ROS-responsive celastrol-nanomedicine alleviates inflammation for dry eye disease.

Dry eye disease (DED) is a major global eye disease leading to severe eye discomfort and even vision impairment. The incidence of DED has been gradually increasing with the high frequency of use of electronic devices. It has been demonstrated that celastrol (Cel) has excellent therapeutic efficacy in ocular disorders. However, the poor water solubility and short half-life of Cel limit its further therapeutic applications. In this work, a reactive oxygen species (ROS) sensitive polymeric micelle was fabricated for Cel delivery. The micelles improve the solubility of Cel, and the resulting Cel loaded micelles exhibit an enhanced intervention effect for DED. Thein vitroresults demonstrated that Cel-nanomedicine had a marked ROS responsive release behavior. The results ofin vitroandin vivoexperiments demonstrated that Cel has excellent biological activities to alleviate inflammation in DED by inhibiting TLR4 signaling activation and reducing pro-inflammatory cytokine expression. Therefore, the Cel nanomedicine can effectively eliminate ocular inflammation, promote corneal epithelial repair, and restore the number of goblet cells and tear secretion, providing a new option for the treatment of DED.

Impact of pH modification of the empirically used tobramycin ophthalmic solution on MIC90 concentration in tears and aqueous humor of donkeys (Equus asinus).

BACKGROUND: Commercial tobramycin ophthalmic solution is frequently used empirically to treat ocular disorders in equines, despite being primarily formulated for use in humans. It has been noted that tobramycin MIC90 concentration (minimal inhibitory concentration to 90% of microbial growth) rapidly declined following topical administration. It is hypothesized that adjustment of the pH of the empirically used tobramycin ophthalmic solution -prepared for human use- with the pH of the tears of donkeys, could increase the bioavailability of the drug and subsequently improve its penetration to the aqueous humor. Therefore, this study aimed to evaluate the impact of pH adjustment of the empirically used tobramycin ophthalmic solution on MIC90 concentration in tears and aqueous humor of donkeys (Equus asinus). The study was conducted on six (n = 6) clinically healthy donkeys. In each donkey, one eye was randomly selected to receive 210 µg tobramycin of the commercial tobramycin (CT) and used as a positive control (C group, n = 6). The other eye (treated eye) received 210 µg of the modified tobramycin ophthalmic solution (MT) (T group, n = 6). Tears and aqueous humor samples were collected 5-, 10-, 15-, 30- min, and 1-, 2-, 4-, and 6 h post-instillation. RESULTS: Modifying the pH of the empirically used commercial tobramycin ophthalmic solution in donkeys at a pH of 8.26 enhanced the drug's bioavailability. The MIC90 of the most hazardous bacteria isolated from equines' eyes such as Pseudomonas aeruginosa (MIC90 = 128 µg/ml) and Staphylococcus aureus (MIC90 = 256 µg/ml) was covered early (5 min post-instillation) and over a longer period in donkey tears (239-342 min) and aqueous humor (238-330 min) with the modified tobramycin solution. CONCLUSIONS: Adjustment of the pH of the commercial tobramycin ophthalmic solution, empirically used by veterinarians to treat donkeys' ophthalmic infections at a pH of 8.26, isotonic with the donkeys' tears pH, resulting in higher concentrations of tobramycin in tears and aqueous humor for a longer time.

Effects of combined intense pulsed light and cyclosporine 0.05% eyedrops in ocular surface matrix metalloproteinase-9 levels in patients with moderate-to-severe MGD.

To investigate the changes in meibomian gland dysfunction (MGD) and tear matrix metalloproteinase-9 (MMP-9) levels in patients with moderate-to-severe MGD after combined treatment with intense pulsed light (IPL) therapy and cyclosporine 0.05%. Thirty-six patients concurrently treated with IPL and cyclosporine 0.05% ophthalmic drops were retrospectively enrolled. Tear break up time (TBUT), corneal and conjunctival staining scores, Schirmer test, and ocular surface disease index (OSDI) questionnaire responses were recorded. Meibum quality, consistency, and eyelid margin telangiectasia were evaluated. MMP-9 levels were examined by the positivity and signal intensity of red lines (scored 0-4). IPL was performed four times with a vascular filter at 2-week intervals, followed by a 1-month follow-up after treatment cessation. Immediately after each IPL treatment, gentle meibomian gland expression was performed in both the upper and lower eyelids using meibomian gland expressor forceps. TBUT (1.88 ± 1.02 s to 3.12 ± 1.08 s, p < 0.001), corneal and conjunctival staining (6.19 ± 2.11 to 3.12 ± 1.89, p < 0.001), Oxford staining grade (2.66 ± 0.89 to 1.35 ± 0.76, p < 0.001), and OSDI (52.97 ± 21.86 to 36.36 ± 22.45, p < 0.001) scores significantly improved after the combined treatment. Meibum quality, consistency and lid margin telangiectasia showed significant post-treatment improvement in both the upper and lower eyelids. MMP-9 positivity showed a significant decrease (97-69%, p = 0.026) with a reduction in signal intensity (2.72 ± 0.87 to 2.09 ± 0.95, p = 0.011). The combination of IPL therapy and 0.05% cyclosporine eye drops effectively treats moderate-to-severe MGD by reducing symptoms and signs of MGD and by decreasing ocular surface MMP-9-associated inflammation.

