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Supplemental Digital Content is Available in the Text., Purpose: Drug delivery directly to the corneal stroma currently relies on microscopic injections that demonstrate low reproducibility and clinician-dependent variability. With use of biological drugs such as adeno-associated viral (AAV) vectors, precise and consistent drug deposition is critical to reduce concerns related to off-target transduction and the host's immune response to the viral capsid and/or transgene-derived product. Therefore, a precise corneal injection (PCI) microneedle was designed to allow accurate depth-specific injections into the corneal stroma in a macroscopic setting. Methods: High-frequency ultrasound and confocal microscopy demonstrated the consistent ability to predetermine the precise injection depth using PCI needles of varying sizes. Next, a comparison between a standard 31-G needle and PCI needles was performed in vivo using AAV vector gene delivery. Results: Intrastromal corneal injections using the PCI microneedle resulted in less vector leakage at the site of injection and fewer anterior chamber penetrations compared with a standard 31-G needle. Although reporter gene expression appeared similar when the vector was administered with either needle type, a trend toward increased vector genomes was noted in the PCI-injected corneas at the experimental conclusion. As hypothesized, corneal perforation resulted in increased detection of AAV vector genomes in nontarget tissues, highlighting the importance of consistency for biological drug applications in the cornea. Conclusions: Further development of the PCI microneedle is warranted especially for AAV corneal gene therapy and offers the potential to enhance transduction while significantly reducing safety concerns and intraclinician and interclinician injection variability.

PURPOSE: The purpose of this study was to report the analysis of reverse transcriptase polymerase chain reaction (RT-PCR) of nasopharyngeal (NP) samples of cornea donors dying because of causes unrelated to severe acute respiratory coronavirus (SARS-CoV-2). METHODS: A retrospective analysis of all cornea donors dying from causes other than SARS-CoV-2 between August 2020 and December 2020 was performed. Informed consent was obtained from the next of kin of the deceased for RT-PCR testing from NP swabs. Rapid antigen testing from all the deceased was performed before in situ cornea excision. In addition, NP samples in viral transport media for RT-PCR were also collected for SARS-CoV-2 analysis. Corneas were released from the eye bank only after a negative RT-PCR report. RESULT: One hundred eighteen corneas from 59 donors were obtained by the eye bank. Eleven donors (18.64%) were positive for SARS-CoV-2 on RT-PCR testing. Six of these 11 donors had a Ct value of E gene less than 25. CONCLUSIONS: NP samples of cornea donors dying due to causes other than coronavirus disease-19 were positive for SARS-CoV-2 on RT-PCR. This implicates that donors could be having asymptomatic/undetected coronavirus disease infection. We recommend adding the routine testing of NP samples of all cornea donors in the eye banking protocol in this ongoing SARS-CoV-2 pandemic.

Purpose Fungal keratitis can be difficult to medically treat. Topical antifungals are usually applied empirically as the initial option in treating fungal keratitis. Natamycin (NAT) and/or voriconazole (VRCZ) have been widely used in the treatment of fungal keratitis. However, Fusarium solani species complex (FSSC), which are the dominant species of fungal keratitis, are resistant to VRCZ. This study investigated in vitro efficacy of luliconazole (LLCZ), a new imidazole antifungal, against FSSC and other filamentous fungi. Methods A total of 18 Fusarium isolates and 7 others were grown on potato dextrose agar at 30 and 37°C. For Fusarium, species identification and phylogenetic tree analysis were performed based on elongation factor-1α (EF-1α) DNA sequencing. The broth microdilution method was used for antifungal susceptibility testing of 11 antifungal drugs including LLCZ. Results The 18 identified Fusarium isolates belonged to FSSC (n = 13), Fusarium oxysporum species complex (FOSC; n = 2), Fusarium chlamydosporum species complex (FCSC; n = 1), Fusarium incarnatum-equiseti species complex (FIESC; n = 1), and Fusarium fujikuroi species complex (FFSC; n = 1). We further divided 13 FSSC isolates into 3 clades, FSSC5 (n = 8), FSSC3 + 4 (n = 4), and FSSC9-a (n = 1), with 8 FSSC strains growing at 37°C. LLCZ showed lowest minimum inhibitory concentrations (MICs) against all tested filamentous fungi, with a MIC90 against the Fusarium species of 0.06 μg/mL, whereas MIC90 for NAT and VRCZ were 4 and 8 μg/mL, respectively. Conclusions LLCZ has the strongest in vitro antifungal activity among all drugs used against broad-range filamentous fungi including FSSC. LLCZ may potentially be a new medical treatment option for fungal keratitis.

