-nucleus transcriptome sequencing dataset to obtain intrinsic renal cells such as podocytes, endothelial cells, mesangial cells, proximal tubular cells, collecting duct cells and immune cells. Podocytes were further divided into four subclusters, among which the proportion of POD_1 podcytes was significantly greater in DKD kidneys than in control kidneys (34.0 % vs. 3.4 %). The CMap database was searched using the identified DEGs in the POD_1 subcluster, and the drugs, including tozasertib, paroxetine, and xylazine, were obtained. Cell-based experiments showed that tozasertib, paroxetine and xylazine had no significant podocyte toxicity in the concentration range of 0.01-50 µM. Tozasertib, paroxetine, and xylazine all reversed the advanced glycation end products (AGEs)-induced decrease in podocyte marker levels, but the effect of paroxetine was more prominent. Animal experiments showed that paroxetine decreased urine ALB/Cr levels in DKD model mice by approximately 51.5 % (115.7 mg/g vs. 238.8 mg/g, P < 0.05). Histopathological assessment revealed that paroxetine attenuated basement membrane thickening, restored the number of foot processes of podocytes, and reduced foot process fusion. In addition, paroxetine also attenuated renal tubular-interstitial fibrosis. Mechanistically, paroxetine inhibited the expression of GRK2 and NLRP3, decreased the phosphorylation level of p65, restored NRF2 expression, and relieved inflammation and oxidative stress. CONCLUSION: This strategy based on single-cell transcriptome sequencing and CMap data can facilitate the identification and aid the rapid development of clinical DKD drugs. Paroxetine, screened by this strategy, has excellent renoprotective effects.

Unraveling the nexus of NAD+ metabolism and diabetic kidney disease: insights from murine models and human data.

Background: Nicotinamide adenine dinucleotide (NAD+) is a critical coenzyme involved in kidney disease, yet its regulation in diabetic kidney disease (DKD) remains inadequately understood. Objective: Therefore, we investigated the changes of NAD+ levels in DKD and the underlying mechanism. Methods: Alternations of NAD+ levels and its biosynthesis enzymes were detected in kidneys from streptozotocin-induced diabetic mouse model by real-time PCR and immunoblot. The distribution of NAD+ de novo synthetic enzymes was explored via immunohistochemical study. NAD+ de novo synthetic metabolite was measured by LC-MS. Human data from NephroSeq were analyzed to verify our findings. Results: The study showed that NAD+ levels were decreased in diabetic kidneys. Both mRNA and protein levels of kynurenine 3-monooxygenase (KMO) in NAD+ de novo synthesis pathway were decreased, while NAD+ synthetic enzymes in salvage pathway and NAD+ consuming enzymes remained unchanged. Further analysis of human data suggested KMO, primarily expressed in the proximal tubules shown by our immunohistochemical staining, was consistently downregulated in human diabetic kidneys. Conclusion: Our study demonstrated KMO of NAD+ de novo synthesis pathway was decreased in diabetic kidney and might be responsible for NAD+ reduction in diabetic kidneys, offering valuable insights into complex regulatory mechanisms of NAD+ in DKD.

Marrow mesenchymal stem cell mediates diabetic nephropathy progression via modulation of Smad2/3/WTAP/m6A/ENO1 axis.

Diabetic nephropathy (DN) is one of the common microvascular complications in diabetic patients. Marrow mesenchymal stem cells (MSCs) have attracted attention in DN therapy but the underlying mechanism remains unclear. Here, we show that MSC administration alleviates high glucose (HG)-induced human kidney tubular epithelial cell (HK-2 cell) injury and ameliorates renal injury in DN mice. We identify that Smad2/3 is responsible for MSCs-regulated DN progression. The activity of Smad2/3 was predominantly upregulated in HG-induced HK-2 cell and DN mice and suppressed with MSC administration. Activation of Smad2/3 via transforming growth factor-ß1 (TGF-ß1) administration abrogates the protective effect of MSCs on HG-induced HK-2 cell injury and renal injury of DN mice. Smad2/3 has been reported to interact with methyltransferase of N6-methyladenosine (m6A) complex and we found a methyltransferase, Wilms' tumor 1-associating protein (WTAP), is involved in MSCs-Smad2/3-regulated DN development. Moreover, WTAP overexpression abrogates the improvement of MSCs on HG-induced HK-2 cell injury and renal injury of DN mice. Subsequently, α-enolase (ENO1) is the downstream target of WTAP-mediated m6A modification and contributes to the MSCs-mediated regulation. Collectively, these findings reveal a molecular mechanism in DN progression and indicate that Smad2/3/WTAP/ENO1 may present a target for MSCs-mediated DN therapy.

