Codetermination of antimicrobial agents in rabbit tear fluid using LC-MS/MS assay: Insights into ocular pharmacokinetic study.

Managing ocular microbial infections typically requires pharmacotherapy using antibiotic eye drops, such as moxifloxacin hydrochloride (MFX), combined with an antifungal agent like amphotericin B (AB). We carried out and validated an LC-MS/MS assay to quantify these compounds in rabbit tear fluid in order to look into the pharmacokinetics of these two drugs. We employed a protein precipitation technique for the extraction of drugs under examination. A Waters Symmetry C18 column was used to separate the analytes and internal standard. The composition of the mobile phase was like (A) 0.1% v/v formic acid in water and (B) methanol. The detection of MFX and AB was accomplished through the utilization of positive ion electrospray ionization under multiple reaction monitoring mode. The linearity curves for both analytes exhibited an acceptable trendline across a concentration range of 2.34-300 ng/mL for MFX and 7.81-1000 ng/mL for AB in surrogate rabbit tear fluid. The lower limit of quantitation for MFX was 2.34 ng/mL, while for AB, it was 7.81 ng/mL. The approach was strictly validated, encompassing tests of selectivity, linearity (with r2 > 0.99), precision, accuracy, matrix effects, and stability. Consequently, we employed this method to evaluate the pharmacokinetics profiles of MFX and AB in rabbit tear fluid following single topical doses.

Antioxidant Carbon Dots Nanozyme Loaded in Thermosensitive in situ Hydrogel System for Efficient Dry Eye Disease Treatment.

Purpose: Dry eye disease (DED) is a multifactorial ocular surface disease with a rising incidence. Therefore, it is urgent to construct a reliable and efficient drug delivery system for DED treatment. Methods: In this work, we loaded C-dots nanozyme into a thermosensitive in situ gel to create C-dots@Gel, presenting a promising composite ocular drug delivery system to manage DED. Results: This composite ocular drug delivery system (C-dots@Gel) demonstrated the ability to enhance adherence to the corneal surface and extend the ocular surface retention time, thereby enhancing bioavailability. Furthermore, no discernible ocular surface irritation or systemic toxicity was observed. In the DED mouse model induced by benzalkonium chloride (BAC), it was verified that C-dots@Gel effectively mitigated DED by stabilizing the tear film, prolonging tear secretion, repairing corneal surface damage, and augmenting the population of conjunctival goblet cells. Conclusion: Compared to conventional dosage forms (C-dots), the C-dots@Gel could prolong exhibited enhanced retention time on the ocular surface and increased bioavailability, resulting in a satisfactory therapeutic outcome for DED.

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.

Tear Proteomics in Infants at Risk of Retinopathy of Prematurity: A Feasibility Study.

Purpose: This feasibility study investigated the practicability of collecting and analyzing tear proteins from preterm infants at risk of retinopathy of prematurity (ROP). We sought to identify any tear proteins which might be implicated in the pathophysiology of ROP as well as prognostic markers. Methods: Schirmer's test was used to obtain tear samples from premature babies, scheduled for ROP screening, after parental informed consent. Mass spectrometry was used for proteomic analysis. Results: Samples were collected from 12 infants, which were all adequate for protein analysis. Gestational age ranged from 25 + 6 to 31 + 1 weeks. Postnatal age at sampling ranged from 19 to 66 days. One infant developed self-limiting ROP. Seven hundred one proteins were identified; 261 proteins identified in the majority of tear samples, including several common tear proteins, were used for analyses. Increased risk of ROP as determined by the postnatal growth ROP (G-ROP) criteria was associated with an increase in lactate dehydrogenase B chain in tears. Older infants demonstrated increased concentration of immunoglobulin complexes within their tear samples and two sets of twins in the cohort showed exceptionally similar proteomes, supporting validity of the analysis. Conclusions: Tear sampling by Schirmer test strips and subsequent proteomic analysis by mass spectrometry in preterm infants is feasible. A larger study is required to investigate the potential use of tear proteomics in identification of ROP. Translational Relevance: Tear sampling and subsequent mass spectrometry in preterm infants is feasible. Investigation of the premature tear proteome may increase our understanding of retinal development and provide noninvasive biomarkers for identification of treatment-warranted ROP.

