Cornea Epithelium Research Articles

Case Report of Benign Diffuse Anterior Scleritis Associated with SARS-CoV-2

COVID-19 is an emerging infectious disease classified as a zoonotic illness caused by the SARS-CoV-2 strain of coronavirus. This enveloped, positive-sense, non-segmented RNA virus belongs to the order Nidovirales and the family Coronaviridae. Its primary mode of transmission is human-to-human through respiratory droplets. Ophthalmologists worldwide have reported various ocular manifestations associated with the infection. Recent studies have shown that SARS-CoV-2 binds to the cellular receptor for angiotensin-converting enzyme 2 (ACE2) and interacts with the transmembrane serine protease 2 (TMPRSS2), both of which are known to be expressed in the human cornea, retina, and conjunctival epithelium [1, 2]. The aim of this work is to present a case of a patient who developed diffuse viral scleritis related to COVID-19, detailing the diagnostic and therapeutic approaches as well as the patient's progression. We report the case of a 63-year-old man with a history of high triglycerides who developed flu-like symptoms and severe tiredness for over a week. A COVID-19 test came back positive. Ten days later, he had a painful, red right eye but no change in vision. An eye exam showed some redness in the conjunctiva, a clear cornea, and normal pressure in the eye. The fellow eye looked normal as well. He was treated with topical steroids and artificial tears, which helped initially. However, after finishing treatment, his symptoms worsened with more redness and pain, and now both eyes were affected. Further examination of the right eye showed swelling of the eyelid (preseptal cellulitis) and widespread redness in the conjunctiva. The Neosynephrine test was negative, and there was pain when moving the eye. The cornea was clear but had some superficial punctate keratitis, and the anterior chamber looked normal. The left eye also showed redness but was otherwise normal. A full lab workup (including blood tests and serologies for HSV1-2) came back normal. Considering his previous C

Characterizing Biomechanics of Limbal Niche Using Vibrational Optical Coherence Elastography.

The limbal niche is an adult source of epithelial stem cells which regenerate the cornea epithelium. The architecture and biomechanical properties of the limbus have previously been demonstrated to change due to aging and disease. This study aims to non-destructively and simultaneously quantify these limbal niche properties, along with their age-related changes. A lab-built vibrational optical coherence elastography (OCE) device consisting of a phase-sensitive optical coherence tomography (OCT) with a vibrational stimulator has been used to collect structural images and the depth-resolved elasticity of human corneoscleral tissues (aged 4-96 years old). The limbal palisades of Vogt (POV) were delineated well in OCT images which were validated by histology. The POVs have been spatially mapped with simultaneous elasticity measurements in cross-sections, showing tissue stiffness distributions across the undulations. A significant influence of age on the dimensions of the POVs was explored. The elastic modulus within the limbal niches for the ≥65-year-old group was significantly higher than that of the <65-year-old group.

Proteomic Characterization of Corneal Epithelial and Stromal Cell-Derived Extracellular Vesicles.

Communication between the different layers of the cornea (epithelium and stroma) is a complex, yet crucial element in the corneal healing process. Upon corneal injury, it has been reported that the bi-directional cross talk between the epithelium and stroma via the vesicular secretome, namely, extracellular vesicles (EVs), can lead to accelerated wound closure upon injury. However, the distinct protein markers of EVs derived from human corneal epithelial (HCE) cells, keratocytes (HCKs), fibroblasts (HCFs), and myofibroblasts (HCMs) remain poorly understood. All EVs were enriched for CD81 and showed increased expression levels of ITGAV and FN1 in HCM-EVs compared to HCE- and HCF-EVs. All EVs were negative for GM130 and showed minimal differences in biophysical properties (particle concentration, median particle size, and zeta potential). At the proteomic level, we show that HCM-EVs are enriched with proteins associated with fibrosis pathways, such as COL6A1, COL6A2, MMP1, MMP2, TIMP1, and TIMP2, compared to HCE-, HCK-, and HCF-EVs. Interestingly, HCE-EVs express proteins involved with the EIF-2 signaling pathway (stress-induced signals to regulate mRNA translation), such as RPS21, RALB, EIF3H, RALA, and others, compared to HCK-, HCF-, and HCM-EVs. In this study, we isolated EVs from cell-conditioned media from HCE, HCKs, HCFs, and HCMs and characterized their biophysical and protein composition by Western blot, nanoparticle tracking analysis, and proteomics. This study supports the view that EVs from the corneal epithelium and stroma have a distinct molecular composition and may provide novel protein markers to distinguish the difference between HCE-, HCK-, HCF-, and HCM-EVs.

