Keratin species in type II pneumocytes in culture and during lung injury

Abstract

A detailed understanding of alveolar epithelial cell transitions during remodeling after lung injury requires the identification of specific markers. We have developed a panel of monoclonal antibodies against species of the intermediate filament protein, keratin. These individual species are recognized markers of the state of differentiation of various epithelial cells. These and complementary protein analytic methods have been applied to studies of isolated, enriched Type II pneumocyte preparations as well as to normal and injured lung tissues. Monoclonal antibody 24A3, initially raised against Morris hepatoma 7777 keratins, decorated a filament network in isolated cultured rat Type II pneumocytes by indirect immunofluorescence; it reacts by 2-dimensional polyacrylamide gel immunoblot procedures with an acidic, 46,000-dalton keratin. Monoclonal antikeratin antibodies AE1 and AE3, raised against human epidermal keratins, reacted poorly with isolated Type II cells; however, AE3 reacted by immunoblot technique with the 55,000-dalton keratin subclass. The bronchial epithelium reacted intensely with 24A3 as well as with a mix of AE1 plus AE3 in ethanol-fixed, paraffin-embedded sections of normal and injured rat lung. Alveolar regions of normal lung reacted poorly with all 3 antibodies, however, as visualized by light microscopy. At the same time, very large, presumptive epithelial cells in the alveolar regions stained intensely with 24A3 3 days after intratracheal instillation of bleomycin, whereas thin cells lining the alveoli in injured regions were intensely reactive 14 days after bleomycin treatment. These elongated cells may represent Type II pneumocytes in the process of converting to Type I cells.