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Y. Cho, X. Cao, L. Parver, F. Rickles, C.-C. Chan; Increased Tissue Factor Expression in ARPE19 Cells After Lipopolysaccharide and Hydrogen Peroxide Treatment. Invest. Ophthalmol. Vis. Sci. 2010;51(13):2781.
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Age-related macular degeneration (AMD) is the leading cause of blindness in the elderly. Tissue factor (TF) promotes inflammation and angiogenesis, which underlie the pathogenesis of AMD. Previous studies reported that inflammation and oxidative stress, the key contributors to AMD, can induce TF expression in various cell types. In this study, ARPE19 cells were stimulated with lipopolysaccharide (LPS) and hydrogen peroxide (H2O2) to represent a microenvironment of AMD and investigate its effect on TF expression.
5mg Salmonella typhimurium LPS was used at concentrations of 1, 5 and 10µg/mL for 24 hours. 100 or 200µM H2O2 was added for 2 hours by dissolving 30% stock solution. ARPE19 cells were also stimulated with 10µg/mL LPS for 24 hours followed by 100 or 200µM H2O2 for 2 hours. Stimulated cells were examined for TF mRNA and protein expression via RT-PCR and immunohistochemistry. Difference in TF mRNA level was evaluated using SPSS one-way analysis of variance with p<0.05.
1, 5 and 10µg/mL LPS increased TF transcript level to 2.01-, 2.1- (p<0.05) and 3.45-fold (p<0.05) compared to control cells. TF protein expression moderately increased with 10µg/mL stimulation. TF mRNA expression increased 1.30- and 1.52-fold when stimulated by 100 and 200µM H2O2, respectively. TF protein did not increase significantly. For the combined stimulation, TF transcript level increased to 3.88-fold after 10µg/mL LPS and further to 4.51 and 4.95-fold after 100 and 200µM H2O2, respectively. TF protein increased moderately after the treatment.
In ARPE19 cells, LPS and H2O2 induceddose-dependent increase of TF mRNA and moderate increase in TF protein expression. Thus, inflammation and oxidative stress in AMD may lead to enhanced TF expression and possibly upregulated TF function.
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