Moringa oleifera hydroalcoholic leaf extracts mitigate valproate-induced oxidative status in the extraorbital lacrimal gland in a rat model

Dysfunction of the extraorbital lacrimal gland (ELG) can lead to loss of vision due to damage to the epithelium of cornea. The broad-spectrum anti-epileptic drug sodium valproate (SV) has numerous side effects. Moringa oleifera (M.oleifera) is widely used as a food and in folk medicine. The effects of orally administered SV and M. oleifera hydroalcoholic leaf extract on rat ELG were investigated in this study by analysing both antioxidant and oxidant parameters. Additionally, boron level and tissue factor (TF) activity were determined. Protein changes were detected by sodium dodecyl sulfate gel electrophoresis (SDS-PAGE). Significantly lower values of glutathione (GSH), superoxide dismutase (SOD), catalase (CAT) and total antioxidant status (TAS) were observed in the SV group compared to the control group. Treatment with Moringa extract significantly increased SOD, CAT and TAS values in the Moringa given SV group (SVM). While no significant differences were observed between the sialic acid values of the groups, lipid peroxidation (LPO), nitric oxide (NO) and total oxidant status (TOS) values were significantly elevated in the SV group compared to the control group. Due to the effect of Moringa extract, LPO, NO and TOS levels were significantly decreased in the SVM group compared to the SV group. TF activity was not meaningfully altered between groups. Compared to control rats, oxidative stress index (OSI) level significantly increased, whereas the boron level decreased in the SV group. Moringa extract treatment noticeably reduced OSI in the SVM group. According to SDS-PAGE, decreases in the density of protein bands with molecular weights of 51, 83, and 90 kDa were observed in SV given rats compared to the other groups. These decreases were reversed by the administration of Moringa extract. Moringa extract has shown protective properties arising from antioxidant potential, especially with its very low OSI value. Individuals undergoing SV treatment and having ELG complications might consider using Moringa extract to mitigate valproate induced damage.

Cyclic RGD conjugated PEG-PLGA nanoparticle enhances preventative effect of bilirubin on the development of ultraviolet-B (UVB)-induced cataract

Age related cataract (ARC) is characterized by protein conformational disorder with aging stands out as an independent risk factor. Therefore, mitigation of oxidative stress and DNA damage within the lens emerges as a potential therapeutic strategy. While topical administration remains the most common treatment modality for ocular disorder, drug bioavailability is hindered by the cornea epithelium barrier. In this study, we developed a novel cyclic Arg-Gly-Asp (cRGD)-conjugated nanoparticle encapsulating bilirubin (cRNM@BR), an endogenous antioxidant, to combat against cellular senescence and lens opacity associated with ARC. Immunofluorescence staining assay confirmed that cRGD modification significantly prolonged nanoparticle retention on the ocular surface by targeting integrin expressed on corneal epithelium. Moreover, through LC-MS/MS analysis, we demonstrated the enhanced transcorneal penetration efficacy of cRGD-modified nanodrugs both in vivo and in vitro. Our mechanistic investigations revealed that cRNM entered human corneal epithelial cells (HCECs) by clathrin and micropinocytosis-mediated pathways, with the ER/Golgi pathway facilitating its exocytosis. In a mouse model of ultraviolet B (UVB)-induced cataract, topical application of cRNM@BR effectively alleviated intracellular reactive oxygen species (ROS) levels and DNA damage in lens epithelial cells (LECs), thereby preventing cellular senescence and lens opacity. Transcriptomics analysis of LECs in UVB-exposed group and cRNM@BR pretreated group indicated significant enrichment of the cGAS/STING pathway. Further experiments determined that cRNM@BR prevented cellular senescence and lens opacity by ameliorating the cGAS/STING pathway. In conclusion, our findings underscore the therapeutic potential of cRNM@BR in ARC management and highlight the efficacy of cRGD-conjugated nanoplatform for intraocular delivery.

