Glycosaminoglycans and collagen fibril distribution at various depths of the corneal stroma of normal and CXL treated rats.

Corneal collagen cross-linking (CXL) is widely used to treat keratoconus and ecstatic corneal disorders. The present studies were carried out to investigate the distribution of glycosaminoglycans (GAGs) and collagen fibril (CF) at different depths of the normal and CXL treated corneal stroma of four week old rats 7 days after standard CXL application. Ten Wistar rats' corneas were used for the study. The epithelium of the cornea from the left eye of each rat was removed and treated with standard CXL application using riboflavin and Ultraviolet-A (UVA) (3 mW/cm2 for 30 min). The cornea from the right eye was used as the control cornea. The cornea was removed from the eye and processed for transmission electron microscopy. A bottom mounted Quemesa camera was used to capture digital images and these images were analysed using iTEM software. In the control cornea, the GAGs area size was not significantly different in the anterior, middle, and posterior stroma. In the CXL treated rats the GAGs area size gradually increased from the anterior to the posterior stroma whereas the spacing between the GAGs gradually decreased. There were very large GAGs present in the posterior stroma of the CXL treated rats. When comparing the control and CXL cornea, the GAGs area in the CXL cornea was significantly higher and inter-GAGs-spacing was smaller than in the control cornea. In the control cornea, the collagen fibrils diameter was higher in the anterior stroma and lowest in the posterior stroma. In the CXL treated cornea, the CF diameter and the interfibrillar spacing gradually decreased from the anterior to the posterior stroma. On comparison between the control and the CXL treated cornea, the interfibrillar spacing was significantly smaller in the CXL treated cornea than the control cornea in the anterior, middle, and posterior stroma but there was no difference in the diameter. The CXL treatment significantly increased the GAGs area and decreased the inter-GAGs-spacing, and inter-CF-spacing. This could be due to the gradual decline in the availability of riboflavin, UVA, and oxygen in the middle and posterior stroma. Further studies are required to investigate the role of keratan sulphate and chondroitin sulphate by using monoclonal antibodies with immunogold technique.

Grapefruit Extract-Mediated Fabrication of Photosensitive Aluminum Oxide Nanoparticle and Their Antioxidant and Anti-Inflammatory Potential.

Aluminum oxide nanoparticles (Al2O3 NPs) were synthesized using a simple, eco-friendly green synthesis approach in an alkaline medium from the extract of grapefruit peel waste. The pre-synthesized, nano-crystalline Al2O3 NPs were characterized by using spectroscopic (UV-vis, FTIR, XRD, and EDX) and microscopic (SEM and TEM) techniques. The formed Al2O3 NPs exhibited a pronounced absorption peak at 278 nm in the UV-vis spectrum. The average particle size of the as-prepared Al2O3 NPs was evaluated to be 57.34 nm, and the atomic percentages of O and Al were found to be 54.58 and 45.54, respectively. The fabricated Al2O3 NPs were evaluated for antioxidant, anti-inflammatory, and immunomodulatory properties. The Al2O3 NPs showed strong antioxidant potential towards all the four tested assays. The anti-inflammatory and immunomodulatory potential of Al2O3 NPs was investigated by measuring the production of nitric oxide and superoxide anion (O2•-), as well as proinflammatory cytokines tumour necrosis factor (TNF-α, IL-6) and inhibition of nuclear factor kappa B (NF- κB). The results revealed that Al2O3 NPs inhibited the production of O2•- (99.4%) at 100 µg mL-1 concentrations and intracellular NO•- (55%), proinflammatory cytokines IL-6 (83.3%), and TNF-α (87.9%) at 50 µg mL-1 concentrations, respectively. Additionally, the Al2O3 NPs inhibited 41.8% of nuclear factor kappa B at 20 µg mL-1 concentrations. Overall, the outcomes of current research studies indicated that Al2O3 NPs possess anti-inflammatory and immunomodulatory properties and could be used to treat chronic and acute anti-inflammatory conditions.

Effect of Ultraviolet-A and Riboflavin treatment on the architecture of the center and periphery of normal rat cornea: 7 days post treatment.

