Label-free quantitative proteomic analysis of aqueous humor in patients with high myopia

Objective

To characterize proteomic profile in aqueous humor of patients with high myopia using quantitative proteomic analysis.

Methods

Sixty-eight age-related cataract patients were divided into high myopic cataract group and simple cataract group according to that they had high myopia or not, with 34 patients (34 eyes) in each group.Aqueous humor samples (100 μl/patient) were collected from each patient using a 1 ml tuberculin syringe during cataract surgery at Tianjin Medical University Eye Hospital from January 2019 to August 2019.Sixteen samples from each group were selected for protein quantification and comparison by BCA method.The differentially expressed proteins between the two groups were analyzed using label-free liquid chromatography tandem mass spectrometry.The function and signal transduction pathways of differentially expressed proteins were further analyzed by Gene Ontology enrichment and Kyoto Encyclopedia of Genes and Genomes.Eighteen aqueous humor samples from each group were selected to verify the results of mass spectrometry by enzyme-linked immunosorbent assay (ELISA). This study protocol adhered to the Declaration of Helsinki.The use of human samples was approved by an Ethics Committee of Tianjin Medical University Eye Hospital (No.2020KY[L]-40). Written informed consent was obtained from each patient prior to surgery.

Results

The mean protein mass concentration of aqueous humor sample in the high myopic cataract group was (1 134.91±104.78) ng/L, which was significantly higher than that in the simple cataract group (706.71±85.43) ng/L, showing statistically significant difference (t=11.977, P<0.01). A total of 463 proteins were identified and 86 proteins were found to be differentially expressed, including 49 up-regulated proteins and 37 down-regulated proteins in the two groups.These differentially expressed proteins were mainly protein-binding activity modulator, extracellular matrix protein, carrier protein, intercellular signal molecule, protein modifying enzyme and so on, accounting for 32.70%, 14.50%, 9.10%, 9.10% and 7.30%, respectively.Bioinformatics analysis showed that 86 differentially expressed proteins were mainly related to biological processes such as complement activation and regulation, acute inflammatory response, and extracellular matrix tissue remodeling.Among them, 21 differentially expressed proteins were enriched in the complement and coagulation cascades pathways, 15 in the extracellular matrix-receptor interaction pathway, and 8 in the PI3K-Akt signaling pathway.ELISA results showed that the expression trends of three randomly selected differentially expressed proteins of the two groups were consistent with the results of label-free quantitative proteomic analysis.

Conclusions

There are significant changes in proteomic profiles of aqueous humor between the high myopia cataract patients and simple cataract patients.High myopia is closely associated with inflammation and immune interactions, and remodeling of extracellular matrix.