ScRNA-seq links dental fibroblasts heterogeneity with mechanoresponsiveness

Abstract

Background and objective: Periodontal ligament (PDL) and dental pulp (DP) share a common origin but have distinct biological and mechanical functions. To what extent the mechanoresponsive property of PDL can be attributed to its unique transcriptional profiles of cellular heterogeneity is unclear. This study aims to decipher cellular heterogeneity and distinct mechanoresponsive characteristics of odontogenic soft tissues and their underlying molecular mechanisms.

Materials and methods: A single-cell comparison of digested human periodontal ligament (PDL) and dental pulp (DP) was performed using scRNA-seq. An in vitro loading model was constructed to measure mechanoresponsive ability. Dual-luciferase assay, overexpression, and shRNA knockdown were used to investigate the molecular mechanism.

Results: Our results demonstrate striking fibroblast heterogeneity across and within human PDL and DP. We demonstrated that a tissue-specific subset of fibroblasts existed in PDL exhibiting high expression of mechanoresponsive extracellular matrix (ECM) genes, which was verified by an in vitro loading model. ScRNA-seq analysis indicated a particularly enriched regulator in PDL-specific fibroblast subtype, Jun Dimerization Protein 2 (JDP2). Overexpression and knockdown of JDP2 extensively regulated the downstream mechanoresponsive ECM genes in human PDL cells. The force loading model demonstrated that JDP2 responded to tension and that knockdown of JDP2 effectively inhibited the mechanical force-induced ECM remodeling.

Conclusions: Our study constructed the PDL and DP ScRNA-seq atlas to demonstrate PDL and DP fibroblast cellular heterogeneity and identify a PDL-specific mechanoresponsive fibroblast subtype and its underlying mechanism.

Keywords: JDP2; human periodontal ligament fibroblast heterogeneity; mechanoresponsive; single-cell RNA-sequencing.