Single-Cell Insights into Cisplatin Resistance Mechanisms in Bladder Cancer Tumor Microenvironment

Abstract This study integrates single-cell RNA sequencing (scRNA-seq) with in vitro experimental validation to elucidate the molecular mechanisms underlying cisplatin resistance in bladder cancer (BC) and to characterize cellular heterogeneity within the tumor microenvironment (TME). By analyzing scRNA-seq datasets from cisplatin-sensitive and cisplatin-resistant BC samples, we identified key resistant cell subpopulations and resistance-associated signaling pathways. Notably, pronounced heterogeneity was observed among resistant epithelial cells and fibroblasts, accompanied by extensive metabolic reprogramming involving glycolysis, DNA damage repair, and drug metabolism pathways. Cell–cell communication analysis revealed intensified interactions between resistant cell subsets and immune cells or fibroblasts within the TME, with significant activation of MIF, THBS, MHC-II, and FN1 signaling pathways. Developmental trajectory analysis further demonstrated the dynamic transition of fibroblasts from cisplatin-sensitive to resistant states. Survival analyses across multiple cancer types confirmed the prognostic relevance of resistance-associated genes, including SPINK1, PHGR1, and APOD. Functional validation using a cisplatin-resistant bladder cancer cell line showed marked upregulation of SPINK1 following resistance induction. SPINK1 knockdown significantly reduced the cisplatin IC50 and suppressed MIF signaling. Moreover, resistant tumor cells enhanced macrophage tolerance to cisplatin via the MIF axis, an effect that was reversed by pharmacological MIF inhibition. Collectively, this integrated single-cell and experimental study reveals critical resistant cell subpopulations, metabolic reprogramming features, and intercellular communication networks driving cisplatin resistance in BC, highlighting potential molecular targets for therapeutic intervention and drug development.