Transcriptomic Identification of Renin-Angiotensin System-Related Candidate Biomarkers and External Testing of a...

This work aimed to identify candidate renin-angiotensin system (RAS)-related blood transcriptomic biomarkers associated with hypertension, construct an externally tested candidate diagnostic model, and validate selected genes in an Ang II-induced endothelial cell model. Two public microarray datasets, GSE75360 and GSE74144, were analyzed. Differential expression analysis was performed using limma, followed by GO/KEGG enrichment, preranked GSEA, CIBERSORT immune infiltration analysis, protein-protein interaction and regulatory network construction, and machine learning-based feature selection using logistic regression and random forest. A logistic regression model based on the selected genes was developed in GSE75360 and externally tested in GSE74144. Experimental validation was performed in Ang II-induced HUVECs using qRT-PCR, western blotting, ELISA, and CST3/FURIN loss- and gain-of-function assays, followed by CCK-8, Transwell, inflammatory, oxidative stress, and endothelial function analyses. In GSE75360, 173 differentially expressed genes were identified, including 18 RAS-related differentially expressed genes. Eight candidate genes, LRP1, CTSD, MTHFR, AUTS2, FURIN, CST3, FCER1G, and TBXAS1, were selected by combined machine learning analyses. Enrichment and immune infiltration analyses indicated that these genes were mainly associated with immune and inflammatory signatures. The eight-gene model showed high discrimination in GSE75360 and retained moderate performance in GSE74144. In Ang II-treated HUVECs, most candidate genes were upregulated at the mRNA level, and CST3, FURIN, and TBXAS1 were further validated at the protein level. CST3 and FURIN modulation altered Ang II-induced endothelial viability, migration, expression of inflammatory/adhesion markers, ROS accumulation, and eNOS/NO-related functional readouts. This study identified candidate RAS-related biomarkers and immune-associated signatures in hypertension and developed an externally tested candidate diagnostic model. The in vitro findings support the functional relevance of CST3 and FURIN in Ang II-induced endothelial responses, warranting further clinical and mechanistic validation.