Integrated analysis of single-cell RNA-seq and bulk RNA-seq reveals distinct cancer-associated fibroblasts in head and neck squamous cell carcinoma

BACKGROUND: The heterogeneity of cancer-associated fibroblasts (CAFs) in head and neck squamous cell carcinoma (HNSCC) has been widely acknowledged, but has not yet been elucidated. The potential roles and clinical relevance of CAFs subclusters in HNSCC progression remain obscure. METHODS: In this study, we combined single-cell and bulk tissue transcriptome profiles of HNSCC with clinical data from The Cancer Genome Atlas (TCGA). The Seurat package was used to perform single-cell RNA-seq analysis to distinguish distinct CAFs subtypes. Prognostic relevance of several CAFs markers was assessed and functional analysis was also performed. RESULTS: We identified eight CAFs subclusters; of these, seven showed enhanced expression levels in HNSCC tumor tissues compared to normal tissue, and three (clusters 0, 3, and 4) were associated with poorer overall survival. Further functional analysis revealed that cluster 0 was characterized by myofibroblasts with high alpha smooth muscle actin (aSMA) expression and enrichment in smooth muscle contraction. The cluster 3 exhibited expression of extracellular matrix (ECM)-related genes and was enriched in epithelial-mesenchymal transition (EMT)-related gene sets. Cluster 4 expressed high levels of the major histocompatibility complex (MHC) class II family, which was characterized as antigen-presenting CAFs. CONCLUSIONS: We determined CAFs heterogeneity in HNSCC. 8 CAFs subclusters were recognized and 3 of which were prognosis related. The 3 CAFs subclusters showed distinct phenotypes enriched in myofibroblast function, ECM remodeling and antigen-presenting function respectively.