软骨调节素调控BMSCs骨和软骨双向分化平衡的研究

The repair of cartilage, bone defects and complex defects perplexed clinical surgical specialties,tissue engineered bone and cartilage constructed by bone marrow stromal cells (BMSCs) provided a method to the challenge.However, tissue engineered cartilage was prone to ossify and could not maintain the stable cartilage phenotype after subcutaneous implantation.The results of our last National Natural Science Foundation of China showed that chondromodulin-1 play an important and key role in the inhibiting osteogenic differentiation and promoting chondrogenic differentiation of BMSCs.On this basis,this study is to further explore the specific mechanisms and related factors of chondromodulin-1 in the osteogenic and chondrogenic differentiation balance of BMSCs. First, to detect the difference of chondromodulin-1 in the tissue engineered bone and different subtypes of cartilage before and after transplantation, then to study the role of chondromodulin-1 in the endochondral ossification by suppression expression of chondromodulin-1 by shRNA on different subtypes of chondrocytes; Second, to construct engineered cartilage by interference and overexpression of chondromodulin-1 in BMSCs, and to detect the differences the extracellular matrix;At last, to regulate the balance of osteogenic and chondrogenic differentiation by the dual gene transfection of chondromodulin-1 and bone morphogenetic protein, so as to maintain the stable phenotype of engineered cartiage after subcutaneous implantation,and provide a theoretical basis for the construction of tissue engineering osteochondral graft.

软骨、骨缺损和复合缺损的修复是临床多个外科专业的难题，骨髓基质干细胞（BMSCs）诱导分化构建组织工程骨和软骨为解决难题提供了答案，但是构建的软骨植入体内后容易发生骨化难以维持稳定的表型。申请人的上一个自然基金结果表明软骨调节素在抑制BMSCs骨向分化和促进其向软骨分化的过程中起着关键的作用，本实验在此基础上进一步研究软骨调节素在BMSCs骨、软骨双向分化平衡中的具体机制和相关因素。首先检测体外构建组织工程化骨和不同类型软骨体内移植后软骨调节素表达的差异，然后通过shRNA干扰不同类型软骨细胞中软骨调节素的表达来研究其在不同类型软骨内骨化过程中的作用；再通过干扰和过表达软骨调节素的BMSCs构建骨和不同类型的软骨，检测各种细胞外基质的变化；最后通过骨形成蛋白和软骨调节素的双基因转染调控骨和软骨双向分化平衡，为组织工程软骨体内表型的维持,以及构建骨、软骨复合组织提供理论依据。

软骨、骨缺损和复合缺损的修复是临床多个外科专业的难题，骨髓基质干细胞（BMSCs）诱导分化构建组织工程骨和软骨为解决难题提供了答案，但是构建的软骨植入体内后容易发生骨化难以维持稳定的表型。申请人的上一个自然基金结果表明软骨调节素在抑制BMSCs 骨向分化和促进其向软骨分化的过程中起着关键的作用，本实验在此基础上进一步研究软骨调节素在BMSCs 骨、软骨双向分化平衡中的具体机制和相关因素。. 首先检测骨和软骨组织中软骨调节素表达的差异，结果表明软骨组织中的软骨调节素的表达明显高于骨组织中骨调节素的表达（p <0.05）.. 其次，我们通过shRNA 干扰骨髓间充质干细胞中软骨调节素的表达，发现其在成骨分化能力增强，而成软骨能力降低，这提示软骨调节素在间充质干细胞在干细胞向骨和软骨分化方向中起着调控作用。. 在体内实验中，我们通过干扰和过表达软骨调节素的BMSCs 构建骨和软骨，检测相关基因表达的变化，研究发现过表达软骨调节素的BMSCs构建的组织工程化软骨.在移植到体内过程中，能够明显抑制软骨血管化的发生，而干扰软骨调节素的BMSCs构建的组织工程化骨在移植到体内过程中，更加容易向骨组织分化。.综上所述，我们通过研究表明软骨调节素能够调控骨和软骨双向分化平衡，为组织工程软骨体内表型的维持,以及构建骨、软骨复合组织前期试验基础。