Integration of Single-Cell and Spatial Transcriptome Sequencing Identifies CDKN2A as a Senescent Biomarker in Endothelial Cells Implicating Hepatocellular Carcinoma Malignancy

Purpose: Hepatocellular carcinoma (HCC) is one of the most malignant tumors worldwide, remaining difficult to manage. The role of cellular senescence in HCC has been gradually recognized. The present study aimed to comprehensively analyze the senescence-related features of HCC in single-cell and spatial dimension. Methods: Integration of single-cell RNA sequencing (scRNA-Seq) and spatial transcriptome sequencing (ST) data enabled identification of senescence-related genes (SRGs) for HCC. A novel prognostic model based on SRGs was constructed, and spatial distribution of senescent endothelial cells (ECs) within HCC tumors was analyzed. Then, the role of senescent ECs was verified through in vitro and in vivo experiments. Results: Significant heterogeneity in senescence profiles of ECs was observed, with specific regulatory alterations existing the communication between senescent ECs and other cell types. Spatial analysis revealed senescent ECs mainly located in the core region of HCC. The interaction of senescent ECs and immune cells implicated their role in tumor progression and immunotherapeutic response. In addition, CDKN2A was identified as an independent risk factor for HCC prognosis by constructing a prognostic model. CDKN2A promoted senescence of ECs which exhibited a secretory phenotype. Furthermore, senescent ECs with CDKN2A overexpression promote the malignancy of HCC both in vitro and in vivo. Conclusion: The present study advances understanding of senescent ECs in HCC pathogenesis, providing an insight into evaluation of ECs senescence as a therapeutic target for HCC.