MATURATION OF UNDIFFERENTIATED LUNG EPITHELIAL-CELLS INTO TYPE-II CELLS IN-VITRO - A TEMPORAL PROCESS THAT PARALLELS CELL-DIFFERENTIATION IN-VIVO

Background: Formation of alveolar-like structures (ALS) by mature feta l rabbit type II pneumocytes (day 29 gestation) and long-term differen tiation on Engelbreth-Holms-Swarm mouse tumor extract or EHS gel (Matr igel) were reported by our group (Blau et al., 1988. J. Cell Physiol., 136:203-214). We now describe structural organization and differentia tion of immature lung epithelial cells, isolated at day 22 gestation, into mature type II cells in vitro. Methods: Peripheral pulmonary tiss ue was pooled and undifferentiated epithelial cells isolated for prima ry culture on Matrigel. Cells were examined 12-16 h after plating and on days 1, 3, 5, and 7 of culture and assessed by phase contrast and b y transmission electron microscopy after fixation in situ. Results: Ce lls formed ALS 12-16 h after plating. Spherule diameter increased abou t four to eight times from day 1-7 in culture. There was rapid transfo rmation of tall columnar cells to cuboidal, normal polarization of cel ls with respect to cell-free lumen of ALS, progressive reduction of gl ycogen zones, apparent gradual increase of cell organelles such as Gol gi apparatus, rough endoplasmic reticulum and mitochondria, and appare nt extrusion of lipidic figures into the lumen. These morphologic tran sformations in vitro temporally paralleled cell differentiation in viv o. The relative increase of C-14-acetate precursor into phosphatidylch oline in contrast to cardiolipin was consistent with these transformat ions. Conclusions: Under the conditions of our culture system, maturat ion of undifferentiated pulmonary epithelial cells is reproduced in vi tro along the same time course and according to the same developmental sequence of fetal lungs in vivo. (C) 1994 Wiley-Liss, Inc.