Single-cell landscape of the mouse HNSCC-related bone invasive lesion

To understand the invasive microenvironment of head and neck squamous cell carcinomas (HNSCCs) at a single-cell resolution, we performed single-cell RNA-sequencing (scRNA-seq) using cells derived from calvaria of tumor-free control mice and tumor-excised calvarial tissues from pre-invasive stage（scratch-negative）and invasive stage（scratch-positive）groups 1 week after the implantation of C57BL/6-derived HNSCC cell line HNM007 cells. Single-cell RNA-sequencing was performed on live cells, mixed CD45+ and CD45– cells in a 1:1 ratio, derived from calvarial tissues of tumour-free control mice and tumour-excised pre-invasive stage（scratch-negative）and invasive stage（scratch-positive）mice 1 week after the implantation of HNM007 cells. The HNM007 cells were implanted onto the calvaria with or without prior scratching of the bone surface using a 21 G needle. This procedure mechanically removes the periosteum.