Effects of Ripasudil Hydrochloride on Epithelial Repair in a Mouse Cornea

Purpose Effect of topical administration of a Rho kinase inhibitor, ripasudil, on epithelial wound healing in a mouse cornea was investigated. Effects of treatment of cultured human corneal epithelial cell (HCEC) line and organ-cultured corneal epithelium with ripasudil on expression of p-ERK was also examined. Methods Epithelial defects with a diameter of 2.0 mm were prepared in the central corneas of C57BL/6 mice with or without 1-week travoprost pre-treatment, to which ripasudil or PBS as a control was instilled every 6 h immediately after preparation. The mice eyes were cultured with or without travoprost for 24-hrs. The expression levels of p-ERK in epithelium of mice eyes were compared by immunostaining after further 24-hrs culture with or without ripasudil for 24-hrs. HCEC were cultured with or without ripasudil and processed for examination for proliferation activity and protein expression of p-ERK by either immunostaining or Western blotting. The cells were also treated with or without travoprost for 24-hrs, and were further cultured with or without ripasudil. Expression levels of p-ERK were examined by Western blotting. Results Ripasudil treatment suppressed post-debridement epithelial healing in association with reduced proliferation activity in peripheral (limbal) epithelium in cornea with or without pre-treatment with travoprost. Ripasudil treatment accelerated p-ERK expression. Ripasudil supplementation upregulated proliferation with increased p-ERK in HCEC. Conclusion Ripasudil treatment promotes wound healing of the mouse corneal epithelium by enhancing cell proliferation on peripheral (limbal) epithelium.

Comparison of the repeatability and reproducibility of corneal thickness mapping using optical coherence tomography according to tear film break-up time

BackgroundTo compare the repeatability and reproducibility of corneal and corneal epithelial thickness mapping using anterior segment optical coherence tomography (AS-OCT) according to tear film break-up time (TBUT).MethodsThe included eyes were divided into three subgroups according to TBUT (group 1: TBUT ≤ 5 s, group 2: 5 s < TBUT ≤ 10 s, and group 3: TBUT > 10 s). All eyes were imaged separately thrice by two operators to obtain the thickness maps (TMs) of the cornea and corneal epithelium based on spatial zones encompassing a 9-mm-diameter area. Each TM consisted of 25 areas. Intraoperator (repeatability) and interoperator (reproducibility) standard deviations (Sws), coefficients of variation (CoVs), and intraclass correlation coefficients (ICCs) among the tests were calculated and compared in all the areas.ResultsAltogether, 132 eyes of 67 subjects were included (50, 47, and 35 eyes in groups 1, 2, and 3; respectively). The ICCs of corneal epithelial thickness and corneal thickness were > 0.75 in most of the areas. Pairwise comparisons showed that AS-OCT exhibited lower repeatability in group 1 than in groups 2 and 3 (P < 0.05). However groups 2 and 3 showed similar results. Sws and CoVs of corneal epithelial thickness exhibited no significant interoperator differences. While no significant differences were observed in corneal thickness in most of the areas.ConclusionsTBUT significantly influences the repeatability of corneal and corneal epithelial thickness measurements. Poor tear film stability requires careful evaluation of corneal epithelial thickness.

SMAD4-Dependent Signaling Pathway Involves in the Pathogenesis of TGFBR2-Related CE-like Phenotype.

