Dysfunctional latent transforming growth factor ß activation after corneal injury in a classical Ehlers-Danlos model.

Patients with classical Ehlers Danlos syndrome (cEDS) suffer impaired wound healing and from scars formed after injuries that are atrophic and difficult to close surgically. Haploinsufficiency in COL5A1 creates systemic morphological and functional alterations in the entire body. We investigated mechanisms that impair wound healing from corneal lacerations (full thickness injuries) in a mouse model of cEDS (Col5a1+/-). We found that collagen V reexpression in this model is upregulated during corneal tissue repair and that wound healing is delayed, impaired, and results in large atrophic corneal scars. We noted that in a matrix with a 50 % content of collagen V, activation of latent Transforming Growth Factor (TGF) ß is dysregulated. Corneal myofibroblasts with a haploinsufficiency of collagen V failed to mechanically activate latent TGF ß. Second harmonic imaging microscopy showed a disorganized, undulated, and denser collagen matrix in our Col5a1+/- model that suggested alterations in the extracellular matrix structure and function. We hypothesize that a regenerated collagen matrix with only 50 % content of collagen V is not resistant enough mechanically to allow adequate activation of latent TGF ß by fibroblasts and myofibroblasts.

Keratocyte-Derived Myofibroblasts: Functional Differences With Their Fibroblast Precursors.

Purpose: In this study, we aim to elucidate functional differences between fibroblasts and myofibroblasts derived from a keratocyte lineage to better understand corneal scarring. Methods: Corneal fibroblasts, derived from a novel triple transgenic conditional KeraRT/tetO-Cre/mTmG mouse strain that allows isolation and tracking of keratocyte lineage, were expanded, and transformed by exposure to transforming growth factor (TGF)-ß1 to myofibroblasts. The composition and organization of a fibroblast-built matrix, deposited by fibroblasts in vitro, was analyzed and compared to the composition of an in vitro matrix built by myofibroblasts. Second harmonic generation microscopy (SHG) was used to study collagen organization in deposited matrix. Different extracellular matrix proteins, expressed by fibroblasts or myofibroblasts, were analyzed and quantified. Functional assays compared latent (TGF-ß) activation, in vitro wound healing, chemotaxis, and proliferation between fibroblasts and myofibroblasts. Results: We found significant differences in cell morphology between fibroblasts and myofibroblasts. Fibroblasts expressed and deposited significantly higher quantities of fibril forming corneal collagens I and V. In contrast, myofibroblasts expressed and deposited higher quantities of fibronectin and other non-collagenous matrix components. A significant difference in the activation of latent TGF-ß activation exists between fibroblasts and myofibroblasts when measured with a functional luciferase assay. Fibroblasts and myofibroblasts differ in their morphology, extracellular matrix synthesis, and deposition, activation of latent TGF-ß, and chemotaxis. Conclusions: The differences in the expression and deposition of extracellular matrix components by fibroblasts and myofibroblasts are likely related to critical roles they play during different stages of corneal wound healing.

Confocal Microscopy, Anterior Segment Optical Coherence Tomography and Clinical Findings in a Non-Granulomatous Uveitis Case for Mpox Infection.

PURPOSE: To report a rare case of non-granulomatous anterior uveitis (clinical, IVCM in vivo confocal microscopy, and anterior optical coherence tomography (OCT) findings) in a non-immunosuppressed patient with mpox infection. METHODS: A 24-year-old male was consulted for bilateral ocular pain, red eye, itchiness, and mucoid discharge. In his left eye, multiple vesicles and papules, some with central ulceration, were found in the superior and inferior eyelids. Mucoid discharge, chemosis, limbitis, and an anterior chamber reaction were also found. A conjunctival PCR swab for mpox was positive. The patient was treated with topical steroids with a good response. RESULTS: OCT and IVCM showed sub-endothelial deposits, stromal edema, hyperreflective multinucleated images in the epithelial layer, activated stromal images, and stromal edema characterized by fusiform hyperreflectivity that was resolved with the proposed treatment. CONCLUSIONS: This is the first report of OCT and IVCM in a non-immunosuppressed patient with mpox infection with a good response to topical steroids.

Stromal matrix directs corneal fibroblasts to re-express keratocan after injury and transplantation.

Every tissue has an extracellular matrix (ECM) with certain properties unique to it - the tissue 'niche' - that are necessary for normal function. A distinct specific population of quiescent keratocan-expressing keratocytes populate the corneal stroma during homeostasis to maintain corneal function. However, during wound healing, when there is alteration of the niche conditions, keratocytes undergo apoptosis, and activated corneal fibroblasts and myofibroblasts attempt to restore tissue integrity and function. It is unknown what the fate of activated and temporary fibroblasts and myofibroblasts is after the wound healing process has resolved. In this study, we used several strategies to elucidate the cellular dynamics of corneal wound healing and the fate of corneal fibroblasts. We injured the cornea of a novel mouse model that allows cell-lineage tracing, and we transplanted a cell suspension of in vitro-expanded corneal fibroblasts that could be tracked after being relocated into normal stroma. These transplanted fibroblasts regained expression of keratocan in vivo when relocated to a normal stromal niche. These findings suggest that transformed fibroblasts maintain plasticity and can be induced to a keratocyte phenotype once relocated to an ECM with normal signaling ECM.

