Vinsamlegast notið þetta auðkenni þegar þið vitnið til verksins eða tengið í það: http://hdl.handle.net/1946/24873
Purpose: Disturbances in the retinal blood supply are thought to play a role in a number of ocular diseases. Activation of the renin-angiotensin system’s (RAS) vasoprotective axis is a promising way of reducing the progression of those diseases by increasing blood flow to the retina. The purpose of this study was to locate the AT1 and MAS receptors in porcine retinal arteries for the first time, both of whom play key but opposite roles in RAS, and examine if it was possible to counter AT1 induced vasoconstriction by MAS activated vasodilation in those same arteries.
Methods: Freshly slaughtered bovine- and porcine eyes were obtained from local abattoirs. For immunofluorescence, whole retinal vascular segments (2-12mm length) were isolated and fixed (N=28), with additional samples embedded into paraffin (n=2). The samples were incubated with primary antibodies against AT1 and MAS and to determine their location in/on the vessels, primary antibodies against smooth muscle, astrocytes and endothelium were used and the vessels viewed under a confocal microscope. For the myograph experiments, both bovine (N=37) and porcine (N=33) eyes were bisected, a retinal artery isolated and mounted in a wire myograph. Ang-II and U-46619 were tested as vasoconstrictors, along with Ang (1-7), DIZE and dorzolamide as potential vasodilators.
Results: Both AT1 (n=16) and MAS (n=16) were detected in all the samples tested and were found to co-localize in the smooth muscle layer (n=15), on astrocytes (n=14) and endothelium (n=4). Their distribution also varied along the length of the arteries, being more concentrated nearer the optic disk. In the paraffin sections, AT1 (n=2) and MAS (n=2) were found to co-localize in both smooth muscle and endothelium. Six different lots of Ang-II were tested on both bovine and porcine retinal arteries. Only 35% of the bovine retinal arteries responded to Ang-II at 10-5 M (n=1), 10-6 M (p=0.012, n=7) and 10-7M (p=0.005, n=5). KCl controls at 0.15M worked every time (p=0.001, n=7). Only 9% of porcine retinal arteries responded to Ang-II (N=33). Neither Ang (1-7) (n=13) nor DIZE (n=9) were able to produce vasodilation when precontracted with U-46619. Whereas Ang (1-7) with DIZE were able to completely reverse the effect of Ang-II (n=1). U-46619 produced contraction in every healthy porcine experiment (p=0.00, n=16), as well as every bovine experiment (p=0.03, n=2). Dorzolamide produced a mean vasodilation of 70% (p=0.002, n=7) when precontracted with U-46619 and completely abolished the effect of Ang-II (n=1).
Conclusion: AT1 and MAS are located in porcine retinal arteries, where they co-localize in the same vascular layers, in the endothelium, smooth muscle layer and astrocytes, in accordance with previous studies in other species and tissues. But despite AT1 being located in the retinal vasculature Ang-II rarely produced vasoconstriction, compared to the vasoconstrictor controls U-46619 and KCl which were consistent. Ang (1-7) and DIZE only produced vasodilation when precontracted with Ang-II which was rare but never with U-46619, which is also in accordance with previous studies. This is the first time that both main receptors for Ang-II and Ang (1-7) have been directly identified in the porcine retinal vasculature. But since Ang-II almost never produced vasoconstriction, their purpose remains unclear. The activation of the RAS vasoprotective axis however remains an intriguing prospect for future studies, with this study taking us one step closer to discovering its potential.
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