(1) Background: Our previous data indicated that disturbance of the Transforming Growth Factor beta (TGFB) signaling pathway via its Type-2 Receptor (TGFBR2) can cause a Corneal Ectasia (CE)-like phenotype. The purpose of this study is to elucidate whether the SMAD4-dependent signaling pathway is involved in the TGFBR2-related CE-like pathogenesis. (2) Methods: Smad4 was designed to be conditionally knocked out from keratocytes. Novel triple transgenic mice, KerartTA; Tet-O-Cre; Smad4flox/flox (Smad4kera-cko), were administered with doxycycline (Dox). Optical Coherence Tomography (OCT) was performed to examine Central Corneal Thickness (CCT), Corneal Radius, Anterior Chamber and CE-like phenotype and compared to the littermate Control group (Smad4Ctrl). (3) Results: The OCT revealed normal cornea in the Smad4Ctrl and a CE-like phenotype in the Smad4kera-cko cornea, in which the overall CCT in Smad4kera-cko was thinner than that of Smad4Ctrl at P42 (n = 6, p < 0.0001) and showed no significant difference when compared to that in Tgfbr2kera-cko. Furthermore, the measurements of the Anterior Chamber and Corneal Radius indicated a substantial ectatic cornea in the Smad4kera-cko compared to Smad4Ctrl. The H&E staining of Smad4kera-cko mimics the finding in the Tgfbr2kera-cko. The positive immunostaining of cornea-specific marker K12 indicating the cell fate of cornea epithelium remained unchanged in Smad4kera-cko and the Proliferating Cell Nuclear Antigen (PCNA) immunostaining further indicated an enhanced proliferation in the Smad4kera-cko. Both immunostainings recapitulated the finding in Tgfbr2kera-cko. The Masson's Trichrome staining revealed decreased collagen formation in the corneal stroma from both Smad4kera-cko and Tgfbr2kera-cko. The collagen type 1 (Col1a1) immunostaining further confirmed the reduction in collagen type 1 formation in Smad4kera-cko. (4) Conclusions: The aforementioned phenotypes in the Smad4kera-cko strain indicated that the SMAD4-dependent signaling pathway is involved in the pathogenesis of the CE-like phenotype observed in Tgfbr2kera-cko.

Absence of the Klotho Function Causes Cornea Degeneration with Specific Features Resembling Fuchs Endothelial Corneal Dystrophy and Bullous Keratopathy.

The Klotho loss-of-function mutation is known to cause accelerated senescence in many organs, but its effects on the cornea have not been published. The present study aims to investigate the effects of the Klotho null mutation on cornea degeneration and to characterize the pathological features. Mouse corneas of Klotho homozygous, heterozygous, and wild-type mice at 8 weeks of age for both genders were subject to pathological and immunohistological examinations. The results show an irregular topography on the corneal surface with a Klotho null mutation. Histological examinations revealed a reduced corneal epithelial cell density, endothelial cell-shedding, and decreased cornea stromal layer thickness in the absence of the Klotho function. Furthermore, guttae formation and the desquamation of wing cells were significantly increased, which was comparable to the characteristics of Fuchs endothelial corneal dystrophy and bullous keratopathy. The mechanism analysis showed multi-fold abnormalities, including oxidative stress-induced cornea epithelium apoptosis and inflammation, extracellular matrix remodeling in the stroma, and a disruption of epithelial repair, presumably through the epithelial-mesenchymal transition. In conclusion, cornea degeneration was observed in the Klotho loss-of-function mutant mice. These pathological features support the use of Klotho mutant mice for investigating age-related cornea anomalies, including Fuchs endothelial corneal dystrophy, bullous keratopathy, and dry eye diseases.

Development and validation of a reliable rabbit model of limbal stem cell deficiency by mechanical debridement using an ophthalmic burr

A simple and reproducible method is necessary to generate reliable animal models of limbal stem cell deficiency (LSCD) for assessing the safety and efficacy of new therapeutic modalities. This study aimed to develop and validate a rabbit model of LSCD through mechanical injury. The corneal and limbal epithelium of New Zealand White rabbits (n = 18) were mechanically debrided using an ophthalmic burr (Algerbrush II) with a 1.0-mm rotating head after 360° conjunctival peritomy. The debrided eyes were serially evaluated for changes in corneal opacity, neo-vascularization, epithelial defect and corneal thickness using clinical photography, slit lamp imaging, fluorescein staining, and anterior segment optical coherence tomography scanning (AS-OCT). Following this, an assessment of histopathology and phenotypic marker expression of the excised corneas was conducted. The experimental eyes were grouped as mild (n = 4), moderate (n = 10), and severe (n = 4) based on the grade of LSCD. The moderate group exhibited abnormal epithelium, cellular infiltration in the stroma, and vascularization in the central, peripheral, and limbal regions of the cornea. The severe group demonstrated central epithelial edema, peripheral epithelial thinning with sparse goblet cell population, extensive cellular infiltration in the stroma, and dense vascularization in the limbal region of the cornea. A significant decrease in the expression of K12 and p63 (p < 0.0001) was observed, indicating the loss of corneal epithelium and limbal epithelial stem cells in the LSCD cornea. This study demonstrates that the Alger brush-induced mechanical debridement model provides a reliable model of LSCD with comprehensive clinic-pathological features and that is well suited for evaluating novel therapeutic and regenerative approaches.