Effects of topical ketorolac tromethamine on tear parameters, meibography, goblet cell density, and conjunctival oxidative stress in healthy dogs.

OBJECTIVES: The objective of the study was to evaluate whether a twice-daily instillation of 0.45% preservative-free ketorolac tromethamine (FKT) or 0.4% benzalkonium chloride-preserved ketorolac tromethamine (BACKT), every 12 h for 30 days may affect tear film parameters and the meibography in healthy dogs. Additionally, we assessed whether the same treatments irritated the ocular surface, affected goblet cell density (GCD), and the levels of oxidative stress biomarkers (OSB) in the conjunctiva of the same dogs. PROCEDURES: Experimental and masked comparison study. In 11 healthy dogs baseline values of the lipid layer thickness, tear meniscus height, non-invasive tear breakup time (NI-TFBT), and the meibomian gland (MG) loss were assessed by OSAvet®. For each dog, one eye received 40 µL of BACKT, while the other received 40 µL FKT, every 12 h for 30 consecutive days. Tear parameters and meibography were repeated 15, 30, and 60 days post-treatments. Conjunctival hyperemia and blepharospasm were monitored at the same time points. At baseline and Day 30, a conjunctival biopsy was collected for GCD and OSB determination. RESULTS: Conjunctival hyperemia and blepharospasm were not observed. At Day 15, the MG loss increased only in FKT-treated eyes (p < .001). On Day 30, both treatment groups showed increased MG loss, shortened NI-TFBT, and reduced GCD and catalase (p < .05). At Day 30, BACKT-treated eyes showed lower levels of superoxide dismutase (SOD) (p = .006) and higher levels of malondialdehyde (MDA) (p = .02). Differences between treatments were not observed for any parameter at any time point (p > .05). 60 days after treatment, OSAvet® parameters tended to return to values assessed at baseline; however, significant differences remained for MG loss (p < .05). CONCLUSIONS: Twice-daily instillation of KT, containing or not BAC, for 30 consecutive days shortened NI-TFBT, decreased GCD, and increased the MG loss in healthy dogs. KT should be used with caution when prescribed for long periods, particularly in patients with tear film abnormalities. However, future controlled studies using KT, BAC, and other topical NSAIDs are indicated to further support this finding.

A Novel Ex Vivo Blinking Method for Comparing the Corneal Residence Time of New Shampoo Formulations.

In the cosmetics sector, many products such as shampoos have a probability of accidental ocular exposure during their routine use. One very specific safety parameter is the residence time of the substance on the corneal surface, as prolonged exposure may cause injury. In this study, we developed a system that simulates corneal exposure to blinking and tear flow, for comparing the corneal clearance times of viscous detergent formulations. The Ex Vivo Eye Irritation Test (EVEIT), which uses corneal explants from discarded rabbit eyes from an abattoir, was used as the basis for the new system. To simulate blinking, we developed a silicone wiping membrane to regularly move across the corneal surface, under conditions of constant addition and aspiration of fluid, to mimic tear flow. Six shampoo formulations were tested and were shown to differ widely in their corneal clearance time. Three groups could be identified according to the observed clearance times (fast, intermediate and slow); the reference shampoo had the shortest clearance time of all tested formulations. With this new system, it is now possible to investigate an important physicochemical parameter, i.e. corneal clearance time, for the consideration of ocular safety during the development of novel cosmetic formulations.

Comparison of the effects of intranasal and intramuscular midazolam-butorphanol-ketamine on intraocular pressure, tear production and sedation in New Zealand White rabbits.

