中医药治疗足细胞胰岛素抵抗的研究进展

We explored the protection of mangiferin monosodium salt (MGM) on kidney injury in rats with streptozotocin (STZ)-induced diabetic nephropathy (DN) by "multiomics" analysis combined with systems pharmacology, with a specific focus on ferroptosis, inflammation, and podocyte insulin resistance (IR) signaling events in kidneys. MGM treatment afforded renoprotective effects on rats with STZ-induced DN by alleviating systemic IR-induced renal inflammation and podocyte IR. These mechanisms were correlated mainly with the MGM treatment-induced inhibition of the mitogen-activated protein kinase/nuclear factor-kappa B axis and activation of the phosphorylated insulin receptor substrate 1(Tyr608)/phosphorylated phosphatidylinositol 3-kinase/phosphorylated protein kinase B axis in the kidneys of DN rats. MGM had an ameliorative function in renal ferroptosis in rats with STZ-induced DN by upregulating mevalonate-mediated antioxidant capacities (glutathione peroxidase 4 and ferroptosis suppressor protein 1/coenzyme Q10 axis) and weakening acyl-CoA synthetase long-chain family member 4-mediated proferroptotic generation of lipid drivers in kidneys. MGM may be a promising alternative strategy for the treatment of DN.

Abstract Background Podocyte injury is a common pathologic mechanism in obesity-related glomerulopathy (ORG). Safflower injection (SFI), scientifically extracted and refined from safflower, is used to treat diabetic kidney disease according to clinical guideline. Our previous study confirmed that the main active compounds of SFI ameliorated high glucose-induced podocyte injury. It is uncertain whether SFI has an effect on ORG-related podocyte injury. Objectives This study aimed to explore the pharmacological effects and related mechanisms of SFI on podocyte injury of ORG mice. Methods First, by combining ultra-high performance liquid chromatography tandem mass spectrometry analysis with online databases, the pathway enrichment, target-pathway analysis, and human protein–protein interaction network were conducted to discover the possible crucial mechanism of SFI against ORG. Then, ORG mice model was established by high-fat diet and biochemical assays, histopathology and western blot were used to explore the effects of SFI on obesity and podocyte injury. Finally, system pharmacology-based findings were evaluated in ORG mice. Results The results of system pharmacology suggested that SFI could alleviate ORG through insulin resistance (IR)-related pathway by regulating insulin receptor (INSR) and endothelial nitric oxide synthase (eNOS) expressions. The in vivo experiment confirmed that SFI ameliorated obesity, lipid metabolism-related indicators, podocyte injury of ORG mice. The mechanism relationships among IR, INSR, and eNOS were further verified in ORG mice. Conclusions Our findings imply that by up-regulating the expression of renal INSR and eNOS, thereby inhibiting IR, SFI may be a promising candidate for the treatment of ORG.

Diabetic nephropathy (DN) is the predominant cause of end-stage renal disease globally. Diosgenin (DSG) has been reported to play a protective role in podocyte injury in DN. The present study aimed to explore the role of DSG in DN, as well as its mechanism of action in a high glucose (HG)-induced in vitro model of DN in podocytes. Cell viability, apoptosis, inflammatory response and insulin-stimulated glucose uptake were evaluated using Cell Counting Kit-8, TUNEL, ELISA and 2-deoxy-D-glucose assay, respectively. In addition, the expression of AMP-activated protein kinase (AMPK)/sirtuin 1 (SIRT1)/NF-κB signaling-related proteins in podocyte cells was measured using western blotting. The results indicated that DSG enhanced the viability of podocytes after HG exposure, but inhibited inflammatory damage and attenuated insulin resistance. Moreover, DSG induced the activation of the AMPK/SIRT1/NF-κB signaling pathway. Furthermore, treatment with compound C, an inhibitor of AMPK, counteracted the protective effects of DSG on HG-induced podocyte cells. Therefore, DSG may be a potential therapeutic compound for the treatment of diabetic nephropathy.

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Summary Insulin signaling to the glomerular podocyte via the insulin receptor (IR) is critical for kidney function. In this study we show that near-complete knockout of the closely related insulin-like growth factor 1 receptor (IGF1R) in podocytes is detrimental, resulting in albuminuria in vivo and podocyte cell death in vitro. In contrast, partial podocyte IGF1R knockdown confers protection against doxorubicin-induced podocyte injury. Proteomic analysis of cultured podocytes revealed that while near-complete loss of podocyte IGF1R results in the downregulation of mitochondrial respiratory complex I and DNA damage repair proteins, partial IGF1R inhibition promotes respiratory complex expression. This suggests that altered mitochondrial function and resistance to podocyte stress depends on the level of IGF1R suppression, the latter determining whether receptor inhibition is protective or detrimental. Our work suggests that the partial suppression of podocyte IGF1R could have therapeutic benefits in treating albuminuric kidney disease.

&NA; Diabetic nephropathy (DN) contributes to end‐stage renal disease and kidney dysfunction with a proverbial feature of podocyte injury. Inflammation and insulin resistance is recently implicated in the pathogenesis of diabetic kidney injury. Celastrol exerts critical roles in inflammatory diseases and injury progression. However, its function and mechanism in DN remains elusive. Here, celastrol dose‐dependently restored podocyte viability under high glucose (HG) conditions, but with little cytotoxicity in podocyte. Preconditioning with celastrol counteracted HG‐evoked cell apoptosis, LDH release, ROS production and podocyte depletion. Additionally, HG‐elevated high transcripts and secretions of pro‐inflammatory cytokines were reversed following celastrol treatment, including IL‐1&bgr;, TNF‐&agr;, IL‐6. Simultaneously, the inhibitory effects of HG on insulin‐triggered glucose uptake and nephrin expression were overturned after celastrol exposure. Intriguingly, celastrol restored HG‐induced deficiency of autophagy pathway. Nevertheless, blocking the autophagy signaling by its antagonist 3‐MA muted celastrol‐protected against HG‐evoked cell injury, inflammation and insulin resistance. Importantly, celastrol enhanced heme oxygenase‐1 (HO‐1) expression in HG‐stimulated podocytes. Notably, HO‐1 cessation depressed autophagy pathway activation and subsequently blunted beneficial effects of celastrol on HG‐exposed podocytes. These finding suggest that celastrol may protect against HG‐induced podocyte injury, inflammation and insulin resistance by restoring HO‐1‐mediated autophagy pathway, implying a promising therapeutic strategy against DN.

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Histological manifestations of diabetic kidney disease (DKD) include mesangiolysis, mesangial matrix expansion, mesangial cell proliferation, thickening of the glomerular basement membrane, podocyte loss and foot process effacement, hyalinosis of the glomerular arterioles, interstitial fibrosis and tubular atrophy. Glomerulomegaly is a typical finding. Histological features of DKD may occur in the absence of clinical manifestations, having been documented in patients with normal urinary albumin excretion and normal glomerular filtration rate. Furthermore, the histological picture progresses over time while clinical data may remain normal. Conversely, histological lesions of DKD improve with metabolic normalization, following effective pancreas transplantation. Insulin resistance has been associated with the clinical manifestations of DKD (nephromegaly, glomerular hyperfiltration, albuminuria, and kidney failure). Likewise, insulin resistance may underlie the histological manifestations of DKD. Morphological changes of DKD are absent in newly diagnosed type 1 diabetes patients (with no insulin resistance), but appear afterwards when insulin resistance develops. In contrast, structural lesions of DKD are typically present before the clinical diagnosis of type 2 diabetes. Several heterogeneous conditions that share the occurrence of insulin resistance, such as aging, obesity, acromegaly, lipodystrophy, cystic fibrosis, insulin receptor dysfunction, and Alström syndrome, also share both clinical and structural manifestations of kidney disease, including glomerulomegaly and other features of DKD, focal segmental glomerulosclerosis, and C3 glomerulopathy, which might be ascribed to reduction in the synthesis of factor H binding sites (such as heparan sulfate) that leads to uncontrolled complement activation. Alström syndrome patients show systemic interstitial fibrosis markedly similar to that present in diabetes.

Insulin resistance (IR) is stated as diminished insulin action regardless of hyperinsulinemia. The usual target organs for insulin activities are the liver, skeletal muscle, and adipose tissue. Hence, the vasculature and kidneys are nonconventional target organs as the impacts of insulin on these are comparatively separate from other conventional target organs. Vasodilation is achieved by raising endothelial nitric oxide (NO) generation by initiating the phosphoinositide 3-kinase (PI3K) pathway. In insulin-nonresponsive conditions, this process is defective, and there is increased production of endothelin-1 through the mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK) pathway, which predominates the NO effects, causing vasoconstriction. Renal tubular cells and podocytes have insulin receptors, and their purposeful importance has been studied, which discloses critical acts of insulin signaling in podocyte survivability and tubular action. Diabetic nephropathy (DN) is a prevalent problem in individuals with hypertension, poor glycemic management, hereditary susceptibility, or glomerular hyperfiltration. DN could be a significant contributing factor to end-stage renal disease (ESRD) that results from chronic kidney disease (CKD). IR and diabetes mellitus (DM) are the constituents of syndrome X and are accompanied by CKD progression. IR performs a key part in syndrome X leading to CKD. However, it is indistinct whether IR individually participates in enhancing the threat to CKD advancement rather than CKD complexity. CKD is an extensive public health problem affecting millions of individuals worldwide. The tremendous spread of kidney disease intensifies people’s health impacts related to communicable and noncommunicable diseases. Chronic disease regulator policies do not include CKD at global, local, and/or general levels. Improved knowledge of the character of CKD-associated problems might aid in reforming diagnosis, prevention, and management.

Podocytes are insulin-sensitive cells, and their loss is critical in diabetic nephropathy (DN) progression that could lead to end-stage kidney disease. We have previously shown that decreased DUSP4 expression caused elevated JNK phosphorylation in the diabetic kidney and worsened DN characteristics. Yet, the role of DUSP4 in diabetic podocyte insulin resistance and the progression of DN remains unclear. Here, we report that HG-exposed podocytes exhibited reduced DUSP4 expression, increased phosphorylation of JNK and serine 307 of IRS1 as well as Nox4 expression, while decreasing insulin signaling actions. DUSP4 overexpression, JNK and Nox1/4 inhibition prevented HG-induced serine 307 phosphorylation of IRS1 and restored insulin actions. Diabetic mice showed renal dysfunction and insulin resistance, characteristics that were exacerbated in diabetic DUSP4 deficient mice due to Nox1/4 upregulation. Thus, our results demonstrated that diabetes-induced reduction of DUSP4 leads to JNK activation and elevated Nox4 expression, which contributes to podocyte dysfunction, insulin resistance and progression of DN.

Diabetic nephropathy (DN) is becoming a research hotspot in recent years because the prevalence is high and the prognosis is poor. Lipid accumulation in podocytes induced by hyperglycemia has been shown to be a driving mechanism underlying the development of DN. However, the mechanism of lipotoxicity remains unclear. Increasing evidence shows that acetyl‐CoA carboxylase 2 (ACC2) plays a crucial role in the metabolism of fatty acid, but its effect in podocyte injury of DN is still unclear. In this study, we investigated whether ACC2 could be a therapeutic target of lipid deposition induced by hyperglycemia in the human podocytes. Our results showed that high glucose (HG) triggered significant lipid deposition with a reduced β‐oxidation rate. It also contributed to the downregulation of phosphorylated ACC2 (p‐ACC2), which is an inactive form of ACC2. Knockdown of ACC2 by sh‐RNA reduced lipid deposition induced by HG. Additionally, ACC2‐shRNA restored the expression of glucose transporter 4 (GLUT4) on the cell surface, which was downregulated in HG and normalized in the insulin signaling pathway. We verified that ACC2‐shRNA alleviated cell injury, apoptosis, and restored the cytoskeleton disturbed by HG. Mechanistically, SIRT1/PGC‐1α is close related to the insulin metabolism pathway. ACC2‐shRNA could restore the expression of SIRT1/PGC‐1α, which was downregulated in HG. Rescue experiment revealed that inhibition of SIRT1 by EX‐527 counteracted the effect of ACC2‐shRNA. Taken together, our data suggest that podocyte injury mediated by HG‐induced insulin resistance and lipotoxicity could be alleviated by ACC2 inhibition via SIRT1/PGC‐1α.

Epidemiological evidence indicates that hyperinsulinemia is an independent risk factor for albuminuria and renal impairment, yet its molecular basis remains unclear. In prediabetic db/db mice, hyperinsulinemia coincided with albuminuria, podocyte injury, and impaired glomerular insulin signaling, characterized by insulin receptor exhaustion and hypoactivity of the IRS1/PI3K/Akt insulin signaling. This led to diminished inhibitory phosphorylation of GSK3βS9 in podocytes, denoting GSK3β hyperactivity. Notably, GSK3β co-localized and physically interacted with IRS1 in glomerular podocytes, phosphorylating IRS1S332 as a direct substrate. In cultured podocytes, prolonged high insulin exposure induced insulin receptor depletion, GSK3β hyperactivity, and increased inhibitory phosphorylation of IRS1S332, forming a self-perpetuating cycle of insulin desensitization and podocyte injury. GSK3β appears to play a key role, as ectopic expression of a constitutively active GSK3β mutant GSK3βS9A enhanced inhibitory phosphorylation of IRS1S332, desensitized insulin signaling, and exacerbated podocyte injury. In contrast, forced expression of a kinase-dead mutant of GSK3β or inhibition of GSK3β with a selective small-molecule inhibitor tideglusib abrogated inhibitory phosphorylation of IRS1S332, restored insulin sensitivity and protected podocytes. In vivo, GSK3βS9Aknock-in mice exhibited impaired insulin signaling and podocyte injury with albuminuria. Conversely, tideglusib treatment in prediabetic db/db mice attenuated podocyte injury and insulin resistance, thereby improving albuminuria. Collectively, hyperinsulinemia directly elicits albuminuria and renal impairment via a cascade of molecular events involving insulin receptor exhaustion, reduced insulin signaling, and GSK3β hyperactivity, which promotes IRS1 inhibition and thereby forms a self-amplifying GSK3β-IRS1 circuit of insulin desensitization and podocyte injury. Targeting GSK3β could disrupt this pathogenic loop and mitigate hyperinsulinemia-induced renal injury.

Podocytes are highly specialized epithelial cells that play a central role in maintaining integrity of the glomerular filtration barrier. Because of their complex architecture and dynamic actin-based cytoskeleton, podocytes have substantial energy requirements, which are predominantly supported by glycolysis. Insulin signaling and glucose uptake are key regulators of cytoskeletal dynamics in these cells. Recent evidence highlights the importance of lactate metabolism in maintaining podocyte metabolic homeostasis, supported by a well-developed system for controlling lactate levels. Monocarboxylate transporter 1 (MCT1), a principal mediator of lactate transport, has emerged as a critical regulator of cellular energy balance. The present study investigated the role of MCT1 in insulin-stimulated glucose metabolism and its impact on podocyte morphology and function. Our findings showed that MCT1 inhibition impaired glucose uptake and suppressed glycolytic flux. This metabolic disruption was accompanied by alterations of the localization of key insulin signaling proteins, disorganization of the actin cytoskeleton, and an increase in permeability of the podocyte layer. Interestingly, MCT1 inhibition also triggered a compensatory shift toward oxidative phosphorylation, potentially linked to an increase in mitochondrial biogenesis. These results underscore the pivotal role of MCT1 in regulating glucose metabolism and actin cytoskeleton organization in podocytes and suggest that lactate transport is essential for preserving their structure and function. Targeting MCT1 and lactate metabolism may offer a novel therapeutic strategy for glomerular diseases that are characterized by insulin resistance and metabolic dysregulation.

