Number Of Keratocytes Research Articles

The Effect of Using Absorbent Carbon on Air Exposed to Air Fresheners on The Histology of The Cornea of The White Rat (Rattus Novergicus)

Air freshener contain hazardous chemical such as formaldehyde. Absorbent carbon is used as an adsorbent to reduce air pollution. The goal of this research is to know the use of absorbent carbon to cornea histology on Rattus novergicus that was induced with air freshener. This research was an experimental study with a post-test only control group. The subjects were 28 one-month old male Rattus novergicus Wistar strains divide into 4 groups: Control (K), air freshener (P1), absorbent carbon (P2) and air freshener plus absorbent carbon (P3).The treatment was performed 8 hours/day for 35 days. Data of anterior epithelial thickness were analyzed by One Way Anova test. Data of the overall thickness of the corneal layer and keratocyte number were analyzed by Kruskal Wallis test. The overall thickness of the corneal layer and anterior epithelial thickness showed that there were not significant difference between subject groups (p>0,05). Anterior epithelial thickness showed that there were not significant difference between subject groups (p>0,05). Keratocyte number showed significant difference between subject groups (p<0,05), Mann Whitney test for Keratocyte number showed results P1>P3>K>P2 Absorbent carbon gives postitive effect to reduce cornea damage on Rattus novergicus that induced by air freshener.

Morphological Changes of Corneal Keratocytes following Surface Ablation Laser Surgery: An Observational Study from the United Kingdom

Introduction: Laser Subepithelial Keratomileusis (LASEK) is a type of surface ablation laser surgery that offers a solution to patients for correcting their ametropia. In rare instances, LASEK has been associated with complications such as the development of postoperative haze, infection, and poor visual outcomes. Imaging features detected within the patient’s cornea using In-Vivo Confocal Microscopy (IVCM) have aided in identifying the cellular basis of complications like these. Aim: To observe and describe the morphological changes seen on IVCM in patients following LASEK surgery. Materials and Methods: This observational study aimed to assess the quantity and morphology of anterior and posterior keratocytes in adults aged 18 years and older. Five participants (9 eyes) who presented for follow-up appointments following LASEK surgery at Manchester Royal Eye Hospital (MREH), United Kingdom, from July to December 2018 were recruited for the study. The cell counts of the anterior and posterior keratocytes were calculated using the Heidelberg Eye Explorer software. The images were analysed with the Image J program (National Institutes of Health, Bethesda, USA). Nerve fibers were assessed using the ACCMetrics program (University of Manchester, UK). The t-test was used to establish the statistical association between variables. Results: Morphological changes, such as zones of hyperreflectivity beneath the epithelium, were observed in all the recruited participants. Immune cells and sub-basal nerve abnormalities were detected in several participants. The number of keratocytes in the anterior stroma of all the participants was found to be much lower compared to that of the control. Only participant number 3 had a greater number of keratocytes in the anterior stromal layer (454 and 514 cells/mm2 ) compared to the other participants, who had a cell count ranging between 156 to 262 cells/mm2. Conclusion: It is evident that visible changes are noted both quantitatively and morphologically in both the anterior and posterior keratocytes postoperatively. More research is required with larger controlled studies to investigate the IVCM imaging biomarkers and morphological features that represent the wound healing process and the factors influencing visual outcomes, ensuring that postoperative complications can be minimised.