Corneal collagen cross-linking (CXL) is a treatment that is widely applied to halt the progression of ectatic diseases such as keratoconus by creating biomechanical strength in the cornea. Most of the studies assessed the effect of the CXL on the cornea without any differentiation of its effect between periphery and the center of the untreated control cornea especially after the 7 days of CXL application. We investigate the ultrastructural changes in the architecture of the center and periphery of rat corneas, 7 days after standard CXL application. Five Wistar rats (10 corneas) were used in the present study. The left eye corneas (5 mm area) were de-epithelialized and irradiated with standard CXL application using riboflavin and Ultraviolet-A (UVA) (3 mW/cm2 for 30 min). The right eye corneas were used as a control. The sclera-cornea button was removed and processed for electron microscopy. Digital images were captured with a bottom mounted Quemesa camera and analyzed using the iTEM software. The ultrastructure of epithelium, hemi-desmosomes, Bowman's layer and stroma were organized in both untreated control and CXL rat cornea in both untreated control and CXL rat cornea. Within the same CXL cornea, both the collagen fibril (CF) diameter and interfibrillar spacing at the center were significantly smaller compared to the peripheral diameter and spacing of the cornea. When comparing the untreated control and CXL cornea, the central interfibrillar spacing of the CXL cornea was significantly smaller than the central spacing the untreated control cornea. In the CXL cornea the peripheral spacing was significantly higher compared to the peripheral interfibrillar spacing of the untreated control cornea. Within the CXL cornea, the proteoglycans (PGs) area and density of the periphery was significantly higher compared to the area and density of the center of the cornea. It suggests that CXL was more effective at the periphery of the cornea. This could be due to the higher amount of leucine rich PG lumican and higher diffusion of oxygen and riboflavin at the periphery cornea.

Corneal Imaging and Densitometry Measurements in Juvenile and Adult Keratoconus Patients to Evaluate Disease Progression and Treatment Effects After Corneal Cross-Linking.

PURPOSE: In this study densitometry software for the Oculus Pentacam was used to investigate the treatment outcomes of corneal cross linking (CXL) in adult and juvenile keratoconus (KCN) patients. Densitometry measurements were taken before and after treatment and followed up for one year. METHODS: A comparative study was carried out at Manchester Royal Eye Hospital. Corneal densitometry measurements collected before and after CXL treatment for 32 eyes from KC patients, aged between 12 and 39, were divided to 2 groups 13-18 years (juvenile group) and 19-39 years (adult group) and analysed and compared to pre and post treatment at 3, 6 and 12 months for each group and between both groups. RESULTS: Analysis of densitometry measurements found higher corneal densitometry after CXL which peaks at three months post treatment in both groups. There was significant diversity in corneal densitometry measurements in the stromal zone 0-2 and 2-6 mm for all layers except the posterior layer for both groups (P<0.05). Significantly increased densitometry value was found higher in the juvenile group at six months in the central (P=0.006) and posterior (P=0.004) layers for zone 0-2 mm. The same layers differed significantly also in the 2-6 mm zone in all layers (P=0.01). One year post treatment the same significant increased densitometry level was seen in the juvenile group in the 0-2 mm zone of the central (P=0.007) and posterior layers (P=0.01), as was the 2-6 mm zone (P=0.04). However, no significant difference was found between pre and post treatment for best corrected visual acuity (BCVA), central corneal thickness (CCT) and thinnest area between both groups. A significant difference was found between pre and post treatment for best corrected visual acuity (BCVA), in the adult group at 6 and 12 months post-treatment from pre-treatment (P=0.02, P=0.16) respectively. CONCLUSION: Corneal clarity post CXL treatment in the juvenile group differed significantly from the adult group. Both groups showed increased haze at 3 months post treatment but the adults showed improvement over the next 9 months. In contrast, the juvenile group showed higher densitometry readings at both 6 and 12 months post treatment in comparison to adult group. The reasons for this remain unclear.

Evaluation of corneal symmetry after UV corneal crosslinking for keratoconus.

PURPOSE: The purpose of this study was to assess UV corneal crosslinking (CXL) treatment outcomes for keratoconus by evaluating the corneal regularity in patients through follow-up using the Oculus Pentacam. PATIENTS AND METHODS: A total of 18 eyes from CXL patients with keratoconus were studied before and after CXL treatment, and six eyes from six patients who were not treated with CXL served as controls. Treated patients had Pentacam images taken before CXL treatment and regularly 3 months post treatment up to the 12th month. Controls were imaged during their first appointment and after 12 months. Symmetry and asphericity were evaluated and correlated with both best-corrected visual acuity (BCVA) and maximum K-readings. RESULTS: In the CXL-treated group, there was a significant improvement in the index of symmetrical variation (ISV) and keratoconus index (KI) at 3 months and in the index of height asymmetry (IHA) and minimum radius of curvature (Rmin) at 9 months post treatment. On the contrary, the untreated group's indices showed some significant worsening in ISV, KI, central keratoconus index (CKI), and Rmin. A novel finding in our study was a slight positive shift of anterior asphericity in the 6 mm, 7 mm, and 8 mm 3 months after treatment, which had a correlation with BCVA (R2=0.390, p=0.053) and a strong correlation with maximum K-reading (R2=0.690, p=0.005). However, the untreated group had no significant changes after 1 year. CONCLUSION: The corneal asymmetrical shape is associated with the spherical aberration alteration influenced by temporal evolution of surface ablation and increased corneal haze. However, insignificant changes in symmetry attest the stabilization effect on cornea postoperatively as compared with controls.