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Masataka Nakanishi, Imran Ahmed Bhutto, Rhonda Grebe, Malia Michelle Edwards, Scott D McLeod, Gerard A Lutty; Caveolae transport of albumin by choriocapillaris from serum to Bruch’s membrane and RPE. Invest. Ophthalmol. Vis. Sci. 2014;55(13):4460.
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© ARVO (1962-2015); The Authors (2016-present)
The choriocapillaris (CC) is the fenestrated capillary system that adjacent to the retinal pigment epithelium (RPE), separated from RPE by Bruch’s membrane (BM). Recent studies show that dysfunction and eventual death of CC is related to development of choroidal neovascularization (CNV) in aged-macular degeneration (AMD). If CC transport is altered and whether it plays a role in the development of AMD is not known. The aim of this study was to examine the role of CC caveolae in transport of serum macromolecules from the CC lumen to RPE.
Alexa647-conjugated to bovine serum albumin (A647-BSA) and PBS were administrated in normal mice and caveolae-deficient mice (cav1-/-) intravenously. Mice were perfused with PBS and sacrificed at 0.5, 1 and 4 hours post injection. Eyecups were cryopreserved. 8 um thick sections were analyzed with a Zeiss 710 confocal microscope and autofluorescence was assessed in the sections of PBS injected eyes. Five nanometer BSA-conjugated gold nanoparticles (BSA-GNP) were administrated intra-arterially via the carotid artery and mice were then perfused with PBS and sacrificed at 1 hour post injection. Eyecups were fixed with 2.5 % paraformaldehyde/2 % glutaraldehyde, post fixed with 0.5 % OsO4, dehydrated and embedded in LX112 resin. 70 nm ultrathin sections were stained with uranyl acetate and lead citrate and visualized on a Hitachi H7600 TEM.
In eyes of normal mice at 0.5 hour after intravenous injection, A647-BSA was localized to the RPE and appeared to have diffused into the region of photoreceptor outer segments. On the other hand, most of A647-BSA was found in CC in eyes of caveolae-deficient mice, suggesting that albumin was only bound to its receptor but not taken up nor transported by caveolae. The majority of BSA-GNP found in normal mice was observed on the luminal not abluminal side of the CC endothelium, however, some was located in caveolae, some perivascular, and some in RPE. Conversely, BSA-GNP was not observed outside the lumens of CC in caveolae-deficient mice.
BSA transport by CC was regulated by caveolae via albumin receptor. The other transport systems of CC will be assessed using different tracers and materials to determine the normal in-vivo transport potential of CC in mice. Simultaneously, changes in CC caveolae volume are being determined by TEM in human aged and AMD subjects.
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