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Mooren's ulcer (MU) is a chronic and painful ulcerative keratitis that is difficult to diagnose, especially when concealed beneath the pterygium, which is a common, benign, wedge-shaped, fleshy tissue growth of the conjunctiva extending onto the cornea. The coexistence of MU and pterygium is extremely rare. A 41-year-old man presented with a 2-month history of unprovoked redness, pain, and blurred vision in the right eye. Corneal epithelial defects around the pterygium head were noted upon slit-lamp examination and fluorescein staining. The patient was initially misdiagnosed with a corneal epithelial defect and pterygium. The initial treatments with anti-inflammatory and corneal epithelial growth promotion tear agents failed. Anterior segment optical coherence tomography (AS-OCT) showed corneal stromal lysis thinning, and in vivo confocal microscopy (IVCM) revealed marked inflammatory cell infiltration and stromal degeneration. We suspected the pathology was an immune-related or tumor-related corneal ulcer. The MU concealed beneath the pterygium was diagnosed by histopathological examination of a biopsy specimen that presented typical localized loss of the corneal epithelium and Bowman's layer, stromal degeneration, and inflammatory cell infiltration. Finally, we performed lamellar keratoplasty (LKP) combined with pterygium excision surgery. The patient recovered with no complications or recurrence during the 1-year follow-up period. Few cases of MU concealed beneath the pterygium have been reported. It is beneficial to rule out the pathological changes concealed beneath the pterygium, combined with multiple means of examination such as slit-lamp examination, AS-OCT, and IVCM. A histopathological examination should be performed to establish a diagnosis.

Objectives To evaluate the safety and efficacy of repeated corneal collagen crosslinking assisted by transepithelial double-cycle iontophoresis (DI-CXL) in the management of keratoconus progression after primary CXL. Methods A retrospective analysis was conducted in the patients who underwent repeated CXL between 2016 and 2018. These patients were treated with DI-CXL if keratoconus progression was confirmed after primary CXL. Scoring of ocular pain and corneal epithelial damage, visual acuity, corneal tomography, in vivo corneal confocal microscopy (IVCM) was performed before and at 3, 6, 12, and 24 months after DI-CXL. Results Overall, 21 eyes of 12 patients (mean age 17.3 ± 1.9 years) were included in this study. Before DI-CXL, an average increase of 4.26 D in Kmax was detected in these patients with a mean follow-up interval of (23.0 ± 13.7) months. After DI-CXL, corneal epithelial damage rapidly recovered within days. Visual acuity remained unchanged with follow-up of 24 months. When compared to baseline, significant decreases were observed in Kmax (at 3 months) and K2 (at 3 and 6 months) after DI-CXL. Corneal thickness of thinnest point significantly decreased at 3 months postoperatively. When compared to baseline, no significant differences were found in any of the refractive or tomographic parameters at 12 and 24 months. IVCM revealed trabecular patterned hyperdense tissues after DI-CXL in the anterior stroma at the depth of 200 μm or more. No corneal infiltration or persistent epithelial defect was recorded after DI-CXL. Conclusion DI-CXL is safe and effective as a good alternative in stabilizing keratoconus progression after primary CXL.

Background/aimsTo improve the accuracy of pterygium screening and detection through smartphones, we established a fusion training model by blending a large number of slit-lamp image data with a small proportion of smartphone data.MethodTwo datasets were used, a slit-lamp image dataset containing 20 987 images and a smartphone-based image dataset containing 1094 images. The RFRC (Faster RCNN based on ResNet101) model for the detection model. The SRU-Net (U-Net based on SE-ResNeXt50) for the segmentation models. The open-cv algorithm measured the width, length and area of pterygium in the cornea.ResultsThe detection model (trained by slit-lamp images) obtained the mean accuracy of 95.24%. The fusion segmentation model (trained by smartphone and slit-lamp images) achieved a microaverage F1score of 0.8981, sensitivity of 0.8709, specificity of 0.9668 and area under the curve (AUC) of 0.9295. Compared with the same group of patients’ smartphone and slit-lamp images, the fusion model performance in smartphone-based images (F1score of 0.9313, sensitivity of 0.9360, specificity of 0.9613, AUC of 0.9426, accuracy of 92.38%) is close to the model (trained by slit-lamp images) in slit-lamp images (F1score of 0.9448, sensitivity of 0.9165, specificity of 0.9689, AUC of 0.9569 and accuracy of 94.29%).ConclusionOur fusion model method got high pterygium detection and grading accuracy in insufficient smartphone data, and its performance is comparable to experienced ophthalmologists and works well in different smartphone brands.