Background QiDiTangShen granules (QDTS), a traditional Chinese medicine (TCM) compound prescription, have remarkable efficacy in diabetic nephropathy (DN) patients, and their pharmacological mechanism needs further exploration. Methods According to the active ingredients and targets of the QDTS in the TCMSP database, the network pharmacology of QDTS was investigated. The potential active ingredients were chosen based on the oral bioavailability and the drug similarity index. At the same time, targets for DN-related disease were obtained from GeneCards, OMIM, PharmGKB, TTD, and DrugBank. The TCM-component-target network and the protein-protein interaction (PPI) network were constructed with the Cytoscape and STRING platforms, respectively, and then the core targets of DN were selected with CytoNCA. GO and KEGG enrichment analysis using R software. Molecular docking to identify the core targets of QDTS for DN. In vivo, db/db mice were treated as DN models, and the urine microalbuminuria, the pathological changes in the kidney and the protein expression levels of p-PI3K, p-Akt, JUN, nephrin and synaptopodin were detected by immunohistochemistry, immunofluorescence method and Western blotting. After QDTS was used in vitro, the protein expression of mouse podocyte clone-5 (MPC5) cells was detected by immunohistochemistry, immunofluorescence and Western blot. Results Through network pharmacology analysis, 153 potential targets for DN in QDTS were identified, 19 of which were significant. The KEGG enrichment analysis indicated that QDTS might have therapeutic effects on IL-17, TNF, AGE-RAGE, PI3K-Akt, HIF-1, and EGFR through interfering with Akt1 and JUN. The main active ingredients in QDTS are quercetin, β-sitosterol, stigmasterol and kaempferol. Both in vivo and in vitro studies showed that QDTS could decrease the urine microalbuminuria and renal pathology of db/db mice, and alleviate podocyte injuries through the PI3K/Akt signaling pathway. Conclusion Through network pharmacology, in vivo and in vitro experiments, QDTS has been shown to improve the urine microalbuminuria and renal pathology in DN, and to reduce podocyte damage via the PI3K/Akt pathway.

Diabetic kidney disease (DKD) is a common complication of diabetes, often characterized by podocyte injury, proteinuria, and eventual renal failure. Sinomenine hydrochloride (SH), an active component derived from traditional Chinese medicine, is clinically effective in treating kidney diseases. This study investigates the protective effects of SH on podocytes under high‐glucose conditions and its mechanism of action. Mouse podocytes (MPC‐5) were treated with SH at concentrations of 50, 200, and 600 μg/mL under high‐glucose conditions (30 mmol/L) for 24 h to establish a DKD model. Cell viability was assessed using CCK‐8 assays, and apoptosis rates were measured using flow cytometry. Autophagy levels were evaluated by detecting LC3‐II, Beclin‐1, and P62 proteins via Western blot analysis, while the involvement of the PI3K/AKT/mTOR pathway was analyzed by examining phosphorylated AKT and mTOR. Transmission electron microscopy was employed to observe autophagosomes. SH improved podocyte viability, reduced apoptosis, and enhanced autophagic activity by increasing LC3‐II and Beclin‐1 expression while decreasing P62 levels. SH also downregulated p‐AKT and p‐mTOR, indicating inhibition of the PI3K/AKT/mTOR pathway. Electron microscopy confirmed increased autophagosomes in SH‐treated groups. SH protects podocytes in a high‐glucose environment by enhancing autophagy through inhibition of the PI3K/AKT/mTOR pathway. These findings provide insights into SH as a potential therapeutic agent for DKD management.

This study utilized db/db mice and MPC5 cells induced by high glucose as experimental models to examine the protective mechanisms of the traditional Chinese medicine formula TangNaikang (TNK) in mitigating podocyte injury in diabetic nephropathy (DN).

Traditional Chinese Medicine (TCM) has demonstrated promising efficacy in managing and preventing the early-stage diabetic nephropathy (DN). Although the exact mechanisms remain elusive, clinical evidence has suggested that Jinlida granules (JLD) are beneficial in improving renal function among patients with DN. The present study aimed to elucidate the effect of JLD on DN and the underlying molecular mechanism. Therefore, podocyte apoptosis was evaluated using flow cytometry and TUNEL staining, while mitochondrial morphology and function were assessed using transmission electron microscopy, MitoTracker, JC-1 and reactive oxygen species staining. RNA sequencing analysis was performed to elucidate the mechanism underlying the effect of JLD on DN. Additionally, to investigate the role of peroxisome proliferator-activated receptor-γ co-activator-1α (PGC-1α) in mitigating JLD-induced mitochondrial dysfunction and podocyte apoptosis, MPC5 cells were transfected with the corresponding small interfering RNA constructs. The results showed that JLD effectively improved renal function and mitigated podocyte injury, as well as ameliorated mitochondrial dysfunction and inhibited apoptosis in db/db mice. In vitro experiments further revealed that JLD exerted a protective effect via inhibiting mitochondrial fission and apoptosis in high glucose-treated podocytes. Furthermore, JLD enhanced the phosphorylation of adenosine monophosphate-activated protein kinase (AMPK), thus promoting the expression of PGC-1α, eventually improving apoptosis and mitochondrial homeostasis. Overall, the current study revealed that JLD could improve mitochondrial homeostasis and reduce cell apoptosis in podocytes via activating the AMPK/PGC-1α pathway, thus providing a theoretical foundation for the clinical management of DN.

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Diabetic nephropathy (DN) represents a severe microvascular complication of diabetes mellitus. As a Traditional Chinese Medicine (TCM) with extensive clinical applications, Ligustri Lucidi Fructus (LLF) exhibits significant anti-DN activity. However, the underlying pharmacological mechanisms, crucial components, and targets for LLF in DN treatment remain unclear. By integrating network pharmacology, molecular docking, and molecular dynamics simulations, the bioactive compounds, potential therapeutic targets, and underlying mechanisms of LLF in the treatment of DN were elucidated, followed by biological validation in a palmitic acid (PA)-induced MPC5 podocyte injury model. Among the 383 DN-related LLF targets identified, TNF emerged as a pivotal one, demonstrating potential binding interaction with the active components salidroside (Sal), apigenin (Api), and tormentic acid (TA). Moreover, Gene Expression Omnibus (GEO) database and KEGG enrichment analysis collectively highlighted the cytosolic DNA-sensing pathway. Notably, the cGAS-STING pathway is central to this pathway. Experimental studies further demonstrated that LLF-containing serum exerted a protective effect on MPC5 podocytes through cGAS-STING pathway suppression. Overall, these findings elucidate the pleiotropic mechanisms underlying LLF’s protective effects against DN, integrating compound–target–pathway interactions and thus offering a rationale for further investigation.

Diabetic nephropathy (DN) is a major complication of diabetes with limited therapeutic options. Yiqi Huoxue Yangyin Decoction (YHY), a traditional Chinese medicine formula designed to treat the TCM syndrome of “Qi and Yin deficiency with blood stasis” often observed in DN, has shown clinical potential. However, its precise mechanism of action, particularly concerning the regulation of podocyte autophagy, remains unclear. The chemical profile of YHY was characterized using ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). The therapeutic effects and mechanisms were evaluated in a db/db mouse model of DN and in high glucose-stimulated MPC-5 podocytes. Key assessments included renal function, podocyte injury markers, autophagy flux, and the activity of the METTL3-m6A-mTOR pathway. UPLC-MS/MS analysis successfully identified multiple chemical metabolites in YHY, providing a comprehensive phytochemical profile of the formula. Treatment with YHY significantly ameliorated renal dysfunction and attenuated podocyte injury in db/db mice. Both in vivo and in vitro models exhibited impaired autophagy and hyperactivated mTOR signaling under diabetic conditions. Crucially, YHY treatment inhibited METTL3 expression and m6A methylation levels, leading to decreased mTOR mRNA stability and protein expression, which subsequently restored autophagic activity. YHY, a traditional Chinese medicine formula with defined chemical metabolites, ameliorates podocyte damage in DN by activating autophagy via the METTL3-m6A-mTOR signaling pathway. This study provides novel insights into the epigenetic regulation of DN and underscores the potential of YHY as a therapeutic agent for this condition.

BACKGROUND Traditional Chinese medicine (TCM) has shown great promise in treating diabetic nephropathy (DN). However, the key targets and mechanisms underlying the therapeutic effects of the active ingredients of modified prescription Jiawei Qihuangyin (JWQHY) remain unclear. METHODS Network pharmacology analysis was employed to identify potential targets of JWQHY in DN. Protein-protein interaction (PPI) and TCM component-target networks were constructed, and KEGG pathway enrichment analysis was performed to determine key therapeutic targets and signaling pathways. Molecular docking suggested an interaction between the major active compound formononetin (FMN) and the central target silent information regulator 1 (SIRT1), which was experimentally validated using cellular thermal shift assay. SIRT1 expression in podocytes was assessed by qRT-PCR and western blotting (WB). Cell viability (CCK-8), apoptosis (flow cytometry), and proinflammatory cytokine secretion (ELISA) were measured to evaluate podocyte injury. The acetylation level of NF-κB p65 and epithelial-mesenchymal transition (EMT)-related proteins were analyzed by WB. In vivo, a DN rat model was established to assess the therapeutic efficacy of JWQHY through biochemical urine analysis, histopathological examination (HE staining), and WB detection of SIRT1, acetylated NF-κB p65, and EMT markers. RESULTS Network pharmacology identified 52 potential overlapping targets of JWQHY in DN, primarily associated with the NF-κB pathway. Among these, SIRT1 was predicted and experimentally confirmed as the main target of FMN. In a high-glucose-induced podocyte injury model, FMN upregulated SIRT1 expression, promoted NF-κB p65 deacetylation, and inhibited podocyte EMT. Consistently, FMN treatment improved renal function, reduced podocyte injury, and modulated SIRT1/NF-κB signaling in DN rats. CONCLUSION JWQHY exerts therapeutic effects in diabetic nephropathy by modulating the SIRT1/NF-κB signaling axis through its active compound formononetin, thereby inhibiting podocyte EMT. These findings provide mechanistic insight into the pharmacological basis of FMN and support its clinical potential in DN treatment.

ETHNOPHARMACOLOGICAL RELEVANCE Diabetic nephropathy (DN), a leading cause of end-stage renal disease, lacks curative therapies. Podocyte injury plays a central role in DN progression, yet strategies to effectively preserve podocyte integrity remain limited. Shenxiao decoction (SXD) is a classic Traditional Chinese Medicine (TCM) prescription, derived from Buzhong Yiqi Decoction and Xuefu Zhuyu Decoction, widely used clinically for DN. However, the molecular mechanisms of podocyte protection are poorly understood. AIM OF THE STUDY This study aimed to evaluate the protective effects of SXD on podocyte injury in DN and to elucidate its regulation of RUNX3 and the JAK2/STAT3 pathway. MATERIALS AND METHODS The main constituents of SXD were characterized by high-resolution quadrupole time-of-flight liquid chromatography-mass spectrometry (HR-Q-TOF LC-MS). DN was modeled in db/db mice fed a high-sugar/high-fat diet and high-glucose-stimulated MPC-5 cells. The therapeutic effects of SXD were assessed by biochemical assays, renal function tests, and histopathology. Cytokine levels were measured by ELISA, and podocin expression was evaluated by Western blot and immunohistochemistry. The underlying mechanism was investigated by assessing RUNX3 expression and JAK2/STAT3 signaling. RUNX3 was silenced in MPC5 cells using small interfering RNA (siRNA), and STAT3 activity was inhibited in vivo with Stattic. RESULTS SXD improved metabolic disturbances, reduced proteinuria, and alleviated renal histopathological damage in DN mice. It suppressed systemic and renal inflammation by lowering proinflammatory cytokine levels and upregulating podocin expression. Mechanistically, SXD restored RUNX3 expression and inhibited JAK2/STAT3 activation both in vivo and in vitro. Silencing RUNX3 abolished the renoprotective effects of SXD in podocytes, whereas inhibiting STAT3 reproduced its protective effects, confirming the critical role of the RUNX3/JAK2/STAT3 axis. CONCLUSIONS SXD alleviates podocyte injury and DN progression by enhancing RUNX3 and suppressing JAK2/STAT3 signaling. These findings uncover the role of RUNX3 and JAK2/STAT3 pathway in podocyte protection and highlight SXD as a promising therapeutic strategy for DN.

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Highlights What are the main findings? ST32da enhances ATF3/HDAC2 recruitment to inhibit NF-κB–IL-6 signaling. ST32da mitigates podocyte injury and kidney lipotoxicity in diabetic mice. What is the implication of the main finding? ST32da show potential for use in treating diabetic nephropathy. Abstract It is necessary to find novel therapeutic strategies for obesity-related diabetic nephropathy (DN) that target both metabolic dysfunction and renal inflammation. ST32da derived from Salvia miltiorrhiza (a well-recognized Traditional Chinese Medicine) induces activating transcription factor 3 (ATF3), a negative regulator of inflammation and metabolic stress. However, the effects of ST32da on obesity-related DN remain underexplored. We investigated the therapeutic potential of ST32da, a synthetic ATF3 inducer derived from Salvia miltiorrhiza, in mitigating obesity-related DN in both in vivo and in vitro models. The Nephroseq database analysis was performed to explore the relationship between Atf3 expression and DN progression. ST32da was administered to db/db knockout and DBA mice to establish obesity-related DN models, and a high-fat diet (HFD)-induced mouse model of obesity-related DN was used to investigate the effects of Atf3 knockout. Molecular and biochemical analyses were conducted in cultured mesangial cells to elucidate the underlying mechanisms. ATF3 deficiency worsened obesity-related DN, increasing glomerular fibrosis, mortality, and inflammation. ST32da restored ATF3 levels and reduced renal injury, glomerular expansion, and pro-inflammatory cytokine expression (e.g., IL-6, TGFβ, TNFα). ST32da-treated mice exhibited reduced hepatic lipid accumulation and improved serum lipid profiles. In mesangial cells, ST32da localized to the cytoplasm and increased ATF3 activity, which suppressed RARRES1 expression and cytokine signaling. Mechanistically, ATF3 interacted with HDAC2 to repress NF-κB—dependent inflammatory gene expression. The findings suggest ST32da is a promising therapeutic candidate for obesity-related DN and associated metabolic disturbances, acting through ATF3 induction to suppress renal inflammation, lipotoxicity, and fibrosis.

Radix astragali, a medicinal material for tonifying Chinese Qi, has widely been used for the treatment of Kidney disease in China and East Asia, especially in reducing the apoptosis of glomerular podocytes. Paecilomyces Cicadidae is a medicinal and edible fungus. In recent years, the application of traditional Chinese medicine (TCM) in solid-state fermentation of edible and medicinal fungi has become a hot issue. Fermentation is a special method to change the properties of TCM. Therefore, the potential roles and molecular mechanisms on podocytes of solid-state fermentation products of Radix astragali and Paecilomyces cicadidae (RPF) in diabetic nephropathy (DN) were studied. In vivo, the effect of RPF and Radix astragali on DN in mice was evaluated by detecting the biochemical indexes of blood and urine, renal function and podocyte integrity. In vitro, the expression of podocyte marker protein, autophagy marker protein and PI3K/AKT/mTOR signaling pathway protein were detected by Western blotting using a high glucose-induced podocyte injury model. The results showed that RPF had a significant alleviative effect on DN mice. RPF can significantly reduce urine protein, serum creatinine, and blood nitrogen urea in DN mice. Morphological analysis showed that RPF could improve kidney structure of DN and reduce the apoptosis of podocytes, and the effect was better than Radix astragali. In vitro results indicated that RPF could enhance autophagy and protect podocytes by inhibiting the PI3K/AKT/mTOR signaling pathway. In summary, RPF has better effect on delaying the development of DN than Radix astragali. RPF enhances autophagy in podocytes and delays DN probably by inhibiting the PI3K/AKT/mTOR signaling pathway.

