Identification of Early Biochemical Changes in the Diabetic Retina Using an Unbiased Proteomics Approach

Diabetic retinopathy (DR) remains a leading cause of blindness in developed countries. Current treatments target late stages of DR when vision has already been affected. The current study used an unbiased proteomic approach to gain a better understanding of early DR pathogenesis prior to clinically detectable microvascular lesions. During these early DR stages, neurodegeneration occurs and is marked by glial activation throughout the neuroretina. As glial activation increases, glial fibrillary acidic protein (GFAP) is upregulated and can be visualized histologically. Postmortem retinas from diabetic donors were divided according to the presence of abundant (n=5) or minimal (n=5) GFAP immunoreactivity and were compared to retinas from nondiabetic donors with no known ocular pathologies (n=10). Retinal lysates from each group were pooled and run on an SDS-PAGE gel. Bands were analyzed sequentially by LC/MS using an Orbitrap Mass Spectrometer. A total of 2,190 proteins were identified across all groups. Pathway activation analysis was performed via Ingenuity Pathway Analysis. Pathways were subsequently grouped into broader functional categories based on the existing literature. Differential pathway analysis revealed significant differences between diabetic retinas with abundant vs. minimal GFAP immunoreactivity. Of particular interest are those pathways that function in both neuroprotective/neurodegenerative and angiogenic pathways, as they may represent a link between early and late diseases stages. Antioxidant/anti-inflammatory pathways, cell survival and apoptosis pathways, and autophagy also appear to play a role. Future studies will be required to confirm and extend these findings. Close analysis of these signaling changes may permit the development of preventive and interventional strategies for use in the early stages of DR, before overt microvascular damage or vision loss occurs.