Heterogeneity of T cells in periapical lesions and in vitro validation of the proangiogenic effect of GZMA on HUVECs

Abstract Aim T cells are key immunomodulatory cells in periapical lesions. This study aimed to explore the roles of T cells in chronic apical periodontitis (CAP) using single‐cell RNA sequencing and to further investigate Granzyme A (GZMA) in angiogenesis regulation. Methodology A total of five CAP samples were collected for single‐cell RNA sequencing. We performed subcluster and lineage‐tracing analyses for T cells. According to differential gene expression, distinct biological functions enriched in T cells of CAP were presented by gene set enrichment analysis (GSEA) and compared with healthy gingiva (data obtained from the GEO database). CellChat was used to explore potential ligand–receptor interactions between T cells and endothelial cells in CAP. The coculture of primary human umbilical vein endothelial cells (HUVECs) and Jurkat T cells, as well as the addition of GZMA recombinant protein, was used to validate the predicted pair of GZMA and coagulation factor II thrombin receptor (F2R) by RT‐PCR, angiogenesis and migration assays. Results A transcriptomic atlas of 44 746 individual cells was constructed from the periapical lesions of five patients with CAP by single‐cell RNA‐seq, and eight cell types were identified. We identified nine subsets of T cells and deciphered the cellular heterogeneity of T cells in CAP at the functional level by subclustering and GSEA. Lineage tracing revealed a distinct lineage of T cells in CAP and predicted the transition of the T cellular state upon CAP. GSEA revealed multiple biological processes and relevant angiogenesis genes upregulated in CAP T cells. GZMA‐F2R pairs were predicted by cell–cell interactions in CAP. High expression of GZMA and F2R was observed in the coculture of HUVECs and Jurkat T cells, and the proangiogenic capacity of the GZMA recombinant protein was emphasized by in vitro experiments. Conclusions Our study provides novel insights into the heterogeneity of T cells in periapical lesions and reveals the potential role of GZMA in T cells in regulating angiogenesis in HUVECs.