Supplemental Digital Content is Available in the Text., Purpose: Loss of corneal endothelial cells (CECs) bears disastrous consequences for the patient, including corneal clouding and blindness. Corneal transplantation is currently the only therapy for severe corneal disorders. However, the worldwide shortages of corneal donor material generate a strong demand for personalized stem cell–based alternative therapies. Because human mesenchymal stem cells are known to be sensitive to their mechanical environments, we investigated the mechanotransductive potential of Descemet membrane–like microtopography (DLT) to differentiate human mesenchymal stem cells into CEC-like cells. Methods: Master molds with inverted DLT were produced by 2-photon lithography (2-PL). To measure the mechanotransductive potential of DLT, mesenchymal stem cells were cultivated on silicone or collagen imprints with DLT. Changes in morphology were imaged, and changes in gene expression of CEC typical genes such as zonula occludens (ZO-1), sodium/potassium (Na/K)-ATPase, paired-like homeodomain 2 (PITX2), and collagen 8 (COL-8) were measured with real-time polymerase chain reaction. At least immunofluorescence analysis has been conducted to confirm gene data on the protein level. Results: Adhesion of MSCs to DLT molded in silicone and particularly in collagen initiates polygonal morphology and monolayer formation and enhances not only transcription of CEC typical genes such as ZO-1, Na/K-ATPase, PITX2, and COL-8 but also expression of the corresponding proteins. Conclusions: Artificial reproduction of Descemet membrane with respect to topography and similar stiffness offers a potential innovative way to bioengineer a functional CEC monolayer from autologous stem cells.

Purpose: Fungal keratitis is a major cause of corneal ulcers, resulting in significant visual impairment and blindness. Fenretinide, a derivative of vitamin A, has been shown to suppress inflammation in a multitude of diseases. In this study, we aimed to characterize the effect of fenretinide in Aspergillus fumigatus keratitis of the eye in a mouse model. Methods: In vivo and in vitro experiments were performed in mouse models and THP-1 macrophage cell cultures infected with A. fumigatus, respectively. Experimental subjects were first pretreated with fenretinide, and then the effect of the compound was assessed with clinical evaluation, neutrophil staining, myeloperoxidase assay, quantitative polymerase chain reaction (qRT-PCR), and western blot. Results: We confirmed that fenretinide contributed to protection of corneal transparency during early mouse A. fumigatus keratitis by reducing neutrophil recruitment, decreasing myeloperoxidase (MPO) levels and increasing apoptosis. Compared with controls, fenretinide impaired proinflammatory cytokine interleukin 1 beta (IL-1β) production in response to A. fumigatus exposure with contributions by lectin-type oxidized LDL receptor 1 (LOX-1) and c-Jun N-terminal kinase (JNK). Conclusions: Together, these findings demonstrate that fenretinide may suppress inflammation through reduced neutrophil recruitment and inflammatory cytokine production in A. fumigatus keratitis.

