EMBRYONIC LUNG BRANCHING MORPHOGENESIS, CELL PROLIFERATION, AND DIFFERENTIATION ARE ALTERED IN EPIDERMAL GROWTH FACTOR RECEPTOR (EGF-R) -/- TRANSGENIC 129/SV × C57BL/6 MICE. • 406

EGF-R activity influences lung growth and cellular differentiation but its requirement in lung development is not understood. We used an EGF-R knockout model (129/Sv × C57BL/6 genetic background) to investigate the functional importance of the EGF-R in lung development. In this knockout model the E. coli Lac Z reporter gene is driven by the EGF-R promoter(Sibilia and Wagner, Science 269: 234-238, 1995). We studied in vivo the EGF-R expression pattern, and in culture the effect of EGF-R knockout on lung morphogenesis, cell proliferation and Hoxb-5 expression. EGF-R +/- and EGF-R-/- embryos (d10.5 to d14.5) were stained for β-galactosidase and lung histology prepared. Embryonic lung (d11.5) from EGF-R +/- matings were cultured for 72 hrs in serum- and hormone-free medium. Cultures were evaluated for branching morphogenesis, cell proliferation, and immunohistology of Hoxb-5. β-galactosidase staining of embryonic lungs showed that EGF-R is expressed in fibroblasts underlying epithelia that appear to be invading the mesenchyme to form new airways. In cultured embryonic lung, branching morphogenesis of EGF-R -/- lungs was 70% and ³H-Thymidine incorporation into DNA was 79% of EGF-R +/+ and EGF-R +/- lungs (Controls).³ H-Thymidine autoradiography of EGF-R -/- cultured lungs showed no effect of EGF treatment (20 ng/ml) on the intermittent epithelial and mesenchymal labelling seen in untreated Controls. Hoxb-5 immunostaining of Control lung cultures was present in mesenchyme and epithelia, with more prominent staining under branch points, especially after EGF treatment. In contrast, Hoxb-5 in EGF-R -/- lungs was decreased in both mesenchyme and epithelia, despite EGF treatment. In summary, EGF-R is expressed in embryonic lung next to airway branch points. Absence of the EGF-R in this mouse background leads grossly to decreased branching morphogenesis, decreased cell proliferation, and altered Hoxb-5 expression, effects not reversed by EGF. We conclude that EGF-R signaling is an important regulator of embryonic lung growth and cell differentiation. (Supported by HL 37930)