In this study, we explored the role of Lhx6 in RA-induced cleft palate by a series of in vivo and in vitro experiments, and elucidated for the first time that Lhx6 affects the proliferative and migratory function of mesenchymal cells by regulating mitochondrial function, including increased rod-shaped mitochondria, decreased mtDNA copy number and ATP levels, and increased mtROS levels. The gene discussed is LHX6; the disease is cleft palate.