Purpose To evaluate the validity and reliability of a novel handheld osmolarity system (I-PEN Osmolarity System; I-MED Pharma Inc, Dollard-des-Ormeaux, Quebec, Canada) for measurement of the osmolarity of a National Institute of Standards and Technology (NIST) traceable solution at a variety of ambient temperatures. Methods A total of 65 measurements of an NIST solution with a verified osmolarity of 290 ± 2 mOsmol/L were taken using 3 separate handheld osmolarity systems, 65 unique single-use sensors (SUSs) from 3 different lots, and 2 adaptors. Mean values were calculated using the device, SUS, and adaptor. Measurements were taken using a handheld osmolarity system, an adaptor, and 56 individual SUSs at 6 different ambient temperatures ranging from 17.7 to 26.5°C. Results Overall, the mean osmolarity measured was 294.06 mOsmol/L (SD ±2.29; percent coefficient of variation 0.78), ranging from 286.60 to 298.18 mOsmol/L. This fell within a prespecified acceptable variability of ±4 mOsmol/L (SD ±7). Mean values did not vary across devices, adaptors, or single-use sensors used. Mean osmolarity measurements increased with rising ambient temperatures, with an R = 0.88. The temperature correction factor was calculated to be 2.01 mOsmol/L per °C. Conclusions The osmolarity system reliably and accurately measured the osmolarity of an NIST solution in a laboratory setting, using an adaptor to correct for differences in resistance between a laboratory NIST solution measurement and direct measurements on the palpebral conjunctiva of the eyelid. The handheld osmolarity system represents a rapid and accurate instrument for measurement of tear osmolarity in a simulated testing setting.

Purpose: To correlate rheumatologic with ophthalmic and laboratory findings in patients with rheumatoid arthritis (RA) to identify what effect these have on development of ocular disease. Methods: This is a cross-sectional study of 172 eyes of 86 patients with RA. Patients were examined by a group of rheumatologists. Sociodemographic, clinical, and laboratory data were collected. All patients underwent complete ophthalmologic examination including corneal topography and endothelial cell count. Results: There was no significant correlation between RA-negative prognostic indicators (NPIs) and pathologic corneal findings. Patients using disease-modifying antirheumatic drugs (DMARDs) and antimalarial drugs had greater corneal volumes (mean difference 8.51 mm3, 90% confidence interval [CI], 3.98-13.04, P = 0.004; and 2.24, 90% CI, 0.32-4.54, P = 0.048, respectively). Patients using azathioprine had lower endothelial cell counts compared with those using other drugs (mean difference 180 cells/mm2, 90% CI, 69-291, P = 0.008). Patients using biologic DMARDs had better tear osmolarity values (between 280 and 300 mOsm/L) than patients not using them (mean difference 14.3 mOsm/L, P = 0.022). There was no correlation between NPIs of RA and positive keratoconus screening indices (Spearman correlation OD -0.013, P = 0.91; OS -0.033, P = 0.76). Conclusions: There was no clear correlation between RA-NPIs and pathologic corneal findings in our study. DMARDs treatment may help maintain corneal integrity in our patients and prevented collagenolytic manifestations of RA. Other medications such as azathioprine should be used carefully, as endothelial damage may potentially occur. Copyright © 2018 Wolters Kluwer Health, Inc. All rights reserved.

Supplemental Digital Content is Available in the Text., Purpose: To evaluate the efficacy of sodium hyaluronate (HA) eye drops for the treatment of diabetic ocular surface diseases in mice. Methods: Male 6- to 8-week-old C57BL/6 mice underwent induction of type 1 diabetes with intraperitoneal injections of streptozotocin, with normal mice as the control. Topical 0.3% HA, 0.1% HA, 0.4% polyethylene glycol eye drops, and normal saline were administered to diabetic mice with an intact or debrided corneal epithelium. Normal saline was applied in the controls. Corneal epithelial wound healing rate, corneal sensation, nerve fiber density, conjunctival goblet cell number, and MUC-5AC content were measured and compared. Results: Compared with the controls, topical 0.3% HA use in diabetic mice showed significant improvements in the corneal epithelial wound healing rate (48 hours: 91.5% ± 4.8% vs. 79.8% ± 6.1%; P < 0.05), corneal sensitivity (4.1 ± 0.3 cm vs. 3.5 ± 0.3 cm; P < 0.05), nerve fiber density (12.9% ± 2.3% vs. 6.6% ± 2.4%; P < 0.05), conjunctival goblet cell number (31.0 ± 8.4/100 μm vs. 19.6 ± 7.1/100 μm; P < 0.05), and MUC-5AC content (12.5 ± 1.4 ng/mg vs. 7.8 ± 1.5 ng/mg protein; P < 0.05). The beneficial effects of 0.3% HA were better than those of 0.1% HA and 0.4% polyethylene glycol. Conclusions: Topical 0.3% HA treatment promoted corneal epithelial regeneration, improved corneal sensation, and increased density of corneal nerve fibers and conjunctival goblet cells in mice with diabetic ocular surface diseases.

