| Abstract |
We isolated lung epithelial cells from an adult male Wistar rat by pronase
digestion and cultured them in Ham's F12 medium containing 5% FBS,
insulin, transferrin, EGF, aFGF and cholera toxin. Out of a mixed cell
population containing type II pneumocytes and ciliated cells as the major
cell types, one cell type was established as a spontaneously immortalized
cell line, This adherent cell line expresses different cytokeratins (CK)
as intermediate filaments, which is specific for epithelial cells. The
predominant appearance of CK 14 and 15, determined by western blotting at
passage 42, points to a possibly bronchial basal cell origin. During
ageing the postconfluent monolayers produce a squamous cell layer that
reacts with a desmoplakin antibody, which recognizes desmosomal structures
that are typical for epithelial cells. In addition, markers for type II
cells, Clara cells and fibroblasts are lacking. The cell line was passaged
more than 40 times within 9 months with population doubling times that
decreased from 31 h to 19 h with increasing passage number. These rat
bronchial epithelial cells (RaBE) are able to grow at cloning density on a
plastic surface with a CFE of 36 +- 3.6% (mean&SD, n=10, 200 cells/100 mm
dish), Chromosome analysis revealed a shift from 62 to 75 chromosomes
(median, n=20-50) between passage 9 and 19. Although immortalized, the
cell line did not change to a transformed phenotype, as indicated by the
strong dependency on serum and growth factors. Whether these cells
represent pluripotent stem cells that are able to differentiate under
complex culture conditions is part of our current investigation. This rat
bronchial epithelial (RaBE) cell line is now well established and might be
useful for cell communication or toxicity tests, and possibly for mutation
assays, which remains a subject for future studies.
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