Circulating Cytokeratin 5+ Progenitor Epithelial Cells Also Express Other Progenitor Cell Markers and Are Necessary for Normal Airway Repair.

Background: Human lung transplants have demonstrated recipient epithelial cells in donor organs, supporting the notion of a circulating epithelial progenitor cell. Resident progenitor epithelial cells in the proximal airway are located in the submucosal gland ducts and basal epithelium and express cytokeratin (CK) 5/14. We have identified circulating progenitor CK5/14 positive epithelial cells in the bone marrow and buffy coat of mice. These cells appear to traffic via the CXCR4/CXCL12 biologic axis. Purpose of study: To further determine additional markers of these circulating progenitor epithelial cells and to disrupt the CXCR4/CXCL12 axis to prevent recruitment of these circulating progenitor cells after airway injury. Methods used: Flow cytometric analysis of cytokeratin 5 (CK5), CXCR4, CD45, CD34, Sca-1, c-kit expressing cells in the buffy-coat and bone marrow of naive mice and CK5-GFP transgenic mice. Subcutaneous heterotopic implantation of dissected female wild type tracheas into male GFP recipient mice. Immunohistochemical and dual immunofluorescence analysis of GFP and CK5 as well as p63 in tracheal transplants. Summary of results: FACS analysis of bone marrow and buffy coat from CK5-GFP transgenic mice and wild type mice revealed that these circulating CK5+ progenitor epithelial cells also express Sca-1,CD34 and c-kit. In the heterotopic tracheal transplant model, wild type tracheal transplantation in CK5-GFP recipient mice demonstrated that the recipient circulating CK5-GFP cells contributed to repair of the airway epithelium. Passive immunization of animals bearing tracheal transplants with specific neutralizing anti-CXCL12 F(Ab)2 antibodies resulted in the unexpected phenotype of the airway epithelium demonstrating squamous metaplasia at day 21 post-transplant.. In contrast, animals bearing tracheal transplants passively treated with control antibodies demonstrated normal pseudostratified epithelium at day 21 post-transplantation. Immunofluorescence for p63 in the tracheal transplants demonstrated that all of the cells of the squamous metaplasia were p63 positive in tracheal transplants from animals that had CXCL12 depleted. Analysis of these tracheal transplants with immunohistochemistry and immunofluorescence for GFP and CK5 demonstrated that the squamous metaplasia was solely derived from resident progenitor epithelial cells. Conclusions: There is a population of CK5+ cells in the bone marrow and circulation of naive mice that also express Sca-1, CD34 and c-kit, which is compatible with a stem cell. These circulating CK5 positive cells contribute to regeneration of the airway epithelium and their recruitment to the progenitor cell niche in the airway appears to be critical for the development of normal pseudostratified columnar epithelium.