Limbal Stem Cell Dysfunction Induced by Severe Dry Eye via Activation of the p38 MAPK Signaling Pathway

Severe dry eye (SDE) can cause grievous damage of the ocular surface and result in vision impairment and even blindness. To investigate the fate of limbal stem cells in SDE and the underlying mechanism, we established an SDE rat model by removing the extraorbital and infraorbital lacrimal glands and maintaining them in a low-humidity environment. One month after the surgery, aqueous tear secretion was reduced dramatically, blood vessels invaded into the central cornea, and inflammatory cells infiltrated into the limbal stroma. The expression of K12 and Pax6 were down-regulated dramatically, while those of K10, Sprr1b, and MUC5AC were up-regulated in the corneal epithelium of the SDE rats. Cell proliferation in the limbal epithelium was up-regulated, while the stem/progenitor marker adenosine 5'-triphosphate-binding cassette member 2 and the limbal epithelial colony-forming efficiency were decreased in the SDE condition. Furthermore, the p38 mitogen-activated protein kinase signaling pathway was activated in the limbal corneal epithelium of SDE rats. The abnormal differentiation and stemness loss in the cornea epithelium could be reversed upon treatment with a p38 inhibitor in a SDE invivo model and invitro hyperosmolar corneal epithelial culture conditions. In conclusion, SDE can lead to limbal stem cell dysfunction, and p38 mitogen-activated protein kinase signaling pathway activation plays an essential role in this process.

Impact of Corneal Toricity on the Distribution of Corneal Epithelial Thickness.

To investigate the impact of corneal toricity on the distribution characteristics of corneal epithelial thickness (CET). A total of 330 eyes in 330 healthy participants were included in this study. They were divided into two groups based on the median of the corneal toricity value: low-toricity group (corneal toricity < 1.50 diopters) and high-toricity group (corneal toricity ≥ 1.50 diopters). The CET within a 9-mm-diameter area of the central cornea was obtained using optical coherence tomography. The difference of CET value between flat and steep meridians (F-S CET) was defined to evaluate the CET distribution. The F-S CET between the two groups was compared, and the correlations between F-S CET and the corneal toricity were analyzed. The CET was thinner in the superior-peripheral area than in other areas. A slight intergroup difference was noted in terms of the F-S CET at the paracentral (0.11 ± 0.93 vs 0.32 ± 0.92, P = .038), midperipheral (0.45 ± 0.78 vs 0.77 ± 0.89, P = .001), and peripheral (3.11 ± 2.18 vs 4.10 ± 2.38, P < .001) zone. In each zone, the difference in F-S CET between the two groups was less than 1 μm. As the area expanded, the F-S CET continued to increase (F = 850.303, P < .001). A weak correlation was observed between F-S CET and corneal toricity (r = 0.103 to 0.240); however, this correlation was not significant in the paracentral zone. Covariance analysis demonstrated that F-S CET was slightly correlated with age, refractive state, and intraocular pressure. The corneal toricity did not significantly affect the distribution of the corneal epithelium in normal corneas. [J Refract Surg. 2023;39(7):482-490.].

Induction of Innate Inflammatory Pathways in the Corneal Epithelium in the Desiccating Stress Dry Eye Model.

