Maintenance of the differentiated type II cell characteristics by culture on an acellular human amnion membrane

We have developed a culture system for guinea pig alveolar type II cells us ing an epithelium-denuded human amnion membrane as a substratum. The differ entiated morphology was maintained for 3 wk by both air-interface feeding a nd immersion feeding when type II cells were cultured on the basement membr ane side of the amnion with fibroblasts on the opposite side (coculture). F unctionally, high levels of surfactant protein B (SP-B) and C (SP-C) messen ger ribonucleic acids (mRNAs) were expressed even after the 3-wk cultivatio n and surfactant protein A mRNA was detected on day 10 of the culture. The differentiation was also maintained when fibroblasts were cultured on lower chambers of the culture plates (separate culture). In contrast, culture of type II cells without fibroblasts (monoculture) could not preserve the mat ure morphology. When the monoculture was supplemented with keratinocyte gro wth factor or hepatocyte growth factor, a monolayer of rather cuboidal type II cells with apical microvilli was maintained. However, the percent area of lamellar bodies in these cells was significantly less than that in fresh ly isolated type II cells, and mRNA expressions of SP-B and SP-C were also considerably suppressed. These findings suggest that other growth factors o r combinations of these factors are necessary for the maintenance of the di fferentiated phenotype. As substratum, a permeable collagen membrane or a t hin gel layer of Engelbreth-Holm-Swarm mouse sarcoma extracts did not prese rve the mature characteristics. This culture system using an acellular huma n amnion membrane may provide novel models for research in type II cells.