Melatonin Rescues Enamel Defects Induced by Maternal Circadian Disruption via Targeting the BMAL1-JNK3 Axis

INTRODUCTION AND AIMS: Developmental defects of enamel (DDE) are highly prevalent, yet effective preventive measures remain elusive. The process of amelogenesis exhibits intrinsic circadian rhythmicity. While the photoperiod-regulated hormone melatonin is a recognized modulator of bone formation, its potential role in coordinating the circadian aspects of enamel biomineralization remains incompletely understood. This study aimed to decipher the impact of aberrant photoperiods on enamel development and to identify melatonin's regulatory targets. METHODS: We assessed the impact of maternal circadian disruption on offspring enamel mineralization using 3D X-ray microscopy (3DXRM), ATR-FTIR, SEM, Western blot, and qRT-PCR. Plasma melatonin concentration was measured by ELISA. Melatonin was administered to pregnant mice under circadian disruption to investigate its effects on enamel mineralization in their offspring. To investigate the molecular mechanisms downstream of BMAL1, we performed Bmal1 knockdown in ameloblast-lineage cells (ALCs) and used RNA sequencing and siRNA transfection. RESULTS: Circadian disruption impaired enamel mineralization (reduced thickness/density, increased CO₃²⁻/PO₄³⁻ ratio, rough surface with disorganized prisms), which was partially rescued by melatonin. Bmal1 knockdown in ALCs disrupted mitochondrial respiration, increased JNK3 phosphorylation, and reduced mineralization; melatonin restored mitochondrial function and suppressed JNK3 overactivation. CONCLUSION: Mechanistically, melatonin inhibits JNK3 phosphorylation to restore enamel mineralization. Bmal1 knockdown impairs the mitochondrial respiratory electron transport chain, while melatonin restores its function, thereby suppressing JNK3 overactivation and ultimately promoting mineralization. CLINICAL RELEVANCE: The BMAL1-JNK3 axis is a therapeutic target for enamel mineralization, providing a novel theoretical basis for perinatal circadian regulation.