Increasing cell density down-regulates the expression of acidic FGF by human RPE cellsin vitro
作者:
KitaokaTakashi,
BostLaurie M.,
IshigookaHitoshi,
AotakiAmy E.,
HjelmelandLeonard M.,
期刊:
Current Eye Research
(Taylor Available online 1993)
卷期:
Volume 12,
issue 11
页码: 993-999
ISSN:0271-3683
年代: 1993
DOI:10.3109/02713689309029225
出版商: Taylor&Francis
关键词: retinal pigment epithelium;acidic fibroblast growth factor (aFGF);gene expression;cell culture;growth factors;human
数据来源: Taylor
摘要:
Previous studies have reported the expression of acidic fibroblast growth factor (aFGF) by rat, bovine, and human retinal pigment epithelium (RPE)in vivo.To critically examine the expression of aFGF by RPE cells, we studied the density dependence of steady-state levels of mRNA and protein expressionin vitro.Northern blot analysis demonstrated 5 transcripts ranging from 4.5 kB to 1 kB. Steady-state levels of all the transcripts decreased as a function of culture density. A polyclonal antibody was raised against recombinant human aFGF and affinity purified on aFGF coupled to AffiGel-10. The resulting antibody crossreacted with bFGF but not FGF-5, but this crossreactivity could be eliminated by absorption of the antibody on bFGF coupled to AffiGel-10. The final antibody preparation recognized only a single band at approximately 18.5 kD in lysates of RPE. Immunohistochemical staining with this antibody preparation demonstrated a marked dependence on cell density after 3 days in culture. Low culture density yielded cells staining moderately for aFGF, while confluent cells exhibited little or no staining. The reduction of aFGF from RPE cells in culture in a density-dependent fashion could also be demonstrated by Western blot analysis.
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