Zuogui Wan (ZGW), a well-known traditional Chinese medicine (TCM), has been used to nourish “Kidney-Yin” for a long time in China, implying a protective effect on the kidney. The aim of the present study is to investigate the effect of ZGW on high glucose-induced podocyte apoptosis and diabetic nephropathy (DN) in db/db mice. ZGW (1 g/kg−1/day−1) was administered intragastrically to db/db mice for 8 weeks. HPLC was used for identifying the components of ZGW, biochemical and histopathological approaches were used for evaluating its therapeutic effects, and cultured mouse podocytes were used for further exploring its underlying mechanism in vitro. ZGW improved renal function and podocyte loss and also normalized kidney reactive oxygen species production in db/db mice. The cytotoxicity of ZGW on mouse podocytes was assessed by the LDH assay. The effect of ZGW on podocyte viability and apoptosis was determined with CCK-8 and Annexin-V/PI staining by treatment with high glucose. ZGW attenuated podocyte apoptosis, and oxidative stress was detected by the peroxide-sensitive fluorescent probe 2′,7′-dichlorodihydrofluorescein diacetate (DCF-DA) staining in a dose-dependent manner. Furthermore, ZGW decreased the expression of caspase-3 and phospho-p38 in both the kidney cortex and high glucose-treated podocytes. Thus, our data from in vivo and in vitro studies demonstrated that ZGW improved renal injury in diabetes by inhibiting oxidative stress and podocyte apoptosis.

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Background QiHuangYiShen granules (QHYS), a traditional Chinese herbal medicine formula, have been used in clinical practice for treating diabetic kidney disease for several years by our team. The efficacy of reducing proteinuria and delaying the decline of renal function of QHYS has been proved by our previous studies. However, the exact mechanism by which QHYS exerts its renoprotection remains largely unknown. Emerging evidence suggests that lncRNA MALAT1 is abnormally expressed in diabetic nephropathy (DN) and can attenuate renal fibrosis by modulating podocyte epithelial-mesenchymal transition (EMT). Objective In the present study, we aimed to explore whether QHYS could modulate lncRNA MALAT1 expression and attenuate the podocyte EMT as well as the potential mechanism related to the Wnt/β-catenin signal pathway. Methods SD rats were fed with the high-fat-high-sucrose diet for 8 weeks and thereafter administered with 30 mg/kg streptozotocin intraperitoneally to replicate the DN model. Quality control of QHYS was performed using high-performance liquid chromatography. QHYS were orally administered at 1.25, 2.5, and 5 g/kg doses, respectively, to the DN model rats for 12 weeks. Body weight, glycated haemoglobin, blood urea nitrogen, serum creatinine, 24-h proteinuria, and kidney index were measured. The morphologic pathology of the kidney was evaluated by Hematoxylin-eosin and Masson's trichrome staining. The expression level of lncRNA MALAT1 was determined by quantitative real-time polymerase chain reaction. In addition, the expression levels of podocyte EMT protein markers and Wnt/β-catenin pathway proteins in renal tissues were evaluated by Western blotting and immunohistochemistry. Results The results showed that QHYS significantly reduced 24-h proteinuria, blood urea nitrogen, kidney index, and ameliorated glomerular hypertrophy and collagen fiber deposition in the kidney of DN rats. Importantly, QHYS significantly downregulated the expression level of lncRNA MALAT1, upregulated the expression of nephrin, the podocyte marker protein, downregulated the expression of desmin and FSP-1, and mesenchymal cell markers. Furthermore, QHYS significantly downregulated the expression levels of Wnt1, β-catenin, and active β-catenin. Conclusion Conclusively, our study revealed that QHYS significantly reduced proteinuria, alleviated renal fibrosis, and attenuated the podocyte EMT in DN rats, which may be associated with the downregulation of lncRNA MALAT1 expression and inhibition of the Wnt/β-catenin pathway.

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Traditional Chinese medicine has certain advantages in the prevention and treatment of diabetic nephropathy (DN); thus, Chinese medicine therapy is considered as a promising strategy for treating DN. Here, the diabetic nephropathy model was established and intervened with Tangshen Decoction to explore its repair effect on diabetic kidney injury and the mechanism of autophagy. Different doses (10, 20 g·kg−1) of Tangshen Decoction (so-called Tangshen Jian, TSJ) or metformin were used to intervene for 16 weeks. The body weight (BW) and fasting blood glucose (FBG) of rats in each group were regularly monitored; a urine protein test kit (CBB method) was used to detect changes in urine protein (UP) content. The serum biochemical indicators, including Cr (creatinine), BUN (blood urea nitrogen), TC (total cholesterol), and TG (triglyceride), were detected by an automatic biochemical analyzer. HE (hematoxylin-eosin) staining, PAS, and electron microscopy were used to observe the podocyte damage. We showed that administration of TSJ or metformin prevented the increases in FBG level, serum Cr, BUN, TC, and TG level, and urine protein excretion in diabetic nephropathy. Simultaneously, the foot process fusion and fall-off were partially reversed after TSJ treatment. TSJ or metformin markedly upregulated the level of nephrin and podocin, accompanied by evident enhancement of podocyte autophagy and activation of p-AMPK/p-ULK1 signaling in the diabetic nephropathy. Therefore, TSJ may enhance podocyte autophagy to relieve diabetic nephropathy through modulation of p-AMPK/p-ULK1 signaling, which has important application prospects in the clinical treatment of diabetic kidney damage in the future.

BACKGROUND Salidroside, an active component from Traditional Chinese Medicine Rhodiola rosea L., has various pharmacological functions including anti-inflammatory, anti-cancer and anti-oxidative properties. However, whether salidroside plays a beneficial role in diabetic nephropathy is still unclear. PURPOSE The objective of this work was to investigate the potential roles of salidroside against diabetic nephropathy and the underlying molecular mechanisms. METHODS Streptozocin was given to obese mice to generate diabetic nephropathy animal model. Salidroside was administered to these mice and proteinuria, podocyte integrity, renal morphology and fibrosis, mitochondrial biogenesis were examined. RESULTS Our results showed that salidroside treatment greatly attenuates diabetic nephropathy as evidenced by decreased urinary albumin, blood urea nitrogen and serum creatinine. Morphological analysis indicated that salidroside improves renal structures in diabetic nephropathy. The decreases in nephrin and podocin expression were markedly reversed by salidroside. Moreover, kidney fibrosis in diabetic nephropathy mice was largely prevented by salidroside. Mechanistically, in salidroside-treated mice, the mitochondrial DNA copy and electron transport chain proteins were significantly enhanced. Meanwhile, the reduced Sirt1 and PGC-1α expression in diabetic nephropathy was almost completely counteracted in the presence of salidroside. CONCLUSIONS Our data showed that salidroside plays a beneficial role against diabetic nephropathy in mice, which probably via Sirt1/PGC-1α mediated mitochondrial biogenesis.

Diabetic nephropathy (DN) is a chronic kidney disease that develops in patients with diabetes mellitus (DM). Renal dysfunction and persistent proteinuria are the main clinical features of DN. Podocyte injury is an important cause of persistent proteinuria and diabetic kidney disease (DKD) progression. Traditional Chinese patent medicines can improve renal function by enhancing autophagy and promoting apoptosis. Keluoxin is a Chinese patent medicine that has the effect of invigorating qi and nourishing yin, activating blood, and eliminating blood stasis. Therefore, we hypothesized that Keluoxin may have a protective effect against diabetic nephropathy in rats with type 2 DM. Rats induced with diabetes through streptozocin (STZ) injection and a high-fat and high-sugar diet were treated with Keluoxin (0.63 g/kg/day) for 8 weeks, and renal function, biochemical indicators, and histopathological changes in renal tissues were observed. Immunofluorescence staining and western blot analysis were used to detect the expression of autophagy-related proteins. The results showed that Keluoxin reduced blood glucose and lipid levels, improved renal function, and alleviated renal histopathological changes in rats with DN. The therapeutic effect was similar to that of Irbesartan (15.6 mg/kg/day). It is inferred that the mechanism works through reducing the obstruction of downstream pathways of autophagy by improving the lysosomal degradation function and alleviating podocyte injury. This study demonstrates that Keluoxin could regulate autophagy in podocytes, alleviate kidney injury in rats with DN, and have a protective effect on renal function; its mechanism can thus be a potential therapy for DN.

Diabetic nephropathy (DN), a leading cause of end-stage renal disease, is associated with high morbidity and mortality rates worldwide and the development of new drugs to treat DN is urgently required. Bu-Shen-Huo-Xue (BSHX) decoction is a traditional Chinese herbal formula, made according to traditional Chinese medicine (TCM) theory, and has been used clinically to treat DN. In the present study, we established a high-fat diet/streptozotocin-induced diabetic mouse model and treated the mice with BSHX decoction to verify its therapeutic effects in vivo. Ultraperformance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS) was applied to analyze the chemical composition and active compounds of BSHX decoction. Markers of podocyte epithelial-mesenchymal transition and the Rac1/PAK1/p38MAPK signaling pathway were evaluated to investigate the mechanism underlying function of BSHX decoction. BSHX decoction effectively alleviated diabetic symptoms, according to analysis of the renal function indicators, serum creatinine, blood urea nitrogen, serum uric acid, and urinary albumin excretion rate, as well as renal histopathology and ultrastructural pathology of DN mice. We identified 67 compounds, including 20 likely active compounds, in BSHX decoction. The podocyte markers, nephrin and podocin, were down-regulated, while the mesenchymal markers, α-SMA and FSP-1, were up-regulated in DN mouse kidney; however, the changes in these markers were reversed on treatment with BSHX decoction. GTP-Rac1 was markedly overexpressed in DN mice and its levels were significantly decreased in response to BSHX decoction. Similarly, levels of p-PAK1 and p-p38MAPK which indicate Rac1 activation, were reduced on treatment with BSHX decoction. Together, our data demonstrated that BSHX decoction ameliorated renal function and podocyte epithelial-mesenchymal transition via inhibiting Rac1/PAK1/p38MAPK signaling pathway in high-fat diet/streptozotocin-induced diabetic mice. Further, we generated a quality control standard and numerous potential active compounds from BSHX decoction for DN.

Diabetic nephropathy (DN) is one of the most common causes of end stage renal disease (ESRD) in China, which requires renal replacement therapy. Recent investigations have suggested an essential role of podocyte injury in the initial stage of DN. This study investigated the potential therapeutic role of genipin, an active extract from a traditional Chinese medicine, on progression of DN in diabetic mice induced by intraperitoneally injection of streptozocin (STZ). In diabetic mice, orally administration of genipin postponed the progression of DN, as demonstrated by ameliorating body weight loss and urine albumin leakage, attenuating glomerular basement membrane thickness, restoring the podocyte expression of podocin and WT1 in diabetic mice. The protective role of genipin on DN is probably through suppressing the up-regulation of mitochondrial uncoupling protein 2 (UCP2) in diabetic kidneys. Meanwhile, through inhibiting the up-regulation of UCP2, genipin restores podocin and WT1 expression in cultured podocytes and attenuates glucose-induced albumin leakage through podocytes monolayer. Therefore, these results revealed that genipin inhibited UCP2 expression and ameliorated podocyte injury in DN mice.

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ETHNOPHARMACOLOGICAL RELEVANCE Danggui Buxue Decoction (DBD) is a classic traditional Chinese herbal formulation, composed of Astragali Radix (AR) and Angelica Sinensis Radix (ASR) in a ratio of 5:1. It is a traditional Chinese medicine classic prescription for nourishing Qi and Yin (vital energy and body fluids), and it is effective in treating various clinical diseases. Diabetic nephropathy (DN) is categorized under "thirsting," "edema," and "turbid urine" in Traditional Chinese Medicine (TCM). However, the underlying mechanisms by which DBD ameliorates diabetic nephropathy remain unclear. AIM OF THE STUDY To explore the mechanism by which Danggui Buxue Decoction (DBD) regulates podocyte autophagy in diabetic nephropathy (DN). METHODS Male db/m mice served as controls; db/db mice were divided into the model, dapagliflozin, and high/low-dose DBD groups. After 12 weeks of gavage, body weight, fasting blood glucose, urine albumin-to-creatinine ratio, 24-hour urine volume, and blood urea nitrogen were recorded. Renal autophagy was assessed by Masson staining; mRNA levels were measured by qRT-PCR; and protein expression was analyzed by Western blot. The expression of inflammatory factors in the kidney was measured by ELISA. Human renal podocytes were cultured in NG, HG, HG+Blank serum, and HG+DBD-containing serum groups for 48 hours; cell viability was measured by CCK-8, and autophagy was observed by transmission electron microscopy. Changes in autophagy protein and mRNA expression were observed after miR-27a transfection under high glucose conditions. RESULTS DBD can ameliorate renal function and reduce the degree of renal fibrosis in DN mice, enhance the mRNA expression of Beclin-1 and ULK1, and decrease the mRNA expression of Vimentin and α-SMA. This trend mirrors protein expression, and DBD also lowers renal inflammatory factors. DBD-containing serum boosts human renal podocyte viability under high glucose, protecting cells and modulating mRNA levels of Beclin-1, ULK1, P62, and PI3K, with miR-27a-mimic reversing these effects. DBD-containing serum also enhances Beclin-1, suppresses P62, and reduces the expression of p-PI3K/PI3K and p-AKT/AKT. CONCLUSION DBD Regulates Autophagy to Improve Renal Fibrosis in Diabetes via the miR-27a/PI3K/AKT Pathway.

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Membranous nephropathy (MN), one of the most common glomerular diseases, is a noninflammatory autoimmune podocyte disease. In the body, podocytes play an important role in maintaining the stability of the glomerular filtration barrier, and the degree of podocyte damage is a major determinant of MN prognosis because of the limited ability of podocytes to repair and regenerate. Although several therapeutic modalities for inhibiting podocyte damage are currently available, most lead to adverse, nonspecific systemic effects; therefore, newer drugs are necessary to target podocyte damage in MN. Traditional Chinese medicine (TCM) plays an important role in the treatment of primary MN in China. This study reviews the role of podocytes and the mechanisms of podocyte injury in MN glomeruli and summarizes the current status of complement-mediated therapy, the clinical efficacy of TCM in the treatment of primary MN, and the potential mechanisms regulating podocyte injury to support the development of new therapeutic strategies.

