Purpose. To determine if an exogenous plasma membrane protein expresssed in RPE in vivo is polarized and if the reversal or loss of polarity observed in cultured RPE for Na,K ATPase, N-CAM, and RET-PE2 antigen extends to other apical proteins as well. Methods. The neurotrophin receptor p75-NTR was introduced into cultured RPE-J cells or adult rat RPE in vivo and in vitro by gene transfer with a replication defective adenovirus vector. The polarity and sorting of p75-NTR was examined by biotin polarity assays and laser scanning confocal microscopy (LSCM). Results. Introduction of p75-NTR in RPE in vivo was efficiently accomplished by sub-retinal injection. Inspection en face of whole mounted tissue indicated that nearly the entire RPE was expressing p75-NTR. Examination by LSCM of 10μm cryosections revealed that p75-NTR was localized to the apical surface of the RPE with little or no choroidal staining. Co-localization studies of p75-NTR and RET-PE2 antigen confirmed the apical distribution of p75-NTR. In RPE-J cells p75-NTR was found to be apical by both LSCM and steady state biotin polarity assays. Conclusion. This is the first demonstration that a polarized plasma membrane protein expressed in RPE in vivo via adenovirus mediated gene transfer is sorted to the appropriate cell surface. p75-NTR is apically polarized both in vivo and in vitro. This suggests that some apical proteins (ie. Na,K-ATPase) in RPE either aquire and retain apical polarity through stabilizing contacts with components of the cytoskeleton and/or interphotoreceptor matrix, or, that multiple pathways to the apical surface exist in vivo, at least one of which is altered or lost when RPE are removed from their native environment.
|Original language||English (US)|
|Journal||Investigative Ophthalmology and Visual Science|
|State||Published - Feb 15 1996|
ASJC Scopus subject areas
- Sensory Systems
- Cellular and Molecular Neuroscience