Purpose. To investigate the changes of corneal endothelium under different crosslinking conditions and the protective effect of ripasudil. Methods. Corneal crosslinking groups were infiltrated with riboflavin and subsequently irradiated with 0.54 J/cm2 or 1.08 J/cm2 UVA, while noncrosslinking groups included neither UVA nor riboflavin treatment, only 1.08 J/cm2 UVA and only riboflavin treatment. Corneal opacity, variations in corneal endothelial cells, and corneal thickness of all groups were observed by slit lamp, in vivo confocal microscopy, and optical coherence tomography. Immunofluorescence staining and scanning electron microscopy were performed to evaluate changes in the structure and function of the corneal endothelium. The mice that received a corneal crosslinking dose of 1.08 J/cm2 were instilled with ripasudil to explore its protective effect on the corneal endothelium. Results. Treatment with UVA and riboflavin caused an increase in corneal opacity and corneal thickness and decreased endothelial cell density. Furthermore, treatment with UVA and riboflavin caused endothelial cell DNA damage and destroyed the tight junction and pump function of the endothelium, while riboflavin or the same dose of UVA alone did not affect the endothelium. Ripasudil reduced DNA damage in endothelial cells, increased the density of cells, and protected the endothelium’s integrity and function. Conclusion. Riboflavin combined with UVA can damage the corneal endothelium’s normal functioning. The corneal endothelium’s wound healing is dose-dependent, and the ROCK inhibitor ripasudil maintains the endothelium’s pump and barrier functions.

On-road information, including road boundaries, road markings, and road cracks, provides significant guidance or warning to all road users. Recently, the on-road information extraction from LiDAR data have been widely studied. However, for the LiDAR data with lower accuracy and higher noise, some detailed information, such as road boundary, is difficult to be extracted correctly. Furthermore, most of previous studies lack an exploration of efficiently extracting multiple on-road information from a single framework. In this paper, we propose a new framework that can simultaneously extract multiple on-road information from high accuracy LiDAR data and can also more robustly extract detailed road boundaries from low accuracy LiDAR data. First, we propose a Curb-Aware Ground Filter to extract ground points with rich curb structure features. Second, we transform the vertical density, elevation gradient and intensity features of the ground points into multiple feature maps and extract multiple on-road information from the feature maps by employing a semantic segmentation network. Experimental results on three datasets with different data accuracy demonstrate that our method outperforms other recent competitive methods.

Purpose The aim of this study is to compare the long-term effects of transepithelial corneal crosslinking with two continuous cycles of iontophoresis (EI-CXL) and conventional corneal crosslinking (C-CXL) in adults with progressive keratoconus. Methods A retrospective analysis was conducted in adults who underwent C-CXL or EI-CXL between 2013 and 2015. Visual acuity, corneal tomography, anterior segment optical coherence tomography, in vivo corneal confocal microscopy (IVCM), and endothelial cell count (ECC) were performed preoperatively and 5 years postoperatively. Results Sixty-eight patients with a mean age of (24.3 ± 3.8) years were included, 34 for each group. After CXL, UCVA or BCVA remained stable, while the spherical diopter, cylinder diopter, spherical equivalent, and Kmax significantly decreased at 1, 2, and 3 years in both groups than baseline (P Kmax was slightly higher in EI-CXL group (58.16 ± 6.28) than that of C-CXL group (57.46 ± 4.98). At 3 and 5 years, the minimal corneal thickness in C-CXL group was still significantly lower than baseline (P P Conclusion EI-CXL showed approximate efficacy with C-CXL in stabilizing progressive keratoconus in adults. EI-CXL has the potential to be a preferable transepithelial protocol.

This retrospective study is aimed at determining the correlation between cytokine levels and virus status in the aqueous humor of 38 patients with Fuchs heterochromic iridocyclitis (FHI) with/without a viral presence between May 2017 and January 2020. The levels of cytokines were analyzed in the groups with and without virus-related FHI. Among the patients, 50% had rubella virus, 5.26% had cytomegalovirus, and 2.63% had herpes simplex virus infections. The expression of interleukin-6 (IL-6) and IL-8 was significantly higher, and that of basic fibroblast growth factor (bFGF) was significantly lower in the virus-positive group than in the virus-negative group ( P = 0.015 , P = 0.001 , and P = 0.001 , respectively). Although there was no significant difference in the mean expression of vascular cell adhesion protein 1 (VCAM-1), IL-10, and vascular endothelial growth factor (VEGF), that of VCAM-1 and IL-10 was higher ( M = 1338 and M = 1390 , respectively; M = 6.225 and 10.600, respectively) and that of VEGF was lower ( M = 134.5 and M = 38.70 , respectively) in the virus-positive group than in the virus-negative group. Similar findings were observed for the expressions of IL-6, IL-8, and bFGF in the rubella-positive and rubella-negative groups. Viral presence was highly related to FHI, especially that of the rubella virus. High levels of inflammatory cytokines and low levels of neovascularization-related factors are involved in rubella-related FHI. These study findings could be helpful in the diagnosis and treatment of FHI.

