Conjunctival Goblet Cells Research Articles

A Comprehensive Review about Risks of Type-II Diabetes on Ocular Tear Film

Millions of people worldwide suffer from diabetes mellitus (DM), which can lead to systemic issues in a number of organs. Ocular problems, such as dry eye syndrome (DES), are among its less well-known side effects. This review delves into the interactions between diabetes and the composition of tear films, emphasizing alterations in the mucin, aqueous and lipid layer. Dry eye symptoms are exacerbated by induced changes in the components of the tear film in diabetes DM, which lead in decrement in tear production, increment in tear evaporation and tear film instability. Lipid layer is a lubricant, that reduces friction between the ocular surface and the eyelids which promotes high-quality, smooth refractive surface. The lacrimal function unit shields the tear film, preserves the normal function of the ocular surface. The mucin layer is secreted by the conjunctival goblet cells, in hyperglycemia the functionality of the cells are reduced thus, the mucin secretion is also altered which causes instability of the tear film. Diabetic patients can have their tear film integrity assessed with the help of diagnostic methods like Schirmer's Test and Tear Break- Up Time (TBUT). In order to relieve symptoms and maintain ocular health, there should be a complete management of diabetes and the induced tear film disorders.

Efficacy of Topical Cyclosporine Combined with Punctal Plugs in Treating Dry Eye Disease and Inflammation

Purpose To evaluate the effect of punctal plugs combined with cyclosporine eye drops on dry eye disease (DED) and ocular surface inflammation. Methods In a clinical trial, 73 patients were randomly allocated into three groups: punctal plug group, combination therapy group, and cyclosporine group. At the baseline and four weeks after treatment, the Schirmer I test score, fluorescein tear film break-up time (FBUT), ocular surface staining score and dry eye symptoms were assessed. Tear samples were collected to detect the level of inflammatory factors (interleukins, matrix metalloproteinase 9 (MMP-9) and tumor necrosis factor alpha (TNF-α)). In an animal experiment, a New Zealand rabbit dry eye model was induced. The rabbits were randomly divided into control group, punctal plug group, and combination therapy group (n = 6). Conjunctival goblet cell density, protein level of MMP-9 in conjunctiva and mRNA levels of inflammatory factors in conjunctiva and cornea were measured before and after treatment. Results In combination therapy group of the clinical trial, the following results were observed: significant improvement in Schirmer I test scores and FBUT compared to the cyclosporine group and punctal plug group, respectively; a decrease in the tear levels of IL-6, IL-1, and MMP-9 compared to the punctal plug group; and a decrease in the tear levels of IL-1α, IL-6, and IL-17 compared to the baseline (all p < 0.05). In the animal experiment, rabbits in combination therapy group had a higher goblet cell density (p < 0.01) and lower mRNA levels of IL-16 (p < 0.05), IL-17 (p < 0.05), and MMP-9 (p < 0.01) in conjunctiva and that of MMP-9 (p < 0.01) in cornea compared to punctal plug group. Conclusion Cyclosporine eye drops combined with degradable punctal plugs is a more optimized clinical treatment strategy for DED compared with degradable punctal plugs or cyclosporine eye drops alone, considering the influence of comprehensive clinical efficacy and ocular surface inflammation.

IL-37 Inhibits Inflammation of Lacrimal Gland in Dry Eye Mice via the IL-37-PTEN-NFκB Signaling Pathway