Deciphering the molecular nexus of BTG2 in periodontitis and diabetic kidney disease.

OBJECTIVE: To investigate the role of BTG2 in periodontitis and diabetic kidney disease (DKD) and its potential underlying mechanism. METHODS: Gene expression data for periodontitis and DKD were acquired from the Gene Expression Omnibus (GEO) database. Differential expression analysis identified co-expressed genes between these conditions. The Nephroseq V5 online nephropathy database validated the role of these genes in DKD. Pearson correlation analysis identified genes associated with our target gene. We employed Gene Set Enrichment Analysis (GSEA) and Protein-Protein Interaction (PPI) networks to elucidate potential mechanisms. Expression levels of BTG2 mRNA were examined using quantitative polymerase Chain Reaction (qPCR) and immunofluorescence assays. Western blotting quantified proteins involved in epithelial-to-mesenchymal transition (EMT), apoptosis, mTORC1 signaling, and autophagy. Additionally, wound healing and flow cytometric apoptosis assays evaluated podocyte migration and apoptosis, respectively. RESULTS: Analysis of GEO database data revealed BTG2 as a commonly differentially expressed gene in both DKD and periodontitis. BTG2 expression was reduced in DKD compared to normal conditions and correlated with proteinuria. GSEA indicated enrichment of BTG2 in the EMT and mTORC1 signaling pathways. The PPI network highlighted BTG2's relevance to S100A9, S100A12, and FPR1. Immunofluorescence assays demonstrated significantly lower BTG2 expression in podocytes under high glucose (HG) conditions. Reduced BTG2 expression in HG-treated podocytes led to increased levels of EMT markers (α-SMA, vimentin) and the apoptotic protein Bim, alongside a decrease in nephrin. Lower BTG2 levels were associated with increased podocyte mobility and apoptosis, as well as elevated RPS6KB1 and mTOR levels, but reduced autophagy marker LC3. CONCLUSION: Our findings suggest that BTG2 is a crucial intermediary gene linking DKD and periodontitis. Modulating autophagy via inhibition of the mTORC1 signaling pathway, and consequently suppressing EMT, may be pivotal in the interplay between periodontitis and DKD.

Molecular mechanism of renal lipid accumulation in diabetic kidney disease.

Diabetic kidney disease (DKD) is a leading cause of end stage renal disease with unmet clinical demands for treatment. Lipids are essential for cell survival; however, renal cells have limited capability to metabolize overloaded lipids. Dyslipidaemia is common in DKD patients and renal ectopic lipid accumulation is associated with disease progression. Unveiling the molecular mechanism involved in renal lipid regulation is crucial for exploring potential therapeutic targets. In this review, we focused on the mechanism underlying cholesterol, oxysterol and fatty acid metabolism disorder in the context of DKD. Specific regulators of lipid accumulation in different kidney compartment and TREM2 macrophages, a lipid-related macrophages in DKD, were discussed. The role of sodium-glucose transporter 2 inhibitors in improving renal lipid accumulation was summarized.

Glycolytic lactate in diabetic kidney disease.

Lactate elevation is a well-characterized biomarker of mitochondrial dysfunction, but its role in diabetic kidney disease (DKD) is not well defined. Urine lactate was measured in patients with type 2 diabetes (T2D) in 3 cohorts (HUNT3, SMART2D, CRIC). Urine and plasma lactate were measured during euglycemic and hyperglycemic clamps in participants with type 1 diabetes (T1D). Patients in the HUNT3 cohort with DKD had elevated urine lactate levels compared with age- and sex-matched controls. In patients in the SMART2D and CRIC cohorts, the third tertile of urine lactate/creatinine was associated with more rapid estimated glomerular filtration rate decline, relative to first tertile. Patients with T1D demonstrated a strong association between glucose and lactate in both plasma and urine. Glucose-stimulated lactate likely derives in part from proximal tubular cells, since lactate production was attenuated with sodium-glucose cotransporter-2 (SGLT2) inhibition in kidney sections and in SGLT2-deficient mice. Several glycolytic genes were elevated in human diabetic proximal tubules. Lactate levels above 2.5 mM potently inhibited mitochondrial oxidative phosphorylation in human proximal tubule (HK2) cells. We conclude that increased lactate production under diabetic conditions can contribute to mitochondrial dysfunction and become a feed-forward component to DKD pathogenesis.

Bioinformatic analyses reveal lysosomal-associated protein transmembrane 5 as a potential therapeutic target in lipotoxicity-induced injury in diabetic kidney disease.

