Maturation Status of Rat Lung Epithelial Cells in Interspecies Chimeras Generated by Blastocyst Complementation

Abstract

This study investigates the maturation status of rat lung epithelial cells in interspecies chimeras generated via blastocyst complementation (BC) using Fgfr2b-knockout mouse embryos. In our previous study, we succeeded in generating rat-derived lung epithelium in interspecies chimeras using tetraploid complementation; however, the resulting cells remained immature and failed to support respiratory function. In this study, we established two Fgfr2b-KO mouse lines via CRISPR/Cas9 and injected GFP-labeled rat embryonic stem (ES) cells into blastocysts, which were then transferred to pseudopregnant female mice. Comparative histological analysis of airway spaces between wild-type rat lungs (E19.5–E21.5) and BC chimeras revealed that BC lungs reached a maturation stage comparable to E20.5–E21.5 rat lungs. Quantitative Polymerase Chain Reaction of key epithelial maturation markers—including ENaC subunits, Aqp5, and surfactant protein genes—demonstrated late-gestational upregulation in wild-type rat lungs, while expression levels in BC lungs exhibited significant inter-individual variability, corresponding to stages between E19.5 and E21.5 in wild-type rats. These findings suggest that the mouse host environment may partially promote maturation of rat-derived lung epithelial cells; however, additional mechanisms are likely required to achieve functional respiratory capacity.