Lumican from DESC spheroids enhances odontoblastic differentiation of SCAPs.

AIM: During the tooth-root elongation phase, dental epithelial stem cells (DESCs) cluster together and release various signaling molecules, facilitating the differentiation of stem cells from the apical papilla (SCAPs) into odontoblasts. This study aimed to investigate the regulatory mechanism of secretory proteins from DESCs cultured in a spherical system mimicking the in vivo microenvironment on the odontoblastic differentiation of SCAPs during root development, providing theoretical foundations and potential targets for dentin regeneration therapy based on epithelial-mesenchymal interactions. METHODOLOGY: DESCs were isolated from the root tips of rodent incisors and cultured using low-attachment surface to mimic their in vivo clustering. The effects of spheroid culture (versus monolayer culture) on SCAPs viability and odontoblastic differentiation potential were systematically compared. Label-free mass spectrometry and bioinformatics analyses were employed to characterize the secretory protein profiles of DESCs under different culture conditions. In vitro and in vivo experiments validated the role of a key differential protein Lumican and its downstream signaling mechanisms. RESULTS: Compared to monolayer culture, spheroid culture significantly enhanced the secretion of Lumican by DESCs. Lumican demonstrated the ability to significantly promote dentin differentiation of SCAPs in vitro and in vivo. Mechanistic analysis revealed that Lumican binds specifically to integrin β1 (ITGβ1) on the SCAPs membrane, thereby activating the downstream ERK signaling pathway to mediate its pro-differentiation effects. CONCLUSION: This study illustrates that spheroid culture can restore the epithelial microenvironment of DESCs, leading to the release of Lumican, which targets SCAPs. This research uncovers the crucial roles of Lumican and ITGβ1 in DESCs-SACPs interactions and paves the way for potential therapeutic strategies in dentin regeneration.