重组人骨形态发生蛋白2促进髁状突囊内骨折山羊骨髓间充质干细胞的成骨分化

Recombinant human bone morphogenetic protein 2 promotes osteogenic differentiation of bone marrow mesenchymal stem cells from intracapsular condylar fracture goats

背景:国内外学者均认为髁状突囊内骨折后形成颞下颌关节强直的原因主要是大量成骨的过程。评估不同类型髁状突囊内骨折后骨髓间充质干细胞的成骨潜能,为创伤性颞下颌关节强直提供细胞生物学及分子学证据。目的:探讨重组人骨形态发生蛋白2对不同类型髁状突囊内骨折山羊骨髓间充质干细胞成骨分化的影响。方法:16只健康山羊双侧关节随机分为4组,8只山羊为左侧A型髁突囊内骨折组(A组)+右侧B型髁突囊内骨折组(B组),另外8只为左侧C型髁突囊内骨折组(C组)+右侧对照组(D组),分别在术后1,2,3,6个月各时间点A、B组处死2只,C、D组处死2只,即每个时间点共处死4只,分离、提取第3代骨髓间充质干细胞,根据是否进行重组人骨形态发生蛋白2诱导分为诱导组和非诱导组,按照试剂盒说明检测培养7,10 d时的碱性磷酸酶活性及骨钙素含量,诱导培养1周后进行碱性磷酸酶染色。结果与结论:与非诱导组比较,4组骨髓间充质干细胞使用重组人骨形态发生蛋白2诱导后,碱性磷酸酶活性及骨钙素含量明显升高(P<0.05);B组诱导后碱性磷酸酶活性及骨钙素含量较A、C、D组明显升高(P<0.05)。结果表明,体外分离培养的山羊骨髓间充质干细胞在重组人骨形态发生蛋白2诱导作用下表现出明显的成骨活性,且B型髁突囊内骨折后的骨髓间充质干细胞成骨活性明显高于A型、C型髁突囊内骨折。

BACKGROUND:Intracapsular fracture of the condyle may lead to traumatic temporomandibular joint ankylosis,which is mainly related to massive bone formation.Herein,we evaluated the osteogenic potential of bone marrow mesenchymal stem cells after different types of intracapsular condylar fractures,to provide cell biology and molecular evidence for traumatic temporomandibular joint ankylosis.OBJECTIVE:To induce the osteogenesis of bone marrow mesenchymal stem cells after different types of goat intracapsular condylar fractures in vitro by recombinant human bone morphogenetic protein-2.METHODS:Bilateral mandibular condyles of 16 healthy goats were randomly divided into 4 groups:8 goats were used to make models of type A intracapsular condylar fractures of the left condyle(group A)and models of type B intracapsular condylar fractures of the right condyle(group B),and the other 8 goats were used to make models of type C intracapsular condylar fractures of the left condyle(group C)and models of type D intracapsular condylar fractures of the right condyle(group D).One rat from each group was sacrificed at the following intervals,1,2,3 and 6 months postoperatively.Passage 3 bone marrow mesenchymal stem cells were isolated,extracted and induced by recombinant human bone morphogenetic protein-2(induction group)or not(non-induced group),and cultured according to the kit’s instruction.The alkaline phosphatase activity and osteocalcin content were measured at 7 and 10 days after induced culture,and alkaline phosphatase staining was performed at 1 week after induced culture.RESULTS AND CONCLUSION:Compared with the corresponding non-induced group,the alkaline phosphatase activity and the osteocalcin content in groups A,B,C,and D after induction with recombinant human bone morphogenetic protein-2 were significantly increased(P<0.05).The alkaline phosphatase activity and osteocalcin content in group B were significantly higher than those in groups A,C,and D after induced by recombinant human bone morphogenetic protein-2(P<0.05).To conclude,the use of recombinant human bone morphogenetic protein-2 can trigger marked osteoblast differentiation of goat bone marrow mesenchymal stem cells.Moreover,the osteogenic activity of bone marrow mesenchymal stem cells after type B intracapsular condylar fractures was superior to that after type A and C intracapsular condylar fractures.