Role of aquaporins in corneal healing post chemical injury

Aquaporins (AQPs) are transmembrane water channel proteins that regulate the movement of water through the plasma membrane in various tissues including cornea. The cornea is avascular and has specialized microcirculatory mechanisms for homeostasis. AQPs regulate corneal hydration and transparency for normal vision. Currently, there are 13 known isoforms of AQPs that can be subclassified as orthodox AQPs, aquaglyceroporins (AQGPs), or supraquaporins (SAQPs)/unorthodox AQPs. AQPs are implicated in keratocyte function, inflammation, edema, angiogenesis, microvessel proliferation, and the wound-healing process in the cornea. AQPs play an important role in wound healing by facilitating the movement of corneal stromal keratocytes by squeezing through tight stromal matrix and narrow extracellular spaces to the wound site. Deficiency of AQPs can cause reduced concentration of hepatocyte growth factor (HGF) leading to reduced epithelial proliferation, reduced/impaired keratocyte migration, reduced number of keratocytes in the injury site, delayed and abnormal wound healing process. Dysregulated AQPs cause dysfunction in osmolar homeostasis as well as wound healing mechanisms. The cornea is a transparent avascular tissue that constitutes the anterior aspect of the outer covering of the eye and aids in two-thirds of visual light refraction. Being the outermost layer of the eye, the cornea is prone to injury. Of the 13 AQP isoforms, AQP1 is expressed in the stromal keratocytes and endothelial cells, and AQP3 and AQP5 are expressed in epithelial cells in the human cornea. AQPs can facilitate wound healing through aid in cellular migration, proliferation, migration, extracellular matrix (ECM) remodeling and autophagy mechanism. Corneal wound healing post-chemical injury requires an integrative and coordinated activity of the epithelium, stromal keratocytes, endothelium, ECM, and a battery of cytokines and growth factors to restore corneal transparency. If the chemical injury is mild, the cornea will heal with normal clarity, but severe injuries can lead to partial and/or permanent loss of corneal functions. Currently, the role of AQPs in corneal wound healing is poorly understood in the context of chemical injury. This review discusses the current literature and the role of AQPs in corneal homeostasis, wound repair, and potential therapeutic target for acute and chronic corneal injuries.

Effect of preservative-free and preserved prostaglandin analogues on the histology of cornea of adult male guinea pigs following repeated exposure.

Glaucoma is a group of eye diseases that can cause vision loss. Prostaglandin analogues (PGAs) are known to be first-line treatment for patients with glaucoma. Latanoprost is a good, efficient and well-tolerated PGA that is currently available as latanoprost with benzalkonium chloride (BAC) (Xalatan) and preservative-free (PF) prostaglandin analogue latanoprost (Monopost). Lately, using PF anti-glaucoma agents has been considered an essential procedure for enhancing glaucoma care. This study aimed to analyse the histological changes within the corneal tissue with the use of currently available preserved prostaglandins-derived eye drops and PF prostaglandin analogue. In this study, 40 male guinea pigs were divided into four equal groups. Control group, Latanoprost with 0.02% BAC-treated group, Recovery group and PF latanoprost-treated group. After 2 months, the corneal tissues of guinea pigs were prepared for light and electron microscopic studies; morphometric and statistical studies were performed. Our results indicated that guinea pigs treated with latanoprost with BAC exhibited ocular surface changes: there was epithelial thinning with desquamation, the stroma showed irregularly arranged collagen fibres and small keratocytes. Morphometrically, there was a marked decrease in the thickness of epithelium and number of keratocytes. Negative periodic acid Schiff (PAS) reaction was observed in some parts of the epithelial basement membrane. The epithelium gave a strong positive immunoreactivity for Bcl-2-associated X protein (BAX). Guinea pigs left to recover exhibited improvement, while treatment of animals with PF latanoprost resulted in nearly normal corneal structure. In conclusion, PF prostaglandin anti-glaucoma medication seems to be better and have protective effect on cornea of male guinea pigs than prostaglandins with BAC preservative.

Comparison of porcine corneal decellularization methods and importance of preserving corneal limbus through decellularization.

