Vitronectin receptor expression and distribution at the photoreceptor‐retinal pigment epithelial interface

Abstract

Laser scanning confocal microscopy was employed to map the distribution of integrin immunoreactivity at the photoreceptor‐retinal pigment epithelial (RPE) interface of the primate retina, and to determine its relationship to the actin cytoskeleton. Immunolabeling using a polyclonal antibody to the human vitronectin receptor (VnR), a heterodimer containing the αv subunit in combination with either the β3 or β5 subunits, is detected primarily on the apical surface of the retinal pigment epithelium (RPE) in vivo and in vitro. It is also associated with the photoreceptor inner and outer segment cell surfaces. In contrast, immunolabeling using a polyclonal antibody to the human fibronectin receptor (FnR), a heterodimer containing the α5 and β1 subunits, is detected principally on the basolateral surface of the RPE and is virtually absent in photoreceptors. A partial three‐dimensional reconstruction of the anti‐VnR labeling pattern in cone photoreceptors reveals cell surface labeling that originates at the level of the myoid just distal to the outer limiting membrane. It extends distally toward the ellipsoid and terminates at the level of the cone outer segment. Approximately 20–22 immunoreactive foci are distributed evenly around the perimeter of the cone ellipsoid. These foci correspond in number and location to the calycal processes that protrude from the distal portion of the ellipsoid. A double‐labeling procedure, employing VnR antibody and a fluorescently labeled phallotoxin (phalloidin), was used to identify regions of VnR co‐distribution with filamentous actin (F‐actin). One such region includes the VnR‐immunoreactive foci at the margins of the cone inner segments and the actin cables that course through the photoreceptor ellipsoid and terminate within the calycal processes. A second zone of co‐distribution coincides with the actin‐containing, circumferential bundle at the lateral borders of the RPE cells, and a third zone is associated with the apical microvilli of the RPE that ensheath cone outer segments. In order to help identify the specific subunits underlying VnR (αvβ3/5) immunoreactivity, Northern blots of retinal‐RPE RNA were probed with α³²P‐cDNAs to the human αv, β3, and β5 subunits and additional immunolocalization studies were performed using integrin human α or β subunit‐specific antisera. The results from these studies strongly suggest that one or more integrins, containing the αv and/or β5 subunits, are expressed by the photoreceptors and RPE. It thus appears likely that an integrin(s) from the cytoadhesin family located on the plasma membranes of these cells participates in some adhesion‐related event(s) at the interface between the neurol retina and the RPE.