Tear biomarkers.

An extensive exploration of lacrimal fluid molecular biomarkers in understanding and diagnosing a spectrum of ocular and systemic diseases is presented. The chapter provides an overview of lacrimal fluid composition, elucidating the roles of proteins, lipids, metabolites, and nucleic acids within the tear film. Pooled versus single-tear analysis is discussed to underline the benefits and challenges associated with both approaches, offering insights into optimal strategies for tear sample analysis. Subsequently, an in-depth analysis of tear collection methods is presented, with a focus on Schirmer's test strips and microcapillary tubes methods. Alternative tear collection techniques are also explored, shedding light on their applicability and advantages. Variability factors, including age, sex, and diurnal fluctuations, are examined in the context of their impact on tear biomarker analysis. The main body of the chapter is dedicated to discussing specific biomarkers associated with ocular discomfort and a wide array of ocular diseases. From dry eye disease and thyroid-associated ophthalmopathy to keratoconus, age-related macular degeneration, diabetic retinopathy, and glaucoma, the intricate relationship between molecular biomarkers and these conditions is thoroughly dissected. Expanding beyond ocular pathologies, the chapter explores the applicability of tear biomarkers in diagnosing systemic diseases such as multiple sclerosis, amyotrophic lateral sclerosis, Alzheimer's disease, Parkinson's disease, and cancer. This broader perspective underscores the potential of lacrimal fluid analysis in offering non-invasive diagnostic tools for conditions with far-reaching implications.

A surface-engineered contact lens for tear fluid biomolecule sensing.

The eyes provide rich physiological information and offer diagnostic potential as a sensing site, and probing tear constituents via the wearable contact lens could be explored for healthcare monitoring. Herein, we propose a novel adhesive contrast contact lens platform that can split tear film by natural means of tear secretion and blinking. The adhesive contrast is realized by selective grafting of a lubricant onto a polydimethylsiloxane (PDMS)-based contact lens, leading to high pinning zones on a non-adhesive background. The difference in contact angle hysteresis facilitates the liquid splitting. Further, the method offers control over the droplet volume by controlling the zone dimension. The adhesive contrast contact lens is coupled with fluorescent spectroscopic as well as colorimetric techniques to realize its potential as a diagnostic platform. The adhesive contrast contact lens is exploited to detect the level of lactoferrin in tear by sensitizing split droplets with Tb3+ ions. The adhesive contrast contact lens integrated with a fluorescence spectrometer was able to detect the lactoferrin level up to a concentration of 0.25 mg mL-1. Additionally, a colorimetric detection based on the fluorescence of the lactoferrin-terbium complex is demonstrated for the measurement of lactoferrin, with a limit of detection in the physiological range up to 0.5 mg mL-1.

In-depth correlation analysis between tear glucose and blood glucose using a wireless smart contact lens.

Tears have emerged as a promising alternative to blood for diagnosing diabetes. Despite increasing attempts to measure tear glucose using smart contact lenses, the controversy surrounding the correlation between tear glucose and blood glucose still limits the clinical usage of tears. Herein, we present an in-depth investigation of the correlation between tear glucose and blood glucose using a wireless and soft smart contact lens for continuous monitoring of tear glucose. This smart contact lens is capable of quantitatively monitoring the tear glucose levels in basal tears excluding the effect of reflex tears which might weaken the relationship with blood glucose. Furthermore, this smart contact lens can provide an unprecedented level of continuous tear glucose data acquisition at sub-minute intervals. These advantages allow the precise estimation of lag time, enabling the establishment of the concept called 'personalized lag time'. This demonstration considers individual differences and is successfully applied to both non-diabetic and diabetic humans, as well as in animal models, resulting in a high correlation.

Comparing meibomian gland visibility on optical coherence tomography and Keratograph 5M images using objective and subjective grading methods.