ABSTRACT Purpose This study aims to investigate the role of Interleukin-37 (IL-37) in mouse models of dry eye. Methods Two murine models of dry eye were employed in this investigation. The evaluation of the anti-inflammatory impact of IL-37 (200 μl, 10 μg/ml) on dry eye mice involved intraperitoneal injections administered once daily for 7 days. Additionally, intraperitoneal injection of VO-Ohpic trihydrate (VO, 0.25 mg/kg) in dry eye mice was performed to investigate the role of PTEN in the IL-37 anti-inflammatory signaling pathway. Tear production was assessed using phenol red cotton thread, while corneal damage was examined through sodium fluorescein staining using a slit lamp. Histological alterations in the lacrimal gland were observed through H&E staining. PAS staining was used to assess conjunctival goblet cells. The levels of NFκB-P65, p-NFκB-P65, IL-1β, IL-6, TNF-α, CD3, AQP5, α-SMA and PTEN proteins were determined via Western blotting or immunofluorescence. Results Following IL-37 treatment, both dry eye models exhibited reduced corneal fluorescence staining scores and enhanced tear production. In lacrimal gland, the expression of p-NFκB-P65, IL-1β, IL-6, CD3 and TNF-α was diminished, while PTEN, AQP5, α-SMA expression increased after IL-37 treatment in both dry eye mice. However, the intraperitoneal injection of VO significantly attenuated the anti-inflammatory effect of IL-37 on dry eye mice. Conclusion IL-37 emerges as an anti-inflammatory mediator within the lacrimal gland of dry eye mice, exerting its effects through the IL-37-PTEN-NFκB signaling pathway.

Development of human amniotic epithelial cell-derived extracellular vesicles as cell-free therapy for dry eye disease

PurposeThis study aimed to investigate the therapeutic potential of extracellular vesicles (EVs) derived from human amniotic epithelial cells (hAEC-EVs) for Dry Eye Disease (DED) treatment. MethodsHighly purified EVs were isolated from the culture supernatants of hAECs, which obtained from term placenta and characterized. Proteomic contents were analyzed for assessing its biological function related to the therapeutic potentials for DED. Subsequently, we examined the therapeutic efficacy of hAEC-EVs on human corneal epithelial cells exposed to hyperosmotic stress and in an experimental DED mouse model induced by desiccation stress. ResultsProteomic analysis of hAEC-EVs revealed proteins linked to cell proliferation and anti-inflammatory responses. We demonstrated efficient uptake of hAEC-EVs by ocular surface cells. Under DED conditions, EV treatment increased corneal epithelial cell proliferation and migration, and concurrently reducing inflammatory cytokines. In the DED mouse model, hAEC-EVs showed significant improvements in corneal staining score, tear secretion, corneal irregularity, and conjunctival goblet cell density. Additionally, hAEC-EVs exhibited a mitigating effect on ocular surface inflammation induced by desiccation. ConclusionsThese findings suggest that hAEC-EVs hold potential as a cell-free therapy for corneal epithelial defects and ocular surface diseases, presenting a promising treatment option for DED.

Sex-dependent differential increase of specialized pro-resolving mediators in extracellular vesicles secreted by human primary conjunctival goblet cells during allergic inflammation

AimsConjunctival epithelium lines the inside of the eyelids and covers the sclera, thus providing stability to the eye surface. Goblet cells in conjunctival epithelium (CjGCs) are well known for their mucin-secretion function, which wet and protect the ocular surface, but other aspects are still not well understood. To expand our understanding beyond their mucin-secreting function, we investigated CjGC-secreted extracellular vesicles (EVs) and lipid mediators therein. Materials and methodsUsing histamine-mediated allergic inflammation in human primary CjGCs (HCjGCs) as a disease model, we quantified using ELISA a proinflammatory mediator PGE2 and two specialized pro-resolving mediators (SPMs) LXA4 and RvD1 in EVs secreted during allergic inflammation. Key findingsAt 18 h post histamine stimulation, the amount of LXA4 and RvD1 in EVs was notably higher compared to those in unstimulated. Interestingly, this increase was only observed in female EVs but not in males. The mean fold increase of LXA4 and RvD1 in female EVs was 3.9 and 3.4, respectively, but it was only 0.9 and 1.0 in male EVs. Supplying docosahexaenoic acid (DHA, the source of RvD1 and other SPMs) to the culture medium during the allergic inflammation resulted in even higher mean fold increase of 5.3 and 6.9 for LXA4 and RvD1 in female EVs, respectively, but it was only 0.5 and 0.8 in male EVs. SignificanceWe conclude that HCjGCs show a clear sex difference in allergic response. Our results may also provide a new insight into the male predisposition to severe forms of allergic conjunctivitis and potential improvement in disease care in the clinic.

Purinergic agonists increase [Ca2+]i in rat conjunctival goblet cells through ryanodine receptor type 3.