BackgroundAllergic conjunctivitis (AC) is one of the reported potential risk factors of progression in keratoconus patients after corneal cross-linking surgery; however, the causal relationship is still inconclusive. Recent studies have indicated that various inflammatory cytokines play a vital role in the development of primary keratoconus. It is still unclear whether these inflammatory mediators also trigger CXL failures. This study aimed to investigate the impact of AC on the rabbit corneas after trans-epithelial corneal cross-linking (TCXL).MethodsA total of six rabbits were kept untreated as the normal control (NC) group. A total of 18 rabbits were treated by TCXL and divided into three groups (six in each group), namely, no treatment (TCXL group); induction of AC (TCXL + AC group); and induction of AC plus topical prednisolone acetate (TCXL + AC + PA group), according to additional treatment. AC was induced by topical application of ovalbumin after intraperitoneal pre-sensitization with ovalbumin. Rabbits were evaluated by slit lamp, in vivo laser scanning confocal microscopy, anterior segment optical coherence tomography, and measurement of corneal biomechanics. The cornea specimens were collected for the transmission electron microscope, the collagenase I digestion test, and PCR assay for TNF-α, IL-6, IL-1β, matrix metalloproteinase 9 (MMP-9), lysyl oxidase (LOX), and tissue inhibitor of metalloproteinases 1 (TIMP-1) on the day (D) 28.ResultsOn D28, the TNF-α, IL-6, IL-1β, MMP-9, and LOX levels were significantly increased while the TIMP-1 was decreased in the TCXL + AC group when compared with the TCXL and TCXL + AC + PA groups. In vivo confocal microscopy revealed that at a depth of 150–210 μm, a trabecular patterned hyperdense structure surrounded by elongated needle-like processes could be observed in the TCXL and TCXL + AC + PA groups, but hardly seen in the TCXL + AC group. The demarcation lines were indistinct and blurred in the TCXL + AC group. An electron microscope demonstrated less interlacing fibril lamellae and higher interfibrillar spacing in the TCXL + AC group. The stability of corneal biomechanics and resistance to collagenase were decreased in the TCXL + AC group.ConclusionThe corneal microstructures induced by TCXL and biomechanical stability were diminished in rabbits with AC but could be maintained by topical anti-inflammatory treatment. Our results supported the causal relationship between altered cytokine profiles and corneal microstructure after primary corneal cross-linking.

The change in hyaluronidase (HAase) is related to specific changes in the structure of vitreous, and it is necessary to develop simple but sensitive methods for HAase detection. In this work, a thin film fabricated from a hyaluronic acid (HA)-polyethyleneimine (PEI) hydrogel has been covered on a mixed cellulose microporous membrane (MCEM) to form a HA-PEI-MCEM firstly and it was then applied in a filtration system. The permeability of the filter membrane greatly affects the amount of water passing through within a certain time and the water can be collected and quantitatively measured with a simple electronic balance easily. The low permeability of the HA-PEI-MCEM allows a small amount of water to be drained. But after the addition of HAase, which can hydrolyze HA in the hydrogel, the permeability of the membrane increased. Therefore, the amount of water passing through the HA-PEI-MCEM composite membrane increased accordingly. The composite of the membrane, and the reaction conditions after the addition of HAase were optimized. Under the optimized conditions, the amount of water collected within 5 min showed a linear relationship with the HAase concentration in a range of 1.0-36 U mL-1 with a limit of detection of 0.35 U mL-1.The proposed method has been applied to detect HAase in vitreous samples with satisfactory results.

Background: Infectious keratitis after refractive surgery is uncommon, and mycobacterium abscess has never been observed in corneal infections in China. Purpose: To propose an unusual presentation of two cases about Mycobacterium abscess infection after laser in situ keratomileusis (LASIK). Participants: Both cases 1 and 2 were females who exhibited a corneal ulceration after LASIK in the same hospital. Methods: The first patient in the study was a 28-year-old woman who underwent Epi-LASIK surgery in both eyes. She developed an infectious corneal ulcer in one eye after 50 days post-surgery. She was referred to our out-patient clinic 3 months post-surgery with corneal perforation, and was treated with a therapeutic penetrating keratoplasty. The second patient was a 27-year-old woman who developed infectious keratitis in one eye after 2 months after bilateral LASIK and was referred to us 3 months later. Results: Cultures in both the cases were identified as a Mycobacterium abscess, and case 1 was treated with penetrating keratoplasty eventually, while case 2 recovered after sensitive treatment. Conclusions: Infectious keratitis after refractive surgery is uncommon, and Mycobacterium abscess has never been observed in corneal infections. An accurate diagnosis and identification of the causative agent is very important.

Purpose. The present study investigated the relationship between dry eye and the disease activity in patients with rheumatoid arthritis (RA). Methods. Patients with RA were divided by the Ocular Surface Disease Index (OSDI) into the symptomatic group (score ≥ 12) and the asymptomatic group (score 2.6) and the stable group (score ≤ 2.6). In the control group, 20 healthy adults with matched sex and age were selected. RA patients and healthy adults were inspected for the tear film break time (TBUT), tear meniscus height (TMH), corneal fluorescein staining (CFS), meibomian scan (MS), meibomian gland secretion score (MSS), and eyelid margin assessment (EMS). Results. The TBUT of the RA group was significantly less than that of the control group, while the CFS, MS, EMS, and MSS were higher. The TBUT of the symptomatic RA group was significantly less than that of the asymptomatic group, and the CFS was higher. In the active RA group, only the CFS was higher than that of the stable group, and there was no significant difference between the two groups for other parameters. Furthermore, there was no significant correlation between the course of RA and the dry eye (P>0.05). Conclusion. The rheumatoid activity does not necessarily lead to an aggravation of dry eye. Regardless of the duration, RA was not found to exhibit relation with the severity of dry eye. Translational Relevance. RA patients with disease active period cannot be ignored for the existence of dry eye, since patients with dry eye often lack the signs and symptoms.

