Impact of intercellular adhesion molecule 1-positive dental pulp stem cells in deep caries progression

Dental pulp stem cells (DPSCs) regulate immune responses; however, their heterogeneity in deep caries remains unclear. This study aimed at investigating the role of intercellular adhesion molecule 1-positive DPSCs (ICAM1⁺ DPSCs) within the immune microenvironment of deep carious pulp tissue to develop therapeutic strategies.

Single-cell sequencing was used to compare cellular profiles between deep caries and healthy pulp tissues. ICAM1⁺ DPSCs were quantified using immunofluorescence/flow cytometry in human/mouse models and sorted for functional analyses. Odontogenic differentiation was assessed using alkaline phosphatase/Alizarin Red staining, while inflammatory mediator production was assessed using RT-qPCR, Western Blot, ELISA, SCENIC and RNA-seq. THP-1 was cultured in conditioned media from ICAM1⁺ DPSCs and ICAM1^- DPSCs. RT-qPCR, Western Blot and flow cytometry were used to assess the proportion of proinflammatory to reparative THP-1. Macrophage-derived cytokines (IL-1β/4/6/10 and TNF-α) were tested for DPSCs to ICAM1⁺ differentiation induction.

Cellular profiling showed a significant increase in ICAM1⁺ DPSCs and proinflammatory monocytes in deep carious dental pulp tissue, with ICAM1⁺ DPSCs closely interacting with mononuclear macrophages. Immunofluorescence and flow cytometry confirmed the increase in ICAM1⁺ DPSCs in deep caries in the affected human and mouse pulp tissue. Alkaline phosphatase and Alizarin Red staining, SCENIC, RT-qPCR, Western Blot and ELISA revealed decreased odontogenic differentiation in ICAM1⁺ DPSCs and increased expression of CEBPD, IL-6, CCL2 and CXCL10 in ICAM1⁺ DPSC cells. RT-qPCR, Western Blot and flow cytometry indicated an elevated proinflammatory to reparative THP-1 ratio for THP-1 that was cultured in ICAM1⁺ DPSC-conditioned media for 1-3 days.

During deep caries progression, TNF-α drives the transformation of DPSCs into inflammatory ICAM1⁺ DPSCs. This subcluster exhibits impaired odontogenic differentiation capacity, secretes proinflammatory cytokines and chemokines, and enhances macrophage inflammatory activity, contributing to the advancement of deep caries lesions.