OBJECTIVE: To compare the effects of intranasal (IN) and intramuscular (IM) midazolam-butorphanol-ketamine on intraocular pressure (IOP), tear production (TP) and sedation in rabbits. STUDY DESIGN: Prospective, randomized, crossover experimental study. ANIMALS: Fourteen male New Zealand White rabbits, aged 1-2 years, body mass 3.1 ± 0.8 kg (mean ± standard deviation). METHODS: Rabbits were administered midazolam (1 mg kg-1), butorphanol (1.5 mg kg-1) and ketamine (5 mg kg-1) via IN and IM routes. IOP, TP and sedation scores were assessed at 0 (before drug administration), 5, 15, 30, 45 and 60 minutes after drug administration. Heart rate (HR), respiratory rate (fR), rectal temperature (RT), noninvasive mean arterial blood pressure (MAP) and peripheral hemoglobin oxygen saturation (SpO2) were simultaneously recorded until 45 minutes after drug administration. The onset and duration of sedation and sedation scores were recorded. RESULTS: Drug delivery route had no significant impact on mean IOP (p = 0.271) or TP (p = 0.062), and there were no significant changes over time for IOP (p = 0.711) or TP (p = 0.372). Similarly, delivery route had no significant impact on HR (p = 0.747), fR (p = 0.872), RT (p = 0.379), MAP (p = 0.217) and SpO2 (p = 0.254). Sedation onset was faster with IN (3.0 ± 1.0 minutes) than with IM administration (4.9 ± 0.7 minutes) (p = 0.011), but sedation duration was significantly longer with IM (52.6 ± 7.2 minutes) than with IN delivery (30.7 ± 6.8 minutes) (p = 0.004). There was no significant difference in sedation scores between the two delivery routes at any of the recorded time points. CONCLUSIONS AND CLINICAL RELEVANCE: The combination of midazolam-butorphanol-ketamine had minimal impact on physiological and ocular variables regardless of the route of administration, whereas IN drug administration led to a shorter onset and duration of action than IM administration.

The impact of systemic isotretinoin treatment on the tear film, meibomian glands, and corneal endothelium.

PURPOSE: The study aims to investigate changes in tear function, meibomian glands and corneal endothelium in patients receiving systemic isotretinoin therapy. MATERIALS AND METHODS: This prospective study included 38 eyes from 38 patients (23 females and 15 males) treated with systemic isotretinoin (0.5-1 mg/kg/day) following the diagnosis of acne vulgaris. All patients underwent a comprehensive ophthalmologic examination at baseline, 1st month, and third month of treatment. Subjective complaints were assessed using the Ocular Surface Disease Index (OSDI). Tear functions were evaluated through non-invasive tear break up time (NIBUT) and Schirmer I test. Meibomian gland (MG) changes were examined using meibography. Corneal parameters, including endothelial cell density (ECD), coefficient of variation (CV), the number of cells with a hexagonal shape (6A), average cell area (AVG), and central corneal thickness (CCT) were assessed using non-contact specular microscopy. RESULTS: The mean age of the patients was 19.29 ± 2.83 years. Ocular surface-related discomfort, measured with OSDI scores, significantly worsened at the third month measurements compared to the pre-treatment values (p < 0.001). In the 1st month of treatment, there was a significant decrease in NIBUT (p < 0.05). No statistically significant difference was found in the Schirmer test results at each visit. According to the 1st and third-month analysis, there was a significant increase in MG loss compared to the pre-treatment period (p < 0.001). ECD, CV, 6 A, AVG measurements at the first and third months showed a significant change compared to the pre-treatment values (p < 0.001). No significant difference was observed in the CCT measurements during the treatment. CONCLUSION: Systemic isotretinoin disrupted tear stability, caused MG loss, deterioration in corneal endothelium, and led to symptomatic complaints in patients.

Potential Therapeutic Role of Pituitary Adenylate Cyclase-Activating Polypeptide for Dry Eye Disease.