PURPOSE: To investigate if there is a visible difference in meibomian gland (MG) length between images captured with the Visante optical coherence tomography (OCT; wavelength = 1,310 nm) and the OCULUS Keratograph 5M (K5M; wavelength = 880 nm). METHODS: Adults between 18 and 40 years were recruited. Baseline dry eye disease was evaluated with the Standard Patient Evaluation of Eye Dryness (SPEED) and tear meniscus height and tear breakup time with the K5M. Right upper and lower eyelid MGs were imaged with the K5M and Visante OCT. Each image was graded with the 0 to 3 meiboscore scale. The central 5 MGs were evaluated with ImageJ for percent gland length visibility. RESULTS: Thirty participants were analyzed with a median (interquartile range [IQR]) age of 23.0 (5.0) years (53.3 % female). Overall, participants were asymptomatic and had normal tear films. Meiboscores based on K5M and Visante OCT was significantly different for the lower eyelid (0[1] vs 1[2]; p = 0.007) but not the upper eyelid (0[1] vs 0[1]; p = 1.00). The mean percent gland visibility of the upper eyelid (82.7[9.6] vs 75.2[13.5]; p < 0.001) and the lower eyelid (81.2[12.7] vs 64.1[17.6]; p < 0.001) were significantly greater on the Visante OCT than the K5M images, respectively. CONCLUSION: OCT images had significantly greater percent visible MG lengths than the K5M images. This suggests viable segments of the MGs may be missed with typical imaging, which may explain how it is possible that studies have found less post-treatment MG atrophy.

The impact of dry eye disease on retinal image quality in children.

BACKGROUND AND OBJECTIVES: Dry eye disease (DED) is increasingly prevalent, resultinginhigher morbidityamong children. This study evaluates the impact of DED severity on visual quality using double-pass technology, focusing on dynamic observation of the ocular light scatter in pediatric DED cases. METHOD: In this non-interventional, cross-sectional study, a mild DED group (37 cases, 37 eyes), a moderate DED group (40 cases, 40 eyes), and a control group of healthy children (35 cases, 35 eyes) were examined. Measurements included the Schirmer I test, tear film break-up time (BUT), and vision-related quality of life assessments using the Modified Ocular Surface Disease Index (OSDI) questionnaires. Participants underwent visual quality analysis using double-pass technology, which measured the modulation transfer function cut-off frequency value, Strehl ratio, objective scatter index (OSI), and OQAS-II value (OQAS-II value 100%, OQAS-II value 20%, and OQAS-II value 9%) under natural conditions. Additionally, dynamic changes in OSI post-blinking, Tear film mean-OSI , and the corresponding standard deviation OSI were recorded. RESULTS: Statistically significant differences were observed among the groups in modulation transfer function cutoff, Strehl ratio, OSI, OQAS-II value 100 %, OQAs-II value 20 %, OQAs-II value 9 %, tear film mean OSI, and standard deviation OSI (P < 0.05). As DED severity increased, tear film mean OSI significantly rose, while modulation transfer function cutoff, strehl ratio, OQAS-II value 100 %, OQAS-II value 20 %, OQAS-II value 9 % notably declined. All optical quality parameters were correlated with BUT, with no association observed with age, sex, or Schirmer I test. CONCLUSION: Dual-channel technology objectively assesses visual quality in pediatric DED, demonstrating that tear film scattering significantly affects retinal imaging and visual quality in children with DED.

Raman spectroscopy assisted tear analysis: A label free, optical approach for noninvasive disease diagnostics.

