Changes in Human Meibum Lipid Composition Related to the Presence and Severity of Meibomian Gland Dysfunction.

Purpose: Changes in meibum composition and quantity in meibomian gland dysfunction (MGD) result in tear film instability and dry eye. This exploratory study aimed to identify changes in (O-acyl)-ω-hydroxy fatty acid (OAHFA) and hydrocarbon chain (HC) unsaturation levels in meibum related to the presence and severity of MGD. Methods: Meibum samples were collected from 3 cohorts of adults with no MGD, mild-to-moderate MGD, and severe MGD in a noninterventional clinical trial (NCT01979887). OAHFAs, cholesterol esters (CE), HC unsaturation, and HC length in the meibum samples were quantified with 1H-nuclear magnetic resonance spectroscopy using 2 methods of normalization. Results: Meibum samples from 62 subjects were analyzed: 21 non-MGD, 21 mild-to-moderate MGD, and 20 severe MGD. Meibum OAHFA and CE levels and HC unsaturation were reduced with increasing severity of MGD, with most pairwise comparisons significant (P < 0.05, t-tests), following the order non-MGD > mild-to-moderate MGD > severe MGD. Regardless of the resonances used for normalization, each pairwise comparison of OAHFA, CE, and HC unsaturation levels in MGD (combined severities) versus non-MGD samples was significant (P < 0.01, t-test). Analysis using various normalization equations showed reductions of 20%-22% for OAHFAs, 51%-57% for CE, and 36%-66% for HC unsaturation in MGD (combined severities) compared with non-MGD. HC length was not altered in MGD (combined severities) compared with non-MGD samples (t-test). Conclusions: Meibum OAHFA, CE, and HC unsaturation levels were reduced in MGD and were lowest in the severe MGD cohort. These findings may contribute to the understanding of the pathophysiology of MGD.

Meibum Lipidomic Analysis in Evaporative Dry Eye Subjects.

Meibomian Glands (MG) are sebaceous glands responsible for the production of meibum, the main component of the Tear Film Lipid Layer (TFLL). The TFLL facilitates the spread of the tear film over the ocular surface, provides stability and reduces tear evaporation. Alterations in meibum composition lead to different ocular alterations like Meibomian Gland Dysfunction (MGD) and subsequent Evaporative Dry Eye (EDE). The aim of the present study was to investigate the composition and abundance of meibum lipids and their relationship with eyelid margin abnormalities, lipid layer patterns and MG status. The study utilizes a lipidomic approach to identify and quantify lipids in meibum samples using an Elute UHPLC system. This system considered all four dimensions (mass/charge, retention time, ion mobility and intensity) to provide the accurate identification of lipid species. Samples were categorized as healthy or low/no signs of alteration (group 1) or severe signs of alteration or EDE/MGD (group 2). The current investigation found differences in Variable Importance in Projection lipid abundance between both groups for the MGD signs studied. Changes in meibum composition occur and are related to higher scores in eyelid margin hyperaemia, eyelid margin irregularity, MG orifice plugging, MG loss and lipid layer pattern.

Comparative Characterization of Human Meibomian Glands, Free Sebaceous Glands, and Hair-Associated Sebaceous Glands Based on Biomarkers, Analysis of Secretion Composition, and Gland Morphology.