The aim of this study is to compare the three previously applied, conventional porcine corneal decellularization methods and to demonstrate the importance of preserving the corneal limbus through decellularization. Fresh, wild-type (with or without) limbus porcine corneas were decellularized using three different methods, including (i) sodium dodecyl sulfate (SDS), (ii) hypertonic saline (HS), and (iii) N2 gas (NG). Post-treatment evaluation was carried out using histological, residual nuclear material, and ultrastructural analyses. Glycerol was used to help reduce the adverse effects of decellularization. The corneas were preserved for two weeks in cornea storage medium. All three decellularization methods reduced the number of keratocytes at different rates in the stromal tissue. However, all methods, except SDS, resulted in the retention of large numbers of cells and cell fragments. The SDS method (0.1% SDS, 48h) resulted in almost 100% decellularization in corneas without limbus. Low decellularization capacity of the NG method (<50%) could make it unfavorable. Although HS method had a more balanced damage-decellularization ratio, its decellularization capacity was lower than SDS method. Preservation of the corneoscleral limbus could partially prevent structural damage and edema, but it would reduce the decellularization capacity. Our results suggest that SDS is a very powerful decellularization method, but it damages the cornea irreversibly. Preserving the corneoscleral limbus reduces the efficiency of decellularization, but also reduces the damage.

The Effect of Mitomycin C on the number of Keratocytes on the Posterior Wall of the Stroma of Cornea after Photorefractive Keratectomy

The Effect of Mitomycin C on the number of Keratocytes on the Posterior Wall of the Stroma of Cornea after Photorefractive Keratectomy

Histological Corneal Alterations in Keratoconus After Crosslinking-Expansion of Findings.

To investigate histopathologic, immunohistochemical, and electron microscopic findings in 8 keratoplasty specimens with a history of corneal collagen crosslinking (CXL) for keratoconus. Five new (hitherto unreported) and 3 previously published specimens were analyzed. Corneal buttons of 8 keratoconus corneas (5-114 months after CXL) were compared with 5 keratoconus specimens without CXL and 5 normal corneas for morphological alterations. Corneal buttons were evaluated by light microscopy and immunohistochemistry using antibodies against CD34, PGP 9.5, nestin, telomerase reverse transcriptase, and Ki67 as well as by transmission electron microscopy. Keratoconus corneas after CXL showed a significant keratocyte loss (except 1 specimen with an increased keratocyte number), whereas keratoconus corneas without CXL revealed a higher keratocyte density compared with healthy controls. Keratocyte loss could be clinically correlated with corneal opacification and corneal perforation. In corneas after CXL, the remaining keratocytes appeared more polymorphic and revealed a different expression of surface markers similar to keratocytes in corneal scars. The presence of proteoglycans, nerves, and endothelial cells was unaffected by CXL. CXL may cause permanent keratocyte loss or repopulation of altered keratocytes, resulting in clinical complications such as corneal opacification or perforation. Despite its good safety profile and high effectiveness in progressive keratoconus, CXL should be performed in accordance with current guidelines strictly adhering to protocol and safety standards.

In vivo confocal microstructural analysis of corneas presenting Kayser-Fleischer rings in patients with Wilson's disease.

To evaluate microstructural differences between corneas with and without Kayser-Fleischer rings in age-matched subjects with Wilson's disease with neurological symptoms, using confocal laser scanning microscopy. The study included 12 subjects with Wilson's disease with neurological symptoms. Twelve corneas presented clinically with classic Kayser-Fleischer rings, visible on slit lamp examination; the other 12 served as controls. The subjects underwent a comprehensive clinical examination. Microstructural analysis using confocal laser scanning microscopy evaluated increased corneal thickness, decreased number of cells, increased debris or specific deposits, and unusual microstructures. Clinically, the subjects with Kayser-Fleischer rings had similar corneal findings and normal intraocular pressure; two had typical sunflower cataracts and decreased visual acuity. The control eyes all presented normal visual acuity, intraocular pressure, and corneal appearance. The microstructural analysis demonstrated similar findings in all the affected corneas. Compared with the control corneas, there were fewer keratocytes in the anterior stroma (17.380 vs. 22.380/mm3). Round, "hollow" dark areas were observed between the keratocytes; these were universal and similar in appearance in all affected corneas and all cornea layers. In the peripheral posterior stroma, there were dust-like, bright, granular deposits that tended to increase in number and density toward Descemet's membrane, masking the peripheral endothelium. The control corneas presented a normal microstructure apart from dust-like granular deposits in the periphery. In vivo confocal microscopy is a useful tool for evaluating the corneal microstructure when a Kayser-Fleischer ring is clinically present. The ring consists of granular, bright particles that increase in density toward Descemet's membrane, and is associated with a decreased number of keratocytes and peculiar dark, round areas in all stromal layers, probably a sign of corneal damage. When the ring is not visible in subjects with Wilson's disease, changes to the corneal microstructure are insignificant.

