PACS-2 deficiency in tubular cells aggravates lipid-related kidney injury in diabetic kidney disease.

BACKGROUND: Lipid accumulation in tubular cells plays a key role in diabetic kidney disease (DKD). Targeting lipid metabolism disorders has clinical value in delaying the progression of DKD, but the precise mechanism by which molecules mediate lipid-related kidney injury remains unclear. Phosphofurin acidic cluster sorting protein 2 (PACS-2) is a multifunctional sorting protein that plays a role in lipid metabolism. This study determined the role of PACS-2 in lipid-related kidney injury in DKD. METHODS: Diabetes was induced by a high-fat diet combined with intraperitoneal injections of streptozotocin (HFD/STZ) in proximal tubule-specific knockout of Pacs-2 mice (PT-Pacs-2-/- mice) and the control mice (Pacs-2fl/fl mice). Transcriptomic analysis was performed between Pacs-2fl/fl mice and PT-Pacs-2-/- mice. RESULTS: Diabetic PT-Pacs-2-/- mice developed more severe tubule injury and proteinuria compared to diabetic Pacs-2fl/fl mice, which accompanied with increasing lipid synthesis, uptake and decreasing cholesterol efflux as well as lipid accumulation in tubules of the kidney. Furthermore, transcriptome analysis showed that the mRNA level of sterol O-acyltransferase 1 (Soat1) was up-regulated in the kidney of control PT-Pacs-2-/- mice. Transfection of HK2 cells with PACS-2 siRNA under high glucose plus palmitic acid (HGPA) condition aggravated lipid deposition and increased the expression of SOAT1 and sterol regulatory element-binding proteins (SREBPs), while the effect was blocked partially in that of co-transfection of SOAT1 siRNA. CONCLUSIONS: PACS-2 has a protective role against lipid-related kidney injury in DKD through SOAT1/SREBPs signaling.

GBP2 promotes M1 macrophage polarization by activating the notch1 signaling pathway in diabetic nephropathy.

Background: Diabetic nephropathy (DN) is one of the most common diabetic complications, which has become the primary cause of end-stage renal disease (ESRD) globally. Macrophage infiltration has been proven vital in the occurrence and development of DN. This study was designed to investigate the hub genes involved in macrophage-mediated inflammation of DN via bioinformatics analysis and experimental validation. Methods: Gene microarray datasets were obtained from the Gene Expression Omnibus (GEO) public website. Integrating the CIBERSORT, weighted gene co-expression network analysis (WGCNA) and DEGs, we screened macrophage M1-associated key genes with the highest intramodular connectivity. Subsequently, the Least Absolute Shrinkage and Selection Operator (LASSO) regression was utilized to further mine hub genes. GSE104954 acted as an external validation to predict the expression levels and diagnostic performance of these hub genes. The Nephroseq online platform was employed to evaluate the clinical implications of these hub genes. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) were performed to elucidate the dominant biological functions and signal pathways. Finally, we conducted experiments to verify the role of GBP2 in M1 macrophage-mediated inflammatory response and the underlying mechanism of this role. Results: Sixteen DEGs with the highest connectivity in M1 macrophages-associated module (paleturquoise module) were determined. Subsequently, we identified four hub genes through LASSO regression analysis, including CASP1, MS4A4A, CD53, and GBP2. Consistent with the training set, expression levels of these four hub genes manifested memorably elevated and the ROC curves indicated a good diagnostic accuracy with an area under the curve of greater than 0.8. Clinically, enhanced expression of these four hub genes predicted worse outcomes of DN patients. Given the known correlation between the first three hub genes and macrophage-mediated inflammation, experiments were performed to demonstrate the effect of GBP2, which proved that GBP2 contributed to M1 polarization of macrophages by activating the notch1 signaling pathway. Conclusion: Our findings detected four hub genes, namely CASP1, MS4A4A, CD53, and GBP2, may involve in the progression of DN via pro-inflammatory M1 macrophage phenotype. GBP2 could be a promising prognostic biomarker and intervention target for DN by regulating M1 polarization.

PACS-2 deficiency aggravates tubular injury in diabetic kidney disease by inhibiting ER-phagy.