In recent times, tear fluid analysis has garnered considerable attention in the field of biomarker-based diagnostics due to its noninvasive sample collection method. Tears encompass a reservoir of biomarkers that assist in diagnosing not only ocular disorders but also a diverse list of systemic diseases. This highlights the necessity for sensitive and dependable screening methods to employ tear fluid as a potential noninvasive diagnostic specimen in clinical environments. Considerable research has been conducted to investigate the potential of Raman spectroscopy-based investigations for tear analysis in various diagnostic applications. Raman Spectroscopy (RS) is a highly sensitive and label free spectroscopic technique which aids in investigating the molecular structure of samples by evaluating the vibrational frequencies of molecular bonds. Due to the distinct chemical compositions of different samples, it is possible to obtain a sample-specific spectral fingerprint. The distinctive spectral fingerprints obtained from Raman spectroscopy enable researchers to identify specific compounds or functional groups present in a sample, aiding in diverse biomedical applications. Its sensitivity to changes in molecular structure or environment provides invaluable insights into subtle alterations associated with various diseases. Thus, Raman Spectroscopy has the potential to assist in diagnosis and treatment as well as prognostic evaluation. Raman spectroscopy possesses several advantages, such as the non-destructive examination of samples, remarkable sensitivity to structural variations, minimal prerequisites for sample preparation, negligible interference from water, and the aptness for real-time investigation of tear samples. The purpose of this review is to highlight the potential of Raman spectroscopic technique in facilitating the clinical diagnosis of various ophthalmic and systemic disorders through non-invasive tear analysis. Additionally, the review delves into the advancements made in Raman spectroscopy with regards to paper-based sensing substrates and tear analysis methods integrated into contact lenses. Furthermore, the review also addresses the obstacles and future possibilities associated with implementing Raman spectroscopy as a routine diagnostic tool based on tear analysis in clinical settings.

Another Brick in the Wall of Tear Film Insights Added Through the Total Synthesis and Biophysical Profiling of anteiso-Branched Wax and Cholesteryl Esters.

The tear film lipid layer (TFLL) plays a vital part in maintenance of ocular health and represents a unique biological barrier comprising unusual and specialized lipid classes and species. The wax and cholesteryl esters (WEs and CEs) constitute roughly 80-90% of the TFLL. The majority of species in these lipid classes are branched and it is therefore surprising that the synthesis and properties of the second largest category of species, i.e., the anteiso-branched species, remain poorly characterized. In this study, we have developed a total synthesis route and completed a detailed NMR spectroscopic characterization of two common anteiso-branched species, namely: (22S)-22-methyltetracosanyl oleate and cholesteryl (22'S)-22'-methyltetracosanoate. In addition, we have studied their structural properties in the bulk state by wide-angle and small-angle X-ray scattering and their behavior at the aqueous interface using Langmuir monolayer techniques. A comparison to the properties displayed by iso-branched and straight-chain analogues indicate that branching patterns lead to distinct properties in the CE and WE lipid classes. Overall, this study complements the previous work in the field and adds another important brick in the tear film insights wall.

Lipid-based eye drop formulations for the management of evaporative dry eyes.

Dry eye disease is a progressive prevalent ocular surface disorder that arises from various factors and is characterized by insufficient quality and/or quantity of tears. The underlying pathophysiology is intricate and can progress to chronic, difficult-to-treat conditions. Multiple strategies and therapeutic approaches are utilized in its management that target one or more etiopathological components of dry eyes, which may include aqueous tear deficiency or evaporative dry eyes. The primary focus of this paper is on treatment alternatives that utilize lipids for the treatment of evaporative dry eyes. This may arise from either abnormal lipid production or inadequate lipid spreading caused by meibomian gland dysfunction. The hypothesis behind the development of these lipid-containing eye drops is that if they can imitate the lipid layer, they may be able to help in the management of the signs and symptoms of evaporative dry eyes. The lipids used in commercial formulations for dry eyes are mineral oil, castor oil, phospholipids, omega-3 fatty acid, and medium-chain triglycerides. The literature suggests the potential of lipid-containing eye drops to alleviate some of the signs and symptoms and enhance the quality of life for individuals suffering from evaporative dry eyes.

Characterizing the Robustness of Distinct Clinical Assessments in Identifying Dry Eye Condition of Animal Models.

