iTRAQ-Based Proteomics Investigation of Aqueous Humor from Patients with Coats' Disease

Abstract

Background: Coats' disease is an uncommon form of retinal telangiectasis, and the identification of novel proteins that contribute to the development of Coats' disease is useful for improving treatment efficacy. Proteomic techniques have been used to study many eye diseases; however, few studies have used proteomics to study the development of Coats' disease.

Methods: Isobaric tagging for relative and absolute protein quantification (iTRAQ) was employed to screen differentially expressed proteins (DEPs) in the aqueous humor (AH) between stage 3A patients (n = 8), stage 3B patients (n = 14), stage 4 patients (n = 2) and control patients (n = 20). Differentially co-expressed proteins (DCPs) were present in all three stages of Coats' disease and were considered disease-specific proteins. These proteins were further analyzed using Gene Ontology (GO) functional annotations.

Results: A total of 819 proteins were identified in the AH, 222 of which were significantly differentially expressed (fold change > 2 and P < 0.05) in the samples from at least one stage of Coats' disease. Of the DEPs, 46 were found among all three stages of Coats' disease and the controls; therefore, they were considered Coats' disease-specific proteins (DCPs). A GO classification analysis indicated that the DCPs were closely related to structural molecule activity, cell adhesion molecule binding and receptor binding. Western blotting confirmed the expression levels of haptoglobin and apolipoprotein C-I were significantly up-regulated in Coats' disease.

Conclusions: The 46 Coats' disease-specific proteins may provide additional insights into the mechanism of Coats' disease and represent potential biomarkers for identifying individuals with Coats' disease.

Fig 1. Schematic representation of the workflow of the iTRAQ experiment.

Fig 2. Representative fundus views of the different stages of Coats' disease, including stage 3A, stage 3B, and stage 4, and the controls.

Fig 3. Heat map analysis of the differentially expressed proteins between the CK group and three groups of patients with different stages of Coats' disease.

The color scale shown at the top illustrates the relative protein expression level across all the samples. Red represents an expression level lower than the mean, whereas green represents an expression level above the mean.

Fig 4. Venn diagram indicating the differentially co-expressed proteins (DCPs) from the patients with the three stages of Coats' disease.

The numbers in parentheses indicate the total number of DCPs in stage 3A, stage 3B and stage 4, which are represented by purple, yellow and green, respectively.

Fig 5. GO annotation and functional classification of the differentially expressed aqueous humor proteins using GO terms for cellular components, molecular functions and biological processes.

Fig 6. Western blotting analyses of 2 DCPs confirmed that the proteins accumulated in three groups of patients.