Meibomian gland dysfunction (MGD) is one of the main causes of dry eye disease. To better understand the physiological functions of human meibomian glands (MGs), the present study compared MGs with free sebaceous glands (SGs) and hair-associated SGs of humans using morphological, immunohistochemical, and liquid chromatography-mass spectrometry (LCMS)-based lipidomic approaches. Eyelids with MGs, nostrils, lips, and external auditory canals with free SGs, and scalp with hair-associated SGs of body donors were probed with antibodies against cytokeratins (CK) 1, 8, 10, and 14, stem cell markers keratin 15 and N-cadherin, cell-cell contact markers desmoglein 1 (Dsg1), desmocollin 3 (Dsc3), desmoplakin (Dp), plakoglobin (Pg), and E-cadherin, and the tight junction protein claudin 5. In addition, Oil Red O staining (ORO) was performed in cryosections. Secretions of MGs as well as of SGs of nostrils, external auditory canals, and scalps were collected from healthy volunteers, analyzed by LCMS, and the data were processed using various multivariate statistical analysis approaches. Serial sections of MGs, free SGs, and hair-associated SGs were 3D reconstructed and compared. CK1 was expressed differently in hair-associated SGs than in MGs and other free SGs. The expression levels of CK8, CK10, and CK14 in MGs were different from those in hair-associated SGs and other free SGs. KRT15 was expressed differently in hair-associated SGs, whereas N-cadherin was expressed equally in all types of glands. The cell-cell contact markers Dsg1, Dp, Dsc3, Pg, and E-cadherin revealed no differences. ORO staining showed that lipids in MGs were more highly dispersed and had larger lipid droplets than lipids in other free SGs. Hair-associated SGs had a smaller number of lipid droplets. LCMS revealed that the lipid composition of meibum was distinctively different from that of the sebum of the nostrils, external auditory canals, and scalp. The 3D reconstructions of the different glands revealed different morphologies of the SGs compared with MGs which are by far the largest type of glands. In humans, MGs differ in their morphology and secretory composition and show major differences from free and hair-associated SGs. The composition of meibum differs significantly from that of sebum from free SGs and from hair-associated SGs. Therefore, the MG can be considered as a highly specialized type of holocrine gland that exhibits all the histological characteristics of SGs, but is significantly different from them in terms of morphology and lipid composition.

Modeling meibum secretion: Alternatives for obstructive Meibomian Gland Dysfunction (MGD).

PURPOSE: While changes in meibum quality are correlated with severity of meibomian gland dysfunction (MGD) and dry eye disease, little is known regarding the mechanics of meibum secretion. The purpose of this study was to develop a finite element model of meibum secretion and evaluate the effect of various factors that might impact meibum delivery to the ocular surface. METHODS: A finite element analysis in COMSOL 6.0 was used to simulate the flow of meibum within the gland's terminal excretory duct. Historical normal human meibum rheology data taken over the meibum melting range from fluid (35-40 °C) to solid (25-30 °C) were then used to calculate the minimum yield stress and plastic viscosity of meibum. The effects of meibum melting state, eyelid pressure and terminal duct diameter on meibum flow rates were then systematically investigated. RESULTS: The melting state of meibum from liquid to solid was associated with an increase in the minimum yield stress and plastic viscosity that caused an exponential decrease in meibum flow. Modeling also established that there was a linear correlation between meibum flow rate and eyelid pressure needed to express meibum and the 4th power of the terminal duct radius. CONCLUSIONS: Our results suggest that changes in the melting state of meibum from fluid to solid, as well as changes in the radius of the terminal excretory duct and the force exerted by the eyelid can lead to dramatic decreases in the flow of meibum. Together these findings suggest alternative mechanisms for meibomian gland obstruction.

Therapeutic Efficacy and Safety of Intense Pulsed Light for Meibomian Gland Dysfunction in Patients with Chronic Ocular Graft-Versus-Host Disease.

PURPOSE: To evaluate the efficacy and safety of intense pulsed light combined with meibomian gland expression (IPL-MGX) for treating meibomian gland dysfunction (MGD) associated with chronic ocular graft-versus-host disease (oGVHD). METHODS: This retrospective study included 18 patients (18 eyes) with Fitzpatrick skin type ≤ IV, who underwent 3 to 8 sessions of IPL-MGX. Dry eye symptomology, ocular surface parameters, and adverse events were evaluated. RESULTS: Of 18 eyes, 83.3% and 66.7% showed severe oGVHD and severe MGD, respectively. At 4 weeks after the final session, significant improvements in the OSDI (P < 0.001), SPEED (P = 0.001), meibum expressibility (P < 0.001), and meibum quality (P = 0.016) were observed. At 12 weeks after, the OSDI (P = 0.009), SPEED (P = 0.002), and meibum expressibility (P = 0.008) significantly improved. No adverse events owing to IPL were reported. CONCLUSION: IPL-MGX may improve the ocular symptoms, ameliorate meibomian gland secretion, and is considered as a safe treatment for MGD in oGVHD patients.