Supplemental Digital Content is Available in the Text., Purpose: To investigate toxicity associated with buffers commonly used in topical ocular drug formulations using a human corneal–limbal epithelial (HCLE) and a human conjunctival epithelial (HCjE) cell model. Methods: HCLE and HCjE cells were incubated for 10, 30, or 60 minutes with 4 different buffers based on borate, citrate, phosphate, and Tris-HCl at 10, 50, and 100 mM concentrations. To detect possible delayed effects on cell viability, after 60 minutes of buffer incubation, cells were further incubated for 24 hours with a cell medium. Cell viability was determined using a colorimetric XTT–based assay. The morphology of cells was also investigated. Results: HCjE cells showed more sensitivity to buffer incubation than HCLE cells. The 100 mM phosphate buffer displayed significant delayed effects on cell viability of HCLE 16.8 ± 4.8% and HCjE 39.2 ± 6.1% cells after 60 minutes of exposure (P < 0.05). HCjE cell viability was reduced after 60 minutes incubations with 50 and 100 mM citrate buffer to 42.8 ± 6.5% and 39.3 ± 7.9%, respectively, and even lower percentages at the delayed time point (both P < 0.05). HCLE cell morphology was distinctly altered by 100 mM phosphate and Tris buffers after 30 minutes, whereas HCjE cells already showed marked changes after 10 minutes of exposure to 100 mM citrate and phosphate buffers. Conclusions: We observed a time-dependent decrease of viability in both HCLE and HCjE cells exposed to higher buffer concentrations. Therefore, we propose further in vivo studies to translate these finding to humans to discern the real effects of the buffer concentration in eye drops on the ocular surface.

Purpose: To understand soft contact lens (SCL) and gas-permeable (GP) lens wearers' behaviors and knowledge regarding exposure of lenses to water. Methods: The Contact Lens Risk Survey (CLRS) and health behavior questions were completed online by a convenience sample of 1056 SCL and 85 GP lens wearers aged 20 to 76 years. Participants were asked about exposing their lenses to water and their understanding of risks associated with these behaviors. Chi-square analyses examined relationships between patient behaviors and perceptions. Results: GP lens wearers were more likely than SCL wearers to ever rinse or store lenses in water (rinsing: 91% GP, 31% SCL, P < 0.001; storing: 33% GP, 15% SCL P < 0.001). Among SCL wearers, men were more likely to store (24% vs. 13%, P = 0.003) or rinse (41% vs. 29%, P = 0.012) their lenses in water. Showering while wearing lenses was more common in SCL wearers (86%) than GP lens wearers (67%) (P < 0.0001). Swimming while wearing lenses was reported by 62% of SCL wearers and 48% of GP lens wearers (P = 0.027). Wearers who rinsed (SCL; P < 0.0001, GP; P = 0.11) or stored lenses in water (SCL; P < 0.0001, GP P = 0.007) reported that this behavior had little or no effect on their infection risk, compared with those who did not. Both SCL (P < 0.0001) and GP lens wearers (P < 0.0001) perceived that distilled water was safer than tap water for storing or rinsing lenses. Conclusions: Despite previously published evidence of Acanthamoeba keratitis' association with water exposure, most SCL, and nearly all GP lens wearers, regularly expose their lenses to water, with many unaware of the risk.