To provide the general information on corneal transplantation (CT) in China, China Cornea Society designed a questionnaire on CT from 2014 to 2018 and entrusted it to 31 committee members for implementation of the survey nationwide. This article presents the results of the survey and compares the indicators used in the survey and those in the annual statistical report released by the Eye Bank Association of America (EBAA). The number of corneal transplantations completed by the 64 hospitals from 2014 to 2018 was respectively 5377, 6394, 7595, 8270 and 8980, totally 36,616 (22,959 male and 13,657 female). The five largest hospitals by the number of corneal transplantations completed 15,994 surgeries in total, accounting for 43.68% of all the surgeries performed in the 64 hospitals. The most common indication for corneal transplantations was corneal leukoma (7683, 20.98%), followed by bacterial keratitis (4209, 11.49%), corneal dystrophies (4189, 11.44%), keratoconus (3578, 9.77%) and corneal perforation (2839, 7.75%). The main surgical techniques were penetrating keratoplasty (PK) (19,896, 54.34%), anterior lamellar keratoplasty (ALK) (13,869, 37.88%). The proportion of PK decreased from 57.97% in 2014 to 52.88% in 2018 while the proportion of ALK increased from 36.04% in 2014 to 37.92% in 2018. The geographical distribution of keratoplasties performed in China is unbalanced. PK and ALK were the main techniques of CT and corneal leukoma, bacterial keratitis and corneal dystrophies were the main indications for CT in China.

Purpose To investigate the effects and mechanisms of the peroxisome proliferator-activated receptor alpha (PPAR-α) agonist fenofibrate on the formation of ocular surface squamous metaplasia induced by topical benzalkonium chloride (BAC) in a mouse model. Methods Ocular surface squamous metaplasia was induced in 16 days by topical BAC application in mice. During the period of induction, mice were divided into four groups: no additional treatment (BAC+UT), topical vehicle (BAC+Vehicle), topical fenofibrate (BAC+Feno), or topical fenofibrate plus intraperitoneal injection of MK886 (BAC+Feno+MK886). The parameters of tear film were evaluated on day 16, and eye specimens were collected. Histologic investigation; PAS assays; immunostaining for cytokeratin 10 (K10), Ki67, and F4/80; and PCR assays for TNF-α and IL-6 were performed. Cell Counting Kit 8 (CCK-8) assays were performed to evaluate the inhibitory effects of fenofibrate on RAW264.7 cells. Results Fenofibrate suppressed the formation of BAC-induced instable tear film. In the BAC+Feno group, the expression of K10 and Ki67 was lower than in the other three groups. The number of goblet cells was reduced in eyes of the BAC+UT and BAC+Vehicle groups but was maintained in eyes of the BAC+Feno group. The number of F4/80-positive cells and the levels of TNF-α and IL-6 mRNA were significantly reduced in the cornea of the BAC+Feno group. These effects of fenofibrate could be attenuated by MK886. The cell viability of RAW264.7 cells could be significantly inhibited by fenofibrate in a dose-dependent pattern. Conclusions Topical application of fenofibrate suppressed the formation of ocular surface squamous metaplasia, which might be mediated through the PPAR-α signaling pathway.

Background: Proparacaine hydrochloride (PH), as a a local anesthetic, is used regularly in ophthalmic surgery and optometry. However, few pieces of research on the ocular surface toxicity of PH eye drops have so far been reported. Purpuse: To evaluate the effect of PH on the ocular surface of mice. Methods: Male C57/BL6 mice were divided into four groups: normal, vehicle-treated, 0.05% PH-treated and 0.5% PH treated 7 days. The clinical indications were tear production, corneal sensitivity, and fluorescein staining. The structure and morphology of the cornea were examined by confocal microscopy; the thickness of the corneal center was examined by anterior segment optical coherence tomography (AS-OCT); and the corneal epithelial microvilli morphology was examined using scanning electron microscopy. Hematoxylin-eosin staining was used to characterize the central corneal morphology. Immunofluorescence staining for cytokeratin 10 (K10) was employed to evaluate squamous metaplasia in the corneal epithelium. Results: The results show that topical PH treatment diminished tear production and corneal sensitivity, and increased corneal fluorescein staining scores. Moreover, PH altered the corneal epithelial microvilli morphology, disrupted the epithelial barrier of the cornea, and promoted apoptosis of ocular surface cells. Furthermore, the expression of K10 in the corneal epithelium was found to be increased. Conclusion: Treatment with 0.5% PH caused instability of the tear film, and changes in corneal sensitivity and ocular surface homeostasis.

