干细胞技术构建具组织学结构的工程化食道及其在动物体内原位修复的机制

Stem cell has developed to become a promising biotechnology. It is also becoming a perspective cell source for tissue engineering and regenerative medicine. Based on our previous study on separately construct of tissue engineered esophageal mucosa and muscle, in this proposal, we will combine these two architectures to constitute a whole esophagus organ with the bridge of decellularized esophageal mucosa-submucosa extracellular matrix (DECM), using bone marrow mesenchymal stem cells (MSC) as the cell source. DECM is used as the matrix, on which biocompatible polymer or/and protein will be electrospun to form nanofibrous lumen. Basement membrane major proteins like collagen (IV), laminin, entactin and proteoglycan extracted from an esophageal mucosa will be coated to simulate a normal epithelium "niche", aiming at promoting the epithelium regeneration. On the other hand, tubular scaffold with endo-circular and ex-longitudinal micro-channel patterns has been fabricated to guide the growth and differentiation of muscle tissue with the normal esophageal architecture. MSC will be seeded on the various layers of the scaffold and studied the mechanism of differentiation into epithelial cells and muscle cells individually and simultaneously, in vitro and in vivo. We believe that this kind of composite scaffold and mesenchymal stem cell will be a promising pathway for esophagus remodeling and regeneration with histological structure and biological functions. These studies will provide invaluable experimental bases for the future clinical attempt.

干细胞技术发展至今已成为生物技术领域最具有发展前景和后劲的前沿技术，也是组织工程和再生医学领域中最具可操作性的种子细胞之一。本申请基于本团队前期在食道组织工程中粘膜层和肌层分别成功构建的基础上，首次提出以骨髓间充质干细胞（MSC）为种子细胞，以脱细胞的食道粘膜细胞外基质为"Bridge"， 以"层/层对接"的方法，将上皮层和具有内环外纵的复层肌组织支架分别与原食道粘膜层和肌层组织缝接。其中上皮层采用电纺丝技术，以生物可降解聚合物和蛋白质为基材，将从动物食道粘膜中提取的基膜蛋白Coating于电纺丝纤维，营造类似体内的"Niche" 以获得具功能的上皮层组织；而肌层是通过具天然消化道肌组织的内环形外纵形复层图案化支架以引导肌细胞的生长和分化而制成。在体外充分研究MSC分别和同时向上皮细胞和肌细胞诱导分化机制的基础上，重点在动物体内进行原位诱导再生，以最终构建具有组织学结构和生物功能的再生食道

干细胞技术发展至今已成为生物技术领域最具有发展前景和后劲的前沿技术，也是组织工程和再生医学领域中最具可操作性的种子细胞之一。本申请基于本团队前期在食道组织工程中粘膜层和肌层分别成功构建的基础上，提出以骨髓间充质干细胞（MSC）为种子细胞，以脱细胞的食道粘膜细胞外基质为“Bridge”， 以“层/层对接”的方法，将上皮层和具有内环外纵的复层肌组织支架分别与原食道粘膜层和肌层组织缝接。其中上皮层采用电纺丝技术，以生物可降解聚合物和蛋白质为基材，将从动物食道粘膜中提取的基膜蛋白Coating于电纺丝纤维，营造类似体内的“Niche” 以获得具功能的上皮层组织；而肌层是通过具天然消化道肌组织的内环形外纵形复层图案化支架以引导肌细胞的生长和分化而制成。在体外充分研究MSC分别和同时向上皮细胞和肌细胞诱导分化机制的基础上，重点在动物体内进行原位诱导再生，以兔子为主要实验动物，实验结果获得了具功能的食管生物支架，以GFP转染本体动物原代骨髓间充质干细胞，证实了干细胞在体内的分化机制和组织形成及功能发挥的有效性。