ATP and benzoylbenzoyl-ATP (BzATP) increase free cytosolic Ca2+ concentration ([Ca2+]i) in conjunctival goblet cells (CGCs) resulting in mucin secretion. The purpose of this study was to investigate the source of the Ca2+i mobilized by ATP and BzATP. First-passage cultured rat CGCs were incubated with Fura-2/AM, and [Ca2+]i was measured under several conditions with ATP and BzATP stimulation. The following conditions were used: 1) preincubation with the Ca2+ chelator EGTA, 2) preincubation with the SERCA inhibitor thapsigargin (10-6 M), which depletes ER Ca2+ stores, 3) preincubation with phospholipase C (PLC) or protein kinase A (PKA) inhibitor, or 4) preincubation with the voltage-gated calcium channel antagonist nifedipine (10-5 M) and the ryanodine receptor (RyR) antagonist dantrolene (10-5 M). Immunofluorescence microscopy (IF) and quantitative reverse transcription polymerase chain reaction (RT-qPCR) were used to investigate RyR presence in rat and human CGCs. ATP-stimulated peak [Ca2+]i was significantly lower after chelating Ca2+i with 2 mM EGTA in Ca2+-free buffer. The peak [Ca2+]i increase in CGCs preincubated with thapsigargin, the PKA inhibitor H89, nifedipine, and dantrolene, but not the PLC inhibitor, was reduced for ATP at 10-5 M and BzATP at 10-4 M. Incubating CGCs with dantrolene alone decreased [Ca2+]i and induced CGC cell death at a high concentration. RyR3 was detected in rat and human CGCs with IF and RT-qPCR. We conclude that ATP- and BzATP-induced Ca2+i increases originate from the ER and that RyR3 may be an essential regulator of CGC [Ca2+]i. This study contributes to the understanding of diseases arising from defective Ca2+ signaling in nonexcitable cells.NEW & NOTEWORTHY ATP and benzoylbenzoyl-ATP (BzATP) induce mucin secretion through an increase in free cytosolic calcium concentration ([Ca2+]i) in conjunctival goblet cells (CGCs). The mechanisms through which ATP and BzATP increase [Ca2+]i in CGCs are unclear. Ryanodine receptors (RyRs) are fundamental in [Ca2+]i regulation in excitable cells. Herein, we find that ATP and BzATP increase [Ca2+]i through the activation of protein kinase A, voltage-gated calcium channels, and RyRs, and that RyRs are crucial for nonexcitable CGCs' Ca2+i homeostasis.

Clinical ocular surface characteristics and expression of MUC5AC in diabetics: a population-based study.

To investigate the clinical characteristics and the expression of mucin 5AC (MUC5AC) in in diabetic and non-diabetic subjects with or without dry eye disease. A total of 399 participants (796 eyes) aged 50-80 years were enrolled in this study. Participants were divided into 4 groups: non-diabetic without dry eye group (normal group), non-diabetic with dry eye group, diabetic without dry eye group and diabetic with dry eye group. Demographic information, fasting plasma glucose (FBG), and glycated haemoglobin A1C (HbA1C) data were collected. Additionally, ocular surface disease index (OSDI) questionnaire, signs of dry eye, tear osmolarity, and meibomian glands were evaluated. Tear MUC5AC expression and conjunctival goblet cells density (GCD) were tested. Compared with non-diabetic with dry eye group, diabetic with dry eye group showed significantly lower tear film osmolarity (TFO), but higher corneal fluorescein and conjunctival lissamine green staining scores. In comparison with diabetic without dry eye group, diabetic with dry eye group showed significantly higher TFO, corneal fluorescein and conjunctival lissamine green staining scorers. The MUC5AC concentration and GCD of diabetic with dry eye group was significantly lower than those of the non-diabetic with dry eye group. Diabetic subjects with higher HbA1c levels (≥7.8%) showed higher TFO and shorter fluorescein tear break time. Diabetics with dry eye exhibited notably higher corneal fluorescein and conjunctival lissamine green staining scores. Conjunctival goblet cells and MUC5AC were significantly reduced in diabetics. Higher TFO was associated with the duration of diabetes and HbA1c levels.

