人牙囊细胞与胶原凝胶复合煅烧骨三维培养的实验研究

目的:建立人牙囊细胞(dentalfolliclecells,DFCs)的体外三维立体培养模型。方法:取第4代DFCs分别 接种于煅烧骨(sinteredbovinebone,SBB)(A组)、胶原凝胶(B组)、胶原复合煅烧骨三维支架上(C组),并以二维 培养的DFCs(D组)作对照,1周、2周取材,扫描电镜观察细胞形态以及与材料的贴附情况,细胞计数观察细胞在 材料上的增殖情况,组织化学方法检测DFCs的碱性磷酸酶(ALP)活性。结果:扫描电镜见A组、B组和C组细胞 均完全伸展,但C组细胞数量明显增多,细胞外基质分泌增加,优于A组、B组和对照组D组,而对照组细胞在二维 培养条件下复层生长呈膜状结构,膜表面部分细胞脱落死亡,细胞外基质分泌较实验组少。A组与C组的ALP活 性无差异(P>0.05),但显著高于对照组B组和对照组(P<0.05)。1周时3组细胞Ⅰ型胶原合成情况无明显差 异,但2周时C组细胞的Ⅰ型胶原平均灰度值明显高于其他2组(P<0.01)。结论:A、B、C组对DFCs的贴附、增 殖,分化均有影响,但C组作支架最优。胶原凝胶与煅烧骨的复合支架更有利于牙囊细胞的增殖贴附和分化。

An in Vitro Study of Three Dimensional Culture of Human Dental Follicle Cells

Objective: To investigate the effect of sintered bovine bone combined with gel on differentiation and proliferation of human dental follicle cells in vitro. To establish three dimensional cultured model of human dental follicle cells. Methods: Human dental follicle cells were collected and seeded on three-dimensional sintered bone, collagen gel and sintered bone combined with collagen gel in vitro. One and two weeks after set up , dental follicle cells attachment and morphology were observed by scanning electronic microscope(SEM), the proliferation of the cells was evaluated by cell counting, Alkaline phosphatase(ALP) activity was tested; bone sialoprotein (BSP) synthesis were examined by immunohistechemistry staining. Results: SEM(scanning electronic microscope) showed the porous structure of the two materials and the eugenic growth of cells in the scaffolds. DFCs adhered and proliferated on the three-dimensional scaffolds. Conclusion: It is feasible to establish three -dimensional cultured model of human DFCs by seeding the cells on the scaffolds of sintered bone combined with collagen gel. This may be useful for further study as the scaffolds of periodontal tissue engineering.