Dry eye disease (DED) is caused by a reduction in the volume or quality of tears. The prevalence of DED is estimated to be 100 million in the developed world. As aging is a risk factor for DED, the prevalence of DED is expected to grow at a rapid pace in aging populations, thus creating an increased need for new therapies. This review summarizes DED medications currently in clinical use. Most current medications for DED focus on stimulating tear secretion, mucin secretion, or suppressing inflammation, rather than simply replenishing the ocular surface with moisture to improve symptoms. We recently reported that the neuropeptide PACAP (pituitary adenylate cyclase-activating polypeptide) induces tear secretion and suppresses corneal injury caused by a reduction in tears. Moreover, it has been reported that a PACAP in water and a 0.9% saline solution at +4 °C showed high stability and achieved 80-90% effectiveness after 2 weeks of treatment. These results reveal PACAP as a candidate DED medication. Further research on the clinical applications of PACAP in DED is necessary.

High-Fat Diet-Induced Functional and Pathologic Changes in Lacrimal Gland.

The lacrimal gland is critical for maintaining the homeostasis of the ocular surface microenvironment through secreting aqueous tears in mammals. Many systemic diseases such as Sjögren syndrome, rheumatoid arthritis, and diabetes can alter the lacrimal gland function, eventually resulting in aqueous tear-deficient dry eye. Here, a high-fat diet (HFD) experimental mouse model was used to clarify how hyperlipidemia affects lacrimal gland function. Aqueous tear secretion fell about 50% after 1 month on a HFD. Lipid droplets accumulated in the matrix and acinar cells of the lacrimal gland after this period, along with changes in the lipid metabolism, changes in gene expression levels, and disruption of fatty acid oxidative activity. Immune cell infiltration and rises in the gene expression levels of the inflammation-related cytokines Il1ß, Tnfα, Tsg6, Il10, Mmp2, and Mmp9 were found. HFD also induced mitochondrial hypermegasoma, increased apoptosis, and decreased lacrimal gland acinar cell proliferation. Replacement of the HFD with the standard diet partially reversed pathologic changes in the lacrimal gland. Similarly, supplementing the HFD with fenofibrate also partially reversed the inhibited tear secretion and reduced lipid accumulation, inflammation, and oxidative stress levels. The authors conclude that a HFD induces pathophysiological changes and functional decompensation of the lacrimal gland. Therefore, ingestion of a HFD may be a causative factor of dry eye disease.

Influence of Different Antidepressants on Ocular Surface in Patients With Major Depressive Disorder.

PURPOSE: Several studies have previously reported the association between dry eye and depression along with the treatment of depression. The aim of this study was to investigate the effects of different antidepressant drugs on tear parameters in patients with major depressive disorder. METHODS: We recruited 132 patients who were using different antidepressants and 58 healthy controls. Venlafaxine, duloxetine, escitalopram, and sertraline were used by 34, 28, 36, and 34 patients, respectively. The participants filled out and completed the Beck Depression Scale. We recorded Schirmer test, tear breakup time (TBUT) and corneal staining values of the participants. The Ocular Surface Disease Index was completed by the participants. In addition, we evaluated the tear meniscus parameters by using anterior segment optical coherence tomography. RESULTS: All conventional dry eye tests and tear meniscus parameters were significantly lesser in the depression group than in the control group (Schirmer test, 11.41 ± 6.73 mm and 22.53 ± 4.98 mm; TBUT, 5.29 ± 2.92 seconds and 13.38 ± 1.72; Corneal staining, tear meniscus area, 0.026 ± 0.012 mm2 and 0.11 ± 0.025 mm2; tear meniscus depth, 182.75 ± 78.79 µm and 257.48 ± 90.1 µm; tear meniscus height, 290.3 ± 133.63 µm and 459.78 ± 180.26 µm, in patients and controls, respectively). The tear parameters of the duloxetine group were lowest among the drug groups and Schirmer test, and TBUT of the venlafaxine group was statistically significantly different from the duloxetine group (P = 0.028 and P = 0.017, respectively). Ocular Surface Disease Index score of the depression group was significantly higher than the control group (31.12 ± 21.15 and 17.43 ± 11.75 in depression and control group, respectively.). CONCLUSIONS: We found that the usage of selective serotonin reuptake inhibitors and serotonin noradrenaline reuptake inhibitors affects the ocular surface by a mechanism other than the anticholinergic system. Besides serotonin blockage, the noradrenaline blockade of serotonin noradrenaline reuptake inhibitors may increase the dry eye findings on the ocular surface.

The artemisinin analog SM934 alleviates dry eye disease in rodent models by regulating TLR4/NF-κB/NLRP3 signaling.

