Aims/hypothesis: Diabetic retinopathy is the most common complication of diabetes and a leading cause of blindness among working-age adults. Anatomical and functional changes occur in the retina and retinal pigment epithelium (RPE) prior to clinical symptoms of the disease. However, the molecular mechanisms responsible for these early changes, particularly in the RPE, remain unclear. To begin defining the molecular changes associated with pre-retinopathic diabetes, we conducted a comparative proteomics study of human donor RPE. Methods: The RPE was dissected from diabetic human donor eyes with no clinically apparent diabetic retinopathy (n=6) and from eyes of age-matched control donors (n=17). Soluble proteins were separated based upon their mass and charge using two-dimensional (2-D) gel electrophoresis. Protein spots were visualised with a fluorescent dye and spot densities were compared between diabetic and control gels. Proteins from spots with significant disease-related changes in density were identified using mass spectrometry. Results: Analysis of 325 spots on 2-D gels identified 31 spots that were either up- or downregulated relative to those from age-matched control donors. The protein identity of 18 spots was determined by mass spectrometry. A majority of altered proteins belonged to two major functional groups, metabolism and chaperones, while other affected categories included protein degradation, synthesis and transport, oxidoreductases, cytoskeletal structure and retinoid metabolism. Conclusions/interpretation: Changes identified in the RPE proteome of pre-retinopathic diabetic donor eyes compared with age-matched controls suggest specific cellular alterations that may contribute to diabetic retinopathy. Defining the pre-retinopathic changes affecting the RPE could provide important insight into the molecular events that lead to this disease.
Bibliographical noteFunding Information:
Acknowledgements This research was supported in part by grants EY014176 (to D. A. Ferrington) and AG025392 (to T. W. Olsen) from the National Institutes of Health, by the Minnesota Lions Macular Degeneration Center, by a career development award from the American Federation for Aging Research and Foundation Fighting Blindness (to D. A. Ferrington), by the Minnesota Medical Foundation and by an unrestricted grant from Research to Prevent Blindness Foundation. The authors thank the Minnesota Lions Eye Bank for their assistance in procuring eyes for this study. The mass spectrometry analysis was performed at the Mass Spectrometry Consortium for the Life Sciences at the University Minnesota.
- 2-D gels
- Human donor eyes
- MALDI-TOF mass spectrometry
- Retinal pigment epithelium