组织工程化三维支气管模型体外再造的实验研究

Reconstruction of Tissue Engineered 3D Bronchial Model in Vitro

目的以人支气管上皮细胞系和人胎肺成纤维细胞系为种子细胞、胶原凝胶复合Matrigel为支架材料,探索采用体外构建组织工程化三维支气管模型的可行性,为研究支气管上皮细胞和成纤维细胞相互作用提供模型。方法将支气管上皮细胞和肺脏成纤维细胞在添加Matrigel的液态Ⅰ型胶原中共培养,应用静态拉伸系统体外构建支气管模型。通过大体观察、相差显微镜观察、HE染色和免疫组织化学染色(广谱CK等)的方法对细胞的生长情况及形成的三维组织结构进行鉴定。结果在体外应用静态拉伸系统能构建出组织工程化三维支气管模型。大体观察可以看到收缩良好的组织片层,并具有一定韧性;相差显微镜下可以观察到网状结构的形成,细胞活性良好;HE染色表明了三维网状结构的形成;免疫组织化学染色广谱CK和Vimentin呈阳性。结论体外成功构建出组织工程化三维支气管模型,种子细胞共培养可在支架材料中完成极性生长过程并形成网状结构。

Objective To reconstruct tissue-engineered 3D bronchial model using human bronchial epithelial cells and human embryo lung fibroblast as seeding cells, and liquid collagen mixed Matrigel as scaffold. Methods Human bronchial epithelial cells and human embryo lung fibroblast were mixed with liquid collagen supplementing with matrigel and casted in 12-wells plate to reconstruct cells-collagen sheet. Macroscopic observation, phase-contrast microscopy observation, routine HE staining and immunohistochemistry staining（CK ets） were employed to assess the engineered 3D model. Results We reconstructed engineered 3D bronchial model successfully in vitro by tissue engineering techniques and exerted static stretch onto the collagen sheet. From Macroscopic observation, we gained contracted well sheet. We also observed network structure in phase-contrast microscopy meanwhile the viability of cells was fine. HE staining showed the formation of 3D network structure. The immunohistoehemistry staining of CK and Vimentin were positive. Conclusion We reconstructed engineered 3D bronchial model successfully in vitro and seeding cells could implement polarity growing in the scaffold materials then gained the network structure.