PURPOSE: The study aims to characterize the robustness of distinct clinical assessments in identifying the underlying conditions of dry eye disease (DED), with a specific emphasis on the involvement of conjunctival goblet cells. METHODS: Seven rabbits receiving surgical removal of the lacrimal and Harderian glands were divided into two groups, one with ablation of conjunctival goblet cells by topical soaking of trichloroacetic acid (TCA) to the bulbar conjunctiva (n = 3) and one without (n = 4), and the conditions of DED were assessed weekly using Schirmer test, tear breakup time (TBUT), tear osmolarity, and National Eye Institute (NEI) fluorescein staining grading. After 8 weeks, the rabbits were sacrificed, and the eyes were enucleated for histopathological examination. RESULTS: Histopathological analysis revealed corneal epithelial thinning in both groups. While TCA soaking significantly decreased the density of conjunctival goblet cells, DED rabbits without TCA also showed a partial reduction in goblet cell density, potentially attributable to dacryoadenectomy. Both groups showed significant decreases in Schirmer test and TBUT, as well as an increase in tear osmolarity. In DED rabbits with TCA soaking, tear osmolarity increased markedly, suggesting that tear osmolarity is highly sensitive to loss and/or dysfunction of conjunctival goblet cells. Fluorescein staining was gradually and similarly increased in both groups, suggesting that fluorescein staining may not reveal an early disruption of the tear film until the prolonged progression of DED. CONCLUSION: The Schirmer test, TBUT, tear osmolarity, and NEI fluorescein grading are distinct, yet complementary, clinical assessments for the evaluation of DED. By performing these assessments in definitive DED rabbit models, both with and without ablation of conjunctival goblet cells, the role of these cells in the homeostasis of tear osmolarity is highlighted. Characterizing the robustness of these assessments in identifying the underlying conditions of DED will guide a more appropriate management for patients with DED.

New Insights into the Molecular Structure of Tear Film Lipids Revealed by Surface X-ray Scattering.

The tear film lipid layer (TFLL) is a unique biological membrane that serves a pivotal role in the maintenance of ocular surface health. Reaching an overarching understanding of the functional principle of the TFLL has been hampered by a lack of insights into the structural and functional roles played by individual lipid classes. To bridge this knowledge gap, we herein focus on studying films formed by principal lipid classes by surface scattering methods. Through grazing incidence X-ray diffraction and X-ray reflectivity studies, we reveal quantitative data about the lattice distances, molecular tilt angles, and mono/multilayer thickness and density profiles for central TFLL lipid classes under close to simulated physiological conditions. In addition, we discuss the correlation of the results to those obtained previously with the natural lipid composition of meibum.

Multi-biomarker combination detection system for diagnosis and classification of dry eye disease by imaging of a multi-channel metasurface.

Dry eye disease (DED) is one of the most common ocular surface diseases, characterized by unstable tear film and ocular inflammation, affecting hundreds of millions of people worldwide. Currently, the clinical diagnosis of DED mainly relies on physical methods such as optical microscopy and ocular surface interferometric imaging, but classifying DED is still difficult. Here, we propose a compact and portable immune detection system based on the direct imaging of a nanophotonic metasurface with gradient geometry, for fast and ultra-sensitive detection of multiple biomarkers (i.e. Matrix metalloproteinase-9 (MMP-9), Lipocalin-1 (LCN-1), Lactoferrin (LTF)) in tears for the diagnosis and classification of DED. This centimeter-scale concentric nanophotonic metasurface, which consists of millions of unique metallic nanostructures, was fabricated through a cost-effective nanoimprint lithography (NIL) process. The immune detection system based on the antibody-modified metasurface shows favorable detection selectivity, an ultra-high sensitivity (3350 pixels/Refractive Index Unit (RIU)) and low limit of detection (LOD) (0.3 ng/mL for MMP-9, 1 ng/mL for LTF, and 0.5 ng/mL for LCN-1). Further clinical sampling and detection results demonstrated that this multi-biomarker detection system enabled accurate determination and symptom classification of DED, manifesting high correlation and consistency with clinical diagnosis results. The advantages such as low sample consumption, one-step detection, simple operation, and simultaneous detection of multiple biomarkers make the platform promising for screening and detecting a broader range of biomarker combinations in clinical practice.

