Integrative transcriptomic and single-cell analysis identifies anoikis-related molecular signatures in vascular calcification
Abstract Background Vascular calcification (VC) is a life-threatening complication of chronic kidney disease (CKD) driven by vascular smooth muscle cell (VSMC) osteogenic transdifferentiation. Anoikis, a form of adhesion-dependent apoptosis, is involved in cardiovascular remodeling, yet its regulatory role in CKD-associated VC remains unexplored. Methods We performed an integrative transcriptomic and single‑cell analysis using a human in vitro VSMC calcification model and a rat in vivo CKD‑VC model. Differentially expressed genes were identified under a unified threshold (FDR < 0.05, |log2FC| > 1). Strict one‑to‑one ortholog mapping was applied. Differentially expressed anoikis‑related genes were screened, followed by functional enrichment, three machine‑learning algorithms, and external validation. Results We identified 48 differentially expressed anoikis‑related genes, with the signal derived mainly from human VSMCs rather than cross‑species conservation. The cell adhesion molecule pathway was significantly dysregulated. Seven hub genes were identified: BDNF, CRYAB, CYP1B1, DAPK1, HAS2, PDGFRB, and PLAU. The seven‑gene model achieved an AUC of 0.811 in the independent validation cohort. Single‑cell analysis revealed cell‑type‑specific expression patterns, with the highest anoikis module scores in osteoblast‑like cells and macrophages. Conclusions This study identifies novel anoikis‑related molecular signatures associated with VC. These genes are involved in cell adhesion, VSMC homeostasis, chaperone‑mediated cytoprotection, and matrix remodeling. Our exploratory findings provide new insights into VC pathogenesis and require further functional and clinical validation in CKD‑specific cohorts.