Corneal confocal microscopy and dry eye findings in contact lens discomfort patients

To evaluate the corneal confocal microscopy and dry eye findings in patients with contact lens discomfort. The study included 3 groups of participants: Contact lens wearers using silicone hydrogel soft contact lenses who are symptomatic (CLD, n=15) or asymptomatic (ACL, n=11) and non-wearers as controls (n=14). Duration of contact lens wear, Ocular Surface Disease Index (OSDI) questionnaire responses, fluorescein tear break-uptime (FBUT), and corneal confocal microscopy findings were recorded. Mean age was 25.7±8.2 years and male/female ratio was 7/33. Demographic findings were similar regarding the groups. CLD patients had a longer lens use history than ACL (median 5 vs 2 years, p<0.001). OSDI scores were higher in CLD group than ACL or controls (p<0.001, p=0.002). FBUT was significantly lowest in CLD group, compared to controls and ACL (p<0.001, p=0.039). FBUT was also lower in ACL patients compared to controls (p=0.036). There was no difference between basal epithelium cell counts between all 3 groups. Anterior stromal activated keratocyte numbers were similar between contact lens using groups but was lower in controls (p=0.005). However, dendritiform cells in the sub-basal nerve layer were higher in CLD group compared to controls but similar to ACL (p<0.001, p=0.058). Graded sub-basal nerve tortuosity was more prominent in CLD group than the ACL (p=0.014). Patients with CLD had been wearing contact lenses for longer than those without symptoms. OSDI and FBUT scores were worse in CLD patients. In contact lens discomfort patients, there were increased dendritiform cells, indicating intensified inflammatory status of the cornea.

Retreatment strategies following Small Incision Lenticule Extraction (SMILE): In vivo tissue responses

With any refractive correction, including Small Incision Lenticule Extraction (SMILE), there may be a residual refractive error that requires a retreatment. Here, we investigated the tissue responses following various retreatment procedures in a rabbit model of SMILE. All rabbits underwent a -6.00D correction with SMILE. Two weeks later, they underwent -1.00D enhancement by: (i) VisuMax Circle, followed by excimer ablation (S+C); (ii) secondary SMILE anterior to the primary procedure (S+SE); or (iii) surface ablation (S+P), and were examined for 28 days. S+P induced corneal edema and haze, and more CD11b- (23±6 cells) and TUNEL-positive (36±4 cells) cells in the central stromal superficial layers early post-operatively (p<0.001 compared to other procedures). The corneas appeared normal on day 28 after S+P, but had a lower number of keratocytes near the laser ablated plane compared to other procedures. S+SE and S+C did not induce corneal haze and resulted similar level of fibronectin. However, S+C resulted in more inflammatory (10±2 cells; p = 0.001) and apoptotic cells (25±2 cells; p<0.001) compared to S+SE (7±1 inflammatory cells and 21±3 apoptotic cells) early post-operatively. In conclusion, each SMILE retreatment method resulted in unique tissue responses. S+SE offers advantages, such as minimal inflammation and cell death, as well as maintaining a ‘flap-less’ surgery, over other procedures. However, depending on the degree of enhancement, the lenticule may become too thin to be extracted and the procedure becomes more difficult to perform than S+C and S+P. S+P can maintain corneal integrity by avoiding flap creation and is technically more simple to perform than the others, but the surgery needs to be supplemented with mitomycin-C in order to reduce inflammation and modulate better wound healing.