Purpose: To demonstrate that vasoactive intestinal peptide (VIP), a corneal endothelial (CE) cell autocrine factor, maintains the integrity of corneal endothelium in human donor corneoscleral explants precut for endothelial keratoplasty. Methods: Twelve paired human donor corneoscleral explants used as control versus VIP-treated explants (10 nM, 30 minutes, 37°C) were shipped (4°C) to the Lions Eye Institute for Transplantation and Research for precutting (Moria CBM-ALTK Keratome), shipped back to the laboratory, and cultured in ciliary neurotrophic factor (CNTF, 0.83 nM, 37°C, 24 hours). Trephined endothelial discs (8–8.5 mm) were analyzed for differentiation markers (N-cadherin, CNTF receptor α subunit [CNTFRα], and connexin 43) by Western blot after a quarter of the discs from 4 paired explants were cut away and stained with alizarin red S for microscopic damage analysis. Two additional paired explants (6 days in culture) were stained for panoramic view of central CE damage. Results: VIP treatment increased N-cadherin and CNTFRα levels (mean ± SEM) to 1.38 ± 0.11-fold (P = 0.003) and 1.46 ± 0.22-fold (P = 0.03) of paired controls, respectively, whereas CE cell CNTF responsiveness in upregulation of connexin 43 increased to 2.02 ± 0.5 (mean ± SEM)-fold of the controls (P = 0.04). CE damage decreased from (mean ± SEM) 10.0% ± 1.2% to 1.6% ± 0.3% (P < 0.0001) and 9.1% ± 1.1% to 2.4% ± 1.0% (P = 0.0006). After 6 days in culture, the damage in whole CE discs decreased from 20.0% (control) to 5.5% (VIP treated). Conclusions: VIP treatment before precut enhanced the preservation of corneal endothelium.

Background: Epithelial neoplasm is an important global health-care problem, with high morbidity and mortality rates. Early diagnosis and appropriate treatment are essential for increased life survival. Prediction of occurrence of malignancy in a disease-free individual by any means will be a great breakthrough for healthy living. Aims and Objectives: The aims and objectives were to predict the genetic predisposition and propose a prediction protocol for epithelial malignancy of various systems in our body, in a disease-free individual. Methods: We have searched databases both manually and electronically, published in English language in Cochrane group, Google search, MEDLINE and PubMed from 2000 to 2019. We have included all the published, peer-reviewed, narrative reviews; randomized controlled trials; case–control studies; and cohort studies and excluded the abstract-only articles and duplicates. Specific words such as “etiological factors,” “pathology and mutations,” “signs and symptoms,” “genetics and IHC marker,” and “treatment outcome” were used for the search. A total of 1032 citations were taken, and only 141 citations met the inclusion criteria and were analyzed. Results: After analyzing various articles, the etiological factors, clinical signs and symptoms, genes and the pathology involved and the commonly used blood and tissue markers were analyzed. A basic investigation strategy using immunohistochemistry markers was established. Conclusion: The set of proposed biomarkers should be studied in future to predict genetic predisposition in disease-free individuals.

Purpose: To search for gold-standard histology indicators using alternative imaging modalities in keratoconic corneas. Methods: Prospective observational case–control study. Fourteen keratoconic corneas and 20 normal corneas (10 in vivo healthy subjects and 10 ex vivo donor corneas) were examined. Images of corneas were taken by spectral domain optical coherence tomography (SD-OCT) and in vivo confocal microscopy (IVCM) before keratoplasty. The same removed corneal buttons were imaged after keratoplasty with full-field optical coherence microscopy (FFOCM) and then fixed and sent for histology. Controls consisted of normal subjects imaged in vivo with IVCM and donor corneas imaged ex vivo with FFOCM. Corneal structural changes related to pathology were noted with each imaging modality. Cell density was quantified by manual cell counting. Results: Keratoconus indicators (ie, epithelial thinning/thickening, cell shape changes, ferritin deposits, basement membrane anomalies, Bowman layer thinning, ruptures, interruptions, scarring, stromal modifications, and appearance of Vogt striae) were generally visible with all modalities. Additional features could be seen with FFOCM in comparison with gold-standard histology, particularly in the Bowman layer region, whereas the combination of SD-OCT plus IVCM detected 76% of those features detected in histology. Three-dimensional FFOCM imaging aided interpretation of two-dimensional IVCM and SD-OCT data. Basal epithelial cell and keratocyte densities were significantly lower in patients with keratoconus than those in normals (P < 0.0001). Conclusions: Structural and cellular assessment of the keratoconic cornea by means of either in vivo SD-OCT combined with IVCM or ex vivo FFOCM in both cross-sectional and en face views can detect as many keratoconus indicators as gold-standard histology.

