Growth and differentiation of tracheal epithelial progenitor cells

J. Y. Liu, P. Nettesheim and S. H. Randell Laboratory of Pulmonary Pathobiology, National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina 27709. The purpose of these studies was to determine whether both basal and secretory rat tracheal epithelial (RTE) cells served as multipotent epithelial progenitors and whether both cell types gave rise to a similar "poorly differentiated" cell during the early phase of epithelial regeneration in denuded tracheal grafts. Griffonia simplicifolia I (GSI) lectin and flow cytometry were used for cell sorting. More than 98% of GSI-positive cells expressed plasma membrane alpha 1-3 terminal galactose (Gal), and 95% contained keratin 14 (K14), phenotypic markers for basal cells; < 1% were secretory or ciliated cells. Less than 2% of the GSI-negative cells expressed Gal or K14, but this fraction contained 16% ciliated cells and 54-79% secretory cells, dependent on whether periodic acid-Schiff staining or binding of an anti-secretory cell monoclonal antibody (RTE 12) was used as the criterion. Equal numbers of viable cells from either fraction were inoculated into denuded tracheal grafts, which were studied on days 1-14. At 24 h, greater numbers of GSI-negative than -positive cells were found attached to the graft wall; the keratin staining pattern of the attached cells was similar to that of the parent cell populations, but monoclonal antibody-detectable secretory and ciliated cell epitopes, originally present in the GSI-negative fraction, were lost. 5-Bromo-2'-deoxyuridine uptake was not seen at 24 h, but by 48 h all epithelial cells from both fractions entered the cell cycle. From 48 to 96 h, cells derived from either fraction were ultrastructurally indistinguishable; they were poorly differentiated and highly proliferative, and all expressed Gal and K14. A mature epithelium evolved from the poorly differentiated cells in both sets of grafts, but secretory and ciliated cells appeared earlier in grafts inoculated with GSI-negative cells. The results strongly suggest that in this model of tracheal epithelial regeneration both basal and secretory cells "dedifferentiated" into a similar highly proliferative phenotype from which a mucociliary epithelium "redifferentiated."