Quantitative Analysis of Collagen Produced by Rabbit Keratocytes using Second Harmonic Generation Microscopy

ABSTRACTPurpose: To quantitatively analyze the collagen produced by rabbit keratocytes treated with mitomycin C (MMC) and/or transforming growth factor-beta (TGF-β1), and to investigate the collagen arrangement in situ, using second harmonic generation (SHG) microscopy.Methods: Keratocytes were divided into four groups: The keraotocytes in group rabbit keratocytes were treated with serum free medium containing 1% methylcellulose as controls. The cells in group TGF were received with TGF-β1 (at 2 ng/ml) and the same medium as the control. After exposure to MMC at 0.02 mg/ml for 5 minutes, the cells in group MMC were received with the same treatment as controls. In group TGF-MMC, the cells were exposed to MMC at 0.02 mg/ml for 5 minutes first, and then treated as group TGF. The signals of collagen were detected by SHG at 2 and 4 weeks post-treatment. The collagen content, as well as the keratocyte numbers, was calculated by MetaMorph Offline.Results: The collagen arrangement could be detected clearly by SHG. The cell number of group TGF was higher than other groups at 2 and 4 weeks post-treatment. The collagen content in group MMC was the least among all the groups at 2 weeks, while the collagen content in group TGF was higher than in group rabbit keratocytes and MMC at 4 weeks. No significant SHG side effects were found in cell proliferation and collagen deposition.Conclusions: SHG can provide a highly sensitive method to quantitatively measure collagen content, as well as to analyze its arrangement in situ.

Corneal Structural Changes in Nonneoplastic and Neoplastic Monoclonal Gammopathies.

To investigate corneal confocal microscopic changes in nonneoplastic and neoplastic monoclonal gammopathies. Three groups of subjects were considered: group 1, twenty normal subjects; group 2, fifteen patients with monoclonal gammopathy of undetermined significance (MGUS); group 3, eight patients with smoldering multiple myeloma and eight patients with untreated multiple myeloma. After hematologic diagnosis, patients underwent ophthalmologic exam and in vivo confocal microscopic study. The statistical analysis was performed using ANOVA and Student-Newman-Keuls tests and receiver operating characteristic (ROC) curve analysis. Epithelial cells of gammopathic patients showed significantly higher reflectivity than controls, demonstrated by optical density (P < 0.001). Subbasal nerve density, branching, and beading were significantly altered in gammopathic patients (P = 0.01, P = 0.02, P = 0.02, respectively). The number of keratocytes was significantly reduced in neoplastic patients (P < 0.001 versus both normal and MGUS) in the anterior, medium, and posterior stroma. The ROC curve analysis showed good sensitivity and specificity for this parameter. Group 2 and 3 keratocytes showed higher nuclear and cytoplasmatic reflectivity in the medium and posterior stroma. Endothelial cells were not affected. Patients with neoplastic gammopathies showed peculiar alterations of the keratocyte number, which appeared significantly reduced. A follow-up with corneal confocal microscopy of patients with MGUS is suggested as a useful tool to identify peripheral tissue alterations linked to possible neoplastic disease development.

Evaluation of short-term ex vivo culture with air/liquid system of porcine cornea

Regarding therapies for treatment of corneal wounds, ex vivo corneal culture is the most effective for minimizing expensive animal studies. Eighteen porcine enucleated eyes were soaked in 0.2% povidone iodine solution for disinfection prior to cornea excision. Subsequently, corneas were excised from whole eyes and filled with an agar/medium mixture. Corneas were transferred into culture dishes, after which culture medium was added until the limbus was covered. Cultures were then placed onto a plate rocker to mimic blinking action, followed by incubation at 37°C and 5% CO2. Corneas were harvested on Days 0, 3, and 7 after incubation, and optical coherence tomography (OCT) was performed on Day 7. Two eyes from each group were fixed in 2% glutaraldehyde/4% paraformaldehyde for low-vacuum scanning electron microscopy (LV-SEM), and four eyes from each group were fixed in 10% neutral-buffered formalin for histological analysis. OCT results showed that central corneal thickness significantly increased by Day 7 compared to Day 0 (p&lt;0.05). Using LV-SEM, gaps between endothelial cells were detected on Day 7 of ex vivo culture. In the histological evaluation, four to five stratified squamous cell layers, wing cells, and basal cells in the epithelium as well as flat-shaped keratocytes in the stroma were found on Day 0. By Day 7, stratified squamous cells and basal cells had decreased in number, and slightly round-shaped keratocytes were observed; however, the number of keratocytes was similar to that on Day 0. In this short-term ex vivo culture, epithelium and endothelium were sensitive to culture, whereas stroma and keratocytes were well maintained. An additional deswelling method will be needed to obtain more successful results in porcine corneal ex vivo culture.