Emerging data have revealed that damage to tubular epithelial cell is a driving force in the progression of diabetic kidney disease (DKD). However, the specific mechanisms by which lipotoxicity contributes to the injury of these cells, thereby influencing the development of DKD, are yet to be fully understood. Here, we analyzed the GSE 30529 microarray datasets of human tubulointerstitial tissue samples from the Gene Expression Omnibus database (GEO). Concurrently, we conducted RNA-sequencing on palmitic acid (PA)-treated human renal proximal tubule epithelial cells (HK2 cells). After normalization, the differentially expressed genes (DEGs) were screened by R software and gene ontology (GO) enrichment analysis was conducted, and lysosomal-associated protein transmembrane 5 (LAPTM5) was finally selected. Our findings indicate that the expression of LAPTM5 was obviously increased in DKD patients, and the correlation between LAPTM5, and other clinical parameters of DKD was analyzed using the Spearman correlation analysis. The potential of LAPTM5 as a prognostic biomarker for DKD was further consolidated through receiver operating characteristic (ROC) analysis. To further verify the function of LAPTM5, we established mouse or in vitro systems mimicking DKD. The results showed that a consistent upregulation of LAPTM5, which was also found to be linked with inflammatory mediators within the context of DKD. Additionally, LAPTM5 silencing significantly downregulated mRNA expression of inflammatory factors in PA-treated HK2 cells. These results indicate that LAPTM5 is a potential biomarker and therapeutic treatment target for DKD. This discovery paves the way for future research and development of targeted interventions aimed at mitigating the progression of this prevalent condition.

Renal Tissue-Derived Exosomal miRNA-34a in Diabetic Nephropathy Induces Renal Tubular Cell Fibrosis by Promoting the Polarization of M1 Macrophages.

Background: Diabetic nephropathy (DN) is the leading cause of chronic kidney disease, and the activation and infiltration of phagocytes are critical steps of DN. This study aimed to explore the mechanism of exosomes in macrophages and diabetes nephropathy and the role of miRNA-34a, which might provide a new path for treating DN. Materials and Methods: The DN model was established, and the success of the model establishment was confirmed by detecting general indicators, HE staining, and immunohistochemistry. Electron microscopy and NanoSight Tracking Analysis (NTA) were used to see the morphology and size of exosomes. MiRNA-34a inhibitor, miRNA-34a mimics, pc-PPARGC1A, and controls were transfected in macrophages with or without kidney exosomal. A dual-luciferase reporter gene experiment verifies the targeting relationship between miRNA-34a and PPARGC1A. After exosomal culture, macrophages are co-cultured with normal renal tubular cells to detect renal tubular cell fibrosis. Q-PCR and western blot were undertaken to detect related RNA and proteins. Results: An animal model of diabetic nephropathy was successfully constructed. Macrophages could phagocytose exosomes. After ingesting model exosomes, M1 macrophages were activated, while M2 macrophages were weakened, indicating the model mice's kidney exosomes caused the polarization. MiRNA-34a inhibitor increased PPARGC1A expression. MiRNA-34a expressed higher in diabetic nephropathy Model-Exo. MiRNA-34a negatively regulated PPARGC1A. PPARGC1A rescued macrophage polarization and renal tubular cell fibrosis. Conclusion: Exosomal miRNA-34a of tubular epithelial cells promoted M1 macrophage activation in diabetic nephropathy via negatively regulating PPARGC1A expression, which may provide a new direction for further exploration of DN treatment.

Atorvastatin ameliorates diabetic nephropathy through inhibiting oxidative stress and ferroptosis signaling.

Clinically, statins have long been used for the prevention and treatment of chronic renal diseases, however, the underlying mechanisms are not fully elucidated. The present study investigated the effects of atorvastatin on diabetes renal injury and ferroptosis signaling. A mouse model of diabetes was established by the intraperitoneal injection of streptozotocin (50 mg/kg/day) plus a high fat diet with or without atorvastatin treatment. Diabetes mice manifested increased plasma glucose and lipid profile, proteinuria, renal injury and fibrosis, atorvastatin significantly lowered plasma lipid profile, proteinuria, renal injury in diabetes mice. Atorvastatin reduced renal reactive oxygen species (ROS), iron accumulation and renal expression of malondialdehyde (MDA), 4-hydroxynonenal (4-HNE), transferrin receptor 1 (TFR1), and increased renal expression of glutathione peroxidase 4 (GPX4), nuclear factor erythroid 2-related factor (NRF2) and ferritin heavy chain (FTH) in diabetes mice. Consistent with the findings in vivo, atorvastatin prevented high glucose-induced ROS formation and Fe2+ accumulation, an increase in the expression of 4-HNE, MDA and TFR1, and a decrease in cell viability and the expression of NRF2, GPX4 and FTH in HK2 cells. Atorvastatin also reversed ferroptosis inducer erastin-induced ROS production, intracellular Fe2+ accumulation and the changes in the expression of above-mentioned ferroptosis signaling molecules in HK2 cells. In addition, atorvastatin alleviated high glucose- or erastin-induced mitochondria injury. Ferroptosis inhibitor ferrostatin-1 and antioxidant N-acetylcysteine (NAC) equally reversed the expression of high glucose-induced ferroptosis signaling molecules. Our data support the notion that statins can inhibit diabetes-induced renal oxidative stress and ferroptosis, which may contribute to statins protection of diabetic nephropathy.

