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M. Kengatharan, M. Verghese, K. Kiuchi, J. P. Cooke, P. Campochiaro; Nicotinic Receptor Antagonist Mecamylamine Reduces Laser-Induced Choroidal Neovascularization in C57bl/6j Mice. Invest. Ophthalmol. Vis. Sci. 2007;48(13):6016.
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Assess the effect of mecamylamine (ATG003, MEC), a non-selective nicotinic acetylcholine (nACh) receptor antagonist with anti-angiogenic properties on the development of laser-induced choroidal neovascularization (CNV) in a mouse model of neovascular AMD.
Four to five week-old female C57BL/6J mice were divided into 4 groups (10 animals per group). Groups were: (1) 50 mg/kg/day (s.c.) MEC using Alzet® osmotic pumps implanted 2 days prior to laser lesion, (2) Pegaptanib (Macugen®) by intra-vitreous (i.vt) injection (17.7 µg/eye/week), (3) saline (PBS, Sterile, s.c.) using Alzet® osmotic pump as well as intravitreal injection (5µL), 4) no treatment (controls). Laser-photocoagulation-induced rupture of Bruch’s membrane (at 3 locations) was used to generate CNV according to a method of Lima e Silva, et al 2005. Two weeks after rupture of the Bruch’s membrane, mice were anesthetized and perfused with fluorescein-labeled dextran (MW 2 x 106) and choroidal flat mounts were prepared. These were later examined by fluorescence microscopy, and images were digitized and the area of each CNV lesion analyzed using image analysis software. In a separate experiment, the effect of MEC (1-100µM) on VEGF (100ng/mL) induced endothelial cell (HMVEC, EC) tube formation in cell culture was investigated. Data are expressed as mean ± SEM. Statistical comparisons were made using one-way analysis of variance (ANOVA-Dunnett’s).
In control animals the average CNV lesion area calculated from the total number of rupture sites was 18.0±0.9 mm2 x 10-3. MEC (s.c.) reduced CNV lesion area by 36±6% which was statistically significant compared to a reduction of 2±6% caused by vehicle (PBS) (p<0.001, n=10). Pegaptanib (i.vt) reduced CNV lesion area by 29±5% which was statistically significant compared to vehicle (PBS) (p<0.01, n=10). These results suggest that inhibiting nACh receptors leads to a reduction in laser-induced CNV. Additionally, MEC, dose-dependently inhibited VEGF induced EC tube formation in vitro. These data are consistent with the observation that activation of the nACh receptors on EC leads to proliferation, migration and tube formation (angiogenesis) and inhibiting this process leads to a reduction in CNV which is a hall mark of neovascular AMD.
MEC, a non-selective nACh receptor antagonist, inhibits VEGF induced angiogenesis in vitro and reduces CNV in a mouse model of neovascular AMD.Reference: Lima e Silva R, et al., (2005). IOVS. Sep;46(9):3323-30.
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