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BACKGROUND: ANCA-associated vasculitis (AAV), and nephrotic syndrome encompassing diseases including minimal change disease (MCD), focal and segmental glomerulosclerosis (FSG), membranous nephropathy (MN), remain a challenge due to their varied immunological characteristics. Recent therapeutic advancements have highlighted the importance of understanding these diseases' immunological landscapes. METHODS: This study analyzed transcriptomics data from renal glomerular tissues of patients with AAV, FSG, MCD, MN, and normal controls. Utilizing an immune-related gene set of 883 genes, methods including Gene Set Variation Analysis (GSVA), LASSO regression, and Weighted Correlation Network Analysis (WGCNA) were used. Predictions of immune cell compositions were made through CIBERSORT, TIMER, MCPcounter, and quanTIseq algorithms. RESULTS: The study revealed distinct immunogenetic pathways enriched in each disease: hematopoietic cell lineage in ANCA, linoleic acid metabolism in FSG, PPAR signaling in MCD, and drug metabolism in MN. Classifiers based on immune gene expression showed high accuracy (AUC: ANCA 0.812, FSG 0.99, MCD 1, MN 0.888). Co-expression modules and PPI networks highlighted unique pathways for each disease. Predictions of immune cell composition showed elevated macrophages in FSG and MN, with Treg levels elevated across all four diseases compared to normal controls and highest in FSG. Correlation analyses demonstrated significant associations between classifier scores and immune cell types. CONCLUSION: This study offers accurate classifiers for AAV, FSG, MCD, and MN, and reveals distinct immunological pathways. These findings advance personalized treatments and highlight potential therapeutic targets in AAV and nephrotic syndrome. Further research should validate these results for clinical applications.
RESUMEN
OBJECTIVE: To explore the effects of core-binding factor α1 (Cbfα-1) gene silenced by siRNA on osteogenic differentiation and calcification of vascular smooth muscle cells (VSMC) induced by high phosphate in vitro. METHODS: VSMC were cultured in vitro and passaged 3 to 8 times. Four pairs of Cbfα-1 siRNA were designed and synthesized. Transfection was performed with cationic lipid vectors (Lipofectamine 2000). Transfection conditions were optimized by the FAM fluorescent labeling-siRNA to screen effective siRNA sequences by reverse transcription-polymerase chain reaction (RT-PCR). After transfection with effective siRNA sequences, VSMCs were divided into 4 groups: (1) normal phosphate (Pi 1.3 mmol/L); (2) high phosphate (Pi 2.6 mmol/L); (3) siRNA transfection: high phosphate+Cbfα-1-siRNA; (4) negative transfection control: high phosphate+negative control siRNA. Cbfα-1 and osteopontin (OPN) mRNA and protein expression were detected by RT-PCR and Western blotting. Calcium deposition was visualized by Alizarin stain method. RESULTS: The transfection efficiency was around 55% with a concentration of Cbfα-1 siRNA 100 nmol/L and Lipo 8 µl/ well. Cbfα-1 siRNA 1952 was chosen as the effective sequence with a suppression ratio up to 81.8%. At 24 and 48 h post-transfection, the expression of Cbfα-1 mRNA was significantly lower in siRNA transfection group than that in high phosphate group (0.335 ± 0.059 vs 0.714 ± 0.106, 0.574 ± 0.036 vs 0.726 ± 0.086, all P < 0.01) . At 48 and 72 h post-transfection, the expression of Cbfα-1 protein in siRNA transfection group was significantly lower than that in high phosphate group (both P < 0.01) . While Cbfα-1 gene was silenced by siRNA in siRNA transfection group, the mRNA and protein expression of OPN significantly declined (all P < 0.05) and calcium deposition in cell layers decreased. CONCLUSIONS: Cbfα-1 siRNA can effectively inhibit the expression of Cbfα-1 mRNA and protein in VSMC and thus suppress the transformation of VSMC into osteoblast-like cells and calcification induced by high phosphate. Cbfα-1 may become a potential therapeutic target in vascular calcification of chronic kidney disease.