Apoptosis and NETotic cell death affect diabetic nephropathy independently: An study integrative study encompassing bioinformatics, machine learning, and experimental validation.

OBJECTIVE: Although programmed cell death (PCD) and diabetic nephropathy (DN) are intrinsically conneted, the interplay among various PCD forms remains elusive. In this study, We aimed at identifying independently DN-associated PCD pathways and biomarkers relevant to the related pathogenesis. METHODS: We acquired DN-related datasets from the GEO database and identified PCDs independently correlated with DN (DN-PCDs) through single-sample Gene Set Enrichment Analysis (ssGSEA) as well as, univariate and multivariate logistic regression analyses. Subsequently, applying differential expression analysis, weighted gene co-expression network analysis (WGCNA), and Mfuzz cluster analysis, we filtered the DN-PCDs pertinent to DN onset and progression. The convergence of various machine learning techniques ultimately spotlighted hub genes, substantiated through dataset meta-analyses and experimental validations, thereby confirming hub genes and related pathways expression consistencies. RESULTS: We harmonized four DN-related datasets (GSE1009, GSE142025, GSE30528, and GSE30529) post-batch-effect removal for subsequent analyses. Our differential expression analysis yielded 709 differentially expressed genes (DEGs), comprising 446 upregulated and 263 downregulated DEGs. Based on our ssGSEA as well as univariate and multivariate logistic regressions, apoptosis and NETotic cell death were appraised as independent risk factors for DN (Odds Ratio > 1, p < 0.05). Next, we further refined 588 apoptosis- and NETotic cell death-associated genes through WGCNA and Mfuzz analysis, resulting in the identification of 17 DN-PCDs. Integrating protein-protein interaction (PPI) network analyses, network topology, and machine learning, we pinpointed hub genes (e.g., IL33, RPL11, and CX3CR1) as significant DN risk factors with expression corroborating in subsequent meta-analyses and experimental validations. Our GSEA enrichment analysis discerned differential enrichments between DN and control samples within pathways such as IL2/STAT5, IL6/JAK/STAT3, TNF-α via NF-κB, apoptosis, and oxidative phosphorylation, with related proteins such as IL2, IL6, and TNFα, which we subsequently submitted to experimental verification. CONCLUSION: Innovatively stemming from from PCD interactions, in this study, we discerned PCDs with an independent impact on DN: apoptosis and NETotic cell death. We further screened DN evolution- and progression-related biomarkers, i.e. IL33, RPL11, and CX3CR1, all of which we empirically validated. This study not only poroposes a PCD-centric perspective for DN studies but also provides evidence for PCD-mediated immune cell infiltration exploration in DN regulation. Our results could motivate further exploration of DN pathogenesis, such as how the inflammatory microenvironment mediates NETotic cell death in DN regulation, representing a promising direction for future research.

Mitochondrial metabolic reprogramming in diabetic kidney disease.

Diabetic kidney disease, known as a glomerular disease, arises from a metabolic disorder impairing renal cell function. Mitochondria, crucial organelles, play a key role in substance metabolism via oxidative phosphorylation to generate ATP. Cells undergo metabolic reprogramming as a compensatory mechanism to fulfill energy needs for survival and growth, attracting scholarly attention in recent years. Studies indicate that mitochondrial metabolic reprogramming significantly influences the pathophysiological progression of DKD. Alterations in kidney metabolism lead to abnormal expression of signaling molecules and activation of pathways, inducing oxidative stress-related cellular damage, inflammatory responses, apoptosis, and autophagy irregularities, culminating in renal fibrosis and insufficiency. This review delves into the impact of mitochondrial metabolic reprogramming on DKD pathogenesis, emphasizing the regulation of metabolic regulators and downstream signaling pathways. Therapeutic interventions targeting renal metabolic reprogramming can potentially delay DKD progression. The findings underscore the importance of focusing on metabolic reprogramming to develop safer and more effective therapeutic approaches.