Experimental Study On Replacement Of Canine Cervical Esophagus With Tissue-engineered Esophagus Constructed By Seeding Autologous Cells On PLGA Scaffolds

Objects: To isolate, culture and proliferate autologous seed-cells. To induce autologous bone marrow mesenchymal stem cells(BMSCs) to differentiate into epithelial cells(ECs). To design and manufacture artificial biodegradable scaffolds and construct tissue-engineered esophagus(TEE) by seeding autologous cells on them. To replace a segment of canine cervical esophagus with TEE. Methods: EECs, myofibroblasts(MFbs) and BMSCs were cultured and proliferated in vitro. BMSCs induced to differentiate by epidermal growth factor(EGF) were identified by immunocytochemistry. Scaffolds made of poly(D,L-lactic-co-glycolic) acid(PLGA) by the method of particulate leaching were assessed by histology and electron microscope. Cells were seeded on scaffolds statically or dynamically and cultured in vitro or in vivo to construct TEE. The cervical esophagus of canis familiaris was partly replaced with TEE and gastrostomy feeding was conducted after operation. The dogs were sacrificed at different time points and the specimens were assessed by histology, immunohistochemistry and electron microscope. Results: Autologous seed-cells were isolated and proliferated successfully. The BMSCs could differentiate into cells which were cytokeratin and epithelial membrane antigen positive. The PLGA scaffold had suitable mechanical strength. It's pore size was 180~280μm and porosity was about 90%. The cells could adhere to the scaffold tightly and grow into TEEs. Collagen IV could promote aderence between seed-cells and PLGA scaffolds. The dogs could survive long term after operation. The transplatation site expanded in the short term and tended to be normal afterward. In one month, glands could be observed and the muscular layer could be observed in two months in the operative site. Conclutions: Autologous seed-cells can be harvested and proliferated in vitro conveniently. The porous PLGA scaffolds are suitable for TEE. Seeding autologous seed-cells on PLGA scaffolds can construst TEEs suitable for substituting a segment of the autologous cervical esophagus.