ETHNOPHARMACOLOGICAL RELEVANCE Diabetic nephropathy (DN) is a typical chronic microvascular complication of diabetes, characterized by proteinuria and a gradual decline in renal function. At present, there are limited clinical interventions aimed at preventing the progression of DN to end-stage renal disease (ESRD). However, Chinese herbal medicine presents a distinct therapeutic approach that can be effectively combined with conventional Western medicine treatments to safeguard renal function. This combination holds considerable practical implications for the treatment of DN. AIM OF THE STUDY This review covers commonly used Chinese herbal remedies and decoctions applicable to various types of DN, and we summarize the role played by their active ingredients in the treatment of DN and their mechanisms, which includes how they might improve inflammation and metabolic abnormalities to provide new ideas to cope with the development of DN. MATERIALS AND METHODS With the keywords "diabetic nephropathy," "Chinese herbal medicine," "clinical effectiveness," and "bioactive components," we conducted an extensive literature search of several databases, including PubMed, Web of Science, CNKI, and Wanfang database, to discover studies on herbal formulas that were effective in slowing the progression of DN. The names of the plants covered in the review have been checked at MPNS (http://mpns.kew.org). RESULTS This review demonstrates the superior total clinical effective rate of combining Chinese herbal medicines with Western medicines over the use of Western medicines alone, as evidenced by summarizing the results of several clinical trials. Furthermore, the review highlights the nephroprotective effects of seven frequently used herbs exerting beneficial effects such as podocyte repair, anti-fibrosis of renal tissues, and regulation of glucose and lipid metabolism through multiple signaling pathways in the treatment of DN. CONCLUSIONS The potential of herbs in treating DN is evident from their excellent effectiveness and the ability of different herbs to target various symptoms of the condition. However, limitations arise from the deficiencies in interfacing with objective bioindicators, which hinder the integration of herbal therapies into modern medical practice. Further research is warranted to address these limitations and enhance the compatibility of herbal therapies with contemporary medical standards.

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Background Diabetes is a prevalent chronic metabolic disorder, and the rising rates of this condition, along with its complications, significantly threaten public health. Traditional treatments for diabetes have certain limitations in practical applications, and it is particularly important to find new, effective treatments with fewer side effects. With a long history and rich experience, traditional Chinese medicine (TCM) effectively treats diabetes. Methods Data from randomized controlled trials concerning TCM and its effects on diabetes were gathered and analyzed from various databases. A meta-analysis was conducted on the 58 selected articles, and the potential mechanisms of action of the active ingredients in TCM were examined using network pharmacology techniques. Results Meta-analysis of 58 randomized trials (n=7,318) demonstrated significant improvements in fasting glucose (MD=-0.53 mmol/L [-0.67,-0.39], P<0.00001), HbA1c (MD=-0.40% [-0.61,-0.20], P = 0.0001), and insulin resistance (HOMA-IR: MD=-0.90 [-1.51,-0.29], P = 0.004), alongside favorable lipid modulation (LDL: MD=-0.14 mmol/L, P = 0.0002). Network pharmacology revealed six core herbs (Astragalus membranaceus, Coptis chinensis, etc.) targeting 32 hub genes (AKT1, IL1B, PPARG, etc.) through three key pathways: insulin signaling (PI3K-AKT), inflammatory regulation (TNF/IL-17), and oxidative stress response (HIF-1/NRF2 axis). The polypharmacological effects were mediated by multi-component interactions involving quercetin, kaempferol, and stigmasterol. Conclusion TCM has demonstrated considerable effectiveness in managing diabetes. Through meta-analysis and network pharmacology research, this translational study establishes Level 1a evidence for TCM’s antidiabetic efficacy while decoding its systems-level mechanisms. The integrated methodology provides a paradigm for evaluating complex herbal interventions in metabolic disorders. Systematic Review Registration https://www.crd.york.ac.uk/PROSPERO, identifier CRD42024572433.

ETHNOPHARMACOLOGICAL RELEVANCE Membranous nephropathy (MN) is a primary glomerular disease and a major cause of nephrotic syndrome in adults. According to traditional Chinese medicine (TCM) theory, "dampness" is considered a key pathogenic factor. Sanqi Qushi Formula (SQQS), developed by adding dampness-eliminating herbs to Sanqi Oral Liquid, has been shown to improve remission rates in idiopathic MN, although its pharmacological mechanisms are not yet fully clarified. AIM OF THE STUDY This study investigated whether SQQS attenuates podocyte injury in MN by inhibiting endoplasmic reticulum stress (ERS)-induced ferroptosis. METHODS The main components of SQQS were identified and quantified by widely targeted metabolomics. A passive Heymann nephritis (PHN) rat model was established via intravenous injection of anti-Fx1A serum, followed by oral SQQS intervention. Renal function and histopathology were evaluated, and underlying mechanisms were investigated by integrating transcriptomic and proteomic analyses. Structural changes in the endoplasmic reticulum and mitochondria were assessed by transmission electron microscopy. The regulatory effect of SQQS on ERS-induced ferroptosis was further examined using immunohistochemistry, flow cytometry, oxidative stress assays, and related molecular analyses. In vitro, immortalized mouse podocytes MPC-5 were injured with zymosan-activated serum and treated with SQQS-containing serum, the ferroptosis inhibitor Fer-1, the ERS inhibitor 4-PBA, or the ERS activator tunicamycin. The protective effects of SQQS were validated by Western blotting and immunofluorescence. RESULTS SQQS treatment significantly improved renal function, reduced proteinuria, and alleviated glomerular injury in PHN rats. Histopathological and ultrastructural analyses revealed that SQQS reduced glomerular IgG deposition, basement membrane thickening, fibrosis, and podocyte injury. SQQS dose-dependently restored the expression of key podocyte structural proteins (Nephrin, Synaptopodin, Podocin, α-actinin-4, WT-1) and decreased Desmin levels. Integrated transcriptomic and proteomic analyses identified ERS and ferroptosis as central pathways regulated by SQQS. SQQS suppressed ERS markers (GRP78, p-PERK, p-eIF2α, ATF4, CHOP) and pro-ferroptotic proteins (ACSL4, CHAC1), upregulated antioxidant proteins (GPX4, xCT), reduced iron deposition, and attenuated oxidative stress. In vitro, SQQS-containing serum protected MPC-5 podocytes from ZAS-induced ferroptosis by inhibiting ERS, restoring podocyte marker expression, and enhancing antioxidant defenses. CONCLUSION SQQS ameliorates MN by inhibiting the ERS-ferroptosis axis, providing experimental and theoretical support for the application of TCM in the treatment of MN.

ETHNOPHARMACOLOGICAL RELEVANCE Traditional Chinese medicine (TCM) has been applied to diabetic kidney disease (DKD). A large number of animal trials each year focus on TCM for DKD, but the evidence for these preclinical studies is not clear. AIM OF THE STUDY The aim of this study was to study the therapeutic effect of Jiedu Tongluo Baoshen formula (JTBF) on DKD proteinuria and renal protection. At the same time, it is verified that JTBF can reduce podocyte injury by enhancing autophagy function, and then achieve the effect of proteinuria. MATERIALS AND METHODS We use high performance liquid chromatography to detect and analyze the fingerprint of JTBF to find the chemical composition. Subsequently, we constructed a DKD rat model induced by high-fat diet and streptozocin (HFD + STZ). Urine and blood biochemical automatic analyzer were used to detect 24-hour urine protein quantification (24h-UP) and renal function. The renal pathological changes were observed by H&E and transmission electron microscopy (TEM), and the levels of autophagy-related proteins and mRNA in podocytes were detected by immunohistochemistry, RT-qPCR and Western Blot. The chemical composition of JTBF was screened from traditional Chinese medicine systems pharmacol (TCMSP) and PubChem databases, and the potential targets and associated pathways of JTBF were predicted using kyoto encyclopedia of genes and genomes (KEGG) and protein-protein interaction (PPI) network analysis in network pharmacology, and confirmed in animal experiments and histopathological methods. RESULTS We discovered 77 active ingredients of JTBF. Through animal experiments, it was found that JTBF reduced 24h-UP and promoted the expression of podocin, nephrin, and WT-1 in podocytes, thereby reducing podocyte damage. At the same time, JTBF activates the expression of podocyte autophagy-related proteins (beclin-1, LC3 and P62). Subsequently, through network pharmacology predictions, 208 compounds were obtained from JTBF, and phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin (PI3K/Akt/mTOR) was a potential signal pathway. JTBF was obtained in DKD rat kidney tissue to inhibit the expression of PI3K, Akt and mTOR related proteins. CONCLUSIONS JTBF enhance podocyte autophagy to reduce podocyte damage, thereby effectively treating DKD proteinuria and protecting kidney function.

ETHNOPHARMACOLOGICAL RELEVANCE Sanqi Qushi Formula (SQQS), a clinically validated derivative of the Sanqi oral solution, integrates principles of traditional Chinese medicine (TCM) to treat membranous nephropathy (MN). Its efficacy in reducing proteinuria and preserving renal function has been observed in clinical practice. AIM OF THE STUDY This study aims to elucidate the therapeutic mechanisms, active components, and pathway-specific effects of SQQS in MN, providing a scientific foundation for its clinical use. MATERIALS AND METHODS The components of SQQS were analyzed using UHPLC-MS/MS. A passive Heymann nephritis (PHN) rat model was induced by intravenous injection of anti-Fx1A serum. Rats received oral SQQS for 3 weeks, and urine/serum samples were collected to evaluate renal function and chemokine levels. Renal histopathology was assessed via immunofluorescence, PASM staining, and CD68 immunostaining. Network pharmacology integrated target prediction for SQQS compounds and differentially expressed genes from the Gene Expression Omnibus (GEO) database (MN patient glomeruli). Mitogen-activated protein kinase (MEK)/extracellular signal-regulated kinase (ERK) pathway proteins and epithelial-mesenchymal transition (EMT) markers were analyzed by western blotting (WB). Molecular docking and molecular dynamics simulations evaluated compound-MEK interactions. Human glomerular podocytes were treated with SQQS-derived compounds; viability and migration were assessed using cell counting kit-8 assay and scratch assays. RESULTS UHPLC-MS/MS identified 129 compounds in SQQS. SQQS treatment significantly reduced renal injury markers, glomerular IgG deposition, and basement membrane thickening in PHN rats. GEO database analysis revealed 839 upregulated and 166 downregulated genes in MN glomeruli. Network pharmacology implicated the tumor necrosis factor (TNF) pathway, with 10 upregulated targets (e.g., MAP2K1, MMP3, CXCL10). WB confirmed SQQS suppressed MEK/ERK phosphorylation and decreased MMP3 and α-SMA levels. Renal CD68+ macrophages and associated chemokines (CXCL10, CCL20) were reduced by SQQS. Methylnissolin-3-O-glucoside, a flavonoid from Astragalus mongholicus Bunge, dose-dependently inhibited TNF-α-induced MEK/ERK activation and migration. MEK agonists reversed methylnissolin-3-O-glucoside-mediated MEK/ERK suppression. CONCLUSION SQQS ameliorates MN progression by inhibiting the MEK/ERK pathway, suppressing EMT, and reducing macrophage recruitment, with methylnissolin-3-O-glucoside as a key bioactive component.

Chronic Renal Failure (CRF) refers to the gradual decline in renal function caused by various chronic kidney diseases, eventually leading to end-stage renal failure. Yangshen Paidu Decoction (YPD) is a Traditional Chinese medicine (TCM) formula utilized in CRF treatment. This work has analyzed the effects of YPD on CRF and the specific mechanism. Network pharmacology was performed to screen effective components and targets of YPD, from which key targets and signaling pathways contributing the most to the treatment effects on CRF were determined. Subsequently, we validated the therapeutic role of YPD and the underlying pathological mechanisms using the 5/6-nephrectomy rat model. Network pharmacology analysis showed the mTOR pathway to be a pivotal mechanism underlying the effectiveness of YPD in CRF treatment. YPD significantly suppressed urine protein levels, blood urea nitrogen, and serum creatinine in 5/6 nephrectomized rats. Furthermore, YPD remarkably improved renal pathological injuries. Western blot analysis revealed that YPD enhanced autophagy and upregulated the expression of nephrin, podocin, beclin1, p-AMPK/AMPK, and p- ULK1/ULK1, and attenuated the ratios of p-mTOR/mTOR to its downstream protein phosphorylated eIF4E-binding protein (p-4EBP1). However, these effects were notably reversed by the AMPK inhibitor compound C. Our findings have demonstrated YPD to suppress the mTOR pathway and stimulate autophagy by modulating AMPK pathways, thereby mitigating podocyte injury and enhancing renal function. Our study has confirmed autophagy and the AMPK/mTOR pathway as potential targets for YPD in CRF treatment.

The increased incidence of membranous nephropathy (MN) has made it the most common pathological type of primary nephrotic syndrome in adults in China. According to the theory of Traditional Chinese Medicine (TCM), Mahuang Fuzi (Chinese ephedra and Radix Aconiti Lateralis Preparata) and Shenzhuo Decoction (MFSD) could be used to treat such diseases. We treated patients of MN with MFSD, and observed comparable efficacy to glucocorticoid and/or immunosuppressants. In this study, we observed the therapeutic effect of MFSD on the rat model of passive Heymann nephritis (PHN), a classical MN model. Our results showed that MFSD treatment significantly reduced urinary protein level and podocyte injury in PHN rats, and correspondingly improved renal pathology, with the improvement effect on MN comparable to that of Cyclosporine A (CsA) alone. To explore the potential therapeutical mechanism of MFSD, the main chemical components of MFSD were determined by High-performance liquid chromatography-mass spectrometry (HPLC-MS). There were about 30 active components of MFSD. Next, based on network pharmacology methods, we screened related targets of MSFD on MN, which provided a preliminary understanding of the MFSD bioactive compounds. The clustering analysis showed that its active site might be in the autophagy-related protein and Wnt/β-catenin pathway, which was related to podocyte injury. Finally, we observed an improvement in renal autophagy and a down-regulation of the Wnt/β-catenin pathway after MSFD treatment in a PHN rat model. According to this study, autophagy and Wnt/β-catenin pathway may be potential targets for MFSD in the treatment of MN.

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Diabetic nephropathy (DN) is a common microvascular complication of diabetes mellitus (DM). Radix Astragali (RA), a frequently used Chinese herbal medicine in the Leguminosae family, Astragalus genus, with its extracts, has been proven to be effective in DN treatment both in clinical practice and experimental studies. RA and its extracts can reduce proteinuria and improve renal function. They can improve histopathology changes including thickening of the glomerular basement membrane, mesangial cell proliferation, and injury of endothelial cells, podocytes, and renal tubule cells. The mechanisms mainly benefited from antioxidative stress which involves Nrf2/ARE signaling and the PPARγ-Klotho-FoxO1 axis; antiendoplasmic reticulum stress which involves PERK-ATF4-CHOP, PERK/eIF2α, and IRE1/XBP1 pathways; regulating autophagy which involves SIRT1/NF-κB signaling and AMPK signaling; anti-inflammation which involves IL33/ST2 and NF-κB signaling; and antifibrosis which involves TGF-β1/Smads, MAPK (ERK), p38/MAPK, JNK/MAPK, Wnt/β-catenin, and PI3K/AKT/mTOR signaling pathways. This review focuses on the clinical efficacy and the pharmacological mechanism of RA and its representative extracts on DN, and we further document the traditional uses of RA and probe into the TCM theoretical basis for its application in DN.

Glomerular diseases are major components of kidney disease, mainly manifested as podocyte disease, immune complex-associated glomerulonephritis, and renal autoimmune disease. They are the third leading cause of end-stage renal disease, especially in younger populations. Podocyte death or shedding is a key factor in the progression of glomerular diseases, and autophagy plays an important role in maintaining podocyte homeostasis. Dysfunction of autophagy has been associated with the progression of various glomerular diseases. Modulation of autophagy has been shown to play a protective role in experimental models of glomerular diseases, suggesting that autophagy is a potential therapeutic target for glomerular diseases. Traditional Chinese medicine (TCM) has unique advantages in the treatment of kidney disease. In vivo and in vitro studies have shown that TCM compounds can reduce podocyte apoptosis, inhibit inflammation, improve endoplasmic reticulum stress and oxidative stress responses, and play an active role via autophagy. This review summarizes the roles of autophagy in glomerular diseases and provides evidence that TCM compounds can regulate autophagy through different signaling pathways to ameliorate glomerular diseases.