Dissociation Kinetics and Antimicrobial Activity of Ofloxacin Antibiotic in Artificial Tears Via 1H-NMR, Raman, and UV-Vis Spectroscopic Analysis.

Introduction: The hydrogen-bonded networks play a significant role in influencing several physicochemical properties of ofloxacin in artificial tears (ATs), including density, pH, viscosity, and self-diffusion coefficients. The activities of the ofloxacin antibiotic with Ats mixtures are not solely determined by their concentration but are also influenced by the strength of the hydrogen bonding network which highlight the importance of considering factors such as excessive tear production and dry eye conditions when formulating appropriate dosages of ofloxacin antibiotics for eye drops. Objectives: Investigating the physicochemical properties of ofloxacin-ATs mixtures, which serve as a model for understanding the impact of hydrogen bonding on the antimicrobial activity of ofloxacin antibiotic eye drops. Determine the antimicrobial activities of the ofloxacin-Ats mixture with different concentration of ofloxacin. Methods: The ofloxacin-ATs mixtures were analyzed using 1H-NMR, Raman, and UV-Vis spectroscopies, with variation of ofloxacin concentration to study its dissociation kinetics in ATs, mimicking its behavior in human eye tears. The investigation includes comprehensive analysis of 1H-NMR spectral data, self-diffusion coefficients, Raman spectroscopy, UV-Vis spectroscopy, liquid viscosity, and acidity, providing a comprehensive assessment of the physicochemical properties. Results: Analysis of NMR chemical shifts, linewidths, and self-diffusion coefficient curves reveals distinct patterns, with peaks or minima observed around 0.6 ofloxacin mole fraction dissociated in ATs, indicating a strong correlation with the hydrogen bonding network. Additionally, the pH data exhibits a similar trend to viscosity, suggesting an influence of the hydrogen bonding network on protonic ion concentrations. Antibacterial activity of the ofloxacin-ATs mixtures is evaluated through growth rate analysis against Salmonella typhimurium, considering varying concentrations with mole fractions of 0.1, 0.4, 0.6, 0.8, and 0.9. Conclusions: The antibiotic-ATs mixture with a mole fraction of 0.6 ofloxacin exhibited lower activity compared to mixtures with mole fractions of 0.1 and 0.4, despite its lower concentration. The activities of the mixtures are not solely dependent on concentration but are also influenced by the strength of the hydrogen bonding network. These findings emphasize the importance of considering tear over-secretion and dry eye problems when designing appropriate doses of ofloxacin antibiotics for eye drop formulations.

Association of ocular surface and meibomian gland alterations with silicone hydrogel contact lens wear.

PURPOSE: To evaluate silicone hydrogel contact lens (SH-CL) effects on the meibomian glands, corneal structure, and ocular surface parameters. METHODS: Fifty SH-CL wearers for at least 6 months, and 50 sex and age-matched control subjects were recruited for this cross-sectional study. Visual display terminal (VDT) work and CL wear duration were questioned, ocular surface and tear functions were evaluated using OSDI questionnaire, tear break-up time (TBUT), corneal fluorescein staining, and Schirmer test. Corneal sensitivity was measured with Cochet-Bonnet aesthesiometry. Meibography and in vivo confocal microscopy (IVCM) were performed to evaluate meibomian glands and corneal structure. Intergroup comparisons were made using the Chi-square test, Wilcoxon test, or Kruskal-Wallis test. RESULTS: In the CL group, TBUT was shorter (P = 0.01), corneal fluorescein staining (P = 0.04), OSDI scores (P < 0.001), and meiboscores (P < 0.001) were higher than the control group. The biomicroscopic evaluation revealed meibomian gland dysfunction (MGD) in 34 % of the CL group and 20 % of the control group, which was not statistically significant (P > 0.05). IVCM showed that endothelial cell density was lower (P = 0.01) and polymegethism was higher (P < 0.001) in the CL group. Subbasal nerve density and corneal sensitivity measurements were similar in the two groups (P > 0.05). The longer VDT work duration was associated with increased CFS in the CL group (P = 0.05). CONCLUSION: The results showed that SH-CL wear increased dry eye symptoms and ocular discomfort, especially in longer VDT work duration. Meibography revealed significantly worse results in SH-CL wearers. SH-CL-related ocular discomfort seems to be more associated with MGD rather than neurosensorial alterations.

