Human retinal pigment epithelium proteome changes in early diabetes

Abstract

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.

The effects of the Chinese medicine ZiBu PiYin recipe on the hippocampus in a rat model of diabetes-associated cognitive decline: a proteomic analysis

“…First, although DIGE permits accurate quantitative comparisons, some technical constraints remain for use of twodimensional gels. For example, membrane proteins, extremely low-abundance proteins, proteins outside the pH range of 3 to 10, and very high or low molecular mass proteins are likely to be under-represented using this method [30,31]. Second, some spots were analysed by MS, but failed to reach the standards required for successful identification.…”

Impacts of Hypoxia-Inducible Factor-1 Knockout in the Retinal Pigment Epithelium on Choroidal Neovascularization

“…47 A number of independent studies have demonstrated outer BRB breakdown in diabetes and ischemia. 48,49 Overexpression of VEGF has been shown to play a key pathogenic role in breakdown of the barrier and contributes to impaired tight-junction integrity in the outer BRB. 50 Recent studies indicate that HIF-1a may be regulated by multiple factors besides hypoxia, such as nitric oxide (NO), reactive oxygen species (ROS), Sirt1, nuclear factor kappa B 51 ; HIF-2a, the isoform of HIF-1a, may be another important transcription factor up-regulated in CNV.…”

RPE barrier breakdown in diabetic retinopathy: seeing is believing

“…Significance and potential applications of fluorescent microscopy for RPE barrier leakage Alterations of the RPE barrier in diabetes and ischemia are not a new discovery [23,24,30,33,[36][37][38][39][40]82]. However, the extent and significance of the diabetes-induced RPE barrier breakdown have not been widely acknowledged.…”

“…An alteration of the RPE barrier in DR also affects the c wave of electroretinography, a signature of RPE integrity [37][38][39]. In a proteomic study comparing human donors of pre-DR and age-matched controls, 62% of the RPE proteins with significantly altered expression were those reported to have diabetes-induced change in nonretinal tissues, including proteins responsible for metabolism, chaperones in mitochondria, proteins in the cytosol and the endoplasmic reticulum (ER), and oxidative stress releasers [40]. The upregulation of proteins related to metabolism, ER stress, and oxidative stress indicates that the RPE may play a similar role as other tissues that respond to diabetic stress actively.…”