Comparative analysis of human tear fluid and aqueous humor proteomes

Abstract

Purpose: Technological advancements allowing for the analysis of low-volume samples have led to the investigation of human tear fluid and aqueous humor (AH) as potential biomarker sources. However, acquiring AH samples poses significant challenges, making human tear fluid a more accessible alternative. This study aims to compare the protein compositions of these two biofluids to evaluate their suitability for biomarker discovery.

Methods: Paired tear and AH samples were collected from 20 patients undergoing cataract surgery. Tear samples were collected using Schirmer strips prior to surgery, and AH samples were collected from the anterior chamber immediately after corneal incision. Proteins were extracted and analyzed using liquid chromatography-tandem mass spectrometry (LC-MS/MS).

Results: A total of 481 proteins were identified in greater than 50% of the tear samples, and 191 proteins were detected in greater than 50% of the AH samples. Of these proteins, 82 were found to be common between the two biofluids, with ALB, LTF, TF, LCN1, and IGKC being the most abundant.

Conclusion: Although tear fluid and the AH are functionally independent and physically separated, many of the proteins detected in AH were also detected in tears. This direct comparison of the proteomic content of tear fluid and AH may aid in further investigation of tear fluid as a source of readily accessible biomarkers for various human diseases.

Keywords: Aqueous humor; Biomarkers; Proteomics; Tear fluid.

Fig. 1.

Number of unique proteins (A) and unique peptides (B) detected in AH and tear fluid samples. Data is represented as mean ± SEM, n = 20/group, ****p-value <0.0001 (unpaired T-test).

Fig. 2.

Venn diagram depicting the overlap between the number of unique proteins in AH (blue) and tear fluid (red) as identified by LC-MS/MS analysis.

Fig. 3.

Volcano plot comparing the protein content of AH (blue) and tear fluid (red).

Fig. 4.

Scatter plot comparing the normalized expression of proteins in AH (x-axis) and tear fluid (y-axis). Proteins with a mean PSM greater than 8 were considered highly abundant. Proteins highly abundant in both AH and tear fluid are colored purple.

Fig. 5.. Ingenuity Pathway Analysis (IPA) of proteins abundant in human aqueous humor and tear fluid.

IPA visualized major interaction hubs, biological functions, canonical pathways, and diseases associated with proteins present in human AH (blue) and tear fluid (red). Lines of predicted activation (orange) or inhibition (blue) are shown to visualize connections between pathway nodes. Relevant nodes within the network are displayed in orange. (A) a network displaying the connections with high abundance proteins in AH but not tear fluid. (B) a network of highly abundant proteins in tear fluid but not in AH. (C) a network of proteins with the high abundance in both AH and tear fluid.