杜仲黄酮类单体槲皮素、芦丁及金丝桃苷促SD大鼠骨髓间充质干细胞成骨分化的实验研究-外科学(骨外)专业论文.docxVIP

摘 要 目的： 通过对杜仲黄酮类单体的深入研究筛选出杜仲黄酮类单体中具有促进成骨 分化作用的单体并对其作用的分子机制做初步研究。 方法： 实验选取杜仲黄酮类单体槲皮素、芦丁、金丝桃苷协同经典成骨诱导剂诱 导培养第 3 代 SD 大鼠骨髓间充质干细胞，依次分为 7 组分别为 50ug/ml、10ug/ml、 2ug/ml、0.4ug/ml、0.08ug/ml 单体+成骨诱导液组、阳性对照组（诱导剂为 10mmol/l 的β-甘油磷酸钠，10-7mmol/l 地塞米松，50mg/l 维生素 C）、阴性组(加入与诱 导液等体积的 PBS)；各组于诱导分化后 24 天停止培养，并分别于第 7 天、14 天、24 天进行碱性磷酸酶染色及碱性磷酸酶活力测定、实时荧光定量 PCR（real time PCR）检测、茜素红染色，分别鉴定成骨细胞标志性分泌物碱性磷酸酶、检 测成骨分化关键转录因子 Runx2 及 Osterix 表达水平及成骨分化终产物矿化结 节，最后通过 SPASS 软件统计分析各单体促成骨分化的作用。 结果： 结合染色、碱性磷酸酶活力测定及成骨分化关键转录因子表达水平得知杜 仲黄酮类单体槲皮素、芦丁及金丝桃苷在一定浓度范围内协同经典成骨诱导剂 能够促进 SD 大鼠 BMSCs 成骨分化。 结论： 杜仲黄酮类单体槲皮素、芦丁、金丝桃苷协同经典成骨诱导剂能够增强 SD 大鼠 BMSCs 成骨分化。 关键词：杜仲 槲皮素 芦丁 金丝桃苷 成骨分化 诱导 大鼠 II ABSTRACT Objective： Through in-depth study of the flavonoids monomers selects flavonoids monom -ers which promote osteogenic differentiation and do a preliminary study of the molecular mechanism of action. Methods： Experimental monomers flavonoids quercetin、 rutin and hyperin are selected which is designed to collaborate with classic osteogenic inducers induce the P3 SD rBMSCs ，Turn divided into seven groups，respectively 0.08ug/ml、0.4ug/ml、 2ug/ml、10ug/ml、50ug/ml + classic osteogenic inducers, the positive control group (inducer of 10mmol / l of β-glycerol sodium、10-7mmol / l dexamethasone、50mg / l vitamin C), negative group (joined with an equal volume of fluid induced PBS)； Each group stopped at 24 days after the induction of differentiation culture， respectively， in the first 7 days，14 days，24 days alkaline phosphatase staining and activity assay， real-time quantitative PCR (real time PCR)， Alizarin red staining were identified osteoblast marker alkaline secretions detect osteogenic differentiation and Osterix key transcription factor Runx2 expression and osteogenic differentiation of the final product mineralized nodules，finally，statistical analysis of the role and contribute to the differences of each monomer osteogenic differentiation through SPASS software. Result： Combined staining and two key transcription factors expressi