Anti-angiogenic Effects Of Doxazosin On Experimental Choroidal Neovascularization In Mice

Background and purpose: Age-related macular degeneration(AMD)is one of the major causes of severe visual impairment among elderly individuals.Clinically,AMD is further divided into two types,dry and wet.The latter one,which is characterized by the formation of choroidal neovascularization(CNV),is highly responsible for vision loss.In recent years,Doxazosin,usually been prescribed to treat patients with benign prostatic hyperplasia and hypertension,has been found playing a critical role in suppressing the formation of tumor vascularization.This dissertation aims to investigate the anti-angiogenic effects of Doxazosin on laser-induced experimental choroidal neovascularization in mice,and explore its potential mechanism.We hope that this study may provide a new train of thoughts and tactics for clinical treatment of CNV.Contents: 1.The inhibition effects of Doxazosin on leakage,area and thickness of choroidal neovascularization tissues on a laser-induced mouse model.2.The possible inhibition mechanism of Doxazosin on experimental choroidal neovascularization in mice,and its influence on the expression of multiple proteins,such as HIF-1α,p-Akt,p-S6,VEGF,TGF-β2,COX-2,etc.Methods: 1.Establish the laser-induced choroidal neovascularization in mice;2.Observe the leakage levels of choroidal neovascularization in mice via the method of fluorescein angiography;3.Explore the area changes of choroidal neovascularization in mice via the method of choroidal flatmount;4.Measure the thicknesses of choroidal neovascularization tissues in mice via the method of histological study;5.Detect the expression variation of relevant proteins via the method of western blot analysis to further investigate the possible inhibition mechanism of Doxazosin on experimental choroidal neovascularization in mice.Results: 1.CNV model in mice is established through laser-induction.7 days after photocoagulation,the average fluorescein intensity levels of control group and Doxazosin group were 0.7435 ± 0.0311 and 0.5172 ± 0.0282,respectively,according to the fluorescein angiographic studies.There were statistically significant differences between the two groups(P<0.001).And similar results occurred between control and Doxazosin groups 14 days after photocoagulation,the data were 0.8041 ± 0.0353 and 0.5808 ± 0.0414,also indicating statistically significant differences;2.Lesion areas of experimental CNV were measured by observing the isolectin B4 staining under a fluorescence microscope after performing choroidal flat mounts.On day 7 after laser induction,the average CNV lesion areas of the control group and treated group were 12.43 ± 0.84×103 μm2 and 21.81 ± 1.49×103 μm2,respectively,while on day 14,the average areas were 10.03 ± 0.78×103 μm2 and 20.11 ± 1.72×103 μm2;3.H&E-stained sections of CNV lesions were observed 7 days and 14 days after laser induction.7 days after laser injury,Doxazosin-treated mice exhibited a statistically significant decrease in CNV lesion thickness(29.30 ± 2.15 μm)compared with control mice(42.39 ± 2.18 μm).CNV was notably suppressed by Doxazosin treatment after 14 days as well.The lesion thickness observed for the Doxazosin group and the control group were 28.92 ± 1.11μm and 41.10 ± 2.20 μm,respectively;4.3 days after laser photocoagulation,RPE-chroidal-sclera tissues were extracted.The expression variation of relevant proteins was detected via the method of western blot analysis.Compared to control group,the expression levels of the proteins,including HIF-1α,p-Akt,p-S6,VEGF,TGF-β2 and COX-2,in Doxazosin-treated group were significantly descended.Conclusions: 1.The leakage intensity were distinctly declined in Doxazosin-treated group on day 7 and day 14 after laser injury;2.CNV lesion areas in Doxazosin-treated group were significantly decreased compare to the control group 7 days and 14 days after laser induction;3.CNV lesion thicknesses in Doxazosin-treated group were significantly decreased compare to the control group 7 days and 14 days after laser induction;4.Doxazosin may inhibit HIF-1α protein levels by blocking the PI3K/Akt/m TOR signaling pathway and reducing expressions proangiogenic factors,such as VEGF,TGF-β2 and COX-2,which leads to the reduction of choroidal neovascularization formation.