To determine whether 24-hour exposure to the desiccating stress (DS) dry eye model induces NF-kB and NLRP3 inflammasome pathways in the mouse cornea epithelium. Six- to 8-week-old C57BL/6J mice were housed under normal humidity (nonstressed) or subjected to DS from a drafty, low-humidity environment combined with subcutaneous scopolamine four times/day for one day to suppress tear production (DS1). Cornea whole mounts were prepared for immunofluorescent staining, or the corneal epithelium was scraped for NF-kB p-p65 ELISA, Western blot, or real-time PCR to detect NF-kB and inflammasome pathway proteins and gene transcripts, respectively. NF-kB phospho-p65 protein, nuclear NF-kB p-p65, and expression of the NF-kB inducible cytokines (IL-12a, IL-12b, and lymphotoxin b [Ltb]) and chemokine (CCL-2) genes were significantly increased in DS1 compared to nonstressed control. NLRP3 protein and RNA transcripts significantly increased in DS1. NLRP3 and Caspase-1 immunostaining increased in the cornea epithelium at DS1. At DS1 there was no change in IL-18 and a decrease in IL-1β mRNA transcripts; however, levels of bound and total IL-18 protein increased at DS1, and the level of mature IL-1β increased from DS1 to DS5. These findings indicate innate NF-kB and NLRP3 inflammasome inflammatory pathways are induced in the corneal epithelium within one day in the DS dry eye model. NF-kB activation was associated with increased expression of inflammatory mediators involved in dry eye. Induction of these pathways is accompanied by increased bound/total IL-18 and mature IL-1β.

Preliminary Study on an Alternative Test Method with MCTT HCETM for Ocular Irritation Test of Ophthalmic Medical Devices

The sustained growth of the market for ophthalmic medical devices has increased the demand for alternatives to animal testing for the evaluation of eye irritation. The International Organization for Standardization has acknowledged the need to develop novel in vitro tests to replace animal testing. Here, we evaluated the applicability of an alternative method based on a human corneal model to test the safety of ophthalmic medical devices. 2-Hydroxyethyl methacrylate (HEMA) and Polymethyl methacrylate (PMMA), which are used to fabricate contact lenses, were used as base materials. These materials were blended with eye irritant and non-irritant chemicals specified in the OECD Test Guideline (TG) 492 and Globally Harmonized System (GHS) classification. Then, three GLP-certified laboratories performed three replicates using the developed method using 3D reconstructed human cornea epithelium, MCTT HCETM. OECD TG 492 describes the procedure used to evaluate the eye hazard potential of the test chemical based on its ability to induce cytotoxicity in a reconstructed human cornea-like epithelium (RhCE) tissue. Results: The within-laboratory reproducibility (WLR) and between-laboratory reproducibility (BLR) were both 100%. When a polar extraction solvent was used, the sensitivity, specificity, and accuracy were all 100% in each laboratory. When a non-polar extraction solvent was used, the sensitivity was 80%, the specificity was 100%, and the accuracy was 90%. The proposed method exhibited excellent reproducibility and predictive capacity within and between laboratories. Therefore, the proposed method using the MCTT HCETM model could be used to evaluate eye irritation caused by ophthalmic medical devices.

Novel characterization of CXCR4 expressing cells in uninfected and herpes simplex virus-1 infected corneas

PurposeTo characterize CXCR4-expressing cells in uninfected and herpes simplex virus-1 (HSV-1) infected corneas. MethodsThe corneas of C57BL/6J mice were infected with HSV-1 McKrae. The RT-qPCR assay detected CXCR4 and CXCL12 transcripts in uninfected and HSV-1-infected corneas. Immunofluorescence staining for CXCR4 and CXCL12 protein was performed in the frozen sections of herpes stromal keratitis (HSK) corneas. Flow cytometry assay characterized the CXCR4-expressing cells in uninfected and HSV-1-infected corneas. ResultsFlow cytometry data showed CXCR4 expressing cells in the separated epithelium and stroma of uninfected corneas. In the uninfected stroma, CD11b + F4/80+ macrophages are the predominant CXCR4-expressing cells. In contrast, most CXCR4 expressing cells in the uninfected epithelium were CD207 (langerin)+, CD11c+, and expressed MHC class II molecule, documenting the Langerhans cells (LCs) phenotype. After corneal HSV-1 infection, CXCR4 and CXCL12 mRNA levels increased significantly in HSK corneas than in uninfected corneas. Immunofluorescence staining showed CXCR4 and CXCL12 protein localization in the newly formed blood vessels in the HSK cornea. Furthermore, the infection resulted in LCs proliferation, causing an increase in their numbers in the epithelium at 4 days post-infection (p.i.). However, by 9-day p.i., the LCs numbers declined to the counts observed in naïve corneal epithelium. Our results also showed neutrophils and vascular endothelial cells as the prominent CXCR4-expressing cell types in the stroma of HSK corneas. ConclusionsTogether, our data demonstrate the expression of CXCR4 on resident antigen presenting cells in the uninfected cornea and on infiltrating neutrophils and newly formed blood vessels in the HSK cornea.