Genipin increases extracellular matrix synthesis preventing corneal perforation.

PURPOSE: Corneal melting and perforation are feared sight-threatening complications of infections, autoimmune disease, and severe burns. Assess the use of genipin in treating stromal melt. METHODS: A model for corneal wound healing was created through epithelial debridement and mechanical burring to injure the corneal stromal matrix in adult mice. Murine corneas were then treated with varying concentrations of genipin, a natural occurring crosslinking agent, to investigate the effects that matrix crosslinking using genipin has in wound healing and scar formation. Genipin was used in patients with active corneal melting. RESULTS: Corneas treated with higher concentrations of genipin were found to develop denser stromal scarring in a mouse model. In human corneas, genipin promoted stromal synthesis and prevention of continuous melt. Genipin mechanisms of action create a favorable environment for upregulation of matrix synthesis and corneal scarring. CONCLUSION: Our data suggest that genipin increases matrix synthesis and inhibits the activation of latent transforming growth factor-ß. These findings are translated to patients with severe corneal melting.

Collagen XII regulates stromal wound closure.

The role of collagen XII in regulating injury repair and reestablishment of corneal function is unknown. This manuscript aims to investigate the role(s) of collagen XII in the repair of incisional and debridement injuries in an adult mouse model. Two different types of injury in wild type and Col12a1-/- corneas were created to investigate the effects of collagen XII -in wound repair and scar formation-by using clinical photographs, immunohistology, second harmonic generation imaging and electron microscopy. Results showed that collagen XII is a regulator of wound closure after incisional injuries. Absence of collagen XII retarded wound closure and the wound healing process. These findings show that collagen XII regulates fibrillogenesis, CD68 cell lineage infiltration, and myofibroblast survival following injury. In vitro studies suggest that collagen XII regulates deposition of an early and provisional matrix by interacting with two proteins regulating early matrix deposition: fibronectin and LTBP1(latent transforming growth factor ß binding protein 1). In conclusion, collagen XII regulates tissue repair in corneal incisional wounds. Understanding the function of collagen XII during wound healing has significant translational value.

Expression of oxysterols in human lenses: Implications of the sterol pathway in age-related cataracts.

Lanosterol, an oxysterol molecule, has been proposed to help maintain lens transparency by inhibiting the formation of protein aggregates. This sterol is produced by the enzyme lanosterol synthase and is part of a metabolic pathway that forms cholesterol as a final step. Abnormalities in lanosterol synthase are responsible for congenital cataracts. The αA-crystallin protein, which acts as a molecular chaperone to lanosterol synthase, has been reported to have anti-protein aggregation, anti-inflammatory and anti-apoptotic properties. In this work, we evaluated the correlation of lanosterol synthase and αA-crystallin in human cataractous lenses with the grade of opacity, as well as the expression of lanosterol synthase, farnesyl DPP, geranyl synthase and squalene epoxidase genes. Lanosterol synthase and αA-crystallin were overexpressed in cataractous lenses as well as farnesyl-DP synthase, squalene epoxidase, lanosterol synthase and geranyl synthase genes in cataratous lenses in comparison with normal lenses. Our data confirm that lanosterol synthase and the sterol pathway are upregulated in cataractous lenses. This argues for a functional role of the oxysterol pathway and its products as an important mediator in the pathogenesis of human cataracts.

Downregulation of collagen XI during late postnatal corneal development is followed by upregulation after injury.

Collagen XI plays a role in nucleating collagen fibrils and in controlling fibril diameter. The aim of this research was to elucidate the role that collagen XI plays in corneal fibrillogenesis during development and following injury. The temporal and spatial expression of collagen XI was evaluated in C57BL/6 wild-type mice. For wound-healing studies in adult mice, stromal injuries were created using techniques that avoid caustic chemicals. The temporal expression and spatial localization of collagen XI was studied following injury in a Col11a1 inducible knockout mouse model. We found that collagen XI expression occurs during early maturation and is upregulated after stromal injury in areas of regeneration and remodeling. Abnormal fibrillogenesis with new fibrils of heterogeneous size and shape occurs after injury in a decreased collagen XI matrix. In conclusion, collagen XI is expressed in the stroma during development and following injury in adults, and is a regulator of collagen fibrillogenesis in regenerating corneal tissue.

Collagen XII Regulates Corneal Stromal Structure by Modulating Transforming Growth Factor-ß Activity.