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This study aimed to explore the protective effects of kaempferol against podocyte injury in lupus nephritis (LN) and its underlying mechanism. Human podocytes were co-cultured with immunoglobulin G (IgG) from patients with LN (IgG-LN) to simulate podocyte injury in LN. Levels of oxidative stress and inflammatory factors were measured using the different kits. The expression of podocyte injury marker proteins was detected using immunofluorescence and western blotting. Then, we explored the interactions of kaempferol with PI3K, Akt, and nuclear factor kappa B (NF-κB) by molecular docking. Furthermore, PI3K agonists (740 Y-P) and inhibitors (LY294002) were used to explore the mechanism by which kaempferol alleviates IgG-LN-induced podocyte injury. Autophagy was measured by western blotting and Monodansylcadaverine staining. The results indicated that kaempferol could protect podocyte injury induced by IgG-LN in a dose-dependent manner. Kaempferol significantly reduced oxidative stress, increased the activity of antioxidant enzymes, and reduced the levels of inflammatory factors in IgG-LN-treated podocytes. Kaempferol could bind to the active sites of PI3K, Akt, and NF-κB with high affinity, especially to NF-κB. LY294002 enhanced the ameliorative effect of kaempferol against IgG-LN-induced podocyte injury, whereas 740 Y-P weakened this protective effect. Compared to the model group, kaempferol significantly reduced the p-mTOR/mammalian target of rapamycin (mTOR) ratio and activated autophagy. Kaempferol can inhibit inflammation and oxidative stress through the regulation of PI3K/Akt/NF-κB pathway and promote autophagy by regulating the PI3K/Akt/mTOR pathway to improve IgG-LN induced podocyte injury.

Background Organ fibrosis is a common endpoint of a variety of diseases. Many studies have shown that the pathogenesis of diabetic kidney disease (DKD) is related to the excessive activation of the Wnt/β-catenin signaling pathway on podocytes, so the treatment of DKD starts from this signaling pathway. At the same time, DKD, as a metabolic disease, has many connections related to podocyte autophagy. Objectives We experimented the effects of Mahuang Fuzi and Shenzhuo decoction (MFSD) which is the combination of Mahuang Fuzi decoction and Shenzhuo decoction in traditional Chinese medicine compounds used “The Golden Chamber” in high glucose-induced podocytes, determined whether this effect was related to Wnt/β-catenin signaling pathway, and further investigated the relationship between this effect and autophagy. Methods The mice podocytes were stimulated by using 30 mmol/L of high glucose and serum containing MFSD or Wnt/β-catenin signaling pathway inhibitor DKK1 (100 ng/ml) was used to intervene podocytes before high glucose stimulation. Podocyte injury-related proteins, Wnt/β-catenin signaling pathway-related proteins, and autophagy-related proteins were detected by using western blotting and immunofluorescence analysis. Results Our results showed that DKK1 and MFSD treatment significantly upregulated the protein expressions of nephrin, podocin, podocalyxin, and podoplanin in high glucose-induced podocytes and downregulated the β-catenin protein expression. Furthermore, the protein expressions of beclin1, LC3B, and P62 were also significantly increased in high glucose-induced podocytes. Conclusion Our experiments confirmed that the destruction of podocytes in DKD is related to the excessive activation of Wnt/β-catenin signaling pathway and the inhibition of autophagy after activation. MFSD treatment can inhibit the activation of Wnt/β-catenin signaling pathway in podocytes stimulated by high glucose and helpful in reducing the podocyte injury. This protective mechanism can be related to the enhancement of podocyte autophagy by MFSD treatment.

ETHNOPHARMACOLOGICAL RELEVANCE Shen-Qi-Di-Huang decoction (SQDHD) is a renowned decoction in traditional Chinese medicine, dating back to the Qing Dynasty. SQDHD has been widely applied in treating renal diseases, including Membranous nephropathy (MN), with its proven positive clinical outcomes. Nevertheless, the precise mechanism by which SQDHD exerts its therapeutic effects on MN remains uncertain. AIM OF THE STUDY The present research aimed to observe whether SQDHD promotes podocyte autophagy by inhibiting USP14 to increase the K63 ubiquitination of Beclin1, thereby improving MN. MATERIALS AND METHODS An MN model was established in rats using Passive Heyman Nephritis (PHN) to explore the underlying mechanisms in vivo. The kidney function parameters were evaluated, and the histomorphology of glomerular tissues was examined. Autophagy-related protein expression was assessed using immunofluorescence staining and western blotting assays. Co-immunoprecipitation (Co-IP) was used to detect the K63 ubiquitination of Beclin1. MPC5 cells were treated in vitro with serum obtained from several rat groups. Subsequently, the expression of autophagy-related proteins, formation of autophagosomes, expression of USP14, and K63 ubiquitination of Beclin1 were quantified. RESULTS Our results demonstrated that SQDHD intervention reduced urinary protein levels, mitigated podocyte damage in MN model rats, and improved kidney tissue pathology. Furthermore, in vitro and in vivo data revealed that SQDHD therapy significantly increased podocyte autophagy, decreased USP14 expression, and raised Beclin1's K63 ubiquitination. CONCLUSION These results provided a scientific rationale supporting the ability of SQDHD to substantially alleviate MN progression by inducing podocyte autophagy through the inhibition of USP14 expression.

Background Diabetic kidney disease (DKD) is one of diabetic complications, which has become the leading cause of end-stage kidney disease. In addition to angiotensin-converting enzyme inhibitor/angiotensin II receptor blocker(ACEI/ARB) and sodium-glucose cotransporter-2 inhibitor (SGLT2i), traditional Chinese medicine (TCM) is an effective alternative treatment for DKD. In this study, the effect of Qufeng Tongluo (QFTL) decoction in decreasing proteinuria has been observed and its mechanism has been explored based on autophagy regulation in podocyte. Methods In vivo study, db/db mice were used as diabetes model and db/m mice as blank control. Db/db mice were treated with QFTL decoction, rapamycin, QFTL + 3-Methyladenine (3-MA), trehalose, chloroquine (CQ) and QFTL + CQ. Mice urinary albumin/creatinine (UACR), nephrin and autophagy related proteins (LC3 and p62) in kidney tissue were detected after intervention of 9 weeks. Transcriptomics was operated with the kidney tissue from model group and QFTL group. In vitro study, mouse podocyte clone-5 (MPC-5) cells were stimulated with hyperglycemic media (30 mmol/L glucose) or cultured with normal media. High-glucose-stimulated MPC-5 cells were treated with QFTL freeze-drying powder, rapamycin, CQ, trehalose, QFTL+3-MA and QFTL + CQ. Cytoskeletal actin, nephrin, ATG-5, ATG-7, Beclin-1, cathepsin L and cathepsin B were assessed. mRFP-GFP-LC3 was established by stubRFP-sensGFP-LC3 lentivirus transfection. Results QFTL decoction decreased the UACR and increased the nephrin level in kidney tissue and high-glucose-stimulated podocytes. Autophagy inhibitors, including 3-MA and chloroquine blocked the effects of QFTL decoction. Further study showed that QFTL decoction increased the LC3 expression and relieved p62 accumulation in podocytes of db/db mice. In high-glucose-stimulated MPC-5 cells, QFTL decoction rescued the inhibited LC3 and promoted the expression of ATG-5, ATG-7, and Beclin-1, while had no effect on the activity of cathepsin L and cathepsin B. Results of transcriptomics also showed that 51 autophagy related genes were regulated by QFTL decoction, including the genes of ATG10, SCOC, ATG4C, AMPK catalytic subunit, PI3K catalytic subunit, ATG3 and DRAM2. Conclusion QFTL decoction decreased proteinuria and protected podocytes in db/db mice by regulating autophagy.

Tripterygium glycoside (TG) is a traditional Chinese medicine extract with immunosuppressive, anti-inflammatory and anti-renal fibrosis effects. Epithelial-mesenchymal transition (EMT) and cell apoptosis are considered to be the major cause of podocyte injury in diabetic kidney disease (DKD). However, it remains unknown as to whether TG is able to alleviate podocyte injury to prevent DKD progression. Therefore, the present study aimed to clarify the podocyte protective effects of TG on DKD. TG, Twist1 small interfering RNA (siRNA) and Twist1 overexpression vector were added to DKD mouse serum-induced podocytes in vitro. Autophagic and EMT activities were evaluated by immunofluorescence staining and western blot analysis. Apoptotic activity was evaluated by Annexin V-FITC/PI flow cytometric analysis. The results revealed that after treatment with DKD mouse serum, autophagy was decreased, whereas EMT and apoptotic rate were increased, in podocytes. In addition, Twist1 expression was increased in DKD-induced podocytes. Furthermore, following Twist1-small interfering RNA transfection, the DKD-induced podocyte EMT and apoptotic rate were markedly reduced, indicating that Twist1 may be a promising therapeutic target for DKD. The present results also revealed that overexpression of Twist1 increased podocyte apoptosis, although this was decreased after TG treatment, indicating that TG may exhibit a protective effect on podocytes by inhibiting the Twist1 signaling pathway. After the addition of 3-benzyl-5-((2-nitrophenoxy) methyl)-dihydrofuran-2(3H)-one, an activator of mTORC1, the effects of TG on podocyte EMT, apoptosis and the autophagy were reversed. These findings indicated that TG may alleviate EMT and apoptosis by upregulating autophagy through the mTOR/Twist1 signaling pathway in DKD.

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Introduction Yu Nu compound (YNJ) is a traditional Chinese medicine widely utilized to treat type 2 diabetes possibly through mediating autophagy. Abnormal podocyte autophagy and apoptosis could result in podocyte loss in diabetics nephropathy (DN). The mechanism of Yu Nu compound in DN is still unclear. Therefore, the study aims to investigate the effects of Yu Nu compound and analyze the potential mechanism. Methods Goto-Kakizaki (GK) rats were administered using YNJ with different doses once a day by gavage for 4 weeks. The renal cortex injury was observed by HE staining and electron microscope. Cell apoptosis of renal cortex was analyzed by TUNNEL staining. The mTOR, autophagy-related proteins and apoptosis-related proteins were detected by Western blot or real-time PCR in vivo and vitro. MPC5 cells were exposed to high glucose (HG, 30mM) for 12h to simulate podocyte injury in DN. MPC5 cells were treated by serum containing YNJ with different dosages. Cell activities and apoptosis were, respectively, detected through Cell Counting Kit-8 (CCK8) assay and flow cytometry. Results The results showed that the medium dose of YNJ had better effects on decreasing blood glucose and improving renal injury in GK rats, followed by decreasing mTOR levels. The autophagy levels were enhanced in renal cortex, accompanied with the increase of cell apoptosis in vivo. Besides, the proteins regulating autophagy and apoptosis were significantly modulated by YNJ in GK rats. Then, we found that the decreasing endogenous mTOR could reverse the effects of YNJ on podocyte apoptosis and autophagy in vivo. Discussion The study suggested that YNJ recovered normal autophagy and suppressed apoptosis through regulating mTOR. The maintenance of normal basal autophagic activity possibly based on the effect of YNJ on multiple target was essential for maintaining podocyte function.

Abstract Background Qing-Re-Xiao-Zheng-Yi-Qi formula (QRXZYQF), based on the "Shen-Luo-Zheng-Jia" principles of traditional Chinese medicine, has been reported to reduce 24-hour urinary total protein in diabetic kidney disease (DKD) patients, slow disease progression, and improve podocyte injury. This study aims to explore the mechanisms of QRXZYQF in improving podocyte injury. Methods Diabetes was induced in male C57BL/6J mice by intraperitoneal injection of streptozotocin (STZ). After 12 weeks of QRXZYQF, blood glucose, blood urea nitrogen, serum creatinine, microalbumin, and the urinary albumin-to-creatinine ratio (UACR) were monitored. Renal pathological changes were evaluated using hematoxylin and eosin (H&E), Masson, and periodic acid–Schiff (PAS) staining. RNA sequencing (RNA-Seq) was performed to identify differences in renal mRNA expression and enrichment pathways. The involvement of autophagy–lysosomal and NOD-like receptor pathways was examined by western blotting and immunofluorescence in renal tissues and cultured podocytes. Results Following 12 weeks of QRXZYQF, renal function improved and ECM accumulation and glomerulosclerosis were markedly reduced. Subsequently, RNA-Seq analysis showed that the autophagy–lysosomal and NOD-like receptor signaling pathways were the potential pathways involved in the mechanism of QRXZYQF. Moreover, QRXZYQF reduced the levels of NLRP3, apoptosis-associated speck-like protein (ASC), as well as Caspase-1 in vitro. Furthermore, we performed interventions using the lysosomal membrane-permeabilizing agent (L-leucyl-L-leucine-O-methylester) and found that QRXZYQF inhibits NLRP3 overexpression by protecting lysosomal membranes and preventing the leakage of Cathepsin B (CB) into the cytoplasm. Conclusions QRXZYQF inhibits NLRP3-mediated podocyte pyroptosis by stabilizing lysosomal membranes, providing insights into its protective mechanism and potential therapeutic targets for DKD. Graphical Abstract

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Diabetic kidney disease (DKD) is a leading cause of chronic kidney disease worldwide, and podocyte ferroptosis plays a crucial role in its pathogenesis. Hirsutine (HS) reduces blood glucose levels and improve insulin resistance in diabetic mice, suggesting its potential use in diabetes treatment. Here, we established a db/db mouse model of DKD and administered HS for 8 weeks. We found that HS decreased the concentrations of iron, reactive oxygen species (ROS), and malondialdehyde (MDA) in renal tissues. Furthermore, HS treatment restored mitochondrial morphology, increased Glutathione Peroxidase 4(GPX4) levels, and decreased p53 levels, alleviating podocyte loss in the DKD mouse model. However, the effects of HS were reversed by the p53 activator Nutlin-3. Therefore, we propose HS may mitigate podocyte injury in DKD by regulating the p53/GPX4 pathway, providing a novel strategy for targeting podocyte ferroptosis in DKD.

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Hyperglycemia significantly decreases 3',5'-cyclic guanosine monophosphate (cGMP)-dependent pathway activity in the kidney. A well-characterized downstream signaling effector of cGMP is cGMP-dependent protein kinase G (PKG), exerting a wide range of downstream effects, including vasodilation and vascular smooth muscle cells relaxation. In podocytes that are exposed to high glucose concentrations, crosstalk between the protein deacetylase sirtuin 1 (SIRT1) and adenosine monophosphate-dependent protein kinase (AMPK) decreased, attenuating insulin responsiveness and impairing podocyte function. The present study examined the effect of enhancing cGMP-dependent pathway activity on SIRT1-AMPK crosstalk in podocytes under hyperglycemic conditions. We found that enhancing cGMP-dependent pathway activity using a cGMP analog was associated with increases in SIRT1 protein levels and activity, with a concomitant increase in the degree of AMPK phosphorylation. The beneficial effects of enhancing cGMP-dependent pathway activity on SIRT1-AMPK crosstalk also included improvements in podocyte function. Based on our findings, we postulate an important role for SIRT1-AMPK crosstalk in the regulation of albumin permeability in hyperglycemia that is strongly associated with activity of the cGMP-dependent pathway.

