钟状期成釉细胞的体外生长研究  

Growth pattern of ameloblasts in bell stage of human deciduous tooth germ in-vitro culture system

作　　者：田华[1] 吕平[1] 高岩[1] 陈苹[2] 周春燕[2] 高学军[1] 

机构地区：[1]北京大学口腔医学院.口腔医院牙体牙髓科,北京100081 [2]北京大学基础医学院生物化学与分子生物学,北京100081

出　　处：《牙体牙髓牙周病学杂志》2009年第6期309-313,共5页Chinese Journal of Conservative Dentistry

基　　金：国家自然科学基金资助项目(30572033)

摘　　要：目的:通过对乳牙牙胚成釉细胞的体外培养生长情况的研究,了解成釉细胞的生长和分化过程。方法:解剖分离因疾病原因引产遗弃的19周胎儿下颌骨,半侧固定后拍摄数字X线影像,连续切片HE染色;另半侧未固定的组织分离乳牙牙胚成釉器,胶原酶消化法制成细胞悬液,采用差速贴壁和无血清培养基选择性培养成釉细胞。培养的乳牙胚成釉细胞通过细胞角蛋白14免疫细胞染色鉴定,并以RT-PCR方法检测釉质基质蛋白的表达。结果:X线影像显示牙胚冠部有硬组织形成,组织学乳牙牙胚处于钟状期,可见分化的成釉细胞分泌大量细胞外基质,釉牙本质界处釉质已矿化。无血清培养基中牙源性上皮的生长分化良好,细胞角蛋白14染色阳性,RT-PCR显示有釉蛋白、釉原蛋白和成釉蛋白的表达。结论:采用无血清培养基可以选择性的培养乳牙胚钟状期成釉细胞,能够较好反映成釉细胞的生长分化状态。钟状期分化的成釉细胞进入细胞外基质分泌活跃期,并在釉牙本质界处最早出现釉质的矿化。AIM ： To set up a culture system to observe the growth pattern of human ameloblast - like cells, in order to understand the initiation and differentiation of ameloblast. METHODS ：A mandible of 19 weeks human fe- tus was obtained and studied with approval of Ethic Committee of Peking University. X - ray digital image was taken. Then the consecutive sections were prepared with one half of the mandible, while with the other half, enamel organ epi- thelials were separated and digested with collagenase. The ameloblast - like cells were selectively cultured by differenti- al adhering method and in K - SFM media. The expression of enamel matrix proteins was analyzed by RT - PCR and immunocytochemisty. RESULTS：Bell stage tooth germs were recognized. In this stage, the differentiated ameloblast aligned orderly and secreted large amounts of extracellular matrix. Mineralization initiated from enamel - dentinal junc- tion. Cells isolated from the tooth organ, mostly are ameloblast - like ceils, proliferated and differentiated in K - SFM. The ameloblast-like cells were immunopositive for cytokeratin 14. Enamelin, amelogenin and ameloblastin were de- tected by RT - PCR. CONCLUSION： K - SFM media could be used selectively to culture ameloblast - like cells. This study demonstrated that in bell stage the ameloblast secreted enamel matrix, and the earliest mineralization oc- curred in the enamel - dentinal junction.

关 键 词：成釉细胞 体外培养 釉质矿化 基质蛋白 

分 类 号：R780.2[医药卫生—口腔医学]