Platelet recruitment promotes keratocyte repopulation following corneal epithelial abrasion in the mouse.

Corneal abrasion not only damages the epithelium but also induces stromal keratocyte death at the site of injury. While a coordinated cascade of inflammatory cell recruitment facilitates epithelial restoration, it is unclear if this cascade is necessary for keratocyte recovery. Since platelet and neutrophil (PMN) recruitment after corneal abrasion is beneficial to epithelial wound healing, we wanted to determine if these cells play a role in regulating keratocyte repopulation after epithelial abrasion. A 2 mm diameter central epithelial region was removed from the corneas of C57BL/6 wildtype (WT), P-selectin deficient (P-sel-/-), and CD18 hypomorphic (CD18hypo) mice using the Algerbrush II. Corneas were studied at 6h intervals out to 48h post-injury to evaluate platelet and PMN cell numbers; additional corneas were studied at 1, 4, 14, and 28 days post injury to evaluate keratocyte numbers. In WT mice, epithelial abrasion induced a loss of anterior central keratocytes and keratocyte recovery was rapid and incomplete, reaching ~70% of uninjured baseline values by 4 days post-injury but no further improvement at 28 days post-injury. Consistent with a beneficial role for platelets and PMNs in wound healing, keratocyte recovery was significantly depressed at 4 days post-injury (~30% of uninjured baseline) in P-sel-/- mice, which are known to have impaired platelet and PMN recruitment after corneal abrasion. Passive transfer of platelets from WT, but not P-sel-/-, into P-sel-/- mice prior to injury restored anterior central keratocyte numbers at 4 days post-injury to P-sel-/- uninjured baseline levels. While PMN infiltration in injured CD18hypo mice was similar to injured WT mice, platelet recruitment was markedly decreased and anterior central keratocyte recovery was significantly reduced (~50% of baseline) at 4–28 days post-injury. Collectively, the data suggest platelets and platelet P-selectin are critical for efficient keratocyte recovery after corneal epithelial abrasion.

Assessing microstructures of the cornea with Gabor-domain optical coherence microscopy: pathway for corneal physiology and diseases.

Gabor-domain optical coherence microscopy (GD-OCM) was applied ex vivo in the investigation of corneal cells and their surrounding microstructures with particular attention to the corneal endothelium. Experiments using fresh pig eyeballs, excised human corneal buttons from patients with Fuchs' endothelial dystrophy (FED), and healthy donor corneas were conducted. Results show in a large field of view (1 mm×1 mm) high definition images of the different cell types and their surrounding microstructures through the full corneal thickness at both the central and peripheral locations of porcine corneas. Particularly, an image of the endothelial cells lining the bottom of the cornea is highlighted. As compared to healthy human corneas, the corneas of individuals with FED show characteristic microstructural alterations of the Descemet's membrane and increased size and number of keratocytes. The GD-OCM-based imaging system developed may constitute a novel tool for corneal imaging and disease diagnosis. Also, importantly, it may provide insights into the mechanism of corneal physiology and pathology, particularly in diseases of the corneal endothelium.