Podocyte, the gatekeeper of the glomerular filtration barrier, is a primary target for growth factor and Ca2+ signaling whose perturbation leads to proteinuria. However, the effects of insulin action on store-operated Ca2+ entry (SOCE) in podocytes remain unknown. Here, we demonstrated that insulin stimulates SOCE by VAMP2-dependent Orai1 trafficking to the plasma membrane. Insulin-activated SOCE triggers actin remodeling and transepithelial albumin leakage via the Ca2+-calcineurin pathway in podocytes. Transgenic Orai1 overexpression in mice causes podocyte fusion and impaired glomerular filtration barrier. Conversely, podocyte-specific Orai1 deletion prevents insulin-stimulated SOCE, synaptopodin depletion, and proteinuria. Podocyte injury and albuminuria coincide with Orai1 upregulation at the hyperinsulinemic stage in diabetic (db/db) mice, which can be ameliorated by the suppression of Orai1-calcineurin signaling. Our results suggest that tightly balanced insulin action targeting podocyte Orai1 is critical for maintaining filter integrity, which provides novel perspectives on therapeutic strategies for proteinuric diseases, including diabetic nephropathy. Perturbations of Ca2+ signaling in podocytes may deteriorate kidney function and eventually lead to proteinuria. Here the authors show that insulin can affect the function of the calcium regulator Ora1 in podocytes, which is critical for maintaining kidney filter integrity.

Podocytes are sensitive to insulin, which governs the functional and structural integrity of podocytes that are essential for proper function of the glomerular filtration barrier. Lysosomes are acidic organelles that are implicated in regulation of the insulin signaling pathway. Cathepsin D (CTPD) and lysosome-associated membrane protein 1 (LAMP1) are major lysosomal proteins that reflect the functional state of lysosomes. However, the effect of insulin on lysosome activity and role of lysosomes in the regulation of insulin-dependent glucose uptake in podocytes are unknown. Our studies showed that the short-term incubation of podocytes with insulin decreased LAMP1 and CTPD mRNA levels. Insulin and bafilomycin A1 reduced both the amounts of LAMP1 and CTPD proteins and activity of CTPD, which were associated with a decrease in the fluorescence intensity of lysosomes that were labeled with LysoTracker. Bafilomycin A1 inhibited insulin-dependent endocytosis of the insulin receptor and increased the amounts of the insulin receptor and glucose transporter 4 on the cell surface of podocytes. Bafilomycin A1 also inhibited insulin-dependent glucose uptake despite an increase in the amount of glucose transporter 4 in the plasma membrane of podocytes. These results suggest that lysosomes are signaling hubs that may be involved in the coupling of insulin signaling with the regulation of glucose uptake in podocytes. The dysregulation of this mechanism can lead to the dysfunction of podocytes and development of insulin resistance.

BACKGROUND/AIMS Rehmanniae Radix (RR) and Cornus officinalis (CO) are a typical herbal pair used to treat diabetic nephropathy (DN) in clinical practice. DN can be effectively treated by catalpol (Cat) and loganin (Log), the main active components of RR and CO respectively, through combating apoptosis, oxidative stress and inflammation. Herein, a spontaneous DN and podocyte injury model induced by advanced glycation end products (AGEs), i.e. KK-Ay mice, was used to explore the cooperative effects of Log and Cat on DN and the mechanism targeting the AGEs-RAGE (receptor for AGE) pathway. METHODS AND KEY FINDINGS Log and Cat alone or in combination mitigated diabetic symptoms, decreased the level of fasting blood glucose, and increased that of serum insulin. The two drugs alone or in combination protected renal function from damage, prevented extracellular matrix hyperplasia and glycogen deposition, as well as alleviated the loss of podocytes detected by histological assay and immunohistochemistry. Flow cytometry revealed that Log and Cat alone or in combination relieved the apoptosis of AGEs-induced podocytes in vitro. Silencing RAGE by RNA interference played a protective role in podocyte apoptosis, whereas overexpression of it worked oppositely. Western blot exhibited that Log and Cat alone or in combination inhibited the activation of RAGE/p38 MAPK/p65 NF-κB and RAGE/Nox4/p65 NF-κB pathways in podocytes. The inhibitory effects of drug combination were more evident than those of individual treatments. SIGNIFICANCE Log and Cat cooperatively resisted the apoptosis of podocytes upon DN by targeting AGEs-RAGE and its downstream pathways p38 MAPK and Nox4.

Our prior investigations have established that Inonotus obliquus (Chaga) possesses hypoglycemic effects. Persistent hyperglycemia is known to precipitate renal function abnormalities. The functionality of the kidneys is intricately linked to the levels of cyclic guanosine-3',5'-monophosphate (cGMP), which are influenced by the activities of nitric oxide synthase (NOS) and phosphodiesterase (PDE). Enhanced cGMP levels can be achieved either through the upregulation of NOS activity or the downregulation of PDE activity. The objective of the current study is to elucidate the effects of Chaga on disorders of glucolipid metabolism and renal abnormalities in rats with type 2 diabetes mellitus (T2DM), while concurrently examining the NOS-cGMP-PDE5 signaling pathway. A model of T2DM was developed in rats using a high-fat diet (HFD) combined with streptozotocin (STZ) administration, followed by treatment with Chaga extracts at doses of 50 and 100 mg·kg-1 for eight weeks. The findings revealed that Chaga not only mitigated metabolic dysfunctions, evidenced by improvements in fasting blood glucose, total cholesterol, triglycerides, and insulin resistance, but also ameliorated renal function markers, including serum creatinine, urine creatinine (UCr), blood urea nitrogen, 24-h urinary protein, and estimated creatinine clearance. Additionally, enhancements in glomerular volume, GBM thickness, podocyte foot process width (FPW), and the mRNA and protein expressions of podocyte markers, such as nephrin and wilms tumor-1, were observed. Chaga was found to elevate cGMP levels in both serum and kidney tissues by increasing mRNA and protein expressions of renal endothelial NOS and neural NOS, while simultaneously reducing the expressions of renal inducible NOS and PDE5. In summary, Chaga counteracts HFD/STZ-induced glucolipid metabolism and renal function disturbances by modulating the NOS-cGMP-PDE5 signaling pathway. This research supports the potential application of Chaga in the clinical prevention and treatment of T2DM and diabetic nephropathy (DN), with cGMP serving as a potential therapeutic target.

ETHNOPHARMACOLOGICAL RELEVANCE In traditional Chinese medicine, a long term of improper diet causes the Dampness and disturbs Zang-Fu's functions including Kidney deficiency. Atractylodes lancea (Atr) and Magnolia officinalis (Mag) as a famous herb pair are commonly used to transform Dampness, with kidney protection. AIM OF THE STUDY To explore how Atr and Mag protected against insulin signaling impairment in glomerular podocytes induced by high dietary fructose feeding, a major contributor for insulin resistance in glomerular podocyte dysfunction. MATERIALS AND METHODS Liquid chromatography-tandem mass spectrometry (LC-MS/MS) analyze constituents of Atr and Mag. Rat model was induced by 10% fructose drinking water in vivo, and heat-sensitive human podocyte cells (HPCs) were exposed to 5 mM fructose in vitro. Animal or cultured podocyte models were treated with different doses of Atr, Mag or Atr and Mag combination. Western blot, qRT-PCR and immunofluorescence assays as well as other experiments were performed to detect adiponectin receptor protein 1 (AdipoR1), protein kinase B (AKT), Sirt1, p53 and miR-221 levels in rat glomeruli or HPCs, respectively. RESULTS Fifty-five components were identified in Atr and Mag combination. Network pharmacology analysis indicated that Atr and Mag combination might affect insulin signaling pathway. This combination significantly improved systemic insulin resistance and prevented glomerulus morphological damage in high fructose-fed rats. Of note, high fructose decreased IRS1, AKT and AdipoR1 in rat glomeruli and cultured podocytes. Further data from cultured podocytes with Sirt1 inhibitor/agonist, p53 agonist/inhibitor, or miR-221 mimic/inhibitor showed that high fructose downregulated Sirt1 to stimulate p53-driven miR-221, resulting in insulin signaling impairment. Atr and Mag combination effectively increased Sirt1, and decreased p53 and miR-221 in in vivo and in vitro models. CONCLUSIONS Atr and Mag combination improved insulin signaling in high fructose-stimulated glomerular podocytes possibly through upregulating Sirt1 to inhibit p53-driven miR-221. Thus, the regulation of Sirt1/p53/miR-221 by this combination may be a potential therapeutic approach in podocyte insulin signaling impairment.

Emerging evidence suggests that GSK3β, a redox-sensitive transducer downstream of insulin signaling, acts as a convergent point for myriad pathways implicated in kidney injury, repair, and regeneration. However, its role in diabetic kidney disease remains controversial. In cultured glomerular podocytes, exposure to a milieu of type 2 diabetes elicited prominent signs of podocyte injury and degeneration, marked by loss of homeostatic marker proteins like synaptopodin, actin cytoskeleton disruption, oxidative stress, apoptosis, and stress-induced premature senescence, as shown by increased staining for senescence-associated β-galactosidase activity, amplified formation of γH2AX foci, and elevated expression of mediators of senescence signaling, like p21 and p16INK4A. These degenerative changes coincided with GSK3β hyperactivity, as evidenced by GSK3β overexpression and reduced inhibitory phosphorylation of GSK3β, and were averted by tideglusib, a highly-selective small molecule inhibitor of GSK3β. In agreement, post-hoc analysis of a publicly-available glomerular transcriptomics dataset from patients with type 2 diabetic nephropathy revealed that the curated diabetic nephropathy-related gene set was enriched in high GSK3β expression group. Mechanistically, GSK3β-modulated nuclear factor Nrf2 signaling is involved in diabetic podocytopathy, because GSK3β knockdown reinforced Nrf2 antioxidant response and suppressed oxidative stress, resulting in an improvement in podocyte injury and senescence. Conversely, ectopic expression of the constitutively active mutant of GSK3β impaired Nrf2 antioxidant response and augmented oxidative stress, culminating in an exacerbated diabetic podocyte injury and senescence. Moreover, IRS-1 was found to be a cognate substrate of GSK3β for phosphorylation at IRS-1S332, which negatively regulates IRS-1 activity. GSK3β hyperactivity promoted IRS-1 phosphorylation, denoting a desensitized insulin signaling. Consistently, in vivo in db/db mice with diabetic nephropathy, GSK3β was hyperactive in glomerular podocytes, associated with IRS-1 hyperphosphorylation, impaired Nrf2 response and premature senescence. Our finding suggests that GSK3β is likely a novel therapeutic target for treating type 2 diabetic glomerular injury.

The podocyte is one of the main components of the glomerular filtration barrier in the kidney, and its injury may contribute to proteinuria, glomerulosclerosis and eventually kidney failure. C-peptide, a cleavage product of proinsulin, shows therapeutic potential for treating diabetic nephropathy (DN). The aim of this study was to investigate the effect of C-peptide on high glucose-induced podocyte dysfunction. In the present study, we found that the protective effects of islet transplantation were superior to simple insulin therapy for the treatment of DN in streptozotocin (STZ)-treated rats. And such superiority may due to the function of C-peptide secreted at the implanted site. Based on this background, we determined that the application of C-peptide significantly prevented high glucose-induced podocyte injury by increasing the expression of nephrin and synaptopodin. Meanwhile, C-peptide suppressed high glucose-induced epithelial-mesenchymal transition (EMT) and renal fibrosis via decreasing the expression of snail, vimentin, α-smooth muscle actin (α-SMA) and connective tissue growth factor (CTGF). Moreover, the Notch and transforming growth factor-β (TGF-β) signaling pathways were activated by high glucose, and treatment with C-peptide down-regulated the expression of the Notch signaling molecules Notch 1 and Jagged 1 and the TGF-β signaling molecule TGF-β1. These findings suggested that C-peptide might serve as a novel treatment method for DN and podocyte dysfunction.

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Podocytes, specialized epithelial cells that envelop the glomerular capillaries, play a pivotal role in maintaining renal health. The current description and quantification of features on pathology slides are limited, prompting the need for innovative solutions to comprehensively assess diverse phenotypic attributes within Whole Slide Images (WSIs). In particular, understanding the morphological characteristics of podocytes, terminally differentiated glomerular epithelial cells, is crucial for studying glomerular injury. This paper introduces the Spatial Pathomics Toolkit (SPT) and applies it to podocyte pathomics. The SPT consists of three main components: (1) instance object segmentation, enabling precise identification of podocyte nuclei; (2) pathomics feature generation, extracting a comprehensive array of quantitative features from the identified nuclei; and (3) robust statistical analyses, facilitating a comprehensive exploration of spatial relationships between morphological and spatial transcriptomics features.The SPT successfully extracted and analyzed morphological and textural features from podocyte nuclei, revealing a multitude of podocyte morphomic features through statistical analysis. Additionally, we demonstrated the SPT's ability to unravel spatial information inherent to podocyte distribution, shedding light on spatial patterns associated with glomerular injury. By disseminating the SPT, our goal is to provide the research community with a powerful and user-friendly resource that advances cellular spatial pathomics in renal pathology. The implementation and its complete source code of the toolkit are made openly accessible at https://github.com/hrlblab/spatial_pathomics.

Background: Automated podocyte foot process quantification is vital for kidney research, but the established "Automatic Morphological Analysis of Podocytes" (AMAP) method is hindered by high computational demands, a lack of a user interface, and Linux dependency. We developed AMAP-APP, a cross-platform desktop application designed to overcome these barriers. Methods: AMAP-APP optimizes efficiency by replacing intensive instance segmentation with classic image processing while retaining the original semantic segmentation model. It introduces a refined Region of Interest (ROI) algorithm to improve precision. Validation involved 365 mouse and human images (STED and confocal), benchmarking performance against the original AMAP via Pearson correlation and Two One-Sided T-tests (TOST). Results: AMAP-APP achieved a 147-fold increase in processing speed on consumer hardware. Morphometric outputs (area, perimeter, circularity, and slit diaphragm density) showed high correlation (r>0.90) and statistical equivalence (TOST P<0.05) to the original method. Additionally, the new ROI algorithm demonstrated superior accuracy compared to the original, showing reduced deviation from manual delineations. Conclusion: AMAP-APP democratizes deep learning-based podocyte morphometry. By eliminating the need for high-performance computing clusters and providing a user-friendly interface for Windows, macOS, and Linux, it enables widespread adoption in nephrology research and potential clinical diagnostics.

Diabetic nephropathy (DN) is a severe chronic microvascular complications of diabetes mellitus and the leading cause of end-stage renal disease. Although many therapeutic approaches have been developed for treatment of DN, there is still a risk of disease progression. Wenshen Jianpi Decoction (WSJPD), a traditional Chinese medicine, have demonstrated considerable clinical efficacy in treating DN. However, the therapeutic mechanisms of WSJPD remain unclear. The study aimed to assess the potential anti-renal injury effects of WSJPD as well as the underlying mechanism. Network pharmacology was performed to predict the potential targets of WSJPD in DN. Subsequently, HE, PAS, Masson staining, TEM and ELISA were used to assess the effects of WSJPD on renal injury in DN rats. Transcriptomics, metabolomics, and qRT-PCR were employed to analyze the potential mechanism. Network pharmacology analysis revealed that WSJPD might exert a beneficial impact on renal inflammation via the AGE-RAGE signaling pathway. The results of animal experiments indicated that WSJPD had a renoprotective effect by renal dysfunction improvement, inflammation inhibition, glycogen accumulation and podocyte injury suppression.Transcriptomic measures discovered that 212 genes were up-regulated in the DN model group and down-regulated in the WSJPD group. Conversely, 80 genes were down-regulated in the DN model group and up-regulated in the WSJPD group. Moreover, 14 metabolites from rat serum samples were identified as candidate biomarkers. Further analysis demonstrated that WSJPD ameliorated the metabolic disorders and exerted a protective effect by up-regulation of uridine-mediated pyrimidine metabolism, inhibition of inflammation mediated by the NF-κB and TNF signaling pathways, and inhibition of oxidative stress and inflammation mediated by the AGE-RAGE signaling pathway. Our findings showed that WSJPD ameliorated renal damage in DN model rats by regulating kidney pyrimidine metabolism, inflammation-related pathways, and AGE-RAGE signaling pathway. The potential renoprotective effects were verified by histopathology, ELISA, and qRT-PCR. This study not only support a foundation for interpretation of the therapeutic effects of WSJPD on renal lesions, but also provide data support for further explorations of the novel compound preparations to improve renal function in DN.

