Integrated analysis of cell-in-cell related genes and immune microenvironment in heart failure

Background: Heart failure (HF) is a major global public health challenge, and its pathogenesis involves the regulation of a complex immune microenvironment (IME). Cell-in-cell (CIC), as a non-classical form of cell-cell interaction, has been extensively studied in fields like oncology, but its role in HF remains unclear. This study aimed to systematically analyze the expression patterns and functions of CIC-related genes (CRGs) within the HF immune microenvironment. Methods: Based on transcriptomic data from public databases, CIC-related differentially expressed genes (DEGs) between HF and healthy samples were identified. Three machine learning algorithms-Random Forest, LASSO, and SVM-RFE-were employed to screen diagnostic markers and construct a nomogram model. Consensus clustering analysis was used to stratify HF patients into distinct subtypes based on CRG expression, and their immune infiltration characteristics were compared. Single-cell transcriptomic data were utilized to validate the cellular localization of key genes within the HF microenvironment. Experimental validation of key CRGs was performed using a transverse aortic constriction (TAC)-induced HF rat model. Results: A total of 21 CIC-related DEGs were identified. A diagnostic model comprising 10 core genes demonstrated high predictive performance in both the training and validation sets. Based on CRG expression, HF patients were classified into two subtypes: Subtype A was enriched with regulatory T cells and M2 macrophages, exhibiting an immunosuppressive and fibrotic phenotype; Subtype B was dominated by cytotoxic T cells and NK cell infiltration, displaying an immune-activated phenotype. Single-cell analysis revealed high expression of CTSK in fibroblasts and enrichment of GZMB in T/NK cells. Animal experiments confirmed the upregulation of LPAR2 and GZMB and the downregulation of IL-10 in the TAC model. Conclusion: CIC-related genes possess significant diagnostic value in HF and can distinguish HF subtypes with distinct immune microenvironment features. CRGs may participate in HF progression by regulating immune cell infiltration and fibrotic processes, providing a new perspective for understanding HF heterogeneity and developing targeted immunotherapies.