Sjogren`s syndrome and ocular surface changes by laser-scanning in vivo confocal microscopy

Introduction : Dry eye disease (DED) and ocular allergy are common conditions in the eye practice. They have more than 20% of prevalence and that fact increases healthcare problems and affects life quality. Rheumatologic diseases such as Sjogren’s syndrome (SS) are often associated with ophthalmic findings, and belong to the dry eye but with decreased tear film production. Material and Methods : The laser-scanning confocal microscopy (LSCM) was performed to 16 eyes of 8 patients with subjective complains and objective clinical signs of DED and SS. The results of examinations were compared with control group of 28 eyes of 14 healthy patients. The qualitative analysis was performed to calculate the cell density in all cornea layers. The presence and distribution of Langerhans cells (LCs) in cornea was estimated. The quantitative survey understood monitoring of inflammation signs and corneal changes such as a presence of leukocytes. Changes in subbasal nerve plexus (SNP) and Meibomian glands (MGs) were evaluated over time. Results : The quantitative analysis presented calculations of cell density of all cornea layers. The most important finding was the reduction of epithelium cell density. In healthy patients it was 5048.6±56 cells/mm2 compared to 4603±70 cells/mm2 in DED-SS patients, but also the keratocyte number was significantly lower. The density of corneal LCs was compared to the healthy patients: 71±8 cells/mm2 and 19±3cells/mm2 respectively. The quantitative observation showed visible changes in SNP especially in tortuosity and destiny of sub-basal nerves. Conclusion : The ability of LSCM technology to examine different components of the ocular surface has opened new doors for studying the physiology and pathology. LSCM allows understanding the pathological changes of DED and SS. Our study proved that these diseases affect all structures of the anterior ocular surface.

Let this be lattice? Dendritiform erosion in lattice dystrophy type I, a source of confusion.

Lattice dystrophy type I is an autosomal dominant corneal dystrophy, which appears in the first decade of life [1, 2, 4]. Corneal involvement in lattice dystrophy type I is usually bilateral and symmetric. In the early stages, the lesions can appear as irregular, thin branching refractile lines and/or whitish, ovoid dots in the anterior axial stroma with clear intervening stroma. The lines start centrally and increase in size and number with time. They extend to the periphery and involve both the deeper and the superficial stromal tissue. The typical lattice filaments are radially oriented and branch dichotomously near their central terminations. As they overlap one another at various stromal levels, they create the typical lattice pattern. Indirect or retroillumination shows the lattice lines as refractile filaments with a double contour and a clear core. Refractile, glassy, homogeneous, discrete dots or more irregular opacities are seen with the filaments. With time, the opacities coalesce, and a diffuse groundglass haze may develop in the anterior and mid-stroma [1, 2, 4]. The predominant anterior involvement of this dystrophy commonly leads to recurrent erosions and irregularity of the epithelial surface, with accompanying visual disturbances [1, 2, 4]. Lattice dystrophy type I is a primary, localized corneal amyloidosis. Histologically, the epithelium is of variable thickness, with degeneration of the basal epithelium. The basement membrane is thickened and continuous but without normal hemidesmosomes. A variably thick, eosinophilic layer separating the epithelial basement membrane from Bowman’s layer is present and is composed of amyloid and collagen. Bowman’s membrane may be thicker or thinner than normal and is frequently fragmented in its periphery. The stroma contains large irregular deposits that distort the normal configuration of the corneal lamellae. There is a decrease in the number of keratocytes. Descement’s membrane and the endothelium are usually normal [1, 2, 4]. V. Vanlerberghe (&) S. De Craene Ph. Kestelyn Ghent University Hospital, Ghent, Belgium e-mail: vanlerbergheveerle@gmail.com