Dry eye disease (DED) is a multifactorial disorder of the tears and ocular surface characterized by manifestations of dryness and irritation. Although the pathogenesis is not fully illuminated, it is recognized that inflammation has a prominent role in the development and deterioration of DED. ß-aminoarteether maleate (SM934) is a water-soluble artemisinin derivative with anti-inflammatory and immunosuppressive activities. In this study, we established scopolamine hydrobromide (SCOP)-induced rodent model as well as benzalkonium chloride (BAC)-induced rat model to investigate the therapeutic potential of SM934 for DED. We showed that topical application of SM934 (0.1%, 0.5%) significantly increased tear secretion, maintained the number of conjunctival goblet cells, reduced corneal damage, and decreased the levels of inflammatory mediators (TNF-α, IL-6, IL-10, or IL-1ß) in conjunctiva in SCOP-induced and BAC-induced DED models. Moreover, SM934 treatment reduced the accumulation of TLR4-expressing macrophages in conjunctiva, and suppressed the expression of inflammasome components, i.e., myeloid differentiation factor88 (MyD88), Nod-like receptor protein 3 (NLRP3), apoptosis-associated speck-like protein containing CARD (ASC), and cleaved caspase 1. In LPS-treated RAW 264.7 cells, we demonstrated that pretreatment with SM934 (10 µM) impeded the upregulation of TLR4 and downstream NF-κB/NLRP3 signaling proteins. Collectively, artemisinin analog SM934 exerts therapeutic benefits on DED by simultaneously reserving the structural integrity of ocular surface and preventing the corneal and conjunctival inflammation, suggested a further application of SM934 in ophthalmic therapy, especially for DED.

Efficacy of Artificial Tears Based on an Extract of Artemia salina Containing Dinucleotides in a Rabbit Dry Eye Model.

(1) Background: Artemia salina is a brine shrimp containing high concentrations of dinucleotides, molecules with properties for dry eye treatment. For this reason, the purpose of the study was to evaluate the effect of the artificial tears based on an extract of Artemia salina in a rabbit dry eye model. (2) Methods: A prospective and randomized study was carried out. Twenty rabbits were divided into 4 groups (n = 5, each group): healthy rabbits, dry eye rabbits, dry eye rabbits treated with hypromellose (HPMC), and dry eye rabbits treated with Artemia salina. Dry eye was induced by the topical instillation of 0.2% benzalkonium chloride. The measurements were performed before and after the treatment for 5 consecutive days. (3) Results: The topical instillation of artificial tears containing Artemia salina showed beneficial effects on tear secretion, tear break-up time, corneal staining, the density of Goblet cells, heigh of mucin cloud secreted by these cells, and mRNA levels of IL-1ß and MMP9 in conjunctival cells. Compared with the HPMC, there was a statistically significant improvement (p < 0.05) with the Artemia salina in all the variables under study, except for the conjunctival hyperemia, density of Goblet cells, and mRNA levels of IL-6. (4) Conclusions: The potential of artificial tears based on Artemia salina as a secretagogue agent for dry eye treatment was confirmed, opening the door for future clinical trials and studies to extrapolate the findings for dry eye patients.

The Differential Reactive Oxygen Species Production of Tear Neutrophils in Response to Various Stimuli In Vitro.

A large number of polymorphonuclear neutrophils (PMNs) invade the ocular surface during prolonged eye closure (sleep); these leukocytes are commonly referred as tear PMNs. PMNs contribute to homeostasis and possess an arsenal of inflammatory mediators to protect against pathogens and foreign materials. This study examined the ability of tear PMNs to generate reactive oxygen species (ROS), an essential killing mechanism for PMNs which can lead to oxidative stress and imbalance. Cells were collected after sleep from healthy participants using a gentle eye wash. ROS production in stimulated (phorbol-12-myristate-13-acetate (PMA), lipopolysaccharides (LPS) or N-Formylmethionyl-leucyl-phenylalanine (fMLP)) and unstimulated tear PMNs was measured using luminol-enhanced chemiluminescence for 60 min. A high level of constitutive/spontaneous ROS production was observed in tear PMNs in the absence of any stimulus. While tear PMNs were able to produce ROS in response to PMA, they failed to appropriately respond to LPS and fMLP, although fMLP-stimulated tear PMNs generated ROS extracellularly in the first three minutes. Higher ROS generation was observed in isolated tear PMNs which may be due to priming from the magnetic bead cell separation system. The differential responses of tear PMNs in ROS generation provide further evidence of their potential inflammatory roles in ocular complications involving oxidative stress.

