论文部分内容阅读
目的研究Wnt/胞内β-连锁蛋白(β-catenin)信号通路对脂肪组织特异性丝氨酸蛋白酶抑制剂(vaspin)介导大鼠骨髓间充质干细胞(BMSCs)成骨分化的影响。方法体外培养4周龄雄性SD大鼠的BMSCs,分为4组:对照组,实验1、2、3组,分别加入成骨诱导液、含50 ng/ml、100 ng/ml vaspin的成骨诱导液及含100 ng/ml vaspin成骨诱导液+Dickkopf1(DKK1,Wnt/β-catenin信号通路特异性阻断剂),于成骨诱导第7天酶标仪检测碱性磷酸酶(ALP)活性,逆转录-聚合酶链反应(RT-PCR)检测成骨分化相关基因:ALP、核心蛋白结合因子2(RUNX2)、成骨细胞特异性转录因子Osterix(OSX)mRNA及β-catenin的表达水平。结果体外培养BMSCs,并加入不同浓度vaspin诱导后,实验1、2组ALP活性较对照组增加,并呈vaspin浓度依赖性(P均<0.05);成骨分化基因ALP、RUNX2、OSX mRNA表达水平增加,亦呈vaspin浓度依赖性(P均<0.05);β-catenin表达水平仅在加入100 ng/ml vaspin时升高(P<0.05);加入DKK1后(实验3组),ALP活性及成骨分化各基因表达水平较实验2组(100 ng/ml vaspin)显著下降(P均<0.05),β-catenin表达水平也显著下降(P<0.05)。结论 Vaspin可通过Wnt/β-catenin通路促进大鼠BMSCs成骨分化。
Objective To investigate the effect of Wnt / β-catenin signaling pathway on the osteogenic differentiation of rat bone marrow mesenchymal stem cells (BMSCs) induced by adipose tissue-specific serine protease inhibitor (vaspin). Methods BMSCs of 4-week-old male Sprague-Dawley rats were cultured in vitro and divided into 4 groups: control group, experimental group 1, group 2, and group 2, respectively. Osteoinductive solution containing 50 ng / ml vaspin of 100 ng / ml Induced osteogenic differentiation and Dickkopf1 (DKK1, Wnt / β-catenin signaling pathway-specific blockers), and alkaline phosphatase (ALP) was detected by microplate reader on the 7th day after osteogenic induction. The expression of ALP, RUNX2, Osterix (OSX) mRNA and β-catenin in osteoblast were detected by RT-PCR and RT-PCR. Level. Results After BMSCs were cultured in vitro and induced with different concentrations of vaspin, ALP activity in experimental group 1 and 2 was increased compared with control group (P <0.05), and the expression of ALP, RUNX2 and OSX mRNA (P <0.05). The expression of β-catenin increased only when 100 ng / ml vaspin was added (P <0.05). After addition of DKK1 (group 3), ALP activity and Compared with the experimental group 2 (100 ng / ml vaspin), the gene expression of osteoclasts decreased significantly (all P <0.05) and the expression of β-catenin decreased significantly (P <0.05). Conclusion Vaspin can promote the osteogenic differentiation of BMSCs via Wnt / β-catenin pathway.