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Caffeine has developmental toxicity. Prenatal caffeine exposure (PCE) caused intrauterine growth retardation (IUGR) and multiple organ dysplasia. This study intended to explore the effect and mechanism of PCE on long bone development in female fetal rats. In vivo, the PCE group pregnant rats were given different concentrations of caffeine during the gestational Day 9-20. The mRNA expression of osteogenesis-related genes were significantly reduced in PCE group. In the PCE group (120 mg/kg·d), the length and primary center of fetal femur wer