Growth characteristics,morphology, and phospholipid composition of human type II pulmonary alveolar cells grown in a collagen-free microenvironment |
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Authors: | George E Milo G Adolph Ackerman Ronald L Sanders |
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Institution: | (1) Department of Physiological Chemistry, College of Medicine, The Ohio State University, 410 West 12th Ave., 43210 Columbus, Ohio;(2) Department of Anatomy, College of Medicine, The Ohio State University, 410 West 12th Ave., 43210 Columbus, Ohio;(3) The Ohio State University and Comprehensive Cancer Center, College of Medicine, The Ohio State University, 410 West 12th Ave., 43210 Columbus, Ohio;(4) Department of Anatomy and Cellular Biology, Tufts University School of Medicine, 136 Harrison Ave., 02111 Boston, Massachusetts |
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Abstract: | Summary Human lung epithelial cells have been isolated and maintained in pure culture and characterized during their time in culture.
Any residual fibroblasts were removed by selective trypsinization within the first 48 h in culture and the residual epithelial
cells from the primary culture grew to confluent density. The epithelial cells at Passage 2 or greater were serially subpassaged
when cultures reached ca. 80% confluency. This procedure permitted us to conduct biochemical and structural studies of starting
materials and subsequent population doublings.
Electron microscope evaluation of both initial monolayers and cell suspensions showed cultures to be composed of a single
cell type. These cells had microvilli on their free or apical surface. Subsequent population doubling level 1 up to 5 exhibited
the same structures. They contained lamellar inclusions, which are typical of Type II alveolar epithelial cells. Fetal lung
(age 18 to 20 wk) cell suspensions processed for electron microscopy before culturing showed cells to be undifferentiated,
epithelial-like with small microvilli along cell borders, and with desmosomes at cell junctions. Lamellar inclusions were
not observed in these cells. Ultrastructural studies of the cultured epithelial cells demonstrated that the lamellar inclusions
had a slightly positive reaction when tested for acid phosphatase. Phospholipid analysis of these lung epithelial cells showed
a phospholipid composition consistent with that found in surfactant-containing Type II cells. Cultured epithelial cells stained
with phosphine 3-R demonstrated a green fluorescent cytoplasm and nucleus with brightly fluorescent yellow-orange perinuclear
particles. The preceding characterization of these cells leads us to conclude that they exhibit structural and biochemical
features commensurate with Type II epithelial cells from human lung. Moreover, these selection techniques applied to the isolation
of human lung Type II cells from the tissue permit us to study the differentiative function of these cells routinely under
conditions of growth in vitro.
This work was supported in part by grants from EPA, R 806638-01 and 131-640-1599A1 |
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Keywords: | Type II lung cells phospholipid human lung fibroblasts |
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