A Pilot Study of Changes in the Level of Catecholamines and the Activity of α-2-Macroglobulin in the Tear Fluid of Patients with Parkinson's Disease and Parkinsonian Mice.

Development of differential and early (preclinical) diagnostics of Parkinson's disease (PD) is among the priorities in neuroscience. We searched for changes in the level of catecholamines and α-2-macroglobulin activity in the tear fluid (TF) in PD patients at an early clinical stage. It was shown that TF in patients is characterized by an increased level of noradrenaline mainly on the ipsilateral side of pronounced motor symptoms (72%, p = 0.049), a decreased level of adrenaline on both sides (ipsilateral-53%, p = 0.004; contralateral-42%, p = 0.02), and an increased α-2-macroglobulin activity on both sides (ipsilateral-53%, p = 0.03; contralateral-56%, p = 0.037) compared to controls. These changes are considered as potential biomarkers for differential diagnosis. Similar changes in the TF were found in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated mice when modeling clinical and preclinical stages of PD. These data show the adequacy of models to the pathogenesis of PD along the selected metabolic pathways, and also suggest that the found TF changes can be considered as potential biomarkers for preclinical diagnosis of PD. In Parkinsonian mice, the level of catecholamines also changes in the lacrimal glands, which makes it possible to consider them as one of the sources of catecholamines in the TF.

Effects of eye drops containing a mixture of 3% diquafosol sodium and tocopherol acetate (vitamin E) on the ocular surface of murine dry eye.

PURPOSE: To investigate the efficacy of topical application of 3% diquafosol sodium (DQS) and tocopherol (TCP) acetate mixtures in a mouse model of experimental dry eye (EDE). METHODS: After exposure to desiccating stress for 5 days, eye drops consisting of 3% DQS alone, 0.01% TCP alone, or 3% DQS and 0.005% or 0.01% TCP mixture were applied for the treatment of EDE. Tear volume, tear film break-up time (TBUT), corneal fluorescein staining scores (CFSS), and tear film lipid layer grades (TFLLG) were measured at 0, 5 and 10 days after treatment. The 2',7'-dichlorodihydrofluorescein diacetate assay (DCFDA) for reactive oxygen species (ROS) production, enzyme-linked immunosorbent assay (ELISA) for malondialdehyde (MDA), and flow cytometry for CD4 + interferon (IFN)-γ+ T cells were evaluated on the ocular surface at 10 days after treatment. In addition, levels of tumour necrosis factor (TNF)-α, interleukin (IL)-1ß, IL-6, and chemokine CC motif ligand 4 (CCL4) in the conjunctiva were measured using a multiplex immunobead assay, and conjunctival goblet cells were counted by periodic acid-Schiff staining at 10 days after treatment. RESULTS: Both the TCP mixture groups indicated a significant improvement in TBUT, ROS production, and MDA concentrations compared to those in the DQS alone group. Furthermore, the 0.01% TCP mixture group also showed higher tear film lipid layer grades and conjunctival goblet cell density and lower corneal fluorescein staining scores, number of CD4 + IFN-γ+ T cells, and levels of TNF-α, IL-1ß, and CCL4 than the DQS alone group (P < 0.05). CONCLUSIONS: Application of eye drops containing the mixture of DQS and TCP could stabilize the tear film lipid layer, improve TBUT and corneal epithelial damages, decrease ROS production, inflammatory molecules, and T cells, and increase conjunctival goblet cell density on the ocular surface. Topical DQS and TCP mixtures may have a greater therapeutic effect on clinical signs, oxidative damage, and inflammation of dry eye than DQS eye drops.

The correlation of cytokines and sensory hypersensitivity in mild dry eye patients characterized by symptoms outweighing signs.