Dysregulation of Lipid Metabolism in Aging Meibomian Glands and Its Molecular Markers.

The main function of exocrine Meibomian glands (MGs) is to produce a lipid-rich secretion called meibum which plays a critical role in maintaining the ocular surface homeostasis of humans and most mammals. The chemical composition of meibum, and its quantity produced by MGs, largely determine whether it can fulfill its role successfully. Aging was frequently associated with the onset of various MG-related pathologies. The goal of this study was to determine how aging affects the chemical composition and quantity of meibum in mice, and identify possible molecular markers of aging. Unbiased, untargeted and targeted lipidomic evaluation of mouse MG lipids was conducted using liquid chromatography-high-resolution mass spectrometry, and the results were analyzed using Principal Component, Orthogonal Projections to Latent Structures Discriminant, and Partial Least Square Discriminant Analyses. We found that aging leads to dysregulation of lipid metabolism in MGs, changing the ratios of major classes of MG lipids (such as wax esters, triacylglycerols, and phospholipids) in a progressive manner. Several lipid species that belong to these groups of MG lipids are proposed as clear markers of aging in a mouse model.

Exploring Signs and Symptoms Associated with Meibomian Gland Dysfunction for Use as Clinical Trial Endpoints.

Purpose: Dry eye disease is attributed to impaired tear production and/or evaporative dry eye. Evaporative dry eye is frequently associated with meibomian gland dysfunction (MGD). The objective of this study was to identify clinical study endpoints related to MGD. Methods: This 22-day, noninterventional, case-control clinical study involved three cohorts with increasing MGD severity: no MGD, mild/moderate MGD, and severe MGD. Symptoms were assessed with an ocular symptom questionnaire grading blurred vision, eye burning, eye dryness, eye pain, light sensitivity, eye itching, eye foreign body sensation, and overall ocular discomfort. Sign assessments included the maximum meibum quality score (MMQS), tear breakup time, Schirmer tear tests, biomicroscopy, and corneal staining. Signs and symptoms were compared between cohorts and study visits. Results: Seventy-five study participants were assigned to the cohorts (25 per cohort). MMQS scores increased with increasing MGD severity, reflecting the selection criteria for the cohorts. Between-visit scores showed a weighted kappa statistic of 0.72 indicating substantial agreement. Mean scores of all assessed symptoms increased with increasing MGD severity. Scores for symptoms showed moderate (κ = 0.41-0.60) to substantial (κ = 0.61-0.80) agreement between visits. Overall ocular discomfort demonstrated the strongest correlation with the MMQS. Conclusion: The MMQS was a reproducible sign of MGD showing good agreement with ocular symptoms. Overall ocular discomfort was well correlated with typical dry eye symptoms and could potentially be used as a single measure of MGD symptoms. The findings from this observational study may inform endpoints for future clinical trials. ClinicalTrials.gov NCT01979887.

The Effect of Oral Isotretinoin Therapy on Meibomian Gland Characteristics in Patients with Acne Vulgaris.