Toxicity and efficacy of type I interferons on the ocular surface: in vitro, animal, and clinical studies

PurposeTo investigate the toxicity of type I interferons (IFNs) on the ocular surface and assess their efficacy in ocular surface tumors. MethodsWe examined the effects of IFN-α2a, IFN-α2b and IFN-β on corneal epithelial cells and stromal fibroblasts in vitro as well as the impact of IFN-α2a on the ocular surface in mice. Additionally, we analyzed the therapeutic and adverse effects of topically administered IFN-α2a and IFN-α2b in patients with ocular surface tumors. Risk factors contributing to side effects were explored. ResultsIFN-α2a, IFN-α2b or IFN-β reduced cell viability and induced pro-inflammatory cytokines in corneal epithelial cells and stromal fibroblasts. Furthermore, IFNs enhanced the expression of major histocompatibility complex class II and CD40 in corneal epithelial cells. In mice, subconjunctival IFN-α2a injection did not induce corneal epithelial defects or opacity, nor did it reduce aqueous tears or conjunctival goblet cells. In patients, topical IFN-α2a or IFN-α2b administration decreased tumor size and prevented recurrence; however, it was associated with mild side effects, including corneal epitheliopathy and conjunctival hyperemia. These complications were associated with longer IFN use, the presence of underlying ocular surface disease and concurrent use of mitomycin C or anti-glaucoma eye drops. ConclusionAlthough type I IFNs cause direct toxicity on corneal cells, they do not induce significant side effects on the healthy ocular surface. Considering its therapeutic and preventive effects, topical type I IFN is safe and effective for treating ocular surface tumors. The potential for ocular side effects should be considered in eyes with identified risk factors.

THE RELATIONSHIP BETWEEN RISKS OF OBSTRUCTIVE SLEEP APNEA AND DRY EYE DISEASE

Introduction: Dry Eye Disease (DED) may lead to decreased visual function, chronic tissue changes, eyelid, conjunctival, and corneal abnormality. The prevalence of DED in Asia ranges from 21.6%–37.6%. Several studies pointed out the relationship between DED and Obstructive Sleep Apnea (OSA). In patients with OSA, oxidative stress, hypoxia, and ocular surface inflammation increase, leading to a decrease in the functions of meibomian glands, goblet cells, corneal sensitivity, and tear production as a response to the stimulations from the lacrimal glands. The loss of conjunctival goblet cells and meibomian glands are implications of damage to the tear film quality, which results in DED. Method: This is a population-based cross-sectional study. Data were obtained from the database of Biomarker Smarthealth Research in Mendalanwangi, Sidorahayu, and Cepokomulyo Villages, which are located in Malang Regency, having risks of OSA (based on the STOP-BANG questionnaire), above ≥40 years old, and DED Examination with Tear Break-Up Time (TBUT), MGD (Meibominan Gland Dysfunction), Tear Meniscus (TM) involving a total of 518 respondents. The available data were then processed according to the variables and went through DED examinations. This study employed the purposive sampling method. Result: In this study, moderate risk of OSA suggests a significant influence on the occurrence of DED with an odds ratio (OR) 1.66 (p&lt;0.05). In addition, moderate risk of OSA predisposes ADDE (Aqueous Deficiency Dry Eye)-type and Mix-type DED with OR 2.85 and 1.23 (p&lt;0.05). High risk of OSA correlates with the occurrence of ADDE-type DED with OR 2.37 (p&lt;0.05). The age group &gt;60 years old shows a correlation with Mix-type DED with OR 2.09 (p&lt;0.05). Women have a higher predisposition to ADDE-type with OR 2.58 (p&lt;0.05). Conclusion: Moderate risk of OSA influences the occurrence of DED, both ADDE-type and Mix type; whereas high risk of OSA only correlates with ADDE-type. Older age plays a role in the occurrence of Mix-type DED and women have a higher tendency to present with ADDE- type.