Purpose: To explore the correlation of tear and conjunctival cytokines and sensory hypersensitivity in mild dry eye (MDE) patients characterized by symptoms outweighing signs (DESOS). Methods: The subjects comprised 39 patients with MDE characterized by DESOS, 18 patients with common MDE (CMDE), and 15 healthy controls. The patients with DESOS were randomly subdivided into two groups; the C-DESOS group received artificial tears only, and the G-DESOS group received artificial tears and 0.1% fluorometholone eye drops three times a day. Symptoms were assessed using the Ocular Surface Disease Index (OSDI) and the Neuropathic Pain Symptoms Inventory modified for Eye (NPSI-E) questionnaire. Ocular examinations and in vivo confocal microscopy (IVCM) were also employed. Tear and conjunctival cytokines were measured using Multiplex or RT-PCR on Days 0, 7, and 30. The correlation between the expression of cytokines and hypersensitivity status was analyzed. Results: Compared with the CMDE and control groups, the DESOS groups showed a significant increase in symptom scores and in the ratio of symptoms versus signs. IL-1 ß, IL-2, IL-6, and TNF-α in tears and conjunctiva increased in the DESOS groups compared to the CMDE and control groups, indicating a high correlation with hypersensitivity status in the DESOS groups. Glucocorticoid treatment significantly decreased the level of cytokines in tears and conjunctiva in the G-DESOS group and subsequently ameliorated the symptoms. Conclusions: Tear and conjunctival cytokines, including IL-1 ß, IL-2, IL-6, and TNF-α, were correlated with sensory hypersensitivity status in the DESOS groups, suggesting they play an important role in the discordance of symptoms outweighing signs.

Resolvin D2 elevates cAMP to increase intracellular [Ca2+] and stimulate secretion from conjunctival goblet cells.

Under physiologic conditions, conjunctival goblet cells (CGCs) secrete mucins into the tear film to preserve ocular surface homeostasis. Specialized proresolving mediators (SPMs), like resolvins (Rvs), regulate secretion from CGCs and actively terminate inflammation. The purpose of this study was to determine if RvD2 stimulated mucin secretion and to investigate the cellular signaling components. Goblet cells were cultured from rat conjunctiva. Secretion was measured by an enzyme-linked lectin assay, change in intracellular [Ca2+] ([Ca2+]i) using Fura-2, and cellular cAMP levels by ELISA. RvD2 (10-11-10-8 M) stimulated secretion, increased cellular cAMP levels and the [Ca2+]i. RvD2-stimulated increase in [Ca2+]i and secretion was blocked by Ca2+ chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid tetrakis and the PKA inhibitor N-[2-(p-bromocinnamylamino)ethyl]-5-isoquinolinesulfonamide dihydrochloride but not by the cAMP exchange protein inhibitor α-[2-(3-chlorophenyl)hydrazinylidene]-5-(1,1-dimethylethyl)-b-oxo-3-isoxazolepropanenitrile. Forskolin, 3-isobutyl-1-methylxanthine, and 8-bromo-cAMP (8-Br-cAMP) increased [Ca2+]i. Increasing cAMP with 8-Br-cAMP inhibited the increase in [Ca2+]i stimulated by the cAMP-independent agonist cholinergic agonist carbachol. In conclusion, RvD2 uses both cellular cAMP and [Ca2+]i to stimulate glycoconjugate secretion from CGCs, but the interaction of cAMP and [Ca2+]i is context dependent. Thus RvD2 likely assists in the maintenance of the mucous layer of the tear film to sustain ocular surface homeostasis and has potential as a novel treatment for dry eye disease.-Botten, N., Hodges, R. R., Li, D., Bair, J. A., Shatos, M. A., Utheim, T. P., Serhan, C. N., Dartt, D. A. Resolvin D2 elevates cAMP to increase intracellular [Ca2+] and stimulate secretion from conjunctival goblet cells.

In Vitro Topical Delivery of Chlorhexidine to the Cornea: Enhancement Using Drug-Loaded Contact Lenses and ß-Cyclodextrin Complexation, and the Importance of Simulating Tear Irrigation.

Microbial keratitis is a severe, sight-threatening condition caused by various pathogens. Eyedrops are the standard delivery modality for treating these disorders; however, blinking reflex, elevated tear production, and nasolacrimal drainage eliminate much of the instilled dose within a few seconds. Therefore, eyedrops must be applied repeatedly for prolonged periods. The present study aimed to probe more effective ocular delivery of chlorhexidine based upon drug-loaded hydrogel contact lenses and ß-cyclodextrin (ß-CD), while also determining the effect of constant irrigation with simulated tear fluid (STF) in in vitro experiments. Chlorhexidine digluconate (as 0.2 and 2% solutions, ß-CD inclusion complexes, and loaded hydrogel contact lenses) were applied to enucleated porcine eyes as single or multiple 10 µL doses, or as drug-loaded contact lenses, with and without ß-CD. The corneas were then excised and drug-extracted quantified by high-performance liquid chromatography (HPLC). The effect of constant irrigation by STF was evaluated to test the effect of increased tear production on corneal delivery. Potential antimicrobial activity of the delivered drug was also assessed. Results showed that drug-loaded contact lenses delivered the greatest amount of chlorhexidine into the cornea over a 24 h period, while the eyedrop solution comparator delivered the least. The ß-CD significantly enhanced chlorhexidine delivery to the cornea from eyedrop solution, although contact lenses loaded with chlorhexidine-ß-CD failed to enhance delivery. ß-CD within the hydrogel matrix impeded drug release. Constant irrigation with STF significantly reduced the amount of drug delivered to the cornea in all cases. Chlorhexidine retained antimicrobial activity in all delivery methods. Hydrogel contact lenses loaded with chlorhexidine delivered significantly higher levels to the cornea compared to eyedrops, either multiple hourly doses or a single dose. They also offer reduced application, in particular, to a nonulcerated corneal infection. Finally, the importance of fully accounting for tear production in in vitro ocular delivery experiments was highlighted.