The Effect of Quercetin in Corneal Opacity Induced by Mitomycin-C

This study examined the effects of quercetin on corneal opacity caused by corneal edema by suppressing the damage on corneal endothelial cell, which was induced by mitomycin-C (MMC). In the MMC-treated group, the number of keratocytes was noticeably fewer compared to that of other groups. Although this group showed normal amount of fiber in the corneal stroma, the thickness was shown to be very thick and the alignment of the corneal endothelial cells that worked as the barrier against aqueous humor was irregular. According to such results, it was known that corneal opacity induced by MMC is not caused by proliferation of keratocytes, but by corneal edema triggered by the infiltration of aqueous humor. In the MMC+quercetin and quercetin+MMC-treated groups, the number of keratocytes was higher and polymorphonuclear leukocytes infilteration was lower significantly compared to that of the MMC-treated group. Although the amounts of fiber and endothelioid cell arrangement were normal, there was more space observed in the corneal stroma. Nonetheless, these groups showed significantly lower stromal thickness compared to that of the MMC group. In conclusion, quercetin has the effect on the reduction of corneal opacity caused by corneal edema that work MMC-induced damage to the corneal endothelial cells.

A comparison of three methods of decellularization of pig corneas to reduce immunogenicity.

To investigate whether decellularization using different techniques can reduce immunogenicity of the cornea, and to explore the decellularized cornea as a scaffold for cultured corneal endothelial cells (CECs). Transplantation of decellularized porcine corneas increases graft transparency and survival for longer periods compared with fresh grafts. Six-month-old wild-type pig corneas were cut into 100-200 µm thickness, and then decellularized by three different methods: 1) 0.1% sodium dodecyl sulfate (SDS); 2) hypoxic nitrogen (N2); and 3) hypertonic NaCl. Thickness and transparency were assessed visually. Fresh and decellularized corneas were stained with hematoxylin/eosin (H&E), and for the presence of galactose-α1,3-galactose (Gal) and N-glycolylneuraminic acid (NeuGc, a nonGal antigen). Also, a human IgM/IgG binding assay was performed. Cultured porcine CECs were seeded on the surface of the decellularized cornea and examined after H&E staining. All three methods of decellularization reduced the number of keratocytes in the stromal tissue by >80% while the collagen structure remained preserved. No remaining nuclei stained positive for Gal or NeuGc, and expression of these oligosaccharides on collagen was also greatly decreased compared to expression on fresh corneas. Human IgM/IgG binding to decellularized corneal tissue was considerably reduced compared to fresh corneal tissue. The cultured CECs formed a confluent monolayer on the surface of decellularized tissue. Though incomplete, the significant reduction in the cellular component of the decellularized cornea should be associated with a significantly reduced in vivo immune response compared to fresh corneas.

Can the Effect of Transepithelial Corneal Collagen Cross-linking Be Improved by Increasing the Duration of Topical Riboflavin Application? An In Vivo Confocal Microscopy Study

To evaluate the effect of transepithelial corneal collagen cross-linking (CXL) with prolonged riboflavin application by in vivo confocal microscopy and to compare this effect with that of standard CXL with complete epithelial debridement. In eyes with progressive keratoconus, CXL procedure was performed with standard technique and transepithelial technique after prolonged riboflavin drop application for 2 hr. Patients were evaluated with in vivo confocal microscopic examination preoperatively and at postoperative months 1 and 6. The depth of CXL effect was similar in both groups (i.e., 380.86 ± 103.23 μm in standard CXL group and 342.2 ± 68.6 μm in transepithelial CXL group) (P=0.4). The endothelial cell counts and morphological parameters (i.e., pleomorphism and polymegathism) were not significantly affected in both groups (P>0.05 for all). In the standard CXL group, in vivo confocal microscopy revealed anterior stromal acellular hyperreflective honeycomb edema with posteriorly gradually decreasing reflectivity and increasing number of keratocytes and some sheets of longitudinally aligned filamentary deposits. The keratocytes were seen to repopulate in the posterior-to-anterior direction. In transepithelial CXL group, although the depth of CXL effect was similar, less pronounced keratocyte damage, extracellular matrix hyperreflectivity, and sheets of filamentary deposits at the posterior stroma was observed. Transepithelial CXL with prolonged peroperative riboflavin application can achieve similar depth of effect in the stroma with less pronounced confocal microscopic changes as compared with the standard CXL with complete epithelial debridement.