Many topical eye drop solutions used to treat corneal diseases contain preservatives to prevent contamination by microorganisms. The most commonly used preservative in ophthalmic preparations is benzalkonium chloride (BAC).1 As a soluble antimicrobial agent and a cationic surfactant, BAC enhances the penetration of active compounds. A recent study has revealed that some of the side effects of eye drops are caused by preservatives, such as BAC.2 Cytotoxicity of BAC on the ocular surface has been examined using in vitro cell culture systems, such as human corneal cell lines,3–5 human conjunctival cell lines,3,4,6,7 a 3-dimensional model of human corneal epithelium,8 and rabbit corneal cell lines.6,9 The ocular surface toxicity of BAC has also been investigated in several in vivo experiments by applying BAC to the ocular surface of mice,10,11 rats,12 and rabbits.13,14 Because of its toxicity, the use of BAC in eye drops has been criticized in recent years. BAC may induce ocular surface changes, causing ocular discomfort, tear film instability, loss of goblet cells, inflammation, conjunctival squamous metaplasia, epithelial apoptosis, subconjunctival fibrosis, and the potential risk of failure during future glaucoma surgery.15 Several studies have revealed that BAC is detrimental to ocular surface tissues and cells. Therefore, efforts are being made to reduce the adverse effects of BAC, and novel BAC-free ophthalmic formulations have been developed over the last several years.16,17 In addition, new preservatives with a wide range of activity and optimal safety profiles have been introduced.18 Dry eye syndrome is one of the most common multifactorial ocular surface diseases, affecting millions of people worldwide, and resulting in ocular dryness, discomfort, visual disturbance, and instability of tear film.19 Ocular surface inflammation may play a critical role in the pathogenesis of dry eye syndrome. Disease or dysfunction of tear secretory glands leads to changes in tear composition, such as hyperosmolarity, which stimulates the production of inflammatory mediators on the ocular surface.19,20 In turn, inflammation may cause dysfunction or loss of cells responsible for tear secretion or retention.21 Therefore, antiinflammatory therapy has been adopted as a common therapeutic strategy for the treatment of dry eye syndrome. Commonly used antiinflammatory eye drops, such as topical corticosteroids and cyclosporine A, are available for the treatment of dry eye syndrome.22,23 Therefore, it is desirable to identify and evaluate suitable ophthalmic antiinflammatory agents. Fluorometholone (9a-fluoro-11b,17a-dihydroxy-6a-methylpregna-1,4-diene-3,20-dione; FML) is a corticosteroid used for its glucocorticoid activity. This drug is formulated in the form of an ophthalmic solution for the treatment of allergic conjunctivitis and vernal keratoconjunctivitis.24 In particular, FML is an effective antiinflammatory drug widely used to control eye inflammation.25 This study was designed to evaluate the cytotoxicity and antiinflammatory effect of BAC by comparing BAC-containing FML with BAC-free FML, both of which are commercially available, in human corneal epithelial cells (HCECs) and a dry eye mouse model.

