Immune landscape and molecular regulators in renal interstitial fibrosis

Renal interstitial fibrosis (RIF) represents the final pathological hallmark of chronic kidney disease (CKD) and is a pivotal factor leading to irreversible renal failure in end-stage renal disease (ESRD). It is highly clinically relevant and underpinned by complex immune mechanisms. This study aims to elucidate the molecular underpinnings of renal interstitial fibrosis, identify potential diagnostic biomarkers, and propose novel therapeutic targets. By integrating transcriptomic data from public Gene Expression Omnibus (GEO) datasets related to renal interstitial fibrosis, we uncovered a tight correlation between immune cell infiltration and key gene expression patterns. Differentially expressed genes (DEGs) identified from two datasets underwent Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses, revealing significant involvement in immune responses, cell migration, and cytokine-mediated signaling pathways. Gene Set Enrichment Analysis (GSEA) further highlighted critical roles for chemokine signaling, immunoregulatory interactions, and interferon signaling in fibrotic progression. Construction of protein–protein interaction (PPI) networks using the STRING database and Cytoscape software led to the identification of several immune-related hub genes potentially central to fibrosis pathogenesis. These candidates were validated via merged dataset expression profiling and receiver operating characteristic (ROC) curve analysis. In addition, the significance of six hub genes (CD3D, CCL5, CXCL9, CCR5, IL7R, and CD2) in RIF was further validated through Quantitative Real-Time PCR and Western blot analyses in mouse models, as well as immunofluorescence staining in clinical patient samples. Finally, we performed regulatory network analysis involving microRNAs and transcription factors, revealing potential upstream modulators, and identified candidate therapeutic compounds through drug–gene interaction prediction and molecular docking. This study provides new insights into early diagnosis and immunotherapeutic strategies for renal interstitial fibrosis. • This study integrates transcriptomic data to elucidate the critical role of immune cell infiltration and chemokine signaling in renal interstitial fibrosis (RIF) pathogenesis. • We systematically identify six pivotal immune-related hub genes (CD3D, CCL5, CXCL9, CCR5, IL7R, and CD2) via PPI network analysis, which are rigorously validated as diagnostic biomarkers using ROC curves. • The clinical relevance of these hub genes is further confirmed through multi-level experimental validation, including qPCR and Western blot in mouse models, and immunofluorescence in human patient samples. • Beyond the core genes, we construct comprehensive upstream regulatory networks involving miRNAs and transcription factors, and propose novel therapeutic candidates via drug-gene interaction prediction and molecular docking.