UHPLC-MS Profiling Identifies Distinct Tear Lipid Signatures in Dry Eye Disease

A study conceived to characterize the tear-lipid fingerprints associated with aqueous-deficient dry eye (ADDE) and Meibomian-gland dysfunction (MGD) in women and to assess the impact of intense pulsed-light [IPL] therapy on the tear lipidome in MGD) profiled the tear lipids with ultra high-performance liquid chromatography-mass spectrometry (UHPLC-MS). A paper based on this research was published in BMC Ophthalmology (1).

Dry eye disease (DED) is a multifactorial condition that affects the ocular surface characterized by an instability in the tear film (TF) provoked by hyperosmolarity, in combination with additional symptoms which may include ocular surface inflammation and damage and neurosensory abnormalities (1). According to the 2017 Dry Eye Workshop (DEWS) report, TF homeostasis and stability plays an important role in causing DED, as well as its effects on vision (2). The TF is a comparatively thin layer of fluid which covers the surface of the eye; its structural and functional properties are primarily driven by its chemical composition of predominately lipids, proteins, mucins, and electrolytes (3–5).This layer has several vital functions, including protection against pathogens and lubrication of the ocular surface, in addition to forming a smooth refractive surface ensuring optimal vision due to it being the eye’s first refractory layer (6).

In this study, 52 participants participated in two phases: a discovery cohort (9 ADDE, 15 MGD, 13 controls) and an independent validation cohort (15 additional subjects). As mentioned previously, tear lipids were profiled by UHPLC-MS. Unsupervised principal-component analysis (PCA) explored global variance, while supervised partial least-squares discriminant analysis (PLS-DA) and orthogonal PLS-DA (OPLS-DA) defined group differences and yielded candidate biomarkers, with model robustness confirmed by permutation testing and analysis of variance testing of cross-validated predictive residuals (CV-ANOVA). MGD participants received IPL at baseline, day 15, and day 45; clinical metrics and tear samples were obtained before and after therapy (1).

The analysis identified and quantified 176 lipid species in positive- and negative-ion modes (ESI + and ESI-). Supervised PLS-DA clearly separated ADDE, MGD, and control samples, while OPLS-DA highlighted 48 lipids that differed significantly among groups (p < 0.05). Both dry-eye subtypes were characterized by a distinct reduction of lysophospholipids, specifically lysophosphatidylethanolamines (LPE), lysophosphatidylcholines (LPC), lysophosphatidylglycerols (LPG), and lysophosphatidylinositols (LPI); fold change < 0.5) and an enrichment of (O-acyl)-ω-hydroxy fatty acids (OAHFA; fold change > 2. Cholesteryl esters (ChE) revealed a subtype-specific elevation in the MGD-versus-control comparison (fold change > 2). Permutation testing and CV-ANOVA verified the robustness of the ADDE-versus-MGD discrimination model. While IPL therapy showed a marked improvement in clinical metrics (tear break-up time and lissamine-green staining, for example), the changes shown in the tear lipid profile were not statistically significant (1).

Dry-eye subtypes may have discrete lipidomic signatures. As a result, this lipid panel could provide a set of possible therapeutic targets. The expansion of future studies where a broader range of sexes, ages, and larger cohort sizes are included will strengthen these findings, the scientists wrote. While IPL treatment demonstrates clinical improvements in the subjects participating, the increase in the number of shots and light intensity may be needed to verify if those modifications will produce noticeable changes in tear lipid profiles as well as the enhancement of tear film stability (1).

References

1. Acera, A.; Ibarrondo, O.; Mateo-Orobia, A. J. et al. Identification of Tear Lipid Biomarkers in Women with Dry Eye Disease and the Impact of Intense Pulsed Light Therapy: A Case-Control Study. BMC Ophthalmol. 2025, 25 (1), 496. DOI: 10.1186/s12886-025-04315-1
2. Craig, J. P.; Nichols, K. K,.; Akpek, E. K. et al. TFOS DEWS II Definition and Classification Report. Ocul Surf. 2017, 15 (3), 276-283. DOI: 10.1016/j.jtos.2017.05.008
3. Werkmeister, R. M.; Alex, A.; Kaya, S. et al. Measurement of Tear Film Thickness Using Ultrahigh-Resolution Optical Coherence Tomography. Invest. Ophthalmol. Vis. Sci. 2013, 54 (8), 5578-5583. DOI: 10.1167/iovs.13-11920
4. Green-Church, K. B.; Butovich, I.; Willcox, M. et al. The International Workshop on Meibomian Gland Dysfunction: Report of the Subcommittee on Tear Film Lipids and Lipid-Protein Interactions in Health and Disease. Invest. Ophthalmol. Vis. Sci. 2011, 52 (4), 1979-1993. DOI: 10.1167/iovs.10-6997d
5. Azartash, K.; Kwan, J.; Paugh, J. R. et al. Pre-Corneal Tear Film Thickness in Humans Measured with a Novel Technique. Mol. Vis. 2011, 17, 756-767.
6. Butovich, I. A. The Meibomian Puzzle: Combining Pieces Together. Prog. Retin. Eye Res. 2009, 28 (6), 483-498. DOI: 10.1016/j.preteyeres.2009.07.002