Tear Proteomics in Keratoconus: A Quantitative SWATH-MS Analysis

PURPOSE. To elucidate dysregulated proteins in keratoconus (KC) to provide a better understanding of the molecular mechanisms that lead to the development of the disease using sequential window acquisition of all theoretical mass spectra (SWATH-MS) as a protein quantification tool of the tear proteomic profile. METHODS. Prospective cross-sectional study that includes 25 keratoconic eyes and 25 healthy eyes. All participants underwent a clinical, tomographic, and aberrometric exam. Tear sample was collected using Schirmer strips and analyzed by liquid chromatography with tandem mass spectrometry. SWATH-MS was used as a quantification tool of the tear proteomic profile. The expression of the quantified proteins was compared between groups, and the biological and molecular functions of the dysregulated proteins as well as their functional relationships were studied by in silico analysis. RESULTS. A total of 203 proteins were quantified in tear samples of patients with KC and control participants, of which 18 showed differential expression between groups (P < 0.05). An increase in the expression of 7 proteins and a decrease in the expression of 11 proteins were observed. Protein-protein interactions and gene ontology analysis showed the involvement of these dysregulated proteins in structural, inflammatory-immune, iron homeostasis, oxidative stress, and extracellular matrix proteolysis processes. CONCLUSIONS. Tear protein quantification has revealed the dysregulation of proteins involved in biological processes previously associated with KC. Among them, iron homeostasis should be highlighted as a relevant pathway in the KC pathophysiology, and it should be taken into account in the development of therapeutic targets to cope with tissue damage derived from iron accumulation and toxicity.

Automated summary

Utilizing SWATH-MS for tear protein quantification, dysregulated proteins involved in iron homeostasis, inflammatory-immune response, oxidative stress, and other biological processes were identified in patients with keratoconus. The study sheds light on the molecular mechanisms underlying the disease and highlights the potential therapeutic targets related to iron accumulation and tissue damage in keratoconus pathophysiology.