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Considering absence of invasiveness and side effects, tears emerge as a particularly attractive fluid for biomarker discovery and therefore for daily clinical use. However, to date this fluid remains poorly studied in healthy condition. Here, we present an updated in-depth characterisation of the human healthy tear protein composition using proteomics approach. Both eyes of 8 healthy controls (4 men and 4 women, average age: 38 ± 18) were collected using the Schirmer's strip method. After liquid digestion and off-gel electrophoresis fractionation, three independent proteomics analyses were performed on a LTQ-Orbitrap Velos Pro. Globally, 1351 proteins were identified with 2 unique peptides and 1% FDR. Gene ontology analyses showed up that 39% of the tear proteins were enzymes, with high numbers of dehydrogenases, phosphatases, kinases and ligases. Immunoglobulins, serpins and 14-3-3 domains proteins also emerged as the main tear protein families. The glycolysis and the coagulation and complement cascades, which were already shown in tears as involved in ocular and systemic diseases, were highlighted performing pathway analyses. Our study therefore complements the existing data on healthy tears proteome. Nevertheless, extensive studies for deeply and definitively characterise this promising fluid are required in the near future in order to be able to routinely use this fluid in clinics. A better understanding of its protein content will probably open new avenues in the biomarker discovery and clinical practice in the near future., (Copyright © 2018. Published by Elsevier Ltd.)

The aim was to investigate the levels of cytokines and soluble IL-6R in the tears of patients with thyroid-associated orbitopathy (TAO) disease. Schirmer's test was adopted to collect tears from TAO patients (N = 20, 17 women, mean age (±SD): 46.0 years (±13.4)) and healthy subjects (N = 18, 10 women, 45.4 years (±18.7)). Lacrimal cytokines and soluble IL-6R (sIL-6R) were measured using a 10-plex panel (Meso Scale Discovery Company) and Invitrogen Human sIL-6R Elisa kit, respectively. Tear levels of IL-10, IL-12p70, IL-13, IL-6 and TNF-α appeared significantly higher in TAO patients than in healthy subjects. Interestingly, IL-10, IL-12p70 and IL-8 levels increased in tears whatever the form of TAO whereas IL-13, IL-6 and TNF-α levels were significantly elevated in inflammatory TAO patients, meaning with a clinical score activity (CAS) ≥ 3, compared to controls. Furthermore, only 3 cytokines were strongly positively correlated with CAS (IL-13 Spearman coeff. r: 0.703, p = 0.0005; IL-6 r: 0.553, p = 0.011; IL-8 r: 0.618, p = 0.004, respectively). Finally, tobacco use disturbed the levels of several cytokines, especially in patient suffering of TAO. The differential profile of lacrimal cytokines could be useful for the diagnosis of TAO patients. Nevertheless, the tobacco use of these patients should be taken into account in the interpretation of the cytokine levels.

To date, Thyroid-Associated Orbitopathy (TAO), an autoimmune inflammatory disease affecting the eye, remains poorly characterised and its diagnosis challenging. The aim of this study was to investigate the tears of the TAO patients in order to identify potential biomarkers. Two independent quantitative Tandem Mass Tag™ 6-plex experiments were done. After in-solution digestion and isoelectric fractionation, the 12 fractions were analysed with a LTQ Orbitrap Velos coupled to a liquid chromatography. Raw files were searched against Swiss-Prot-AC database using Proteome Discoverer software, with a false discovery rate of 1% at peptide and protein levels. The differential proteins were then verified using orthogonal approaches in independent patients. Globally, 712 tear proteins were quantified with 2 unique peptides. Interestingly, cystatin c (TAO/controls ratio: 1.53), alpha-1 antichymotrypsin (ratio: 1.70) and retinal dehydrogenase (ratio: 0.68), displaying differential levels in the tears of TAO patients using proteomics experiments emerged as highly promising biomarkers after verification. In conclusion, this proteomics study supports the idea that tears reflect biological modifications occurring in a disease context and can therefore be a promising fluid for biomarker discovery. Moreover, our study identified three candidates that could in the future open new avenues in the diagnosis of TAO disease., Significance: Thyroid associated orbitopathy (TAO) is the most common disease affecting the orbit. Moreover, the later, severe stages of the disease can be sight threating [1]. On the other hand, the early sign and symptoms can be mistaken with other ocular pathologies [2]. Here we explore the modification of the tear content of the TAO patients using proteomics strategies and we proposed three new biomarker candidates, which could allow the early diagnosis of the disease and prompt action to prevent more severe stages. Moreover, our findings could also help to better understand the pathophysiology of the disease., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Purpose: Multiple sclerosis is the first cause of progressive neurological disability among young adults living in Western countries. Its diagnosis is mostly based on clinical evaluation, neuroimaging, and in some cases cerebrospinal fluid (CSF) analysis, but no definitive diagnostic test exists. We proposed here that the exploration of tears from multiple sclerosis patients could lead to the discovery of new biomarkers., Experimental Design: Thirty multiple sclerosis patients (20% men) recruited to the Geneva University Hospitals were included in our study (mean age ± SD [years]: 42.4 ± 15.9). Twenty-five control patients (32% men) were also enrolled (mean age ± SD [years]: 42.7±15.1). Tears, CSF or blood was collected for each patient. Three independent quantitative (tandem mass tag) experiments were carried out between tears from multiple sclerosis and control patients. Protein verification was performed by Western blot on tears and CSF and by ELISA on serum samples., Results: Combined proteomics analyses provided 185 identified tear proteins. Among the differential proteins, alpha-1 antichymotrypsin was the only one to be significantly increased in the three experiments with similar ratios (ratios 1.6 to 2.5, p < 0.05). Its tear, CSF and serum elevation were further confirmed by Western blot and ELISA, respectively., Conclusions and Clinical Relevance: This study supports the concept that modifications of the tear proteome can reflect biological abnormalities associated with multiple sclerosis and perhaps other inflammatory conditions affecting the CNS. In addition, alpha-1 antichymotrypsin elevation in tear fluid emerges as a promising biomarker for the diagnosis of multiple sclerosis., (© 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)