Histological Changes Indused by Monosodium Glutamate on the Growth and Development of the Offsprings Albino Mice Cornea

This research aim to study the effect of different doses of a Monosodium Glutamate (MSG) to cause histopathological changes in the offspring corneal for the ages 7, 15, 21, 30, and 60 days. The mothers, type Mus musculus mice has been orally administrated from 17th day of pregnancy by different concentrations (2, 4, and 8 g/Kg/ body weight) of MSG using stomach tube. Microscopic examination showed no changes in the cornea for a seventh day and administrated concentrations of 2 and 4 g / kg body weight, but observed degeneration of fibre stroma, necrosis and the reduction of the anterior epithelial layer of the cornea at dose of 8 g / kg. In the group of 15 days, these changes are mentioned; hyperplasia in the anterior epithelium of the peripheral cornea of 2 g, deformation in the peripheral part of the cornea and the reduction of thickness and intensify the stroma components at a concentration of 4 g either at a concentration of 8 g / kg has appeared hyperplasia in fibroblast cells in the corneal stroma and wave of the basement membrane of anterior epithelial tissue of central cornea. In a group 21days, these changes are shown; necrosis and rarefaction in stroma of central cornea while the reduction of the anterior epithelial layer of the peripheral cornea notes and wave anterior surface of the cornea in the concentration of 2 g / kg. Deformation appeared and reduction layers and irregular with dissociation in the stromal fibers in some areas in the concentration of 4 g / kg, while in the concentration of 8 g / kg show damage in the anterior part of epithelial of the central corneal tissue. In a group 30 days and concentration of 2 g / kg, these changes were observed; necrosis, swelling in some anterior epithelial cells, condensation of nuclei in endothelial cells, with Keratinized at the surface of the cornea and edema in the stroma. Bulge appeared in the stroma of peripheral cornea and hyperplasia in fiberoblast at a concentration of 4g/kg, meanwhile at a concentration of 8 g/kg show necrosis under the anterior epithelium of central cornea and dissociation of stromal fiberoblast. In the group 60 Days at the concentration of 2 g / kg show damage in the anterior epithelial tissue of the peripheral cornea and hyperplasia in some parts of the anterior epithelial tissue in the central cornea. At the concentration of 4g/kg show damage in the stromal fibers of peripheral corneal epithelium and detachment of endothelial tissue. In concentration 8 g/kg reduction appeared in the anterior epithelial tissue in the certain part of the cornea, hyperplasia in endothelial tissue, condensed layer appeared on the surface of anterior epithelial tissue.

A Rare Case of Bilateral Ocular Surface Squamous Neoplasia in an Immunocompetent Patient

Ocular surface squamous neoplasia is a broad entity that includes dysplastic lesions involving the squamous epithelium of conjunctiva and cornea. It is commonly seen in males between 50-75 years of age and has higher incidence in places close to the equator.

Impact of Exosomes Released by Different Corneal Cell Types on the Wound Healing Properties of Human Corneal Epithelial Cells.