Circadian disruption reduces MUC4 expression via the clock molecule BMAL1 during dry eye development

Circadian disruption, as a result of shiftwork, jet lag, and other lifestyle factors, is a common public health problem associated with a wide range of diseases, such as metabolic disorders, neurodegenerative diseases, and cancer. In the present study, we established a chronic jet lag model using a time shift method every 3 days and assessed the effects of circadian disruption on ocular surface homeostasis. Our results indicated that jet lag increased corneal epithelial defects, cell apoptosis, and proinflammatory cytokine expression. However, the volume of tear secretion and the number of conjunctival goblet cells did not significantly change after 30 days of jet lag. Moreover, further analysis of the pathogenic mechanism using RNA sequencing revealed that jet lag caused corneal transmembrane mucin deficiency, specifically MUC4 deficiency. The crucial role of MUC4 in pathogenic progression was demonstrated by the protection of corneal epithelial cells and the inhibition of inflammatory activation following MUC4 replenishment. Unexpectedly, genetic ablation of BMAL1 in mice caused MUC4 deficiency and dry eye disease. The underlying mechanism was revealed in cultured human corneal epithelial cells in vitro, where BMAL1 silencing reduced MUC4 expression, and BMAL1 overexpression increased MUC4 expression. Furthermore, melatonin, a circadian rhythm restorer, had a therapeutic effect on jet lag-induced dry eye by restoring the expression of BMAL1, which upregulated MUC4. Thus, we generated a novel dry eye mouse model induced by circadian disruption, elucidated the underlying mechanism, and identified a potential clinical treatment.

Preliminary study of cyclosporine A/Lifitegrast subconjunctival sustained-release drug membrane in the treatment of dry eyes

BackgroundDry eyes can cause discomfort. To treat dry eye disease, cyclosporine A (CsA) and Lifitegrast are two eye drugs approved by the U.S. Food and Drug Administration (FDA). However, frequent use of eye drops can be challenging and lead to poor compliance, especially in elderly patients. Therefore, this study aimed to develop a drug sustained-release vector and explore its therapeutic effect in animal models of dry eye.MethodsFirstly, drug membranes loaded with both CsA and Lifitegrast using a carrier called poly(lactate-co-ε-caprolactone) (P(LLA-CL)) were prepared and evaluated for their physicochemical properties, release behavior in vitro, and safety in vivo. Next, a rabbit dry eye model using a 0.1% benzalkonium chloride (BAC) solution was developed and treated by drug-loaded micro membranes. We observed and recorded conjunctival hyperemia, corneal staining, corneal edema, corneal neovascularization, conjunctival goblet cells and hematoxylin and eosin (H&E) staining. Finally, we detected the MUC5AC and MMP-9 by immunofluorescence staining and enzyme-linked immunosorbent assay (ELISA).ResultsThe composite film released both CsA and Lifitegrast for at least one month. Compared to the blank membrane group, conjunctival hyperemia, corneal fluorescein staining, corneal edema, corneal neovascularization and conjunctival goblet cells recovered faster in the drug membrane group, and the difference was statistically significant. At the molecular level, the drug membrane group showed an increase in mucin density and a significant anti-inflammatory effect.ConclusionsThe implantation of CsA/Lifitegrast loaded P(LLA-CL) membrane under the subconjunctival of the rabbit eye is safe. The study suggests that this subconjunctival administration could be developed into a minimally invasive delivery system to help patients with dry eye disease who require multiple daily eyedrops but have poor compliance.

Extended depth-of-field wide-field fluorescence microscopy with a micro-mirror array lens system for versatile cellular examination.

We present a versatile extended depth-of-field (EDOF) wide-field fluorescence microscopy using a new, to the best of our knowledge, active device, micro-mirror array lens system (MALS) for calibration-free and orientation-insensitive EDOF imaging. The MALS changed the focal plane during image acquisition, and the system could be operated in any orientation. Two EDOF imaging modes of high-speed accumulation and low-speed surface sectioning were implemented. The performance was demonstrated in non-contact imaging of conjunctival goblet cells in live mice and depth-resolved cellular examination of ex-vivo human cancer specimens. MALS-based EDOF microscopy has potential for versatile cellular examination.

Topical naltrexone increases aquaporin 5 production in the lacrimal gland and restores tear production in diabetic rats.

