【摘要】 目的 探究咖啡对大鼠骨髓间充质干细胞(bone marrow mesenchymal stem cells,BMMSCs)成骨分化的影响，并初步阐明其与Wnt/β-catenin信号通路的关系。方法 通过全骨髓贴壁法分离并培养大鼠BMMSCs，制作不同浓度咖啡条件培养基，碱性磷酸酶染色筛选最适作用浓度，通过qRT-PCR和Western blot观察成骨相关指标变化，并检测β-catenin基因和蛋白的表达水平变化。结果 成功分离并培养出大鼠BMMSCs。ALP活性检测筛选出0.1 mg/m L为咖啡条件培养基最适作用浓度，进一步实验发现，其在前3 d对大鼠BMMSCs增殖无显著影响，在5～9 d时抑制大鼠BMMSCs增殖。咖啡条件培养基抑制大鼠BMMSCs成骨指标的表达，同时抑制Wnt/β-catenin信号通路有关蛋白的表达。结论 咖啡条件培养基可抑制大鼠BMMSCs成骨分化，其作用机制可能与抑制Wnt/β-catenin信号通路有关。更多还原

【Abstract】 Objective To investigate the effect of coffee on the osteogenic differentiation of rat bone marrow mesenchymal stem cells( BMMSCs) and preliminarily elucidate its relationship with the Wnt/β-catenin signaling pathway. Methods Rat BMMSCs were isolated and cultured using the whole bone marrow adherence method. Coffee-conditioned medium was prepared at different concentrations,and the optimal concentration was selected based on alkaline phosphatase( ALP) staining. Changes in osteogenesis-related markers were observed via qRT-PCR and Western blot,and alterations in the gene and protein expression levels of β-catenin were detected. Results Rat BMMSCs were successfully isolated and cultured. ALP activity assay identified 0.1 mg/m L as the optimal concentration of coffee-conditioned medium. Further experiments showed that this concentration had no significant effect on the proliferation of rat BMMSCs during the first 3 days but inhibited their proliferation from Day 5 to 9. Coffee-conditioned medium suppressed the expression of osteogenic markers in rat BMMSCs and concurrently downregulated the expression of proteins related to the Wnt/β-catenin signaling pathway. Conclusion Coffee-conditioned medium can inhibit the osteogenic differentiation of rat BMMSCs,and its mechanism may be associated with the suppression of the Wnt/β-catenin signaling pathway.更多还原