GSDMD Promotes Epithelial-Mesenchymal Transition and Lymph Node Metastasis in Cervical Cancer in vitro and in vivo.

BACKGROUND: Lymph node metastasis (LNM) is a critical determinant of prognosis in cervical cancer (CC). Tumor-associated macrophages (TAMs) and inflammatory signaling within the tumor microenvironment have been implicated in metastatic progression; however, the molecular mechanisms linking inflammation to lymphatic dissemination remain poorly understood. METHODS: We integrated single-cell RNA sequencing (scRNA-seq) of paired primary tumors and metastatic lymph nodes tissues from a single representative patient with clinical histological validation. Functional roles of gasdermin D (GSDMD) were evaluated using in vitro assays, including proliferation, migration, invasion, ELISA, and RNA sequencing, as well as in vivo xenograft and lymph node metastasis models. RESULTS: Single-cell transcriptomic profiling revealed significant activation of pyroptosis-related pathways in metastatic lymph nodes, accompanied by enhanced tumor-stromal interactions. Clinical tissue validation confirmed that GSDMD protein was significantly enriched in metastatic lesions compared to primary tumors. In vitro functional experiments demonstrated that GSDMD promotes epithelial-mesenchymal transition (EMT), enhances migration and invasion, and upregulates VEGFA and VEGFC expression. Mechanistically, GSDMD facilitated the release of pro-inflammatory cytokines such as IL-1β, contributing to a pro-metastatic microenvironment. Notably, GSDMD knockdown suppressed lymph node metastasis in vivo despite promoting primary tumor growth, indicating a context-dependent role in tumor progression. CONCLUSION: Our findings reveal that GSDMD contributes to the pro-metastatic phenotype of cervical cancer. GSDMD dysregulation correlates with activated EMT and lymphangiogenic signaling, potentially facilitating tumor lymphatic dissemination. This study highlights GSDMD as a promising therapeutic candidate for restraining cervical cancer metastatic progression.