Amniotic membrane (AM) has been widely used as a temporary or permanent graft in the treatment of various ocular surface diseases. In this study, we compared the epithelial wound healing and tissue remodeling after ocular surface reconstruction with intact amniotic membrane (iAM) or denuded amniotic membrane (dAM). Partial limbal and bulbar conjunctival removal was performed on New Zealand rabbits followed by transplantation of cryo-preserved human iAM or dAM. In vivo observation showed that the epithelial ingrowth was faster on dAM compared to iAM after AM transplantation. Histological observation showed prominent epithelial stratification and increased goblet cell number on dAM after 2 weeks of follow up. Collagen VII degraded in dAM within 2 weeks, while remained in iAM even after 3 weeks. The number of macrophages and α-SMA positive cells in the stroma of remodelized conjunctiva in the dAM transplantation group was considerably less. In conclusion, dAM facilitates epithelial repopulation and goblet cell differentiation, further reduces inflammation and scar formation during conjunctival and corneal limbal reconstruction.

Pterygium is a very common disease in an eye clinic characterized by a benign proliferation of local conjunctiva that often crosses the limber of cornea and extends into corneal surface. Variety of studies has showed that pterygium is able to result in ocular discomfort and the change of ocular surface environment, such as dry eye. However, the link between abnormal tear film function and pterygium is controversial. Meibomian gland dysfunction (MGD) is a common cause of dry eye and ocular discomfort but is often neglected, which may be the missing link between dry eye and pterygium. In this study, our data firstly revealed increased abnormality of meibomian gland structure and function in pterygium patients, representing with increased abnormality of MGD parameters such as meibum expression (P P

Endothelial-to-mesenchymal transition (EnMT) is a cell transformation process involved in both morphogenesis and pathogenesis. EnMT of corneal endothelial cells happens after endothelial injury and during ex vivo culture. Previous studies have shown that the transforming growth factor-β signaling pathway is involved in this transition. In this study, we found that rat corneal endothelial cells could spontaneously undergo EnMT during ex vivo culture. This change in rat corneal endothelial cells was associated with Notch signaling pathway activation after the first passage, which was blocked by the Notch inhibitor N-[N-(3,5-Difluorophenacetyl)-L-alanyl]-S-phenylglycine t-butyl ester (DAPT). This inhibitor also prevented transforming growth factor β1-, β2-, and β3-induced EnMT and reversed transformed rat corneal endothelial cells to a normal phenotype. Furthermore, DAPT treatment blocked retrocorneal membrane formation in a rat corneal endothelium damage model. Our study indicates that the Notch signaling pathway is involved in the corneal EnMT process, which may be a novel therapeutic target for treating corneal endothelial fibrogenic disorders.

Squamous metaplasia is a common pathologic condition in ocular surface diseases for which there is no therapeutic medication in clinic. In this study, we investigated the effect of a small molecule, APR-246/PRIMA-1(Met), on squamous metaplasia in human conjunctival epithelium.Human conjunctival explants were cultured for up to 12 days under airlifting conditions. Epithelial cell differentiation and proliferation were assessed by Cytokeratin 10 (K10), K14, K19, Pax6, MUC5AC, and p63 immunostaining patterns. β-catenin and TCF-4 immunofluorescent staining and real-time PCR characterized Wnt signaling pathway involvement. Pterygium clinical samples were cultured under airlifting conditions with or without APR-246 for 4 days. p63, K10, β-catenin, and TCF-4 expression in pterygial epithelium was determined by immunofluorescent staining and real-time PCR.Airlift conjunctival explants resulted in increased stratification and intrastromal epithelial invagination. Such pathology was accompanied by increases in K10, K14, and p63 expression, whereas K19 and Pax6 levels declined when compared to those in freshly isolated tissue. On the other hand, APR-246 reversed all of these declines in K10, K14, and p63 expression. Furthermore, K19 and Pax6 increased along with rises in goblet cell density. These effects of APR-246 were accompanied by near restoration of normal conjunctival epithelial histology. APR-246 also reversed squamous metaplasia in pterygial epithelium that had developed after 4 days in ex vivo culture.Reductions in squamous metaplasia induced by APR-246 suggest it may provide a novel therapeutic approach in different squamous metaplasia-associated ocular surface diseases.