Background: The effects of different modes of mechanical ventilation in the same ventilatory support level on ventilator-induced diaphragm dysfunction onset were assessed in healthy rabbits. Methods: Twenty New Zealand rabbits were randomly assigned to 4 groups (n = 5 in each group). Group 1: no mechanical ventilation; group 2: controlled mechanical ventilation (CMV) for 24 hours; group 3: assist/control ventilation (A/C) mode for 24 hours; group 4: high-level pressure support ventilation (PSV) mode for 24 hours. Heart rate, mean arterial blood pressure, PH, partial pressure of arterial oxygen/fraction of inspired oxygen and partial pressure of arterial carbon dioxide were monitored and diaphragm electrical activity was analyzed in the 4 groups. Caspase-3 was evaluated by protein analysis and diaphragm ultra structure was assessed by electron microscopy. Results: The centroid frequency and the ratio of high frequency to low frequency were significantly reduced in the CMV, A/C and PSV groups (P < 0.001). The percent change in centroid frequency was significantly lower in the PSV group than in the CMV and A/C groups (P = 0.001 and P = 0.028, respectively). Electromyography of diaphragm integral amplitude decreased by 90% ± 1.48%, 67.8% ± 3.13% and 70.2% ± 4.72% in the CMV, A/C and PSV groups, respectively (P < 0.001). Caspase-3 protein activation was attenuated in the PSV group compared with the CMV and A/C groups (P = 0.035 and P = 0.033, respectively). Irregular swelling of mitochondria along with fractured and fuzzy cristae was observed in the CMV group, whereas mitochondrial cristae were dense and rich in the PSV group. The mitochondrial injury scores (Flameng scores) in the PSV group were the lowest among the 3 ventilatory groups (0.93 ± 0.09 in PSV versus 2.69 ± 0.05 in the CMV [P < 0.01] and PSV versus A/C groups [2.02 ± 0.08, P < 0.01]). Conclusions: The diaphragm myoelectric activity was reduced in the PSV group, although excessive oxidative stress and ultra-structural changes of diaphragm were found. However, partial diaphragm electrical activity was retained and diaphragm injury was minimized using the PSV mode.

Background: Diabetes aggravates brain injury after cerebral ischemia/reperfusion (I/R). Objective: To investigate whether limb I/R causes cerebral injury in a rat diabetes model and whether glycogen synthase kinase-3β (GSK-3β) is involved. Methods: Male adult Sprague-Dawley rats were assigned into streptozotocin-induced diabetes (n = 30; blood glucose ≥16.7 mmol/L) or control (n = 20) groups, further subdivided into diabetes I/R (3-hour femoral artery/vein clamping), diabetes-I/R + TDZD-8 (I/R plus GSK-3β inhibitor), diabetes-sham, control-sham and control-I/R groups (n = 10 each). Cortical and hippocampal morphology (hematoxylin/eosin); hippocampal CA1 apoptosis (TUNEL assay); cleaved caspase-3 (apoptosis), and Iba1 (microglial activation) protein expression (immunohistochemistry); phosphorylated/total GSK-3β and nuclear factor-κB (NF-κB) protein levels (Western blotting); and serum and brain tissue tumor necrosis factor (TNF)-α levels (enzyme-linked immunosorbent assay) were analyzed. Results: The diabetes-I/R group showed greater cortical and hippocampal injury, apoptosis, cleaved caspase-3 expression and Iba1 expression than the control-I/R group; TDZD-8 reduced injury/apoptosis and cleaved caspase-3/Iba1 expressions. The diabetes-I/R group had lower p-GSK-3β and p-NF-κBp65 expression than the control-I/R group (P < 0.05); TDZD-8 increased p-GSK-3β expression but decreased p-NF-κBp65 expression (P < 0.05). The diabetes-I/R group showed higher elevation of serum and brain tissue TNF-α than the control-I/R group (P < 0.05); TDZD-8 reduced TNF-α production. Conclusions: Diabetes exacerbates limb I/R-induced cerebral damage and activates NF-κB and GSK-3β.

