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S. S. Ni Dhubhghaill, M. T. Cahill, M. Campbell, L. Cassidy, M. M. Humphries, P. Humphries; The Changes in Angiogenic Factors Expressed by Cultured Retinal Pigment Epithelial Cells (ARPE-19) in Response to Acute Cigarette Smoke Exposure. Invest. Ophthalmol. Vis. Sci. 2009;50(13):712.
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Age-related macular degeneration (AMD) is the leading cause of visual impairment in people over 65 in the Western world. Cigarette smoking is the most significant environmental risk factor in the development and progression of the disease. We hypothesise that cigarette smoke exerts it pathological effects in AMD by altering the balance of vasoactive growth factors expressed by the retinal pigment epithelium. These changes may promote the growth of new vessels. We have investigated the effects of smoke on vascular endothelial growth factor (VEGF), basic fibroblast Growth Factor (FGF-2) and pigment epithelium-derived factor (PEDF) in a cultured human RPE cell line.
The commercially available RPE cell line (ARPE-19) was cultured using the protocol from the original paper and maintained post confluence prior to experiment . Cigarette smoke was generated, using an established method, from Marlboro Red cigarettes and cells were exposed to concentrations equivalent to light, moderate and heavy smoke consumption. Post exposure cell responses were analysed by MTS assay, Immunohistochemistry, Western blot, ELISA and real time PCR techniques.
Acute cigarette smoke exposure does not alter the morphology of confluent RPE cell layers nor does it decrease cell viability. However, VEGF secretion is significantly suppressed by smoke at 24, 48 and 72hour exposure protocols (Statistical analysis ANOVA P<0.05). Cellular secretion of PEDF did not show a significant change however intracellular levels of PEDF did decrease after smoke exposure. In contrast, our preliminary findings have indicatred and increase in the expresssion of FGF-2 in response to smoke exposure. Initial rtPCR PEDF analyses indicate a reduction in PEDF mRNA expression.
Cigarette smoke exposure, even for short periods, has a significant effect on protein expression from retinal pigment epithelial cells. This loss of balance between angiogenic factors may facilitate the growth of new vessels in neovascular Age-Related Macular Degeneration.
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