Experimental design considerations for studies of human tear proteins

Harkness, Brooke M.; Hegarty, Deborah M.; Saugstad, Julie A.; Behrens, Hannah; Betz, Jason; David, Larry L.; Lapidus, Jodi; Chen, Siting; Stutzman, Ray E.; Chamberlain, Winston; Perez-Blanco, Maricarmen; Aicher, Sue A.

doi:10.1016/j.jtos.2023.02.005

Cited by 8 publications

(17 citation statements)

References 28 publications

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“…As a readily available and non-invasive sample (e.g., Schirmer's strips, glass capillary uptake), 41 the tear film is an ideal medium for biomarker discovery and diagnostic purposes that can be replenished in the blink of an eye. 8 , 42 A review article by Azkargorta et al 7 provides a comprehensive overview that highlights the various advances in the use of proteomics and peptidomics to analyze the human tear fluid, as well as the translation of these techniques into clinical practice. Identifying biomarkers within the proteome of human tears holds the promise of early disease detection and more targeted, personalized treatment plans.…”

Section: Discussionmentioning

confidence: 99%

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Intrinsic Disorder in the Human Tear Proteome

Gonzalez

Djulbegovic

Antonietti

et al. 2023

Invest. Ophthalmol. Vis. Sci.

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Purpose We aimed to characterize the proteome of human tears and assess for the presence of intrinsically disordered proteins (IDPs). IDPs, despite lacking a rigid three-dimensional structure, maintain biological functionality and could shed light on the molecular interactions within tears. Methods We analyzed a dataset of 1475 proteins identified in the tear film of three healthy subjects. We employed several computational tools, including the Compositional Profiler, Rapid Intrinsic Disorder Analysis Online, Search Tool for the Retrieval of Interacting Genes, and Database of Disordered Protein Predictors to evaluate the intrinsic disorder, protein interactions, and functional characterization of the disordered regions within this proteome. Results Our analysis showed a notable inclination toward intrinsic disorder. Two out of 10 order-promoting residues and five out of 10 disorder-promoting residues were found enriched. Using the Predictor of Natural Disordered Regions (PONDR) VSL2 output, 95% of these proteins were classified as highly or moderately disordered. We revealed an extensive protein–protein interaction network with significant interaction enrichment. The most disordered proteins exhibited higher disorder binding sites and diverse posttranslational modifications compared to the most ordered ones. Conclusions To the best of our knowledge, our study is the first comprehensive analysis of intrinsic disorder in the human tear film proteome, and it revealed an abundance of IDPs and their role in protein function and interaction networks. These findings suggest that variations in the intrinsic disorder of a tear film could be impacted by systemic and ocular conditions, offering promising avenues for disease biomarker identification and drug target development. Further research is needed to understand the implications of these findings in human health and disease.

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Section: Discussionmentioning

confidence: 99%

“…In addition, tear film analysis is a non-invasive and readily available sample for biomarker discovery and diagnostics, with the potential for early disease detection and personalized treatment plans. 1 , 7 , 8 …”

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confidence: 99%

Intrinsic Disorder in the Human Tear Proteome

Gonzalez

Djulbegovic

Antonietti

et al. 2023

Invest. Ophthalmol. Vis. Sci.

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“…A manual check of the characteristics of each patient was conducted to ensure that each instrument run contained samples from both groups (pain/no pain), each clinic site (Portland/Miami), a mixture of patient sex, and a range of tear volumes since we have previously shown that these factors can influence the tear proteome. 16 Each instrument run also contained a set of pooled tears from a group of individuals who were not part of this study to allow normalization across samples between runs. 16,25 We have previously described our TMT proteomics workflow for tear fluid in detail.…”

Section: Tandem Mass Tag Proteomics Sample Preparation and Processingmentioning

confidence: 99%

“…Results from the Orbitrap Fusion instrument were processed as we previously described using the Comet search engine, UniProt human protein database, a reversed sequence strategy to maintain peptide false discovery below 1%, and requiring 3 or more peptide spectral matches to each protein. 16 Reporter ion intensities for identified proteins were then summed for each unique peptide from MS3 synchronous precursor selection scans using in-house-developed scripts available at https://github.com/pwilmart. Normalization of protein intensities was implemented via the internal reference standard approach 26 using the pooled tear samples as reference.…”

Section: Tandem Mass Tag Proteomics Sample Preparation and Processingmentioning

confidence: 99%

Tear Proteins Altered in Patients with Persistent Eye Pain after Refractive Surgery: Biomarker Candidate Discovery

Harkness,

Chen,

Kim

et al. 2024

J. Proteome Res.

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“…Schirmer strip collection is associated with re ex tear induction, which may result in sample dilution and the collection of epithelial and cellular/organelle components [145][146][147] . Our review of studies characterizing the tear uid proteome using mass spectrometry identi ed between 1526 -3498 proteins when utilizing Schirmer strips 90,135,[148][149][150][151][152] compared to a maximum of 890 proteins when utilizing glass microcapillary tubes 23,[153][154][155] . However, it is important to note that most recent publications utilize Schirmer strips and, therefore, the greater number of proteins identi ed might be attributable to the improvements in mass spectrometry, such as increased sensitivity, resolution, and computing power.…”

Section: Pitfalls and Challengesmentioning

confidence: 99%

Tear Fluid as a Matrix for Biomonitoring Environmental and Chemical Exposures

Amini,

Okeme

2023

Preprint

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Purpose: Exposures to hazardous chemicals have been linked to many detrimental health effects. It is critical to have effective biomonitoring methods to measure the levels of these chemicals in humans to better evaluate key environmental exposures that increase the risk of chronic disease and death. Traditional biomonitoring utilizing blood and urine is limited due to the specialized skills and invasiveness of collecting these fluid samples. The systematic review focuses on the tear fluid, which is largely under-researched, as promising alternative to these traditional fluids. The objective is to evaluate the practicability of using human tear fluid for biomonitoring environmental exposures, highlighting potential pitfalls and opportunities. Recent finding: Tear fluid biomonitoring represents a promising method for assessing exposures. Tear fluid uniquely interfaces with the external environment at the air-tear interface, providing a surface for airborne chemicals to diffuse into the ocular environment and interact with biomolecules. Tear fluid also contains molecules from the internal environment that have travelled from the blood to tears by crossing the blood-tear barrier. Our review shows that tear fluid can be used to identify hazardous chemicals from the external and environment environments and to differentiate between different exposure groups. Summary: The review provides a critical analysis of the current state of knowledge in tear-based biomonitoring and draws attention to the promise of using the method for noninvasively increasing access to exposure assessment. The method will become acceptable for routine use as more individual and inter lab studies focus on broadly identifying and quantify exposure markers in tears, understanding factors that influence reliability of the method and standardizing the tear fluid method.

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Experimental design considerations for studies of human tear proteins

Cited by 8 publications

References 28 publications

Intrinsic Disorder in the Human Tear Proteome

Intrinsic Disorder in the Human Tear Proteome

Tear Proteins Altered in Patients with Persistent Eye Pain after Refractive Surgery: Biomarker Candidate Discovery

Tear Fluid as a Matrix for Biomonitoring Environmental and Chemical Exposures

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