补骨脂对破骨细胞-成骨细胞耦联功能的影响

Study the Effect of Fructus Psoraleae on the Coupling of Osteoclast-osteoblast

目的:探讨补骨脂水提物在破骨细胞抑制情况下对破骨细胞-成骨细胞耦联功能的影响及其潜在的作用机理。方法:制备补骨脂水提物,利用MTT法测定补骨脂水提物对RAW264.7细胞的毒性;在RANKL诱导的BMMs破骨细胞分化模型中测定补骨脂水提物对破骨细胞分化的抑制作用,并收集上清液制备条件培养基(CM);采用补骨脂干预后的CM作用于MC3T3-E1前体成骨细胞的间接共培养模型评价补骨脂对破骨细胞-成骨细胞耦联功能的影响;利用RT-qPCR法测定破骨细胞分化关键基因以及破骨细胞源性耦联因子的mRNA表达水平。结果:结果显示,补骨脂水提物在无细胞毒性的浓度下剂量依赖性的抑制破骨细胞分化;破骨细胞-成骨细胞耦联功能研究显示补骨脂在破骨细胞分化抑制的情况下能够维持破骨细胞-成骨细胞耦联功能,其CM分化液不影响MC3T3-E1成骨细胞分化能力;RT-qPCR结果表明补骨脂水提物抑制破骨细胞关键基因DC-Stamp、Oscar和Trap的mRNA表达,促进破骨细胞源性耦联因子C3a mRNA的mRNA表达水平。结论:补骨脂水提物能够在抑制破骨细胞分化抑制的情况下维持正常的破骨细胞-成骨细胞耦联功能,其机制可能与促进破骨细胞源性耦联因子的C3a mRNA表达水平有关。

Objective:This study aimed to investigate the effect and potential mechanism of fructus psoraleae water extract on the coupling function of osteoclasts and osteoblasts under the condition of osteoclast inhibition.Methods:The water extract was prepared and evaluated the toxicity against BMMs cells using MMT assay.The inhibitory effect of fructus psoraleae water extract on osteoclast differentiation was determined in a RANKLinduced BMMs osteoclast differentiation model.And the supernatant was collected to prepare conditioned medium(CM).Then an indirect co-culture model of MC3T3-E1 pre-osteoblasts treated with CM was used to evaluate the effect of osteoclast-osteoblast coupling function.RT-qPCR was used to measure the mRNA expression levels of key genes related to osteoclast differentiation and osteoclast-derived coupling factors.Results:MTT assay showed that the cytotoxic concentration of fructus psoraleae was greater than 125μg/mL.Fructus psoraleae inhibited osteoclast differentiation in a dose-dependent manner at non-toxic concentrations,with IC50 value of 7.15±0.98μg/mL.The study of the coupling function of osteoclasts and osteoblasts showed that fructus psoraleae could maintain this function under the condition of osteoclast differentiation inhibition,and its CM differentiation solution did not affect the differentiation and mineralization ability of MC3T3-E1 osteoblasts.RT-qPCR results showed that fructus psoraleae inhibited the mRNA expression of key genes related to osteoclast differentiation,such as DC-Stamp,Oscar and Trap,and promoted the mRNA expression level of osteoclast-derived coupling factor C3a.Conclusion:Fructus psoraleae has the ability to preserve the normal function of osteoclastosteoblast coupling while suppressing osteoclast differentiation.This mechanism may be attributed to the upregulation of C3a mRNA expression in osteoclast-derived coupling factors.