Experimental Study On The Re-epithelization Of Tracheal Allotransplantation

Objective: To investigate the regeneration process of a well-differentiated and functional human airway epithelium of tracheal heterotopic transplantation.Methods: The experiment was divided into experimental and control groups, epithelial tissue has been removed by enzymatic dissociation, we perfused tracheal epithelial cells in the experimental group and normal saline in the control group, grafts were transplanted into a subcutaneous pouch in the dorsal surface of SD rats, two groups received cyclosporin A by intraperitoneal injection from beginning to end. Rats were killed and removed tracheal allografts at 1,2,3,4 weeks after transplantation. Histological analysis has been used to evaluate the integrity of airway epithelium, submucosal infiltration of inflammatory cells, fibrous tissue hyperplasia and the degree of luminal obliteration. Immunohistochemical staining for specific markers (CK-14, CK-18 and CFTR) were performed to characterize airway cell differentiation and Ki-67 was performed to characterize airway cell proliferation; scan electron microscope (SEM) and transmission electron microscopy (TEM) was used to observe the ultrastructural changes of the cells.Results: one week after allografts implantation, most of tracheal inner surface was covered by non-ciliated flate cells, the regional areas of tracheal inner surface had no epithelial cells; two week after allografts implantation, most of tracheal inner surface was covered by non-ciliated flate cells, the regional areas of tracheal inner surface was covered by columnar epithelial cells; three week after allografts implantation, the entire of tracheal inner surface was covered by columnar epithelial cells, ciliogenesis could be observed in the regional areas of tracheal inner surface; four week after allografts implantation, the entire of tracheal inner surface was covered by ciliated columnar cells. Control group showed no epithelial cells and a large number of inflammatory cells in the lumen of trachea. Immunohistochemical staining showed that epithelial cells expressed CK14 and CK18 on all 4 time points; only four week after allografts implantation, epithelial cells expressed CFTR; one week after allografts implantation, Ki-67 expression was the highest, from two to four week after allografts implantation, Ki-67 expression was decreased gradually. Ultrastructural changes of epithelial cells were observed by scanning electron microscope and transmission electron microscopy: one week after allografts implantation, most of tracheal inner surface was covered by non-ciliated flate cells; at four week, the entire of tracheal inner surface was covered by pseudostratified ciliated columnar epithelium and epithelial cells were tight junction.Conclusions: It was feasible to complete the regeneration of a well-differentiated and functional human airway epithelium in rat tracheae allografts by the perfusion of rat tracheal epithelial cells; the process of airway epithelium regeneration was cell episporium, cell proliferation, cell differentiation, the formation of mature epithelium.