Diabetes mellitus is a prevalent disease that is often accompanied by ocular surface abnormalities including delayed epithelial wound healing and decreased corneal sensitivity. The impact of diabetes on the lacrimal functional unit (LFU) and the structures responsible for maintaining tear homeostasis, is not completely known. It has been shown that the Opioid Growth Factor Receptor (OGFr), and its ligand, Opioid Growth Factor (OGF), is dysregulated in the ocular surface of diabetic rats leading to overproduction of the inhibitory growth peptide OGF. The opioid antagonist naltrexone hydrochloride (NTX) blocks the OGF-OGFr pathway, and complete blockade following systemic or topical treatment with NTX restores the rate of re-epithelialization of corneal epithelial wounds, normalizes corneal sensitivity, and reverses dry eye in diabetic animal models. These effects occur rapidly and within days of initiating treatment. The present study was designed to understand mechanisms related to the fast reversal (<5 days) of dry eye by NTX in type 1 diabetes (T1D) by investigating dysregulation of the LFU. The approach involved examination of the morphology of the LFU before and after NTX treatment. Male and female adult Sprague-Dawley rats were rendered hyperglycemic with streptozotocin, and after 6 weeks rats were considered to be a T1D model. Rats received topical NTX twice daily to one eye for 10 days. During the period of treatment, tear production and corneal sensitivity were recorded. On day 11, animals were euthanized and orbital tissues including conjunctiva, eyelids, and lacrimal glands, were removed and processed for histologic examination including immunohistochemistry. Male and female T1D rats had significantly decreased tear production and corneal insensitivity, significantly decreased number and size of lacrimal gland acini, decreased expression of aquaporin-5 (AQP5) protein and decreased goblet cell size. Thus, 10 days of NTX treatment restored tear production and corneal sensitivity to normal values, increased AQP5 expression, and restored the surface area of goblet cells to normal. NTX had no effect on the number of lacrimal gland acini or the number of conjunctival goblet cells. In summary, blockade of the OGF-OGFr pathway with NTX reversed corneal and lacrimal gland complications and restored some components of tear homeostasis confirming the efficacy of topical NTX as a treatment for ocular defects in diabetes.

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

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. 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. 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. 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.

Role of NADPH Oxidase 4 on Dry Eye Syndrome in Mice.

Objective: This study aims to investigate the effect of NADPH oxidase 4 (NOX4)-mediated inflammation on concanavalin A (ConA)-induced dry eye syndrome (DES) in mice. Methods: Thirty-six mice were randomly divided into Control, Model, no-load Control, and NOX4 interference group. Adenovirus was injected (10 μL) into the lacrimal glands of both eyes of mice in no-load Control group and NOX4 interference group. Four days after adenovirus injection, the Control group was injected with phosphate-buffered saline, and the other groups were injected with ConA (200 μg) in the lacrimal glands of mice to establish DES models. The tear secretion rate was estimated by phenol red thread test. Lissamine green eye staining was used to evaluate conjunctival damage. The corneal surface was observed by hematoxylin-eosin (HE) staining and scanning electron microscopy (SEM). The morphology and quantity of conjunctival epithelial cells and goblet cells were observed by Periodic acid-Schiff staining. The expression of NOX4, NOD-like receptor thermal protein domain-associated protein 3 (NLRP3), interleukin-1β (IL-1β), and mucin 5 subtype AC (MUC5AC) was detected by immunohistochemistry. Results: Compared with the Control group, the Model group showed a significant decrease in tear secretion and an upregulation in microscopic image score. The HE staining and SEM showed corneal and conjunctiva damage in the Model group. The protein expression of NOX4, NLRP3, and IL-1β was upregulated, but MUC5AC was downregulated in the Model group. After interfering with NOX4, all these indicators were reversed. Conclusion: The pathological process of concanavalin A-induced DES appears to be related to NOX4.

Imaging assessment of conjunctival goblet cells in dry eye disease.

Dry eye disease (DED) is a widespread, multifactorial, and chronic disorder of the ocular surface with disruption of tear film homeostasis as its core trait. Conjunctival goblet cells (CGCs) are specialised secretory cells found in the conjunctival epithelium that participate in tear film formation by secreting mucin. Changes in both the structure and function of CGCs are hallmarks of DED, and imaging assessment of CGCs is important for the diagnosis, classification, and severity evaluation of DED. Existing imaging methods include conjunctival biopsy, conjunctival impression cytology and in vivo confocal microscopy, which can be used to assess the morphology, distribution, and density of the CGCs. Recently, moxifloxacin-based fluorescence microscopy has emerged as a novel technique that enables efficient, non-invasive and in vivo imaging of CGCs. This article presents a comprehensive overview of both the structure and function of CGCs and their alterations in the context of DED, as well as current methods of CGCs imaging assessment. Additionally, potential directions for the visual evaluation of CGCs are discussed.