A chemical signal in human female tears lowers aggression in males.

Rodent tears contain social chemosignals with diverse effects, including blocking male aggression. Human tears also contain a chemosignal that lowers male testosterone, but its behavioral significance was unclear. Because reduced testosterone is associated with reduced aggression, we tested the hypothesis that human tears act like rodent tears to block male aggression. Using a standard behavioral paradigm, we found that sniffing emotional tears with no odor percept reduced human male aggression by 43.7%. To probe the peripheral brain substrates of this effect, we applied tears to 62 human olfactory receptors in vitro. We identified 4 receptors that responded in a dose-dependent manner to this stimulus. Finally, to probe the central brain substrates of this effect, we repeated the experiment concurrent with functional brain imaging. We found that sniffing tears increased functional connectivity between the neural substrates of olfaction and aggression, reducing overall levels of neural activity in the latter. Taken together, our results imply that like in rodents, a human tear-bound chemosignal lowers male aggression, a mechanism that likely relies on the structural and functional overlap in the brain substrates of olfaction and aggression. We suggest that tears are a mammalian-wide mechanism that provides a chemical blanket protecting against aggression.

Elucidating the Molecular Interactions between Lipids and Lysozyme: Evaporation Resistance and Bacterial Barriers for Dry Eye Disease.

Dry eye disease (DED) is a chronic condition characterized by ocular dryness and inflammation. The tear film lipid layer (TFLL) is the outermost layer composed of lipids and proteins that protect the ocular surface. However, environmental contaminants can disrupt its structure, potentially leading to DED. Although the importance of tear proteins in the TFLL functionality has been clinically recognized, the molecular mechanisms underlying TFLL-protein interactions remain unclear. In this study, we investigated tear protein-lipid interactions and analyzed their role in the TFLL functionality. The results show that lysozyme (LYZ) increases the stability of the TFLL by reducing its surface tension and increasing its surface pressure, resulting in increased TFLL evaporation and bacterial invasion resistance, with improved wettability and lubrication performance. These findings highlight the critical role of LYZ in maintaining ocular health and provide potential avenues for investigating novel approaches to DED treatment and patient well-being.

Detection of pro-inflammatory cytokines in healthy canine tears using Canine Cytokine SpikeMix™ mass spectrometry via multiple reaction monitoring.

PURPOSE: To evaluate the efficacy of the Canine Cytokine SpikeMix™ and MRM-MS for detecting pro-inflammatory cytokines in canine tears from healthy research Beagles. METHODS: A complete ophthalmic examination was performed on 15 healthy research Beagles to verify no ophthalmic diseases. Tears were collected OU by placing a Weck-Cel® cellulose spear in the ventral conjunctival fornix for 1 min. The Weck-Cel® spear was placed in a 2.0 mL tube with a centrifuge filter forcing tears to flow through the filter into the bottom of the tube. The tears were analyzed using the Canine Cytokine SpikeMix™ and MRM-MS. Descriptive data from this study was reported as the normalized total peak area (nTPA) and median (range) using data imported from the online MRM-MS Skyline program. RESULTS: The level of 16 pro-inflammatory cytokines was successfully detected in all 15 dogs. The four cytokines with the highest median amounts in the samples were IL-2 = 0.1243 (0.019-6.7289), IL-6 = 0.964 (0.0036-16.9365), TNFα = 0.1644 (0.0096-0.7138), and CSF-2 = 0.4022 (0.1475-2.6208). CONCLUSIONS: This study revealed that 16 pro-inflammatory cytokines in canine tears from healthy dogs can be detected with Canine Cytokine SpikeMix™ and MRM-MS.