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Shemin Zeng, Robert F. Mullins; Effect Of AREDS Vitamins On Mouse Endothelial Cell Activation: Possible Implications For AMD. Invest. Ophthalmol. Vis. Sci. 2011;52(14):1767.
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The AREDS vitamin regimen is regularly recommended in the clinic to prevent or delay progression of age-related macular degeneration (AMD), but its mechanism is not well understood. This study was performed in order to investigate whether AREDS vitamins affect the expression of some inflammatory factors and/or cell behaviors of interest in AMD in an endothelial cell line.
Confluent cultures of mouse C166 endothelial cells (EC) were treated with a range of concentrations of AREDS vitamins. Lipopolysaccharide (LPS) and phorbol ester (PMA) were employed to provide a pro-inflammatory stimulus. Cytotoxicity of AREDS was evaluated using lactate dehydrogenase (LDH) ELISA assay. The effect of AREDS on cell motility was assessed using a scratch wound assay. The adhesion of peritoneal macrophages to EC monolayers was determined in the presence or absence of AREDS supplements. The expression of inflammatory factors (ICAM-1, ICAM-2, LFA1 and NF-kB) was determined at the mRNA level with real time PCR.
AREDS vitamins over a five-fold range of concentrations showed no cytotoxicity toward c166 cells at up to 56 hours after treatment (P>0.05). Scratch assays showed reduced EC migration in the presence of AREDS vitamins (P<0.05). In cell adhesion assays, LPS treatment increased the number of macrophages bound to the EC monolayer (P<0.05), and this increased binding was partially mitigated by the presence of AREDS components (38% decrease, p<0.05). Quantitative PCR for inflammatory gene products showed that AREDS supplement treatment alone did not significantly alter expression of the target genes. However, AREDS vitamins were found to alleviate the increased expression of ICAM-1 induced by PMA by more than 90%.
Treatment with AREDS vitamins may decrease EC migration, and appears to alleviate the increased binding of macrophages to ECs induced by LPS. AREDS treatment also represses the expression of ICAM-1 induced by PMA. These results suggest that AREDS vitamins may affect the progression of AMD in part through an ICAM-1 mediated mechanism.
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