Dataset Information

Transcriptomic Analysis Reveals the Protection of Astragaloside IV against Diabetic Nephropathy by Modulating Inflammation.

ABSTRACT:

Background

Diabetic nephropathy (DN) is one of the leading causes of end-stage kidney disease. Recently, there is no specific drug available to block the kidney damage. Astragaloside IV (AS-IV) is a major active component of Astragalus membranaceus (Fisch) Bge and has been demonstrated to benefit the kidney functions. This study explores the potential pharmacological action of AS-IV in DN of rats.

Methods

Male Sprague-Dawley rats were fed with high-fat diet and injected with streptozotocin to induce diabetes. The diabetic rats were randomized and treated with vehicle or AS-IV (80 mg/kg) daily by gavage for 12 weeks as the DN or AS-IV group, respectively. The normal control rats were fed with normal chow and injected with vehicles (n = 8 per group). These rats were monitored for diabetes- and kidney function-related measures. The expression profiles of gene mRNA transcripts in the kidney tissues were analyzed by RNA-seq and quantitative RT-PCR. The levels of advanced glycation end products (AGEs), IL-1β, and IL-18 in the serum samples and kidney tissues were quantified by ELISA. The levels of collagen IV (COL-4) and fibronectin (FN) expression in kidney tissues were examined by immunohistochemistry and Western blot.

Results

In comparison with the DN group, AS-IV treatment significantly reduced blood glucose levels, food and water consumption, 24 h urine, renal index values, 24 h urine total proteins, blood urea nitrogen (BUN) levels, and creatinine clearance rates (CCR), accompanied by minimizing the DN-induced early kidney damages, fibrosis, and microstructural changes. Furthermore, AS-IV treatment significantly modulated the DN-altered gene transcription profiles in the kidney of rats, particularly for inflammation-related genes, including the nucleotide-binding oligomerization domain-like receptor signaling, which was validated by quantitative RT-PCR. AS-IV treatment significantly decreased the levels of serum and kidney AGEs, IL-1β, and IL-18 expression and fibrosis indexes in the kidney of rats.

Conclusion

AS-IV treatment ameliorated the severity of DN by inhibiting inflammation-related gene expression in the kidney of rats.

SUBMITTER: Zhang Y

PROVIDER: S-EPMC7443226 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Publications

Transcriptomic Analysis Reveals the Protection of Astragaloside IV against Diabetic Nephropathy by Modulating Inflammation.

Zhang Yudi Y Tao Chunhe C Xuan Chen C Jiang Junyan J Cao Wenfu W

Oxidative medicine and cellular longevity 20200812

<h4>Background</h4>Diabetic nephropathy (DN) is one of the leading causes of end-stage kidney disease. Recently, there is no specific drug available to block the kidney damage. Astragaloside IV (AS-IV) is a major active component of <i>Astragalus membranaceus</i> (Fisch) Bge and has been demonstrated to benefit the kidney functions. This study explores the potential pharmacological action of AS-IV in DN of rats.<h4>Methods</h4>Male Sprague-Dawley rats were fed with high-fat diet and injected wit ...[more]

PMID: 32855769

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Project description:Purpose: Diabetic retinopathy (DR) is a serious complication of diabetes mellitus, which nearly happens to all the diabetic sufferers. This study aims to identify the preliminary molecular regulation involved in the therapeutic efficacy of astragaloside IV (AS- IV) for DR. Methods: Diabetic rat models were established and treated with AS-IV. Optical coherence tomography (OCT) and Hematoxylin-eosin (HE) staining was employed to demonstrate the histopathological changes. The main targets of AS-IV were identified by searching from public databases of traditional Chinese medicine (GeneCards, PharmMapper and Swiss Target Prediction). Besides, disease targets of DR were also obtained by integrated data from GEO datasets and predicted from public databases. Protein-protein interaction (PPI) network was constructed by Cytoscape with overlapping genes and 10 core targets were selected, on which Gene Ontology (GO) along with Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis were conducted. The interaction between AS-IV and these crucial genes were analyzed using molecular docking. RT-qPCR and western blot were used to verify the expression variation of core targets. Results: OCT imaging and HE staining demonstrated that AS-IV administration significantly increased retinal thickness in diabetic rats, obviously alleviating DR induced histopathological changes as well as elevated blood glucose levels. 107 common targets of AS-IV and DR were determined after intersection. PPI network analysis filtered 10 hub genes potentially targeted by AS-IV, including VEGFA, CASP3, HIF1α, STAT3, CTNNB1, SRC, AKT1, EGFR, IL1β and IL6. Enrichment analysis indicated that these genes were mainly enriched in biological processes like T cell activation, epithelial cell proliferation and protein kinase B signaling, and involved in oxidative stress, apoptosis and inflammation-related pathways. The molecular docking prediction suggested that AS-IV exhibited stable binding to these core targets. In addition, mRNA levels of core targets in diabetic rats were differentially expressed before and after AS-IV treatment. Western blot further revealed that AS-IV treatment elevated DR-depressed protein levels of PI3K and AKT. Conclusion: Our study elucidated the effect of AS-IV in attenuating retinopathy induced by diabetes in rats and preliminarily unveiled the therapeutic efficacy of AS-IV in the treatment of DR might be attributed to activation of PI3K-AKT signaling pathway.

