Comparative Proteome Analysis of Macula versus Peripheral Retina in Cynomolgus Monkey

Abstract

Purpose:: Macular diseases including age-related macular degeneration (AMD) is preferentially affect the macular region of the retina. This may be due to the proteins which especially expressed in the macular region. To screen the macular specific proteins, comprehensive proteome analysis of macular versus peripheral retina was performed using by two dimensional (2D) gel electrophoresis and mass spectrometer of cynomolgus monkey eyes.

Methods:: Tissue from macular and peripheral retina including RPE cell and choroidal membrane was obtained from eight normal monkeys. 2D-gel electrophoresis was performed in four different pH ranges (pH3-10, pH4-7, pH5-8, and pH7-10) for protein separation. Gels were stained and scanned for spot comparison between macular and peripheral retina. Identification of protein in gel spot was carried out by ion spray LC-MS/MS. Then, Western blotting and immunohistochemistry were performed for several proteins to confirm the difference between macular and peripheral region.

Results:: From pH3 to pH10, approximately seven hundred spots were detected on the gel image for macular and peripheral retina samples respectively. Sixty percent of these spots were present in both samples. 37 spots detected only macular were identified 25 proteins which involved Arrestin-C. However, except Arrestin-C, other proteins are known as common protein expressed in retina or other organs. Some proteins were confirmed larger amount in macular region than peripheral using western blotting and immunohistochemistry.

Conclusions:: Macular abundant proteins from cynomolgus monkey eyes were identified by 2D-gel and mass spectrometer. Arrestin-C, component of mitochondria, and gamma-synuclein were identified. Some of proteins identified in macula were not detected the difference of amount between macular and peripheral region. These proteins might have the difference in modification between two regions.

Keywords: macula/fovea • proteomics • age-related macular degeneration