Ocular surface changes in patients who have undergone head and neck radiation therapy.

Dry eye syndrome (DES) is a familiar sequelae of radiation therapy (RT) for head and neck cancers (HNC). Ocular surface changes such as DES occur due to injury to the conjunctival epithelium, goblet cells, corneal surface, lacrimal glands, and meibomian glands. This study aimed at the evaluation and early detection of changes in ocular surface parameters in patients receiving RT for extraocular HNC. Forty-two eyes of 21 patients undergoing HNC RT were evaluated. Radiation technique and dose of radiation to the lens and eye were recorded. Subjects were evaluated for meibomian gland changes by meiboscore grading, ocular surface disease index (OSDI) questionnaire, Schirmer's test, tear film break-up time (TBUT), and slit-lamp examination before RT, immediately post RT, and 6 weeks post RT. A comparison of the ipsilateral eye on the irradiated side to the contralateral eye was done. A significant reduction in TBUT was seen immediately post RT and 6 weeks post RT ( P < 0.001 and 0.008, respectively), with an increase in meiboscore at both visits ( P < 0.001). An OSDI score of >13 was seen in 23.80% of patients post RT, with a significant difference from baseline ( P < 0.001). On comparing ipsilateral and contralateral eye groups, a significant difference from baseline was seen in TBUT ( P < 0.001 and 0.033, respectively) and meiboscore ( P < 0.001 for both eyes). A significant change of >1 second in TBUT and >1.7 in meiboscore was seen with a mean dose of around 8 Gy to the lens. All patients undergoing HNC RT should be followed up for ocular surface and meibomian gland changes. The contralateral eye should also be evaluated. Patients receiving lower doses to the ocular structures should also be kept under follow-up.

Therapeutic effect and mechanism of action of pterostilbene nano drugs in dry eye models

Dry eye disease is a multifactorial dysfunction of the tear film and ocular surface, with etiology involving inflammation and oxidative stress on the ocular surface. Pterostilbene (PS) is a secondary metabolite extracted from plants, which possesses remarkable anti-inflammatory and antioxidant effects. However, its application is limited by light instability and very poor water solubility. We modified fat-soluble PS into a biparental pterostilbene-glutaric anhydride-arginine-glycine-aspartic acid (PS-GA-RGD) nanomedicine by prodrug ligation of functional peptides. The aim of this study was to explore the protective effect and potential mechanism of PS-GA-RGD on dry eye disease in vitro and in vivo. We demonstrated good long-term biocompatibility of PS-GA-RGD through rabbit eye stimulation test. Lipopolysaccharide (LPS) was used to induce murine macrophages RAW 264.7 to establish an inflammation and oxidative stress model. In this model, PS-GA-RGD effectively reduced the production of ROS and 8-OHdG, enhancing the expression of antioxidant factor Nrf2 and antioxidant enzyme heme oxygenase-1. In addition, the expression of NF-κB inflammatory pathway significantly increased in LPS-induced RAW 264.7 cells, while PS-GA-RGD could significantly reduce this pathway. Hypertonic saline was utilized to establish a hypertonic model of human corneal epithelial cells. PS-GA-RGD was found to significantly reduce the production of ROS and NLRP3 inflammasomes in this model, exhibiting superior efficacy compared to PS. Experimental dry eye animal models were co-induced with subcutaneous injection of scopolamine and an intelligently controlled environmental system. We demonstrated that PS-GA-RGD nano drugs can prevent and reduce corneal epithelial cell defects and apoptosis, protect conjunctival goblet cells, and have an excellent anti-inflammatory effect. Finally, we demonstrated that RGD sequence in PS-GA-RGD can enhance cellular uptake, corneal retention, and penetration, thereby increasing their bioavailability and efficacy by a cell uptake assay and rabbit corneal drug retention experiment. Overall, this study highlights the potential of PS-GA-RGD nanomedicines in the treatment of dry eyes.