Autophagy of endoplasmic reticulum (ER-phagy) selectively removes damaged ER through autophagy-lysosome pathway, acting as an adaptive mechanism to alleviate ER stress and restore ER homeostasis. However, the role and precise mechanism of ER-phagy in tubular injury of diabetic kidney disease (DKD) remain obscure. In the present study, we demonstrated that ER-phagy of renal tubular cells was severely impaired in streptozocin (STZ)-induced diabetic mice, with a decreased expression of phosphofurin acidic cluster sorting protein 2 (PACS-2), a membrane trafficking protein which was involved in autophagy, and a reduction of family with sequence similarity 134 member B (FAM134B), one ER-phagy receptor. These changes were further aggravated in mice with proximal tubule specific knockout of Pacs-2 gene. In vitro, transfection of HK-2 cells with PACS-2 overexpression plasmid partially improved the impairment of ER-phagy and the reduction of FAM134B, both of which were induced in high glucose ambience; while the effect was blocked by FAM134B siRNA. Mechanistically, PACS-2 interacted with and promoted the nuclear translocation of transcription factor EB (TFEB), which was reported to activate the expression of FAM134B. Collectively, these data unveiled that PACS-2 deficiency aggravates renal tubular injury in DKD via inhibiting ER-phagy through TFEB/FAM134B pathway.

Using network pharmacology to explore the mechanism of Danggui-Shaoyao-San in the treatment of diabetic kidney disease.

Danggui-Shaoyao-San (DSS) is one of traditional Chinese medicine, which recently was found to play a protective role in diabetic kidney disease (DKD). However, the pharmacological mechanisms of DSS remain obscure. This study would explore the molecular mechanisms and bioactive ingredients of DSS in the treatment of DKD through network pharmacology. The potential target genes of DKD were obtained through OMIM database, the DigSee database and the DisGeNET database. DSS-related targets were acquired from the BATMAN-TCM database and the STITCH database. The common targets of DSS and DKD were selected for analysis in the STRING database, and the results were imported into Cytoscape to construct a protein-protein interaction network. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways enrichment analysis and Gene Ontology (GO) enrichment analysis were carried out to further explore the mechanisms of DSS in treating DKD. Molecular docking was conducted to identify the potential interactions between the compounds and the hub genes. Finally, 162 therapeutic targets of DKD and 550 target genes of DSS were obtained from our screening process. Among this, 28 common targets were considered potential therapeutic targets of DSS for treating DKD. Hub signaling pathways including HIF-1 signaling pathway, TNF signaling pathway, AMPK signaling pathway, mTOR signaling pathway, and PI3K-Akt signaling pathway may be involved in the treatment of DKD using DSS. Furthermore, TNF and PPARG, and poricoic acid C and stigmasterol were identified as hub genes and main active components in this network, respectively. In this study, DSS appears to treat DKD by multi-targets and multi-pathways such as inflammatory, oxidative stress, autophagy and fibrosis, which provided a novel perspective for further research of DSS for the treatment of DKD.

ß-Hydroxybutyrate against Cisplatin-Induced acute kidney injury via inhibiting NLRP3 inflammasome and oxidative stress.

Cisplatin, as a commonly used anticancer drug, can easily lead to acute kidney injury (AKI), and has received more and more attention in clinical practice. ß-hydroxybutyric acid (BHB) is a metabolite in the body and acts as an inhibitor of oxidative stress and NLRP3 inflammasome, reducing inflammatory responses and apoptosis. However, the role of BHB in cisplatin-induced AKI is currently not fully elucidated. In this study, C57BL/6 male mice were randomly divided into normal control group, cisplatin-induced AKI group and AKI with BHB treatment group. Compared to the control, cisplatin-treated mice exhibited high level of serum creatinine, blood urea nitrogen and severe tubular injury, which accompanied with significantly increased expression level of NLRP3, IL-1ß, IL-18, BAX, cleaved-caspase 3, as well as aggravated oxidative stress and renal tubular cell apoptosis. However, these changes were significantly improved in that of BHB treatment. In vitro, our study showed that the expression of cleaved-caspase3, IL-1ß and IL-18 were significantly increased in human proximal tubular epithelial cell line (HK-2) treated with cisplatin compared with the control group, while decreased in cells treated with BHB. Furthermore, a significantly increased expression of cGAS and STING in HK-2 cells treated with cisplatin were found, whereas notably decreased in cells treated with BHB. This data indicates that BHB protects against cisplatin-induced AKI and renal tubular damage mediated by NLRP3 inflammasome and cGAS-STING pathway.

