NR2F2 and its contribution to lymph node metastasis in oral squamous cell carcinoma

Abstract

Objectives

To investigate the role of cancer stem cells (CSCs) in lymph node metastasis (LNM) of oral squamous cell carcinoma (OSCC), focusing on the expression and biological significance of nuclear receptor subfamily 2 group F member 2 (NR2F2).

Methods

Single-cell RNA sequencing data from OSCC patients were analyzed using the CytoTRACE algorithm to assess stemness. Gene set scores were calculated with the irGSEA and GSVA R packages. GO and KEGG analyses identified enriched pathways. NR2F2 and CD44 expression in OSCC and lymph nodes (LNs) were validated via immunohistochemistry and immunofluorescence. NR2F2/Nr2f2 overexpression and knockdown cell lines were established, with stemness markers confirmed by Western blot. Functional assays evaluated stemness, proliferation, migration, and invasion capabilities of OSCC cells. In vivo experiments evaluated the ability of NR2F2 to promote tumor growth and metastasis. Bulk RNA sequencing and drug sensitivity analyses explored NR2F2-related mechanisms and drug responses.

Results

CSCs in OSCC were divided into five subgroups, with NR2F2 identified as the key gene in CSC4, the subgroup with the highest stemness, and found to be overexpressed in metastatic LNs. Immunohistochemistry showed NR2F2 overexpression in OSCC, associated with LNM. Immunofluorescence confirmed co-expression of NR2F2 and CD44 in metastatic OSCC and LNs. Overexpression of NR2F2 enhanced stemness, proliferation, and migration of OSCC cells. In vivo experiments showed that NR2F2 promoted the growth and LNM of OSCC. Bulk RNA sequencing revealed that NR2F2 is involved in multiple pathways and plays a role in LNM. Trametinib was identified as a sensitive drug.

Conclusion

NR2F2 is associated with the maintenance of tumor stemness and may influence LNM in OSCC by promoting tumor cell proliferation and migration.