Abstract
Background: Pyroptosis is a form of programmed cell death characterized by inflammasome activation and the release of inflammatory cytokines, which induce a strong immune response. Unlike apoptosis, pyroptosis can elicit potent immune stimulation, potentially playing a crucial role in anti-tumor immunity. However, it may also promote tumor progression by altering the tumor microenvironment and facilitating immune evasion. This study investigates pyroptosis-related gene expression in hepatocellular carcinoma (HCC), with a focus on identifying key genes that influence prognosis and tumor microenvironment dynamics. Methods: Single-cell RNA sequencing (scRNA-seq) data from 10 HCC patients were obtained from the GEO database (GSE149614), along with spatial transcriptomic data and bulk RNA-seq data from TCGA. We performed data processing and quality control using the Seurat package and applied machine learning techniques, including LASSO regression, to identify key pyroptosis-related genes. Functional analyses, including Gene Ontology (GO), KEGG, and GSVA, were conducted to explore biological pathways. Pyroptosis levels were quantified across cell types, and survival analysis was performed to evaluate prognostic impacts. Cell communication and immune infiltration were also assessed to understand the tumor microenvironment. Results: We identified CHMP4B as a key pyroptosis-related gene in HCC, significantly associated with poor prognosis. High CHMP4B expression was correlated with shorter overall survival (OS) and disease-free survival (DFS). Functional enrichment analysis showed that CHMP4B is involved in cell cycle regulation, DNA repair, and cytoskeletal organization. Spatial transcriptomics revealed heterogeneous CHMP4B expression in the tumor microenvironment, with higher levels found in advanced tumor stages. Moreover, high CHMP4B expression was associated with increased infiltration of immunosuppressive cells, such as monocytes and macrophages, and upregulation of immune checkpoint molecules (PD-L1, CTLA4), suggesting its role in promoting immune evasion. Conclusions: Our findings highlight CHMP4B as a critical regulator of pyroptosis in HCC, influencing tumor progression and immune modulation. High CHMP4B expression may facilitate the development of an immunosuppressive microenvironment, enabling immune escape and tumor growth. The study underscores CHMP4B's potential as a prognostic biomarker and therapeutic target in HCC. However, the limited sample size calls for further validation using larger datasets and multi-omics approaches, such as proteomics and metabolomics, to fully elucidate its functional role in HCC pathogenesis.