Purpose: To evaluate and investigate the distribution and repeatability of anterior corneal surface astigmatism measurements (axis and magnitude) using a novel corneal topographer. Methods: Anterior corneal surface astigmatism was investigated in a total of 195 eyes using a novel multicolored spot reflection topographer (Cassini; i-Optics). Two patient groups were studied, a younger-age group A and an older-age group B. Three consecutive acquisitions were obtained from each eye. The repeatability of measurement was assessed using Bland–Altman plot analysis and is reported as the coefficient of repeatability. Results: Group A (average age 34.3 years) had on average with-the-rule astigmatism, whereas the older-age group B (average age 72.3 years) had on average against-the-rule astigmatism. Average astigmatism magnitude measurement repeatability in group A was 0.4 diopters (D) and in group B 0.4 D. Average astigmatism axis measurement repeatability in group A was 5.4 degrees and in group B 5.5 degrees. The axis measurement repeatability improved with increasing magnitude of astigmatism: in the subgroups with astigmatism between 3.0 and 6.0 D, the axis repeatability was 1.4 degrees (group A) and 1.2 degrees (group B), whereas in the subgroups with astigmatism larger than 6.0 D, the repeatability was 1.1 and 0.6 degrees, respectively. Conclusions: This novel corneal topography device seems to offer high precision in reporting corneal astigmatism. This study reaffirms the established trend of a corneal astigmatism shift from an average “with-the-rule” to “against-the-rule” with aging.

Corneal collagen cross-linking (CXL) as introduced in the Dresden protocol1,2 has succeeded in reducing visual loss in an increasing number of people worldwide. The Bunsen–Roscoe Law suggests that the same cross-linking effect could be completed in a shorter time as long as the total ultraviolet (UV) irradiation energy is the same. Higher intensity, shorter duration treatments have been tried, but there have been safety and effectiveness questions with these variations. Kymionis et al3 recently demonstrated that the 30-minute, 3-mW/cm2 UV-A intensity Dresden protocol CXL yielded a deeper corneal effect than a shorter, more intense treatment of a 10-minute, 9-mW/cm2 CXL protocol, casting some doubt on the application of the Bunsen–Roscoe Law to cross-linking. Studies on porcine eyes have shown toxicity to the endothelium at irradiation levels of 0.35 mW/cm2 for 30 minutes.4 Based on the Lambert–Beer law of exponential decline in UV transmission as a function of corneal depth, a recommendation was derived, and it is now standard, that cross-linking at 3 mW/cm2 should not be performed on corneas thinner than 400 μm. Furthermore, to prevent endothelial damage, there was a recommendation that a riboflavin layer be renewed on the cornea during UV-A irradiation, but variations in the thickness of the riboflavin covering layer over the treatment time and over different treatments is a potential cause of variation in the degree of cross-linking effect. Two laboratory experiments, reported here, were conducted to determine if it was possible to produce a shorter duration, lower energy, time-variable UV intensity cross-linking treatment that would accomplish the goal of adequately increasing corneal stiffness while lowering the risk of endothelial cell damage.

Purpose: To evaluate the biomechanics of corneal ring implants by providing a related mathematical theory and biomechanical model for the treatment of myopia and keratoconus. Methods: The spherical dome model considers the inhomogeneity of the tunica of the eye, dimensions of the cornea, lamellar structure of the corneal stroma, and asphericity of the cornea. It is used in this study for calculating a strengthening factor sf for the characterization of different ring-shaped corneal implant designs. The strengthening factor is a measure of the amount of strengthening of the cornea induced by the implant. Results: For ring segments and incomplete rings, sf = 1.0, which indicates that these implants are not able to strengthen the cornea. The intracorneal continuous complete ring (MyoRing) has a strengthening factor of up to sf = 3.2. The MyoRing is, therefore, able to strengthen the cornea significantly. Conclusions: The result of the presented biomechanical analysis of different ring-shaped corneal implant designs can explain the different postoperative clinical results of different implant types in myopia and keratoconus.