Although recent progress provides mechanistic insights into diabetic nephropathy (DN), effective treatments remain scarce. DN, characterized by proteinuria and a progressive decline in renal function, primarily arises from podocyte injury, which impairs the glomerular filtration barrier. Wogonoside, a bioactive compound from the traditional Chinese herb Scutellaria baicalensis, has not been explored for its role in DN. This study aimed to investigate the therapeutic effects of wogonoside on podocyte injury in DN and its molecular mechanisms. The effects of wogonoside were examined using HFD/STZ-induced DN mouse models and high glucose (HG)-induced MPC-5 cells. Oxidative stress and inflammation markers were analyzed via Western blot and RT-qPCR. Wogonoside targets were identified through DARTS-MS and validated by SPR, molecular docking, alanine scanning, and CETSA. RNA-Seq analysis was employed to identify downstream targets, and the p65-MMP28 axis was explored through p65 knockdown and overexpression studies. The regulatory effect of p65 on Mmp28 was confirmed through dual-luciferase reporter assays and ChIP-qPCR. Wogonoside treatment significantly reduced oxidative stress and inflammation in vivo and in vitro. Mechanistic studies identified p65 as a direct target of wogonoside, with SPR confirming a strong binding affinity (K This study highlights wogonoside as a promising candidate for the treatment of podocyte injury in DN by targeting the NF-κB p65-MMP28 signaling axis. These findings provide novel insights into wogonoside's therapeutic potential and its molecular mechanisms, paving the way for its further development as a DN intervention.

Accumulating evidence reveals that both inflammation and lymphocyte dysfunction play a vital role in the development of diabetic nephropathy (DN). Hyperoside (HPS) or quercetin-3-O-galactoside is an active flavonoid glycoside mainly found in the Chinese herbal medicine Tu-Si-Zi. Although HPS has a variety of pharmacological effects, including anti-oxidative and anti-apoptotic activities as well as podocyte-protective effects, its underlying anti-inflammatory mechanisms remain unclear. Herein, we investigated the therapeutic effects of HPS on murine DN and the potential mechanisms responsible for its efficacy. We used C57BLKS/6J

Diabetic nephropathy (DN) is a severe complication of diabetes, characterized by changes in kidney structure and function. The natural product rosmarinic acid (RA) has demonstrated therapeutic effects, including anti-inflammation and anti-oxidative-stress, in renal damage or dysfunction. In this study, we characterized the heterogeneity of the cellular response in kidneys to DN-induced injury and RA treatment at single cell levels. Our results demonstrated that RA significantly alleviated renal tubular epithelial injury, particularly in the proximal tubular S1 segment and on glomerular epithelial cells known as podocytes, while attenuating the inflammatory response of macrophages, oxidative stress, and cytotoxicity of natural killer cells. These findings provide a comprehensive understanding of the mechanisms by which RA alleviates kidney damage, oxidative stress, and inflammation, offering valuable guidance for the clinical application of RA in the treatment of DN.

Elevated levels of plasma free fatty acid (FFA) and disturbed mitochondrial dynamics play crucial roles in the pathogenesis of diabetic kidney disease (DKD). However, the mechanisms by which FFA leads to mitochondrial damage in glomerular podocytes of DKD and the effects of Berberine (BBR) on podocytes are not fully understood.

Disordered lipid metabolism and disturbed mitochondrial bioenergetics play pivotal roles in the initiation and development of diabetic kidney disease (DKD). Berberine is a plant alkaloid, used in Chinese herbal medicine. It has multiple therapeutic actions on diabetes mellitus and its complications, including regulation of glucose and lipid metabolism, improvement of insulin sensitivity, and alleviation of oxidative damage. Here, we investigated the reno-protective effects of berberine. We used samples from DKD patients and experiments with models of DKD (db/db mice) and cultured podocytes, to characterize energy metabolism profiles using metabolomics. Molecular targets and mechanisms involved in the regulation of mitochondrial function and bioenergetics by berberine were investigated, along with its effects on metabolic alterations in DKD mice. Metabolomic analysis suggested altered mitochondrial fuel usage and generalized mitochondrial dysfunction in patients with DKD. In db/db mice, berberine treatment reversed the disordered metabolism, podocyte damage and glomerulosclerosis. Lipid accumulation, excessive generation of mitochondrial ROS, mitochondrial dysfunction, and deficient fatty acid oxidation in DKD mouse models and in cultured podocytes were suppressed by berberine. These protective effects of berberine were accompanied by activation of the peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α) signalling pathway, which promoted mitochondrial energy homeostasis and fatty acid oxidation in podocytes. PGC-1α-mediated mitochondrial bioenergetics could play a key role in lipid disorder-induced podocyte damage and development of DKD in mice. Restoration of PGC-1α activity and the energy homeostasis by berberine might be a potential therapeutic strategy against DKD.

Linggui-Zhugan (LGZG) comprises four herbs and is a classic formula in traditional Chinese medicine. There is strong clinical evidence of its pleiotropic effects in the prevention of diabetes and its related complications. Although several classes of drugs are currently available for clinical management of diabetic kidney disease (DKD), tight glycemic and/or hypertension control may not prevent disease progression. This study evaluated the therapeutic effect of the ethnopharmacological agent LGZG on DKD. This study aimed to investigate the effects of LGZG formula with standard quality control on experimental DKD and its related metabolic disorders in animal model. Meanwhile, the present study aimed to investigate regulatory effects of LGZG on renal proteinase 3 (PR3) to reveal mechanisms underlying renoprotective benefits of LGZG. LGZG decoction was fingerprinted by high-performance liquid chromatography for quality control. An experimental model of DKD was induced in C57 BL/6J mice by a combination of high-fat diet feeding, uninephrectomy, and intraperitoneal injection of streptozocin. The LGZG decoction was administrated by daily oral gavage. Treatment with LGZG formula significantly attenuated DKD-like traits (including severe albuminuria, mesangial matrix expansion, and podocyte loss) and metabolic dysfunction (disordered body composition and dyslipidemia) in mice. RNA sequencing data revealed a close association of LGZG treatment with marked modulation of signaling pathways related to podocyte injury and cell apoptosis. Mechanistically, LGZG suppressed the DKD-triggered increase in renal PR3 and podocyte apoptosis. In-vitro incubation of mouse immortalized podocytes with LGZG-medicated serum attenuated PR3-mediated apoptosis. Our data demonstrated that the LGZG formula protected against DKD in mice and was closely associated with its inhibitory effects on PR3-mediated podocyte apoptosis.

Defective antioxidant system as well as mitochondrial dysfunction contributes to the pathogenesis and progression of diabetic kidney disease (DKD). Nuclear factor erythroid 2-related factor 2 (Nrf2)-mediated signaling is the central defensive mechanism against oxidative stress and therefore pharmacological activation of Nrf2 is a promising therapeutic strategy. In this study, using molecular docking we found that Astragaloside IV (AS-IV), an active ingredient from traditional formula of Huangqi decoction (HQD), exerted a higher potential to promote Nrf2 escape from Keap1-Nrf2 interaction via competitively bind to amino acid sites in Keap1. When podocyte exposed to high glucose (HG) stimulation, mitochondrial morphological alterations and podocyte apoptosis were presented and accompanied by Nrf2 and mitochondrial transcription factor A (TFAM) downregulation. Mechanistically, HG promoted a decrease in mitochondria-specific electron transport chain (ETC) complexes, ATP synthesis and mtDNA content as well as increased ROS production. Conversely, all these mitochondrial defects were dramatically alleviated by AS-IV, but suppression of Nrf2 with inhibitor or siRNA and TFAM siRNA simultaneously alleviated the AS-IV efficacy. Moreover, experimental diabetic mice exhibited significant renal injury as well as mitochondrial disorder, corresponding with the decreased expression of Nrf2 and TFAM. On the contrary, AS-IV reversed the abnormality and the Nrf2 and TFAM expression were also restored. Taken together, the present findings demonstrate the improvement of AS-IV on mitochondrial function, thereby resistance to oxidative stress-induced diabetic kidney injury and podocyte apoptosis, and the process is closely associated with activation of Nrf2-ARE/TFAM signaling.

Diabetic nephropathy (DN) is one of the most severe microvascular complications of diabetes and has become the leading cause of end-stage renal disease formation. The pathogenesis of diabetic nephropathy is very complex and is still not fully understood. Fisetin is a flavonoid polyphenolic compound that is widely found in different fruits, vegetables, and medicinal plants. Many studies have indicated that it has a variety of pharmacological activities. In this study, we investigated the mechanism of action of fisetin in the protection of DN-induced podocyte injury both in vivo and in vitro. Results showed that fisetin could reduce high glucose (HG)-induced podocyte injury and streptozotocin (STZ)-induced diabetic nephropathy in mice. According to the histopathological staining results, fisetin ameliorated DN-induced glomerular injury in a dose-dependent manner. Western blot and immunofluorescence results showed that fisetin effectively promoted the expression of podocyte functional integrity marker proteins and inhibited the expression of podocyte injury marker proteins. In addition, according to the Western blot and RT-qPCR results, fisetin activates the nuclear translocation of Nrf2 to exert antioxidative stress ability and affects the expression of downstream antioxidant enzymes HO-1, GPX4, and other ferroptosis-related markers, thereby protecting against HG-induced podocyte ferroptosis and oxidative stress injury in DN mice. In summary, this study demonstrated that fisetin could enhance the antioxidative stress capacity of DN mice by promoting the activation of the Nrf2/HO-1/GPX4 signaling pathway in renal tissues, and attenuated HG-induced podocytes injury and STZ-induced DN in mice.

Diabetic nephropathy (DN) is one of the most serious complications of diabetes and the main cause of end-stage renal failure. Rhubarb is a widely used traditional Chinese herb, and it has exhibited efficacy in reducing proteinuria, lowering blood sugar levels and improving kidney function in patients with DN. However, the exact pharmacological mechanism by rhubarb improves DN remain unclear due to the complexity of its ingredients. Hence, we systematically explored the underlying mechanisms of rhubarb in the treatment of DN. We adopted a network pharmacology approach, focusing on the identification of active ingredients, drug target prediction, gene collection, Gene Ontology enrichment and Kyoto Encyclopedia of Genes and Genomes enrichment. Molecular docking technology was used to verify the binding ability between the main active compounds and central therapeutic targets, and screen out the core active ingredients in rhubarb for the treatment of DN. Finally, molecular dynamics simulation was performed for the optimal core protein-ligand obtained by molecular docking using GROMACS software. The network analysis identified 16 active compounds in rhubarb that were linked to 37 possible therapeutic targets related to DN. Through protein-protein interaction analysis, TP53, CASP8, CASP3, MYC, JUN and PTGS2 were identified as the key therapeutic targets. By validation of molecular docking, finding that the central therapeutic targets have good affinities with the main active compounds of rhubarb, and rhein, beta-sitosterol and aloe-emodin were identified as the core active ingredients in rhubarb for the treatment of DN. Results from molecular dynamics simulations showed that TP53 and aloe-emodin bound very stably with a binding free energy of - 26.98 kcal/mol between the two. The results of the gene enrichment analysis revealed that the PI3K-Akt signalling pathway, p53 signalling pathway, AGE-RAGE signalling pathway and MAPK signalling pathway might be the key pathways for the treatment of DN, and these pathways were involved in podocyte apoptosis, glomerular mesangial cell proliferation, inflammation and renal fibrosis. Based on the network pharmacology approach and molecular docking technology, we successfully predicted the active compounds and their respective targets. In addition, we illustrated the molecular mechanisms that mediate the therapeutic effects of rhubarb against DN. These findings provided an important scientific basis for further research of the mechanism of rhubarb in the treatment of DN.

Diabetic nephropathy (DN) is one of the primary complications of diabetes. Fisetin is a flavonoid polyphenol that is present in several vegetables and fruits. The present study investigated the mechanisms of fisetin in DN-induced podocyte injury both

Zhen-wu-tang (ZWT) is a blended traditional Chinese medicine specifically used for various kidney diseases. The present study is to evaluate the effects of ZWT on diabetic nephropathy and investigate the potential anti-diabetic mechanisms. Streptozotocin (STZ)-induced diabetic rats were orally administrated ZWT (80,320 mg/kg b.w.) once a day, for a period of 12 weeks. Body weight, urinary volume, urine protein and blood biochemical parameters were measured every 3 weeks. At the end of the observation period, kidneys were isolated for histology, angiotensin II content determination, real time PCR and Western blot analyses. ZWT (320 mg/kg b.w.) prevented body weight loss, reduced polyurea, urinary protein excretion, serum creatinine and blood urea nitrogen, although it did not alter the hyperglycemia. It ameliorated glomerular hypertrophy and expansion of the mesangial area, swell and effacement of foot process. It also suppressed the increased expression of angiotensin II, nephrin and podocin. Data from this study suggest that ZWT possess a protective effect on renal damage of STZ-induced diabetic nephropathy in rats, by suppressing hyperactivity of renal renin-angiotensin system that turns in to modulate renal nephrin and podocin expressions, thereby protecting podocyte from injury.

Wenshen Jianpi recipe (WSJPR), a blended traditional Chinese medicine, is considered to have the possible beneficial effect on the progression of diabetic nephropathy (DN). This present study was designed to elucidate this protective activity in a rat model with streptozotocin (STZ)-induced DN and to explore the possible underlying mechanism. Adult Sprague Dawley (SD) rats were induced to develop DN through intraperitoneal injection of STZ (60 mg/kg). Animals were orally administered saline, WSJPR at 7.5, 15, 30 g/kg, and valsartan (25 mg/kg) daily for 8 weeks. Blood and 24-h urine samples of each rat were collected for biochemical examination at 2-week intervals. Microcirculatory blood flow in the renal cortex and hemorheology index were also measured. At the end of 8 weeks, all rats were sacrificed to obtain the kidney tissues for histological examination and reverse transcription polymerase chain reaction (RT-PCR) was used to analyze the transcriptional levels of nephrin and podocin genes. WSJPR could improve serum total protein (TP) and albumin (ALB), reduce the excretion rates of urine-TP (U-TP), urine-ALB (U-ALB) and urine urea nitrogen (UUN) (P < 0.05), although it did not significantly alter the hyperglycemia. In addition, treatment with WSJPR could strongly reduce blood flow, erythrocyte aggregation index, and ameliorate microcirculation. In histological measurement, WSJPR-treated rats showed a significant amelioration in glomerular hypertrophy and mesangial expansion. By RT-PCR, we found WSJPR up-regulated the nephrin and podocin expression at mRNA levels. This study suggested that WSJPR could effectively relieve renal damage and improve renal function of DN rats by ameliorating metabolism disorder and increasing the gene expression of nephrin and podocin, which might be a useful approach for the treatment of DN.