To evaluate the early clinical results of keratoconic eyes treated with transepithelial iontophoresis corneal collagen cross-linking (i-CXL) within 1 year.Propective nonrandomized study. Twenty-three eyes of 23 patients with progressive keratoconus with minimum corneal thickness from 380 µm to 420 µm (including the epithelium) were included in this prospective, nonrandomized clinical study and treated with i-CXL. Scoring of pain and foreign body sensation, slit lamp examination, uncorrected visual acuity (UCVA) and best corrected distance visual acuity (BCVA), corneal topography, anterior segment optical coherence tomography (AS-OCT), in vivo corneal confocal microscopy and endothelial cell count were assessed before surgery and at 1, 3, 6 and 12 months (m) postoperatively. Paired t test was applied for statistical analysis.Moderate pain and foreign body sensation were reported by most patients on postoperative Day (D) 1, but rapidly decreased and eventually disappeared on D3. Mild epithelial damage was observed on D1, and the epithelium fully recovered on D3. Improvement of UCVA and BCVA were recorded at 3 months and 12 months postoperatively. Orbscan II corneal topography revealed the significant reductions of Kmax and Kmin from 3m to 12m (Kmax, t = 2.912, P0.01, Kmin, t = 2.508, P0.05) postoperatively while the other parameters remained stable. The Kmax and Kmin at 12m was (52.94 ± 4.87) and (46.78 ± 3.71) respectively, while the preoperative values was (54.37 ± 5.56) and (48.53 ± 3.57) respectively. Within 1m postoperatively, AS-OCT exhibited an increase of reflectance with a white line (demarcation line) in the anterior stroma, in vivo confocal microscopy also showed the significant thickening and increased connections of collagen fibers with maximal depth of about 133 µm. The corneal endothelial cell density remained stable (t = 0.915, P0.05). None of the patients showed postoperative complications such as corneal infection, scarring, ulcer, persistent epithelial defect, etc.Priliminary clinical results within 1 year postoperatively demonstrated the efficacy and safety of i-CXL for the management of progressive keratoconus. This technique was applicable for keratoconic eyes with minimum corneal thickness around 400 µm. i-CXL showed the advantage of short time consuming in surgery, rapid recovery and few complication, and has the potential to become a valid alternative for the treatment of keratoconus.

To evaluate the clinical results of keratoconic eyes with thin corneas that were treated by using corneal collagen cross-linking with hypo-osmolar riboflavin solution.Retrospective, nonrandomized study. Fifteen eyes of 15 patients with progressive keratoconus and corneal thickness of less than 400 µm (without the epithelium) were included in this study. Application of hypo-osmolar riboflavin solution to the cornea for 30 minutes after its de-epithelialization was followed by ultraviolet A collagen cross-linking. Corneal thickness was measured with anterior segment OCT before surgery, after epithelial removal, and after hypotonic riboflavin solution application. Before the ultraviolet A application was started, we must be sure that the thinnest cornea was equal to or greater than 400 µm. Examinations comprised an evaluation of uncorrected distance visual acuity and best corrected visual acuity, slit-lamp microscopy, corneal topography, and endothelial cell counting after the procedure.Before surgery, the mean corneal thickness (with the epithelium) was (399.27 ± 17.87) µm, and after the removal of epithelium, the thickness of the cornea was reduced to (354.00 ± 18.57) µm. After the application of the hypo-osmolar riboflavin solution, this value increased to (477.73 ± 20.87) µm. The improvements in visual acuity and keratometry readings occurred during the follow-up. No statistically signiflcant differences were found between preoperative and postoperative endothelial cell counts. No complications such as scarring lesions in the stroma and corneal endothelial damage were observed throughout the study period.The results of our study, using hypo-osmolar riboflavin solution in a cross-linking procedure in thin corneas, show a stability of keratoconus 12 months after cross-linking.

PURPOSE: To investigate the expression pattern and function of survivin in the development of pterygium. METHODS: Primary pterygia at quiescent or advanced clinical stage and normal human conjunctival tissues were used in this study. Pterygium epithelial cells (PECs) were cultured in keratinocyte serum-free defined medium and harvested at different growth stages. Tissue sections and cultured cells were detected with survivin, phosphorylated survivin (Thr43), p63, p57, and p21 on protein, and/or mRNA level. Cell Counting Kit (CCK)-8 assay was performed to measure proliferation status of primary cultured PECs. Small interfering (si) RNA specific for survivin was transfected on PECs at subconfluence stage. RESULTS: Survivin was highly expressed in all pterygium tissues, but not in normal human conjunctiva, at mRNA and protein levels. It was mainly present in the epithelial cytoplasm of pterygium at quiescent stage, while present in the nucleus of pterygium at advanced stage. Phosphorylated survivin was upregulated in pterygium at advanced stage. Pterygium epithelial cells cultured under subconfluence stage showed higher expression of survivin and p63, but lower expression of p57 and p21, compared with PECs reached confluence. Both total and phosphorylated survivin was mainly expressed in the nuclei of PECs under subconfluence, and there was cytoplasmic translocation of survivin when PECs reached confluence. The knockdown of survivin by siRNA inhibited proliferation of PECs, accompanied by downregulation of p63, and upregulation of p57 and p21. CONCLUSIONS: Higher subcellular expression and phosphorylation of survivin may play roles in the development of pterygium. Survivin could be targeted for the treatment of pterygium.

