RESUMEN
In China, the Astragalus membranaceus root is used to treat chronic kidney disease. Astragaloside IV (AS-IV), the primary bioactive compound, exhibits anti-inflammatory and antioxidative properties; however, its renoprotective mechanism in diabetic kidney disease (DKD) remains unclear. The study aimed to investigate the protective effects of AS-IV on DKD revealing the underlying mechanisms. We established an early diabetic rat model by feeding a high-fat diet and administering low-dose streptozotocin. Twelve weeks post-treatment, renal function was evaluated using functional assays, histological analyses, immunohistochemistry, western blotting, and transmission electron microscopy. HK-2 cells exposed to high glucose conditions were used to examine the effect of AS-IV on oxidative stress, iron levels, reactive oxygen species (ROS), and lipid peroxidation. Network pharmacology, proteomics, molecular docking, and molecular dynamics simulation techniques were employed to elucidate the role of AS-IV in DKD. The results revealed that AS-IV effectively enhanced renal function and mitigated disease pathology, oxidative stress, and ferroptosis markers in DKD rats. In HK-2 cells, AS-IV lowered the levels of lipid peroxides, Fe2+, and glutathione, indicating the repair of ferroptosis-related mitochondrial damage. AS-IV reduced mitochondrial ROS while enhancing mitochondrial membrane potential and ATP production, indicating its role in combating mitochondrial dysfunction. Overall, in silico analyses revealed that AS-IV interacts with HMOX1, FTH1, and TFR1 proteins, supporting its efficacy in alleviating renal injury by targeting mitochondrial dysfunction and ferroptosis. AS-IV may play a renoprotective role by regulating mitochondrial dysfunction and inhibiting. HMOX1/FTH1/TFR1-induced ferroptosis. Accordingly, AS-IV could be developed for the clinical treatment of DKD-related renal injury.
RESUMEN
Tubular injury and oxidative stress are involved in the pathogenesis of diabetic kidney disease (DKD). Astragaloside IV (ASIV) is a natural antioxidant. The effects and underlying molecular mechanisms of ASIV on DKD have not been elucidated. The db/db mice and high-glucose-stimulated HK2 cells were used to evaluate the beneficial effects of ASIV in vivo and in vitro. Succinylated proteomics was used to identify novel mechanisms of ASIV against DKD and experimentally further validated. ASIV alleviated renal dysfunction and proteinuria, downregulated fasting blood glucose, and upregulated insulin sensitivity in db/db mice. Meanwhile, ASIV alleviated tubular injury, oxidative stress, and mitochondrial dysfunction in vivo and in vitro. Mechanistically, ASIV reversed downregulated 17beta-hydroxysteroid dehydrogenase type 10 (HSD17B10) lysine succinylation by restoring carnitine palmitoyl-transferase1alpha (Cpt1a or CPT1A) activity in vivo and in vitro. Molecular docking and cell thermal shift assay revealed that ASIV may bind to CPT1A. Molecular dynamics simulations demonstrated K99 succinylation of HSD17B10 maintained mitochondrial RNA ribonuclease P (RNase P) stability. The K99R mutation of HSD17B10 induced oxidative stress and disrupted its binding to CPT1A or mitochondrial ribonuclease P protein 1 (MRPP1). Importantly, ASIV restored the interaction between HSD17B10 and MRPP1 in vivo and in vitro. We also demonstrated that ASIV reversed high-glucose-induced impaired RNase P activity in HK2 cells, which was suppressed upon K99R mutation of HSD17B10. These findings suggest that ASIV ameliorates oxidative stress-associated proximal tubular injury by upregulating CPT1A-mediated K99 succinylation of HSD17B10 to maintain RNase P activity.
RESUMEN
ETHNOPHARMACOLOGICAL RELEVANCE: The limitations of modern medicine in mitigating the pathological process of diabetic kidney disease (DKD) necessitate novel, precise, and effective prevention and treatment methods. Huangqi, the root of Astragalus membranaceus Fisch. ex Bunge has been used in traditional Chinese medicine for various kidney ailments. Astragaloside IV (AS-IV), the primary pharmacologically active compound in A. membranaceus, is involved in lipid metabolism regulation; however, its potential in ameliorating renal damage in DKD remains unexplored. AIM OF THE STUDY: To elucidate the specific mechanism by which AS-IV moderates DKD progression. MATERIALS AND METHODS: A murine model of DKD and high glucose-induced HK-2 cells were treated with AS-IV. Furthermore, multiomics analysis, molecular docking, and molecular dynamics simulations were performed to elucidate the mechanism of action of AS-IV in DKD, which was validated using molecular biological methods. RESULTS: AS-IV regulated glucose and lipid metabolism in DKD, thereby mitigating lipid deposition in the kidneys. Proteomic analysis identified 12 proteins associated with lipid metabolism regulated by AS-IV in the DKD renal tissue. Additionally, lipid metabolomic analysis revealed that AS-IV upregulated and downregulated 4 beneficial and 79 harmful lipid metabolites, respectively. Multiomics analysis further indicated a positive correlation between the top-ranked differential protein heme oxygenase (HMOX)1 and the levels of various harmful lipid metabolites and a negative correlation with the levels of beneficial lipid metabolites. Furthermore, enrichment of both ferroptosis and hypoxia-inducible factor (HIF)-1 signaling pathways during the AS-IV treatment of DKD was observed using proteomic analysis. Validation results showed that AS-IV effectively reduced ferroptosis in DKD-affected renal tubular epithelial cells by inhibiting HIF-1α/HMOX1 pathway activity, upregulating glutathione peroxidase-4 and ferritin heavy chain-1 expression, and downregulating acyl-CoA synthetase long-chain family member-4 and transferrin receptor-1 expression. Our findings demonstrate the potential of AS-IV in mitigating DKD pathology by downregulating the HIF-1α/HMOX1 signaling pathway, thereby averting ferroptosis in renal tubular epithelial cells. CONCLUSIONS: AS-IV is a promising treatment strategy for DKD via the inhibition of ferroptosis in renal tubular epithelial cells. The findings of this study may help facilitate the development of novel therapeutic strategies.
