Deciphering Stem Cell From Apical Papilla–Macrophage Choreography Using a Novel 3-dimensional Organoid System

Immune cell-mesenchymal stem cell crosstalk modulates the process of repair and regeneration. In this study, a novel heterogeneous cell containing a matrix-based 3-dimensional (3D) tissue construct was used to study the interactions between stem cells from apical papilla (SCAPs) and macrophage for a comprehensive understanding on the cellular signaling mechanisms guiding inflammation and repair.SCAPs and macrophages were seeded with collagen in 3D-printed molds to generate self-assembled tissue constructs, which were exposed to 3 conditions: no stimulation, lipopolysaccharide (LPS), and interleukin (IL)-4 from 0 to 14 days. Specimens from each group were evaluated for cellular interactions, inflammatory mediators (IL-1β, tumor necrosis factor [TNF]-α, macrophage-derived chemokine [MDC], macrophage inflammatory protein [MIP]-1β, monocyte chemoattractant protein [MCP]-1, IL-6, IL-8, transforming growth factor [TGF]-β1, IL-1RA, IL-10), expression of surface markers (CD80, 206), transcription factors (pSTAT1, pSTAT6), and SCAP differentiation markers (dentin sialophosphoprotein [DSPP], dentin matrix acidic phosphoprotein 1 [DMP-1], and alizarin red) using confocal laser scanning microscopy and multiplex cytokine profiling from 2 to 14 days.SCAP and macrophages displayed a cytokine-mediated interaction and differentiation characteristics. The increased pro-inflammatory cytokines/chemokines, IL-1β, TNF-α, MDC, and MIP-1β, in the earlier phase followed by the higher ratio of pSTAT6/pSTAT1 and decreased CD206 (P < .05), indicated a distinct polarization behavior in macrophages during repair in the LPS group. Conversely, the equal ratio of pSTAT6/pSTAT1, late increase in CD206, and amplified secretion of IL-1RA, IL-10, and TGF-β1 (P < .05) in the anti-inflammatory environment, directed alternative macrophage polarization, promoting SCAP differentiation and tissue modeling in IL-4 group.The novel 3D organoid system developed in this study allowed a comprehensive analysis of the SCAP-macrophage interactions during inflammation and healing, providing a deeper insight on the periapical dynamics of the immature tooth.