Tear inflammatory cytokines and ocular surface changes in patients with active thyroid eye disease treated with high-dose intravenous glucocorticoids.

PURPOSE: To evaluate high-dose intravenous glucocorticoid treatment on tear inflammatory cytokines and ocular surface parameters in patients with active TED. Correlations between tear inflammatory cytokines and clinical parameters were also investigated. METHODS: This prospective pilot study included 15 moderate-to-severe and active TED patients. Control group consist of 15 sex and age-matched healthy subjects. All TED patients were treated with high-dose intravenous methylprednisolone with cumulative dose of 4.5 g during the therapy subdivided into 12 weekly infusions. Tear concentrations of interleukin (IL)-1ß, IL-6, IL-8, IL-10, IL-17A, tumor necrosis factor (TNF)-α, and vascular endothelial growth factor (VEGF) were measured by multiplex bead analysis in TED patients at baseline and 12 weeks after treatment. Ocular surface disease index (OSDI), tear break-up time (TBUT), corneal fluorescent staining, and Schirmer's test were obtained from TED and controls. RESULTS: All baseline cytokine levels except for IL-17A were significantly elevated in active TED patients compared with controls. Concentrations of IL-1ß, IL-6, IL-8, TNF-α, and VEGF were significantly decreased at 12 weeks compared with baseline. OSDI and TBUT showed significant improvement at 6 and 12 weeks. There were significant positive correlations between IL-6, IL-8, and CAS, and negative correlation was found between IL-6 level and TED duration before methylprednisolone treatment. The reduction of IL-6, IL-8, and VEGF were positive correlated with the reduction in CAS at 12 weeks. CONCLUSIONS: High-dose glucocorticoids treatment improved ocular surface symptom, increased the tear film stability, and decreased tear inflammatory cytokines in active TED. The reduction of the inflammatory cytokines is consistent with the improvement of clinical parameters.

Effects of topical 1% cyclopentolate hydrochloride on quantitative pupillometry measurements, tear production and intraocular pressure in healthy horses.

OBJECTIVE: To evaluate the effect of topical cyclopentolate hydrochloride (CH) on quantitative pupillometric readings (PR), tear production (TP), and intraocular pressure (IOP) in healthy horses. ANIMALS STUDIED: Fourteen client-owned horses. PROCEDURES: In a two-phase design study, each animal received 1% CH ophthalmic solution in the left eye [treated] and 0.9% NaCl in the right eye [control] (0.2 mL each). In the first phase (n = 7), TP, IOP, and PR assessment was performed by Schirmer tear test I, rebound tonometry and static pupillometry, at 1, 8, 24, 48, 72, 96, 120, 148, 172, and 196-hours post-instillation. In the second phase (n = 7), plateau mydriasis was evaluated by assessing PR hourly for 8 hours post-instillation. For PR assessment, pupil area (PA), vertical diameter (VPD), and horizontal diameter (HPD) were recorded. All pupillometries were obtained in a room with fixed light intensity (45-60 lux). Statistical analysis was performed by generalized estimating equations method for the effect on parameters over time. RESULTS: After topical CH, significant differences in pupil dilation were seen from 1 to 172 hours for VPD and from 8 to 24 hours for PA, without significant differences on HPD over time. In the second phase, plateau PA and VPD were reached at 3 hours, while plateau HPD at 2 hours. No significant effects were detected on TP and IOP in both eyes at any time, nor on PR of the nontreated eyes. CONCLUSIONS: Topical 1% cyclopentolate hydrochloride could be considered an effective and safe option when a mydriatic/cycloplegic drug is needed in horses.