Size-Selective and In Vitro Assessment of Inner Blood Retina Barrier Permeability

Abstract

Assessment of tight junction integrity in vitro is fundamental when studying molecular processes that may be implicated in barrier dysfunction. At the blood brain and inner blood retina barrier (BBB and iBRB, respectively) adjacent endothelial cells lining the microvasculature have been shown to have very low rates of fluid phase transcytosis and high electrical resistances, due in part to the expression of tight junction proteins at the apical periphery of these cells. While these high electrical resistances are difficult to achieve in vitro, owing to complex interactions of endothelial cells in vivo with astrocytes and pericytes, it is possible to make an assessment of paracellular permeability when cells are analysed on a number of different fronts. In this regard, we will outline here a method for determining trans-endothelial electrical resistance, tracer molecule diffusion, and tight junction protein localization in primary cultures of bovine retinal microvascular endothelial cells. This system allows for the screening of a wide range of pro- and anti-angiogenic molecules in an in vitro model of the iBRB and can accurately assess the role individual tight junction proteins play in maintaining tight junction integrity in response to various cell stimuli.