The Undifferentiated State Of Alveolar Epithelial Type Ⅱ Cells Maintain In Mouse Embryonic Fibroblast Cells In Vitro

OBJECTIVE:1.To learn the effect that mouse embryonic fibroblast being feederlayer to maintain the undifferentiated state of alveolar epithelial type IIcells.2.To establish the methods of passaging and culturing alveolarepithelial type II cells in vitro with feeders.3.To understand the role of leukemia inhibitory factor in culturingand maintaining AT2cells on the growth of its undifferentiated state.METHODS:With10ug/ml mitomycin C treated primary mouse embryonicfibroblasts to lose proliferative capacity,used feeder cells culturedalveolar epithelial type II.Identified by immunofluorescence primaryAT2cells,passaged AT2cells cultured on feeders，and passaged AT2cells without feeders.By staining AT1marker proteins-aquaporin5andAT2marker of alveolar surfactant protein C,we observed and comparedthe state of staining AT2cells which cultured on feeders and withoutfeeders.RESULTS1.The AT2cells seeded on MEFs forming colonies on day3,and canbe continuously passaged to the third generation,while the AT2cellspassaged without MEFs can not form paving stone-likecolonies,morphology is flat and extravagant. 2.Immunofluorescence shows that passaged AT2cells seeded onfeeders are SPC positive,AQP5negative,but the passaged cells which didnot seed on feeders are SPC negative,AQP5positive.CONCLUSIONAT2cells can be maintained in undifferentiated state mitomycin Ctreated MEFs as feeder layer in a certain extent.