Project description:ObjectiveDeficiencies in klotho are implicated in various kidney dysfunctions including diabetic nephropathy (DN) related to inflammatory responses. Klotho is closely related to inflammatory responses and is a potential target for ameliorating kidney failure. Pyroptosis, an inflammatory form of programmed cell death, is reported to take part in DN pathogenesis recently. This study is aimed at exploring whether and how klotho inhibited podocyte pyroptosis and whether astragaloside IV (AS-IV) protect podocyte through the regulation of klotho.Materials and methodsSD rat model of DN and conditionally immortalized mouse podocytes exposed to high glucose were treated with AS-IV. Biochemical assays and morphological examination, cell viability assay, cell transfection, phalloidin staining, ELISA, LDH release assay, SOD and MDA detection, MMP assay, ROS level detection, flow cytometry analysis, TUNEL staining assay, PI/Hoechst 33342 staining, immunofluorescence assay, and western blot were performed to elucidate podocyte pyroptosis and to observe the renal morphology.ResultsThe treatment of AS-IV can improve renal function and protect podocytes exposed to high glucose. Klotho was decreased, and AS-IV increased klotho levels in serum and kidney tissue of DN rats as well as podocytes exposed to high glucose. AS-IV can inhibit DN glomeruli pyroptosis in vivo. In vitro, overexpressed klotho and treatment with AS-IV inhibited pyroptosis of podocytes cultured in high glucose. Klotho knockdown promoted podocyte pyroptosis, and treatment with AS-IV reversed this effect. Furthermore, the overexpression of klotho and AS-IV reduces oxidative stress levels and inhibited NF-κB activation and NLRP3-mediated podocytes' pyroptosis which was abolished by klotho knockdown. In addition, both the ROS inhibitor NAC and the NF-κB pathway inhibitor PDTC can inhibit NLRP3 inflammasome activation. NLRP3 inhibitor MCC950 can inhibit pyroptosis of podocytes exposed to high glucose.ConclusionAltogether, our results demonstrate that the protective effect of AS-IV in upregulating klotho expression in diabetes-induced podocyte injury is associated with the inhibition of NLRP3-mediated pyroptosis via the NF-κB signaling pathway.

Project description:Hypertensive nephrosclerosis (HN) and Type 2 diabetic nephropathy (T2DN) are the leading causes of chronic kidney disease (CKD). To explore shared pathogenetic mechanisms, we analyzed transcriptomes of kidney biopsies from patients with HN or T2DN. Total RNA was extracted from 10 μm whole kidney sections from patients with HN, T2DN, and normal controls (Ctrl) (n = 6 for each group) and processed for RNA sequencing. Differentially expressed (log2 fold change >1, adjusted p < 0.05) genes (DEG) and molecular pathways were analyzed, and selected results were validated by immunohistochemistry (IHC). ELISA on serum samples was performed on a related cohort consisting of patients with biopsy-proven HN (n = 13) and DN (n = 9), and a normal control group (n = 14). Cluster analysis on RNA sequencing data separated diseased and normal tissues. RNA sequencing revealed that 88% (341 out of 384) of DEG in HN were also altered in T2DN, while gene set enrichment analysis (GSEA) showed that over 90% of affected molecular pathways, including those related to inflammation, immune response, and cell-cycle regulation, were similarly impacted in both HN and T2DN samples. The increased expression of genes tied to interleukin signaling and lymphocyte activation was more pronounced in HN, while genes associated with extracellular matrix organization were more evident in T2DN. Both HN and T2DN tissues exhibited significant upregulation of genes connected with inflammatory responses, T-cell activity, and partial epithelial to mesenchymal transition (p-EMT). Immunohistochemistry (IHC) further confirmed T-cell (CD4+ and CD8+ ) infiltration in the diseased tissues. Additionally, IHC revealed heightened AXL protein expression, a key regulator of inflammation and p-EMT, in both HN and T2DN, while serum analysis indicated elevated soluble AXL levels in patients with both conditions. These findings underline the shared molecular mechanisms between HN and T2DN, hinting at the potential for common therapeutic strategies targeting both diseases.

Dataset Information

Transcriptomic Analysis Reveals the Protection of Astragaloside IV against Diabetic Nephropathy by Modulating Inflammation.

Background

Methods

Results

Conclusion

Publications

Transcriptomic Analysis Reveals the Protection of Astragaloside IV against Diabetic Nephropathy by Modulating Inflammation.

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