PACS-2 Ameliorates Tubular Injury by Facilitating Endoplasmic Reticulum-Mitochondria Contact and Mitophagy in Diabetic Nephropathy.

Mitochondria-associated endoplasmic reticulum membrane (MAM) may have a role in tubular injury in diabetic nephropathy (DN), but the precise mechanism remains unclear. Here, we demonstrate that the expression of phosphofurin acidic cluster sorting protein 2 (PACS-2), a critical regulator of MAM formation, is significantly decreased in renal tubules of patients with DN, and PACS-2 expression is positively correlated with renal function and negatively correlated with degrees of tubulointerstitial lesions. Conditional deletion of Pacs-2 in proximal tubules (PTs) aggravates albuminuria and tubular injury in a streptozotocin-induced mouse model of diabetes. Mitochondrial fragmentation, MAM disruption, and defective mitophagy accompanied by altered expression of mitochondrial dynamics and mitophagic proteins, including Drp1 and Becn1, are observed in tubules of diabetic mice; these changes are more pronounced in PT-specific Pacs-2 knockout mice. In vitro, overexpression of PACS-2 in HK-2 cells alleviates excessive mitochondrial fission induced by high glucose concentrations through blocking mitochondrial recruitment of DRP1 and subsequently restores MAM integrity and enhances mitophagy. Mechanistically, PACS-2 binds to BECN1 and mediates the relocalization of BECN1 to MAM, where it promotes the formation of mitophagosome. Together, these data highlight an important but previously unrecognized role of PACS-2 in ameliorating tubular injury in DN by facilitating MAM formation and mitophagy.

DsbA-L Ameliorates Renal Injury Through the AMPK/NLRP3 Inflammasome Signaling Pathway in Diabetic Nephropathy.

NLRP3-mediated inflammation is closely related to the pathological progression of diabetic nephropathy (DN). DsbA-L, an antioxidant enzyme, plays a protective role in a variety of diseases by inhibiting ER stress and regulating metabolism. However, the relationship of DsbA-L with inflammation, especially the NLRP3 inflammasome, has not been examined. In this study, we note that activation of the NLRP3 inflammasome and exacerbated fibrosis were observed in the kidneys of diabetic DsbA-L-knockout mice and were accompanied by decreased phosphorylation of AMP-activated protein kinase (AMPK). Moreover, correlation analysis shows that the phosphorylation of AMPK was negatively correlated with NLRP3 expression and tubular damage. In addition, the decreased AMPK phosphorylation and NLRP3 activation induced by high glucose (HG) in HK-2 cells could be alleviated by the overexpression of DsbA-L. Interestingly, the protective effect of DsbA-L was eliminated after treatment with compound C, a well-known AMPK inhibitor. Our findings suggest that DsbA-L inhibits NLRP3 inflammasome activation by promoting the phosphorylation of AMPK.

Caveolin-1 Regulates Cellular Metabolism: A Potential Therapeutic Target in Kidney Disease.

The kidney is an energy-consuming organ, and cellular metabolism plays an indispensable role in kidney-related diseases. Caveolin-1 (Cav-1), a multifunctional membrane protein, is the main component of caveolae on the plasma membrane. Caveolae are represented by tiny invaginations that are abundant on the plasma membrane and that serve as a platform to regulate cellular endocytosis, stress responses, and signal transduction. However, caveolae have received increasing attention as a metabolic platform that mediates the endocytosis of albumin, cholesterol, and glucose, participates in cellular metabolic reprogramming and is involved in the progression of kidney disease. It is worth noting that caveolae mainly depend on Cav-1 to perform the abovementioned cellular functions. Furthermore, the mechanism by which Cav-1 regulates cellular metabolism and participates in the pathophysiology of kidney diseases has not been completely elucidated. In this review, we introduce the structure and function of Cav-1 and its functions in regulating cellular metabolism, autophagy, and oxidative stress, focusing on the relationship between Cav-1 in cellular metabolism and kidney disease; in addition, Cav-1 that serves as a potential therapeutic target for treatment of kidney disease is also described.