Isolation and characterization of a spontaneously arising long-lived line of human keratinocytes (NM1) |
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Authors: | Howard P Baden Joseph Kubilus Joseph C Kvedar Mark L Steinberg Sandra R Wolman |
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Institution: | (1) Harvard Medical School, Department of Dermatology, Massachusetts General Hospital, 02114 Boston, Massachusetts;(2) Department of Pathology, New York University Medical Center, 550 First Avenue, 10016 New York, New York |
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Abstract: | Summary The long-lived keratinocyte line, NM1, was isolated from the epidermis of a pool of foreskins obtained from apprently, normal
neonates at the time of circumcision. Cultures were initiated in Dulbecco’s minimal essential medium containing 20% fetal
bovine serum, 0.4 μg/ml hydrocortisone, 10−9
M cholera, toxin, and 10 ng/ml epidermal growth factor using mitomycin C-treated 3T3 cells as a feeder layer. Unlike normal
keratinocytes which survive for only 150 generations these cells have been in culture for more than a year and have been carried
for more than 400 doublings. The cells seem to follow a pathway, of growth and differentiation that is very similar to normal
keratinocytes. Cytokeratin fibrils, intercellular attachments, and cornified envelopes were observed. The keratin polypeptides
isolated from the NM 1 cells were similar to those previously described in normal cultured, cells; the presence of profilaggrin
and involucrin was demonstrated by sodium dodecyl sulfate electrophoresis and immunoblotting with monoclonal antibodies specific
to these proteins. The NM 1 cells showed a reduced dependency on 3T3 feeder cells but did not form tumors when placed into
athymic nude mice. Screening of the cells for SV40, BK, HPV 16, and HPV 18 viruses was negative. The NM1 cells showed trisomy
of chromosome 8. The long-lived nature of these cells makes them a valuable model for studying growth and differentiation
of kerationocytes. |
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Keywords: | keratinocyte long-lived keratinization epidermis |
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