Corneal wound healing involves communication between the different cell types that constitute the three cellular layers of the cornea (epithelium, stroma and endothelium), a process ensured in part by a category of extracellular vesicles called exosomes. In the present study, we isolated exosomes released by primary cultured human corneal epithelial cells (hCECs), corneal fibroblasts (hCFs) and corneal endothelial cells (hCEnCs) and determined whether they have wound healing characteristics of their own and to which point they modify the genetic and proteomic pattern of these cell types. Exosomes released by all three cell types significantly accelerated wound closure of scratch-wounded hCECs in vitro compared to controls (without exosomes). Profiling of activated kinases revealed that exosomes from human corneal cells caused the activation of signal transduction mediators that belong to the HSP27, STAT, β-catenin, GSK-3β and p38 pathways. Most of all, data from gene profiling analyses indicated that exosomes, irrespective of their cellular origin, alter a restricted subset of genes that are completely different between each targeted cell type (hCECs, hCFS, hCEnCs). Analysis of the genes specifically differentially regulated for a given cell-type in the microarray data using the Ingenuity Pathway Analysis (IPA) software revealed that the mean gene expression profile of hCECs cultured in the presence of exosomes would likely promote cell proliferation and migration whereas it would reduce differentiation when compared to control cells. Collectively, our findings represent a conceptual advance in understanding the mechanisms of corneal wound repair that may ultimately open new avenues for the development of novel therapeutic approaches to improve closure of corneal wounds.

Differences in vulnerability to desiccating stress between corneal and conjunctival epithelium in rabbit models of short-term ocular surface exposure

We evaluate the difference in vulnerability to desiccating stress (DS) between the corneal and conjunctival epithelia to understand different ocular surface staining patterns in dry eye patients. We generated a rabbit model of short-term exposure keratopathy. To induce DS in the ocular surface, rabbit right eyelids were opened for 30 min, with blinking once/minute. Corneal staining scores increased from 3-min post-DS exposure, while conjunctival staining increased from 20-min post-DS. At 20 min, the tear MUC5AC level doubled as compared to pre-DS (p = 0.007). In Western blot analysis, conjunctival AQP5, MUC5AC, and CFTR expression increased significantly in response to DS, compared to control (p = 0.039, 0.002, 0.039, respectively). Immunohistochemistry for CD31 and LYVE-1 were performed. CD31-positive cells and lymphatic space surrounded by LYVE-1-positive cells increased significantly in conjunctival tissue post-DS, compared to control (p = 0.0006, p < 0.0001, respectively). Surface damage was worse in the corneal than in the conjunctival epithelium after DS, by scanning electron microscopy. This study showed that the cornea and conjunctival epithelium show differences in vulnerability to DS. Increased blood vessels and dilated lymphatics, accompanied by increased conjunctival epithelial AQP5, MUC5AC, and CFTR expression, underlie the protective mechanism of the conjunctiva to desiccating stress.

Clinical and Functional Results of the Use of Suspension of Autologous Mononuclear Leucocytes in the Surgical Treatment of Bullous Keratopathy

The purpose: to explore the clinical effectiveness of a new cell-based surgical method for the treatment of bullous keratopathy that includes intrastromal injection of autologous mononuclear leukocytes into the posterior third of the cornea stroma.Material and methods. The study included 58 patients with diagnosis of bullous kerathopathy. They were divided into 2 groups — the main group received surgical treatment by the presented method; and the comparison group received a course of conservative therapy.Results. In 10 days the patients in the main group had a normal structure of the cornea epithelium, slight edema of the stroma, thin folds of the Descemet’s membrane and moderate edema of the endothelium. Similar results in the comparison group were achieved in 1 month, and it was the best condition of the cornea. Later corneal edema increased. In 3 months the patients of the main group had a normal structure of corneal epithelium, single thin folds of the Descemet’s membrane, slight edema of the posterior third of the stroma only in the optical zone, and slight edema of the endothelium. The achieved results were preserved for 12 months. The best correct visual acuity (BCVA) in the main group was achieved by the 6th month. It increased 10 times from the initial level, and the corneal thickness decreased by 27 % from the initial level by the 9th month. In the comparison group, the BCVA was achieved by the 1st month. It increased 2.7 times from the initial level; the maximum decrease in the index of pachymetry by 6.7 % was achieved by the 3rd month from the initial level, and then these parameters increased.Conclusion. The using of a surgical method for the treatment of bullous keratopathy, including intrastromal injection of suspension of autologous mononuclear leukocytes, contributes to the relief of corneal syndrome, increases the transparency of the cornea and restores its normal thickness. These changes are accompanied by increase of BCVA in the postoperative period compared to the course of conservative therapy.