Abstract
Purpose:
To elucidate the mechanisms underlying development of structural and vascular abnormalities in the retina of mice lacking Cyp27a1 and Cyp46a1, enzymes important for cholesterol elimination
Methods:
Evaluations by high-resolution spectral-domain optical coherence tomography (SD-OCT), fluorescein angiography (FA), and transmission electron microscopy (TEM); analysis of gene expression by PCR arrays and quantitative RT-PCR; immuno- and histochemical staining; and sterol profiling by mass spectrometry
Results:
Cyp27a1-/-Cyp46a1-/- retinas have multiple hyperreflective spots on SD-OCT and increased vascular permeability and pathologies of retinal blood vessels on FA. The pathologies of retinal blood vessels include micro- and macroaneurysms, pulling and traction, dilated capillaries, capillary dropout and tortuosity. TEM revealed neovascularization in the normally avascular retinal pigment epithelium (RPE). TEM also showed thickening of the basement membrane in these blood vessels. Abnormal lipid accumulation was detected throughout the retina and in retinal blood vessels by histochemistry staining with oil red O, then confirmed by TEM. Areas with pathological blood vessels and structural abnormalities had increased staining with the markers for activated macrophages F4/80 and Iba-1. Retinal blood vessels, the photoreceptor inner segments (PIS) and the outer plexiform layer (OPL) also showed staining for iso[4]levuglandin E2 and carboxyethylpyrrole adducts as well as 7-ketocholesterol, indicators of elevated oxidative stress. TEM also revealed mitochondrial swelling, degeneration and aggregation. These changes were observed in the RPE, PIS, OPL and inner nuclear layers. Significant alterations were found in the retinal expression of a number of genes including the cholesterol efflux transporter Abcg1, the lipoprotein Apob, the cholesterogenic enzyme Hmgcr, the regulatory protein Insig1, and the inflammatory cytokines/chemokines Ccl1, Cxcl1, Cxcl2 and IL-6.
Conclusions:
We propose that combined deficiency of Cyp27a1 and Cyp46a1 decreases activation of Lxrβ leading to decreased expression of target genes like Abcg1 and almost complete blockage of cholesterol elimination from retinal macrophages and vascular endothelium. The latter causes lipid accumulation, inflammation, and oxidative stress that drive the development of the abnormalities in the Cyp27a1-/-Cyp46a1-/- retinas.
Keywords: 700 retinal neovascularization •
583 lipids •
557 inflammation