Document Detail

Density-dependent expression of FGF-2 in response to oxidative stress in RPE cells in vitro.
MedLine Citation:
PMID:  11803485     Owner:  NLM     Status:  MEDLINE    
PURPOSE: The purpose of this study is to demonstrate the effect of culture density on the steady state mRNA levels of fibroblast growth factor-2 (FGF-2) when retinal pigment epithelial (RPE) cells are subjected to oxidative stress in vitro. METHODS: Subconfluent and confluent cultures of the established RPE cell line ARPE-19, were treated with increasing concentrations of tert-butyl hydroperoxide (tBH) or hydrogen peroxide (H(2)O(2)). Cell viability was measured using the WST-1 assay, and intracellular reactive oxygen intermediate (ROI) production was quantified by dichlorofluoroscein (DCF) fluorescence. Steady state changes in heme oxygenase-1 (HO-1) and FGF-2 mRNAs were measured by Northern blot analysis. RESULTS: Confluent cultures of ARPE-19 cells were less susceptible than subconfluent cultures to the toxic effects of the chemical oxidants. The intracellular reactive oxygen intermediate production was higher in subconfluent than confluent cultures with increasing tBH concentration. At nontoxic concentrations of tBH and H(2)O(2), a dose dependent increase in FGF-2 expression was seen as a function of culture density. FGF-2 mRNA expression was induced after tBH treatment in subconfluent, but not confluent cells. On the other hand, FGF-2 mRNA induction was observed after H(2)O( 2) treatment in confluent, but not subconfluent cultures. In contrast, no density dependent induction of HO-1 mRNA was seen after treatment with either tBH or H(2)O(2). CONCLUSIONS: These results suggest that care should be taken to control for cell density in similar types of in vitro experiments.
M Wada; C M Gelfman; H Matsunaga; M Alizadeh; L Morse; J T Handa; L M Hjelmeland
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't; Research Support, U.S. Gov't, P.H.S.    
Journal Detail:
Title:  Current eye research     Volume:  23     ISSN:  0271-3683     ISO Abbreviation:  Curr. Eye Res.     Publication Date:  2001 Sep 
Date Detail:
Created Date:  2002-01-22     Completed Date:  2002-03-07     Revised Date:  2007-11-14    
Medline Journal Info:
Nlm Unique ID:  8104312     Medline TA:  Curr Eye Res     Country:  England    
Other Details:
Languages:  eng     Pagination:  226-31     Citation Subset:  IM    
Department of Ophthalmology, University of California, Davis, CA 95616, USA.
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MeSH Terms
Blotting, Northern
Cell Count*
Cell Survival
Cells, Cultured
Dose-Response Relationship, Drug
Fibroblast Growth Factor 2 / biosynthesis*,  genetics
Heme Oxygenase (Decyclizing) / genetics,  metabolism
Heme Oxygenase-1
Hydrogen Peroxide / pharmacology
Membrane Proteins
Oxidative Stress / physiology*
Pigment Epithelium of Eye / cytology*,  drug effects,  metabolism
RNA, Messenger / metabolism
Reactive Oxygen Species
tert-Butylhydroperoxide / pharmacology
Grant Support
Reg. No./Substance:
0/Membrane Proteins; 0/RNA, Messenger; 0/Reactive Oxygen Species; 103107-01-3/Fibroblast Growth Factor 2; 75-91-2/tert-Butylhydroperoxide; 7722-84-1/Hydrogen Peroxide; EC protein, human; EC Oxygenase (Decyclizing); EC Oxygenase-1

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