Diabetic nephropathy (DN) is the most frequent chronic microvascular consequence of diabetes, and podocyte injury and malfunction are closely related to the development of DN. Studies have shown that corilagin (Cor) has hepatoprotective, anti-inflammatory, antibacterial, antioxidant, anti-hypertensive, anti-diabetic, and anti-tumor activities. To explore the protective effect of Cor against podocyte injury in DN mice and the underlying mechanisms. Streptozotocin and a high-fat diet were combined to generate DN mice models, which were then divided into either a Cor group or a DN group ( Compared with the control group, the DN mice models had increased fasting blood glucose, glycosylated hemoglobin, triglycerides, and total cholesterol, decreased nephrin and podocin expression, increased apoptosis rate, elevated inflammatory cytokines, and enhanced oxidative stress. All of the conditions mentioned above were alleviated after intervention with Cor. In addition, Cor therapy improved SIRT1 and AMPK expression ( Cor alleviates podocyte injury by regulating autophagy

Among the complications of diabetes, diabetic kidney disease (DKD) frequently emerges, typified by the detrimental effects on renal function, manifesting through inflammation, dysregulated lipid metabolism, and harm to podocytes. Existing research underscores the significance of the soluble form of C-X-C chemokine ligand 16 (CXCL16) within the context of renal impairments. However, whether CXCL16 is involved in the pathogenesis of DKD remains elusive. We report that CXCL16 levels in the serum and kidneys of individuals with DKD were elevated and correlated with various lipid parameters. The expression of CXCL16 in human podocytes subjected to high glucose or palmitic acid induction and exogenous CXCL16 administration in these cells were examined. Higher CXCL16 levels were linked to abnormal lipid metabolism. Exogenous CXCL16 administration induced lipid droplets, microfilament disorganization, apoptosis, oxidative stress, and inflammation, inhibited PPAR γ, up-regulated COX2 expression, and inhibited Nrf2 translocation in podocytes. Molecular analysis revealed that Curcumin (Cur), a polyphenolic compound derived from Curcuma longa and an Nrf2 agonist, targets the ATP-binding pocket of CXCL16, inhibiting its kinase activity. Meanwhile, Cur therapy alleviated podocyte injury, lipid accumulation, improved glomerulopathy, and reduced albuminuria. Furthermore, after silencing CXCL16 expression in podocytes using siRNA, the effects of exogenous CXCL16 were nullified, and Cur no longer exhibited any significant impact. Thus, CXCL16 participates in the pathogenesis of DKD. Inhibition of CXCL16 has shown promising results in experimental models, suggesting its beneficial effects in ameliorating DKD.

Diabetic nephropathy (DN) is one of the complications with the highest mortality among diabetes patients and can lead to renal failure. Modified Huangfeng decoction (MHD) has been widely applied in the clinical treatment of kidney diseases. However, the mechanism by which MHD affects DN has not been fully elucidated. To investigate the impact of MHD on DN in mice and the underlying mechanism. The main ingredients of MHD were identified by liquid chromatography‒mass spectrometry. A high-fat diet- and streptozotocin (STZ)-induced DN mouse model was constructed and treated with MHD for 6 weeks. The serum and urine parameters were measured, and the tissue sections were histologically stained. The mRNA and protein levels of metabolism-, inflammation-, fibrosis-, and autophagy-related markers were examined by qPCR and western blotting. The microbial composition and metabolites of cecal contents were analyzed through full-length 16S rRNA sequencing and nontargeted metabolomics. MHD alleviated insulin resistance in DN mice and ameliorated changes in lipid metabolism and inflammation in the liver and fat. In addition, MHD reduced the levels of kidney injury markers in the serum and urine and attenuated inflammation and fibrosis in the kidney. These results were accompanied by enhanced gut barrier function and a markedly altered microbiota composition and metabolites, with an increased abundance of beneficial bacterial species and metabolites. Moreover, MHD itself and the microbial metabolite spermidine reduced podocyte damage by activating autophagy via the PI3K/AKT/mTOR pathway. MHD potentially ameliorated DN by activating podocyte autophagy via the PI3K/AKT/mTOR pathway and modulating the gut microbiota and its metabolites. Our findings provide a more comprehensive understanding of the mechanism of MHD and the involvement of the gut‒kidney interaction in the progression of DN, laying a theoretical foundation for the clinical application of MHD in DN treatment.

To uncover the potential hub targets of Kunkui Baoshen decoction (KKBS) in alleviating diabetic kidney disease (DKD). Targets associated with KKBS and DKD were curated from TCMSP, GeneCards, OMIM, and Dis- GeNET databases. Common targets were identified through intersection analysis using a Venn diagram. Employing the "Drug-component-target" approach and constructing a Protein-protein Interaction (PPI) network, pivotal components and hub targets involved in KKBS's therapeutic action against DKD were identified. Functional enrichment and Gene Set Enrichment Analysis (GSEA) elucidated the potential mechanisms of these hub targets. Molecular docking simulations validated binding interactions. Subsequently, hub targets were validated using independent cohorts and clinical datasets. Immune cell infiltration in DKD samples was assessed using ESTIMATE, CIBERSORT, and IPS algorithms. A nomogram was developed to predict DKD prevalence. Finally, causal relationships between hub targets and DKD were explored through Mendelian randomization (MR) analysis at the genetic level. Jaranol, isorhamnetin, nobiletin, calycosin, and quercetin emerged as principal effective components in KKBS, with predicted modulation of the PI3K/Akt, MAPK, HIF-1, NF-kB, and IL-17 signaling pathways. The hub targets in the PPI network include proteins involved in regulating podocyte autophagy and apoptosis, managing antioxidant stress, contributing to insulin resistance, and participating in extracellular matrix deposition in DKD. Molecular docking affirmed favorable binding interactions between principal components and hub targets. Validation efforts across cohorts and databases underscored the potential of hub targets as DKD biomarkers. Among 20 model algorithms, the Extra Tree model yielded the largest Area Under the Curve (AUC) in receiver operating characteristic (ROC) analysis. MR analysis elucidated that the targets related to antioxidant stress had a positive impact on DKD, while the target associated with renal tubular basement membrane degradation had a negative impact. Integration of Network Pharmacology, Bioinformatics, and MR analysis unveiled the capacity of KKBS to modulate pivotal targets in the treatment of DKD.

Diabetic nephropathy (DN), one of the serious complications in the diabetes, has a high mortality in the diabetic patients. Bilberry ( Streptozocin (STZ) combined with high fat induced DN model was established in rats. Biochemical indicators, histopathology, 16s third generation sequencing and serum metabolomics were used to evaluate the effects of VCE on DN. Subsequently, a cell model of advanced glycation end products (AGEs) induced podocyte injury was established to verify which compounds in VCE played the main anti-diabetic nephropathy function and the mechanism of action. Finally, Insulin resistance index, lipid metabolism, oxidative stress and inflammatory response indexes of DN rats were significantly improved after 8 weeks of VCE treatment. In addition, intake of VCE modulates gut microbiota composition and reverses the abundance of Our findings suggest that the improvement of gut microbiota and metabolic function were related to the anti-DN potential of VCE, and the underlying mechanism may be related to the inhibition of MAPK/NF-κB signaling pathway.

The current treatment for diabetic nephropathy (DN) is still limited. NaoXinTong Capsule (NXT) is a Chinese Medicine prescribed to patients with cardiovascular disease. It can also ameliorate metabolic syndromes in patients indicating its anti-diabetic properties. Herein we report the therapeutic effects of NXT on the developed DN. The db/db diabetic mice at ˜12 weeks old, the age with DN at middle/advanced stages, were treated with NXT for 12 weeks. We found NXT treatment reduced diabetes-induced hyperglycemia and dyslipidemia, thereby substantially reduced DN progress. In the kidney, NXT reduced mesangial matrix expansion and glomerulosclerosis by inhibiting extracellular matrix accumulation through activation of matrix metalloproteinase 2/9 and inactivating transforming growth factor β1 expression. NXT reduced podocyte injury by reducing renal inflammation and expression of adhesion molecules. Mechanically, NXT potently activated AMPKα in multiple tissues thereby enhancing energy metabolism. In the liver, NXT increased glucokinase expression and insulin sensitivity by increasing insulin receptor substrate 1/2 and protein kinase B (AKT) 1/2 expression/phosphorylation. In skeletal muscle, NXT activated expression of glucose transporter type 4, AKT, glycogen synthase and peroxisome proliferator activated receptor α/γ. In adipose tissue, NXT reduced fatty acid synthase while activating hormone-sensitive lipase expression. Taken together, our study demonstrates that NXT reduced progress of the developed DN by ameliorating glucose, lipid and energy metabolism, maintaining renal structural and functional integrity. Our study also indicates the potential application of NXT for DN treatment in clinics.

Diabetic kidney disease (DKD) progression is strongly associated with podocyte mitochondrial dysfunction. The clinically effective Chinese herbal Baoshentongluo formula (BSTL) has demonstrated significant proteinuria reduction in DKD patients. HPLC-ESI-MS analysis identified characteristic bioactive components in BSTL including astragalosides, rehmanniosides, and tanshinones. However, the molecular mechanisms through which BSTL maintains podocyte homeostasis remain incompletely understood. Mouse podocyte clone-5 (MPC-5) cells and db/db mice were used. Db/db mice were randomized into db/db and db/db + BSTL (16.5 g/kg/d, intragastric administration for 12 weeks). A group of m/m mice served as the control. Renal function, urinary albumin-to-creatinine ratio (UACR), histopathological analysis, apoptotic, and mitophagy-related protein levels were evaluated. MPC-5 cells were exposed to high glucose (HG, 30 mM) and BSTL drug-containing serum (8%) for 24 h grouping as control, HG, HG + BSTL, and HG + siPINK1. Podocyte apoptosis, mitophagy levels, and expression of PTEN-induced putative kinase 1 (PINK1) and E3 ubiquitin ligase (Parkin) were assessed. In db/db diabetic mice, oral administration of BSTL significantly lowered urinary albumin-to-creatinine ratio (P<0.05), improved glomerular filtration rate, and ameliorated renal histopathological changes, decreased LC3-II/LC3-I ratio, and downregulated expression of mitophagy-related proteins PINK1, Parkin, ATG5 and Beclin-1. Treatment with 8% BSTL-containing serum significantly attenuated HG-induced podocyte apoptosis ( Our integrated in vitro and in vivo findings establish that BSTL protects against DKD progression by selectively inhibiting PINK1/Parkin-dependent mitophagy in podocytes to inhibit podocyte injury, which provides both mechanistic insights and therapeutic potential for clinical DKD management.

Rubus suavissimus S.Lee (RS), a traditional ethnomedicine Guangxi, China, has long been used to manage diabetes and its complications. Existing studies have demonstrated the antidiabetic activity of RS and its complications, but the pharmacological material basis and molecular mechanism of its efficacy have not been clarified. This study aimed to elucidate the active constituents and specific molecular mechanisms underlying the therapeutic effects of RS in diabetic kidney disease (DKD). Ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) was employed to identify major active components within RS polyphenol extracts. Core signaling pathways and key active components were screened using network pharmacology analysis and enrichment methods. Subsequently, an in vitro MPC-5 podocyte cell model was established under high-glucose and high-lipid conditions. Data-independent acquisition (DIA) mass spectrometry was used to analyze differentially expressed proteins in the cellular secretome. Mitochondrial ultrastructure was assessed using transmission electron microscopy (TEM). Key protein expression changes were validated by Western blotting. Network pharmacology screening identified myricetin (Myr) as the compound exhibiting the highest binding affinity to PIK3R1, suggesting its role as a key active component in the anti-DKD effects of RS polyphenols. In vitro, high-glucose and high-lipid exposed MPC-5 cells exhibited pronounced mitochondrial swelling and cristae disruption. Myr treatment significantly preserved mitochondrial morphology and induced the formation of double-membrane autophagic vesicles encapsulating damaged mitochondria, indicative of activated mitophagy. Proteomic analysis corroborated these findings. This study demonstrates for the first time that Myr, a principal active component of RS polyphenols, exerts its therapeutic potential in DKD by inhibiting PIK3R1. This inhibition promotes XBP1 expression, indirectly activating both the PI3K/Akt and PINK1/Parkin pathways, ultimately enhancing autophagic flux. Myr effectively activated autophagy and mitophagy by targeting the PI3K/Akt and PINK1/Parkin signaling pathways, facilitating the removal of dysfunctional mitochondria and mitigating cellular damage in DKD models. These findings provide a mechanistic foundation for the use of RS-derived polyphenols in chronic kidney disease management and highlight Myr's potential as a natural therapeutic agent for DKD.

Recent researches have reported the extensive pharmacological activities of Ginsenoside Rg1 including antioxidant, anti-inflammatory, and anticancer properties. Furthermore Rg1 was also shown to protect various kinds of cells from self-digestion by its anti-autophagy activity. In previous studies, angiotensin II (Ang II), a key mediator of renin-angiotensin system, has been demonstrated to contribute to the progression of renal injury including abnormal autophagy. However, whether Rg1 can relieve Ang II-induced autophagy in podocyte as well as the underlying molecular mechanism remains to be elucidated. Here, we employed Ang II-treated podocyte as a model to investigate the effect of Rg1 on autophagy and the involved signal pathways. In the present study, we found that Ang II strongly promoted autophagy in immortalized mouse podocyte cells by observing the formation of autophagosomes and detecting the expression of autophagic marker, for example, LC3-II. Notably, compared to the Ang II-treated cells, treatment with Rg1 significantly inhibited the formation of autophagosomes and expression of autophagy-related proteins in Ang II pre-treated podocyte. Meanwhile, Rg1 downregulated the activity of AMPK and GSK-3β and upregulated the activity of P70S6K in Ang II-treated podocyte. In conclusion, these findings demonstrate that Ang II promotes autophagy in podocyte, and Rg1 effectively attenuates this process through AMPK/mTOR/PI3K pathway, suggesting that Rg1 may be beneficial to alleviate podocyte injury.

Diabetic nephropathy (DN) is one of the most frequent complications of diabetes. Early stages of DN are associated with hyperinsulinemia and progressive insulin resistance in insulin-sensitive cells, including podocytes. The diabetic environment induces pathological changes, especially in podocyte bioenergetics, which is tightly linked with mitochondrial dynamics. The regulatory role of insulin in mitochondrial morphology in podocytes has not been fully elucidated. Therefore, the main goal of the present study was to investigate effects of insulin on the regulation of mitochondrial dynamics and bioenergetics in human podocytes. Biochemical analyses were performed to assess oxidative phosphorylation efficiency by measuring the oxygen consumption rate (OCR) and glycolysis by measuring the extracellular acidification rate (ECAR). mRNA and protein expression were determined by real-time polymerase chain reaction and Western blot. The intracellular mitochondrial network was visualized by MitoTracker staining. All calculations were conducted using CellProfiler software. Short-term insulin exposure exerted inhibitory effects on various parameters of oxidative respiration and adenosine triphosphate production, and glycolysis flux was elevated. After a longer time of treating cells with insulin, an increase in mitochondrial size was observed, accompanied by a reduction of expression of the mitochondrial fission markers DRP1 and FIS1 and an increase in mitophagy. Overall, we identified a previously unknown role for insulin in the regulation of oxidative respiration and glycolysis and elucidated mitochondrial dynamics in human podocytes. The present results emphasize the importance of the duration of insulin stimulation for its metabolic and molecular effects, which should be considered in clinical and experimental studies of DN.