The meibomian gland (MG) in the eyelids, which is the largest sebaceous gland throughout the body, synthesize and secrete lipids to form the superficial tear film layer. It plays a key role in maintaining the ocular surface health. Abnormalities in meibomian gland morphology lead to meibomian gland dysfunction, which is the main cause of evaporative dry eye. Study on meibomian gland morphology will contribute significantly to the diagnosis and treatment of meibomian gland dysfunction. This review is just focusing on the current studies about techniques to visualize the morphology of the MG and changes of meibomian gland morphology related to diseases.

There are several animal models illustrating dry eye pathophysiology. Current study would like to establish an ex vivo tissue culture model for characterizing dry eye. Human conjunctival explants were cultured under airlift or submerged conditions for up to 2 weeks, and only airlifted conjunctival cultures underwent increased epithelial stratification. Starting on day 4, the suprabasal cells displayed decreased K19 expression whereas K10 keratin became evident in airlift group. Pax6 nuclear expression attenuated already at 2 days, while its perinuclear and cytoplasmic expression gradually increased. MUC5AC and MUC19 expression dramatically decreased whereas the full thickness MUC4 and MUC16 expression pattern disappeared soon after initiating the airlift condition. Real time PCR showed K16, K10 and MUC16 gene up-regulated while K19, MUC5AC, MUC19 and MUC4 down-regulated on day 8 and day 14. On day 2 was the appearance of apoptotic epithelial and stromal cells appeared. The Wnt signaling pathway was transiently activated from day 2 to day 10. The inflammatory mediators IL-1β, TNF-α, and MMP-9 were detected in the conditioned media after 6 to 8 days. In conclusion, airlifted conjunctival tissue cultures demonstrated Wnt signaling pathway activation, coupled with squamous metaplasia, mucin pattern alteration, apoptosis and upregulation of proinflammatory cytokine expression. These changes mimic the pathohistological alterations described in dry eye. This correspondence suggests that insight into the pathophysiology of dry eye may be aided through the use of airlifted conjunctival tissue cultures.

PURPOSE To develop a mouse model of limbal stem cell deficiency (LSCD) by topical administration of benzalkonium chloride (BAC). METHODS BAC solutions (0%-0.5%) were applied to the mouse ocular surface for 4 weeks. Corneal neovascularization, inflammation, and epithelial status were observed under slit-lamp microscope. The eyeball and ocular surface tissues were collected at 4 and 12 weeks and labeled with a series of antibodies. Limbal structure was evaluated by light and transmission electron microscopy (TEM). Corneal impression cytology was performed at 12 weeks, and specimens were labeled with periodic acid Schiff (PAS) reagents. RESULTS BAC (0.5%) four times per day for 28 days successfully induced the typical manifestations of LSCD, including corneal neovascularization, severe inflammation in the stroma, and diffuse epithelial defect (P < 0.001). Conjunctival epithelium markers K19 and K13 were positive on the corneal surface. Expression of the putative limbal stem cell markers P63 and ABCG2 was abolished in the limbal epithelium. β-catenin was negative in the basal layer. TEM revealed the irregular basement membrane and the loss of stem cell-specific ultrastructure in the limbal basal epithelium. In the 0.5% BAC group, goblet cells could not be observed on day 28 but emerged after the cessation of BAC, and remained over the cornea after 8 weeks. K13-positive cells were still present over the cornea with the loss of K12. CONCLUSIONS Topical administration of BAC at high concentration and frequency in mouse induces ocular surface changes resembling those of LSCD in humans, representing a novel model of LSCD.

We investigated the efficacy of low-temperature airlift preservation of human corneal limbal tissue for ex vivo expansion and allograft keratolimbal transplantation.Human limbal tissue either was submerged or airlifted in Optisol-GS medium and preserved at 4°C for up to eight days. Hematoxylin and eosin, and E-cadherin staining was performed to investigate epithelial structure and cell-cell junction. Epithelial cell differentiation and proliferation were studied using the biomarkers, such as K10, K12, K14, Ki67, and p63. Cell apoptosis was detected with the TUNEL assay. The epithelial progenitor cell pool was evaluated by clonal culture of epithelial cells on 3T3 feeder layers. For clinical application, keratolimbal transplantation was performed in three patients with partial limbal stem cell deficiency, using limbal tissues preserved under the airlift manner. Pre- and postoperative evaluations were conducted by slit-lamp microscopy and fluorescein staining.After eight days, intact epithelia with strong cell-cell junctions were retained only in airlifted tissues. Airlifting maintained a normal corneal differentiation pattern, along with low proliferation activity and increased proliferation potential, but little apoptosis. Epithelial cells harvested from the airlift preservation for up to eight days exhibited stable clonogenicity. Limbal tissues preserved under the airlift manner successfully reconstructed corneal and limbal surfaces in partial limbal stem cell-deficient patients.Limbal tissues preserved under hypothermic airlift conditions maintain the intact structure, normal phenotype, high viability, and stem cell pool of limbal epithelia. Such a method may be used in eye bank tissue processing and corneal epithelial tissue engineering.

