Inhibition of β-secretase Results in a Retinal Phenotype Involving Both the Vasculature and the RPE

Abstract

Purpose: : β-secretase 1 (BACE1) is an aspartyl protease that is strongly expressed in the brain and implicated in Alzheimmer’s disease. BACE2 has also been identified which shares around 68% homology with BACE1. Since BACE inhibitors are in clinical trials for Alzheimer’s disease we wished to determine the expression pattern of BACE1 and BACE2 in the retina and ascertain if BACE1 and/or BACE2 inhibition affects retinal structure and function.

Methods: : The retina and choroid from BACE1-/- and BACE1-/- BACE 2-/- knockout mice were assessed by routine pathology, immunohistochemistry for vascular density and confocal microscopy for lipofuscin accumulation. BACE1 and BACE2 expression in mouse and human retina/choroid were assessed by PCR and immunohostochemistry. In vitro angiogenesis was assessed using bovine retinal microvascular endothelial cell following exposure to VEGF, PEDF and VEGF+PEDF in the presence or absence of a β-secretase inhibitor. Lipofuscin accumulation was assessed by FACS analysis in human retinal pigment epithelial cells maintained for up to 28 days in the presence or absence of either a β-secretase inhibitor or BACE siRNA.

Results: : Assessment of the BACE 1-/- and combined BACE 1-/- BACE2-/- knockout mice demonstrated a significantly reduced retinal and choroidal vascular density, retinal thinning, apoptosis, thinning of Bruch’s membrane and a greater than 2-fold increase in lipofuscin in the RPE. BACE2-/- mice exhibited a relatively normal neural retina although occasional foci of neural retinal hyperplasia are observed. However, these animals exhibited a highly disrupted choroid together with swelling and lipofuscin accumulation in the overlying RPE but these changes are less severe than in BACE1-/- animals. Both BACE1 and 2 were expressed throughout the retina and choroid but BACE1 expression was highest in the neural retina and BACE2 highest in the RPE/choroid. PEDF induced an increase in BACE1 expression in endothelial cells but had no effect on the constitutive expression of BACE1 in RPE cells. PEDF-inhibition of VEGF-induced microvascular endothelial cell migration, proliferation and tubule formation were abolished by BACE1 inhibition. Furthermore, BACE1 inhibition in cultured RPE cells resulted in a greater than 2.5 fold increase in lipofuscin accumulation over a 28 day period which was associated with an increase in lysosomal pH and a decrease in cathepsin D activity.

Conclusions: : Both BACE1 and BACE2 play an important role in retinal physiology and caution should be undertaken when developing BACE inhibitors for Alzheimer’s disease as they may have off target effects in the retina.

Keywords: retinal neovascularization • retinal pigment epithelium • pathobiology