Blue light-emitting diode promotes mineralization of stem cells from the apical papilla via cryptochrome 1/Wnt/β-catenin signaling

Abstract

This study aimed to determine whether low-intensity blue LED light (4 J/cm²) promotes mineralization of stem cells from the apical papilla (SCAPs) by modulating CRY1 expression and to elucidate the underlying molecular mechanisms. SCAPs identity was validated using flow cytometry. In a controlled experimental design, SCAPs were exposed to blue LED light, followed by quantitative assessment of CRY1 and osteogenic markers (Runx2, OSX, DSPP, DMP-1) via qRT-PCR, Western blotting, and osteogenic staining. To investigate the role of CRY1 in SCAPs osteogenic differentiation, CRY1 was overexpressed using lentiviral transfection. Additionally, the Wnt/β-catenin pathway was analyzed using specific inhibitors (XAV-939) to elucidate the underlying molecular mechanisms. Blue LED irradiation reduced CRY1 mRNA expression to 80% (day 7) and 45% (day 14) of control levels. CRY1 overexpression significantly increased CRY1 mRNA and protein levels (P < 0.001) but decreased ALP activity and ARS staining (P < 0.001). Blue LED treatment restored mineralization parameters to 80% of control levels. Key osteogenic genes (DMP-1, DSPP, Runx2, OSX) showed lower mRNA and protein levels in the CRY1 overexpression group compared to controls. Blue LED exposure increased these levels to 60–74% (mRNA) and 45–67% (protein) of control values. In the Wnt/β-catenin pathway, CRY1 overexpression elevated GSK-3β and reduced p-GSK-3β, β-catenin, and nuclear β-catenin levels. Blue LED treatment restored these levels to 33–54% of control values, indicating pathway activation. Inhibition of the Wnt/β-catenin pathway (using XAV-939) abolished differences in osteogenic gene expression and mineralization between CRY1 overexpression and blue LED-treated groups, confirming its critical role. Blue LED light at 4 J/cm² enhances SCAPs mineralization by modulating CRY1 expression and activating the Wnt/β-catenin pathway. These findings provide mechanistic insights into photobiomodulation (PBM) in bone regeneration and highlight its potential for tissue engineering and regenerative medicine.