INTRODUCTION: The aim of the study was to determine the effect of oral isotretinoin therapy on the functional and morphological condition of the anterior segment of the eye, with particular emphasis on the meibomian glands. METHODS: Twenty-four patients (48 eyes) with a diagnosis of acne vulgaris were involved in the survey. All patients underwent a thorough ophthalmological examination at three time points: before therapy, 3 months after the start of therapy, and 1 month after the completion of isotretinoin therapy. The physical examination included the following elements: blink rate, analysis of the lid margin abnormality score (LAS), tear film break-up time (TFBUT) and Schirmer's test, meibomian gland loss (MGL), and the evaluation of the meibum quality score (MQS) and meibum expressibility score (MES). Additionally, the total score of an ocular surface disease index (OSDI) questionnaire was analysed. RESULTS: In comparison with pretreatment values, significant increases in OSDI during and after the treatment (p = 0.003 and p = 0.004, respectively) were observed. Substantial deterioration during the treatment was observed for MGL (p < 0.0001), MQS (p < 0.001) and LAS (p < 0.0001), while an improvement in those parameters after isotretinoin cessation was observed (p = 0.006, p = 0.02 and p = 0.0003, respectively). The frequency of using artificial eye drops was positively associated with MGL during (Spearman's rank correlation coefficient (Rs) = + 0.31; p = 0.03) and after the cessation of the therapy (Rs = + 0.28; p = 0.04). Meibomian gland atrophy correlated significantly with MQS during (Rs = + 0.29; p = 0.04) and after treatment (Rs = + 0.38; p = 0.008). The decrease in TFBUT values correlated with increased LAS (Rs = - 0.31; p = 0.03) during the course of isotretinoin usage. We found no changes in Schirmer's test or blink rates. CONCLUSION: Isotretinoin therapy leads to increased ocular complaints related to lipid tear film component dysfunction. This is due to reversible changes in meibomian gland morphology and function observed during drug usage.

Meibomian gland development: Where, when and how?

The Meibomian gland (MG) is an indispensable adnexal structure of eye that produces meibum, an important defensive component for maintaining ocular homeostasis. Normal development and maintenance of the MGs is required for ocular health since atrophic MGs and disturbances in composition and/or secretion of meibum result in major ocular pathologies, collectively termed as Meibomian gland dysfunction (MGD). Currently available therapies for MGD merely provide symptomatic relief and do not treat the underlying deficiency of the MGs. Hence, a thorough understanding of the timeline of MG development, maturation and aging is required for regenerative purposes along with signaling molecules & pathways controlling proper differentiation of MG lineage in mammalian eye. Understanding the factors that contribute to the development of MGs, developmental abnormalities of MGs, and changes in the quality & quantity of meibum with developing phases of MGs are essential for developing potential treatments for MGD. In this review, we compiled a timeline of events and the factors involved in the structural and functional development of MGs and the associated developmental defects of MGs during development, maturation and aging.

Sdr16c5 and Sdr16c6 control a dormant pathway at a bifurcation point between meibogenesis and sebogenesis.

Genes Sdr16c5 and Sdr16c6 encode proteins that belong to a superfamily of short-chain dehydrogenases/reductases (SDR16C5 and SDR16C6). Simultaneous inactivation of these genes in double-KO (DKO) mice was previously shown to result in a marked enlargement of the mouse Meibomian glands (MGs) and sebaceous glands, respectively. However, the exact roles of SDRs in physiology and biochemistry of MGs and sebaceous glands have not been established yet. Therefore, we characterized, for the first time, meibum and sebum of Sdr16c5/Sdr16c6-null (DKO) mice using high-resolution MS and LC. In this study, we demonstrated that the mutation upregulated the overall production of MG secretions (also known as meibogenesis) and noticeably altered their lipidomic profile, but had a more subtle effect on sebogenesis. The major changes in meibum of DKO mice included abnormal accumulation of shorter chain, sebaceous-type cholesteryl esters and wax esters (WEs), and a marked increase in the biosynthesis of monounsaturated and diunsaturated Meibomian-type WEs. Importantly, the MGs of DKO mice maintained their ability to produce typical extremely long chain Meibomian-type lipids at seemingly normal levels. These observations indicated preferential activation of a previously dormant biosynthetic pathway that produce shorter chain, and more unsaturated, sebaceous-type WEs in the MGs of DKO mice, without altering the elongation patterns of their extremely long chain Meibomian-type counterparts. We conclude that the Sdr16c5/Sdr16c6 pair may control a point of bifurcation in one of the meibogenesis subpathways at which biosynthesis of lipids can be redirected toward either abnormal sebaceous-type lipidome or normal Meibomian-type lipidome in WT mice.