外胚间充质细胞向神经胶质/雪旺细胞的诱导分化及在BAM生物材料的存活研究

Differentiation of ectomesenchymal cells to glial/schwann cells and the survival of cells on BAM biomaterials

目的探讨体外诱导外胚间充质细胞向神经胶质/雪旺细胞分化的潜能,观察细胞在不同材料上的生长状况,利用分化细胞观察在BAM生物材料的复合情况,为进一步构建组织工程神经提供理论依据。方法取妊娠10.5d胎鼠颌突外胚间充质细胞,纯化后分别用8.5mg/L牛脑垂体提取物BPE、50μmol/L弗司扣林(forskolin)及联合应用诱导细胞分化。相差显微镜下观察细胞的形态变化,免疫组化鉴定细胞性质,将诱导后的细胞种植于胶原膜及胶原凝胶,并于培养3、6d后分别进行光镜、HE染色及扫描电镜观察细胞的生长情况。结果培养3d后外胚间充质细胞形态发生变化,细胞形态为梭形,突起变长。而对照组细胞形态未见明显改变;免疫组化显示外胚间充质细胞表达S-100蛋白,但无GFAP的表达。而诱导分化后出现GFAP的表达,其中BPE及Forskolin联合应用后分化效果最好,约为79.2%。而在BAM凝胶和膜上细胞都可伸展并增殖。结论BPE和Forskolin可不同程度诱导外胚间充质细胞向神经胶质细胞分化。在BAM膜上分化细胞生长状态良好,为该细胞与BAM生物材料构建组织工程神经导管提供了理论依据。

Objective To explore the effect of bovine pituitary extraction（BPE） and forskolin on the differentiation of ectomesenchymal cells to glial/sebwann cells and the survival of cells on the BAM biomaterials. Methods 8.5mg/L BPE and 50μmol/L forskolin were used alone and combination to induce differentiation of ectomesencbymal cells in vitro. The morphology was observed by phase-contrast microscopy, lmmunohistochemistry was used to identify the characteristics of cells. Then, these cells were implanted to different type of BAM. The absorptivity,growth and migration of the ectomesenchymal cells were observed with HE staining and scanning electronic microscope. Results There were great changes in the morphology of the cells treated by BPE and forskolin. Immunohistochemistry showed that most of differentiated ectomesenchymal cells were GFAP positive staining cells compared to the GFAP negative staining to normal ectomesenchymal cell. Conclusion The ectomesenchymal cells can be differentiated to glial cells by BPE and forskolin. Three dimension scaffolds with differentiated cells these findings will preparation for the construct of tissue engineering nerve conduit.