Oxygen concentration has been shown to be crucial in the proliferation and differentiation of various types of cells, while the impact of oxygen tension on the lineage commitment of epithelial cells remains elusive. In this study, we investigated the effect of hypoxia on the differentiation of corneal limbal epithelium using an ex vivo squamous metaplasia model. Under normoxic conditions when exposed to air, the hyperproliferation and abnormal epidermal-like differentiation of human corneal limbal epithelium was induced, whereas when exposed to air under hypoxic conditions, although we observed augmented proliferation, the abnormal differentiation was inhibited. The Notch signaling pathway was activated in hypoxic cultures, whereas the p38 MAPK signaling pathway was downregulated. The addition of Notch inhibitor under hypoxic conditions restored the activation of p38 MAPK and resulted in the recidivation of limbal epithelial cells to epidermal-like differentiation. Moreover, the epidermal-like differentiation of rabbit limbal epithelial cells was also blocked under hypoxic conditions in corneal epithelial cell sheets engineered ex vivo. We concluded that hypoxia can prevent abnormal differentiation while enhancing the proliferation of corneal limbal epithelial cells. Hypoxia coupled with air exposure can be used in the tissue engineering of corneal limbal epithelium.

To evaluate the expression of Pax6 in pterygia epithelium.Fifteen patients (15 eyes) with pterygium who underwent simple excision were enrolled in this study. Pax6, K10, K19 and MUC5AC immunostaining were performed in pterygia tissue and normal conjunctiva.A decline or absence of Pax6 expression and K19 keratin and MUC5AC but increased of K10 expression with epidermal differentiation was observed in pterygia epithelium, in comparison with the normal conjunctival tissue.These results indicate that down-regulation of Pax6 is associated with abnormal differentiation of the epithelial cells in pterygium.

Purpose To investigate the effects and possible mechanisms of serine protease inhibitor A3K (SERPINA3K) on the formation of ocular surface squamous metaplasia in a mouse dry eye model induced by topical benzalkonium chloride (BAC). Methods The eye drops containing SERPINA3K were topically administered during the induction of BAC-induced dry eye. The clinical indications of dry eye were evaluated on day (D)16, including tear break-up time (BUT), tear volume, corneal fluorescein staining, and inflammatory index. Global specimens were collected on D16 and the following examinations were performed: histologic investigation, immunostaining of cytokeratin 10 (K10), p63 and Ki67 in the cornea, and Western blot analysis of tumor necrosis factor-α (TNF-α). Results Serine protease inhibitor A3K suppressed the formation of BAC-induced dry eye, presenting with longer BUTs, lower corneal fluorescein staining scores, and inflammatory index, while no significant changes in tear volume. It also reduced the severity of abnormal differentiation and proliferation on ocular surface with lower expressions of K10, p63, and Ki67, and retained the number of goblet cells in the conjunctival fornix. Serine protease inhibitor A3K significantly decreased the levels of TNF-α in the cornea. Conclusions Topical application of SERPINA3K ameliorated the severity of ocular surface squamous metaplasia and suppressed the formation of BAC-induced dry eye.

Objective To explore meibomian gland dysfunction (MGD) may determine the severity of dry eye conditions in visual display terminal (VDT) workers. Methodology Prospective, case-control study carried out in China.106 eyes of 53 patients (VDT work time >4 hour per day) were recruited as the Long time VDT group; 80 eyes of 40 control subjects (VDT work time ≤4 hour per day) served as the Short time VDT group. A questionnaire of Ocular Surface Disease Index (OSDI) and multiple tests were performed. Three dry eye tests: tear film breakup time (BUT), corneal fluorescein staining, Schirmer I test; and three MGD parameters: lid margin abnormality score, meibum expression assessment (meibum score), and meibomian gland dropout degree (meiboscore) using Keratograph 5 M. Principal Findings OSDI and corneal fluorescein score were significantly higher while BUT was dramatically shorter in the long time VDT group than the short time VDT group. However, the average of Schirmer tear volumes was in normal ranges in both groups. Interestingly, the three MGD parameters were significantly higher in the long time VDT group than the short time one (P

To investigate the differentiation of conjunctival epithelium and tear film function in pingueculae.Twelve patients (12 eyes) who underwent simple excision for pingueculae were enrolled in the study. Immunostaining for K14, K19, K10, MUC5AC, Pax6, P63, Ki67, and K16 was performed on the pinguecula epithelium and normal conjunctival epithelium. Schirmer I test results and tear film break-up time (TFBUT) were evaluated just before and 1 month after surgery.Abnormal epidermal differentiation of pinguecula tissue, as evidenced by a decline in or absence of Pax6 expression, was accompanied by a decline in or absence of K19 keratin and MUC5AC, and an increase in K10 and K14 keratin. Furthermore, the pinguecula epithelium was actively proliferating, as evidenced by positive expression of Ki67, P63, and K16 keratin. The Schirmer I test results did not indicate any significant differences before and after surgery. However, the TFBUT was significantly prolonged 1 month after surgery compared with before surgery.Pinguecula is a condition of abnormal epithelial differentiation. It is characterized by squamous proliferation and metaplasia, resulting in instability of tear film with normal basic tear secretion.