Periodontal Ligament Stem Cell-Derived Exosomes Regulate Muc5ac Expression in Rat Conjunctival Goblet Cells via Regulating Macrophages Toward an Anti-Inflammatory Phenotype

ABSTRACT Background Several studies have reported the protective effects of mesenchymal stem cell-derived exosomes (MSC-Exos) in reducing inflammation and decreasing conjunctival goblet cell (CGC) loss in dry eye disease. However, whether MSC-Exos provide anti-inflammatory profiles in macrophages, thus contributing to CGC protection, has remained elusive. Methods Macrophages were incubated with PKH26‐labeled periodontal ligament mesenchymal stem cell-derived exosomes (PDLSC-Exos) for 12 h, and uptake of PDLSC-Exos by macrophages was observed by a confocal fluorescence microscope. The mRNA expression of TNF-α, IL-10, and Arg1 was detected by quantitative real-time polymerase chain reaction (qRT-PCR). The protein expression of TNF-α and IL-10 were quantified using western blotting. Then, CGCs were exposed to different macrophage supernatants and qRT-PCR was used to detect the Muc5ac mRNA expression of CGCs in response to or absence of cholinergic stimulation. ELISA was used to determine the Muc5ac secretion of CGCs in response to cholinergic stimulation. Results The uptake of PDLSC-Exos by M1 macrophages facilitates M2 macrophage polarization with the elevated expressions of IL-10 and Arg1. In macrophage supernatant-treated CGCs systems, PDLSC-Exo-treated M1 macrophage supernatant significantly enhanced the Muc5ac expression of CGCs in response to, or in the absence of, cholinergic stimulation, while the addition of PDLSC-Exos to the control macrophage supernatant did not generate a change in Muc5ac expression. Conversely, the addition of PDLSC-Exos to the diluted control macrophage supernatant induced a significant increase in Muc5ac expression. Conclusion PDLSC-Exos could protect CGCs against M1 macrophage-mediated inflammation, and the protective effects of PDLSC-Exos are partly attributable to their effects on M1 macrophages.

Ocular surface disease in moderate-to-severe atopic dermatitis patients and the effect of biological therapy.

Atopic dermatitis (AD) is a chronic inflammatory skin disease for which new targeted therapies are currently available. Due to the increased rates of ocular surface disease (OSD) reported during treatment with these new targeted treatments, more insight into the occurrence and pathomechanism of OSD in moderate-to-severe AD patients is needed. Therefore, this review's first part highlights that most patients with moderate-to-severe AD already have characteristics of OSD before starting targeted treatment. Remarkably, not all AD patients with OSD report ocular symptoms. OSD in AD is associated with less conjunctival goblet cells (GC) compared to healthy controls. In addition, OSD severity in AD patients is associated with high AD activity, the presence of eyelid and/or facial eczema, and high levels of AD-related severity biomarkers in tear fluid. The second part of this review highlights that pre-existing ocular pathology (e.g. in combination with the use of ophthalmic medication or eyelid eczema) may be associated with the development of dupilumab-associated ocular surface disease (DAOSD). During dupilumab treatment, DAOSD (which can be new-onset OSD or worsening of pre-existing OSD) is observed in approximately one-third of the dupilumab-treated AD patients. Anti-inflammatory ophthalmic treatment improves DAOSD, and dose reduction of dupilumab may also be an effective treatment option. The pathomechanism of DAOSD is still not fully elucidated. In a prospective study low, but stable conjunctival GC numbers were observed in moderate-to-severe AD patients, before and during dupilumab treatment. However, the Mucin 5 AC (MUC5AC) expression of GCs decreased during dupilumab treatment, suggesting an impairment of the GC function by dupilumab treatment. In addition, higher dupilumab tear fluid levels were found in dupilumab-treated AD patients with moderate-to-severe OSD compared to patients with no or mild OSD, whereas the dupilumab serum levels are similar. Clinicians should be aware of the frequent occurrence of OSD in moderate-to-severe AD patients, and a low-threshold referral to an ophthalmologist is recommended.