Asunto(s)
Nefropatías Diabéticas , Células Epiteliales , Ferroptosis , Subunidad alfa del Factor 1 Inducible por Hipoxia , Proteómica , Saponinas , Triterpenos , Animales , Humanos , Masculino , Ratones , Línea Celular , Diabetes Mellitus Experimental/tratamiento farmacológico , Diabetes Mellitus Experimental/metabolismo , Nefropatías Diabéticas/tratamiento farmacológico , Nefropatías Diabéticas/metabolismo , Células Epiteliales/efectos de los fármacos , Células Epiteliales/metabolismo , Ferroptosis/efectos de los fármacos , Hemo-Oxigenasa 1/metabolismo , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Túbulos Renales/efectos de los fármacos , Túbulos Renales/metabolismo , Túbulos Renales/patología , Metabolismo de los Lípidos/efectos de los fármacos , Lipidómica , Ratones Endogámicos C57BL , Simulación del Acoplamiento Molecular , Saponinas/farmacología , Transducción de Señal/efectos de los fármacos , Triterpenos/farmacologíaAsunto(s)
Colitis , Ferroptosis , Homeostasis , Mucosa Intestinal , Ácido Vanílico , Ferroptosis/efectos de los fármacos , Animales , Mucosa Intestinal/metabolismo , Mucosa Intestinal/efectos de los fármacos , Homeostasis/efectos de los fármacos , Colitis/tratamiento farmacológico , Colitis/metabolismo , Humanos , Ácido Vanílico/farmacología , Ácido Vanílico/uso terapéutico , Canales de Calcio/metabolismo , Canales de Calcio/efectos de los fármacosAsunto(s)
Ferroptosis , Factor 2 Relacionado con NF-E2 , Material Particulado , Transducción de Señal , Factor 2 Relacionado con NF-E2/metabolismo , Ferroptosis/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Animales , Material Particulado/toxicidad , Humanos , Lesión Pulmonar/metabolismo , Lesión Pulmonar/inducido químicamente , Lesión Pulmonar/prevención & control , Lesión Pulmonar/tratamiento farmacológicoRESUMEN
ETHNOPHARMACOLOGICAL RELEVANCE: Qizhuyanggan Decoction (QZD), a traditional Chinese medicine formula, is frequently utilized in clinical practice for managing hepatic fibrosis. However, the specific target and mechanism of action of QZD for hepatic fibrosis treatment remain unknown. AIM OF THE STUDY: By combining network pharmacology, serum medicinal chemistry, and experimental validation methods, our study aimed to investigate the therapeutic effects of QZD on hepatic fibrosis, the anti-hepatic fibrosis active ingredients, and the possible mechanism of anti-hepatic fibrosis action. MATERIALS AND METHODS: The study aimed to investigate the therapeutic effect of QZD on hepatic fibrosis induced by CCl4 in SD rats, as well as its mechanism of action. The rats were anesthetized intraperitoneally using 3% pentobarbital and were executed after asphyxiation with high concentrations of carbon dioxide. Several techniques were employed to evaluate the efficacy of QZD, including ELISA, Western blot, HYP reagent assay, and various pathological examinations such as HE, Masson, Sirius Red staining, and immunohistochemistry (IHC). Additionally, serum biochemical assays were conducted to assess the effect of QZD on liver injury. Network pharmacology, UPLC, molecular docking, and molecular dynamics simulation were utilized to explore the mechanism of QZD in treating hepatic fibrosis. Finally, experimental validation was performed through ELISA, IHC, RT-qPCR, and Western blot analysis. RESULT: Liver histopathology showed that QZD reduced inflammation and inhibited collagen production, and QZD significantly reduced HA and LN content to treat hepatic fibrosis. Serum biochemical analysis showed that QZD improved liver injury. Network pharmacology combined with UPLC screened six active ingredients and obtained 87 targets for the intersection of active ingredients and diseases. The enrichment analysis results indicated that the PI3K/AKT pathway might be the mechanism of action of QZD in the treatment of hepatic fibrosis, and counteracting the inflammatory response might be one of the pathways of action of QZD. Molecular docking and molecular dynamics simulations showed that the active ingredient had good binding properties with PI3K, AKT, and mTOR proteins. Western blot, ELISA, PCR, and IHC results indicated that QZD may treat hepatic fibrosis by inhibiting the PI3K/AKT/mTOR pathway and suppressing M1 macrophage polarization, while also promoting M2 macrophage polarization. CONCLUSIONS: QZD may be effective in the treatment of hepatic fibrosis by inhibiting the PI3K/AKT/mTOR signaling pathway and M1 macrophage polarization, while promoting M2 macrophage polarization. This provides a strong basis for the clinical application of QZD.