Collagen XII is a regulator of corneal stroma structure and function. The current study examined the role of collagen XII in regulating corneal stromal transforming growth factor (TGF)-ß activation and latency. Specifically, with the use of conventional collagen XII null mouse model, the role of collagen XII in the regulation of TGF-ß latency and activity in vivo was investigated. Functional quantification of latent TGF-ß in stromal matrix was performed by using transformed mink lung reporter cells that produce luciferase as a function of active TGF-ß. Col12a1 knockdown with shRNA was used to test the role of collagen XII in TGF-ß activation. Col12a1-/- hypertrophic stromata were observed with keratocyte hyperplasia. Increased collagen fibril forward signal was found by second harmonic generation microscopy in the absence of collagen XII. Collagen XII regulated mRNA synthesis of Serpine1, Col1a1, and Col5a1 and deposition of collagens in the extracellular matrix. A functional plasminogen activator inhibitor luciferase assay showed that collagen XII is necessary for latent TGF-ß storage in the extracellular matrix and that collagen XII down-regulates active TGF-ß. Collagen XII dictates stromal structure and function by regulating TGF-ß activity. A hypertrophic phenotype in Col12a1-/- corneal tissue can be explained by abnormal up-regulation of TGF-ß activation and decreased latent storage.

Collagen XIV Is an Intrinsic Regulator of Corneal Stromal Structure and Function.

Collagen XIV is poorly characterized in the body, and the current knowledge of its function in the cornea is limited. The aim of the current study was to elucidate the role(s) of collagen XIV in regulating corneal stromal structure and function. Analysis of collagen XIV expression, temporal and spatial, was performed at different postnatal days (Ps) in wild-type C57BL/6 mouse corneal stromas and after injury. Conventional collagen XIV null mice were used to inquire the roles that collagen XIV plays in fibrillogenesis, fibril packing, and tissue mechanics. Fibril assembly and packing as well as stromal organization were evaluated using transmission electron microscopy and second harmonic generation microscopy. Atomic force microscopy was used to assess stromal stiffness. Col14a1 mRNA expression was present at P4 to P10 and decreased at P30. No immunoreactivity was noted at P150. Abnormal collagen fibril assembly with a shift toward larger-diameter fibrils and increased interfibrillar spacing in the absence of collagen XIV was found. Second harmonic generation microscopy showed impaired fibrillogenesis in the collagen XIV null stroma. Mechanical testing suggested that collagen XIV confers stiffness to stromal tissue. Expression of collagen XIV is up-regulated following injury. This study indicates that collagen XIV plays a regulatory role in corneal development and in the function of the adult cornea. The expression of collagen XIV is recapitulated during wound healing.

Genipin Delays Corneal Stromal Enzymatic Digestion.

Purpose: To evaluate the use of genipin in delaying enzymatic digestion of corneal stroma. Methods: Human corneal stromal tissue was treated with genipin, a known chemical crosslinker, and then along with control tissue was subjected to enzymatic digestion with collagenase. The effects of genipin treatment in retarding stromal digestion were analyzed with phase contrast microscopy, a protein quantification assay, second harmonic generation imaging, and transmission electron microscopy. Results: Genipin increased stromal resistance to enzymatic digestion when compared with untreated stroma. A morphologic analysis and protein quantification showed increased stromal resistance to enzymatic digestion once stromal tissue was treated with genipin. Second harmonic generation imaging revealed persistent fibrillar collagen signaling in genipin-treated tissue in contrast with untreated tissue suggesting that genipin retards collagenolysis. Conclusions: Genipin increases stromal resistance to enzymatic digestion in controlled experiments as demonstrated by protein quantification studies and through morphologic imaging. Translational Relevance: This study explores the novel use of genipin in delaying enzymatic stromal digestion. Delaying stromal melting in the setting of corneal infectious or autoimmune keratitis can potentially decrease clinical morbidity.

Interventions for Demodex blepharitis and their effectiveness: A systematic review and meta-analysis.

PURPOSE: A systematic review and meta-analysis was performed to evaluate the effectiveness of interventions in the treatment ofDemodex blepharitis in adult patients. METHODS: A systematic review and meta-analysis of studies reporting the efficacy of treatments forDemodex blepharitis in the main databases (PubMed / Scopus / Cochrane / EMBASE / Science Direct / WOS / Scielo / Google Scholar / metaRegister of Controlled Trials / ClinicalTrials.gov/ WHO ICTRP) until November 24, 2020 was performed according to the PRISMA statement for meta-analysis. RESULTS: Overall, 18 studies were included for 29 different interventions in 1195 participants with 1574 eyes that were positive for Demodex Spp. Demodex counts, total eradication, clinical improvement, Ocular Surface Disease Index, Tear Break-Up Time, cylindrical dandruff, Schirmer test, osmolarity and adverse reactions were analysed, and stratified sub-analyses conducted. The overall effects for Demodex count (mean difference), total eradication (risk ratio) and adverse reactions (risk difference) were -2.07 (95 % CI -3.99 to -0.15) p = 0.03, 1.84 (95 % CI 1.27-2.66) p = 0.001 and 0.24 (95 % CI 0.08 to 0.41) p = 0.005, respectively. The most frequent interventions evaluated in the included studies were tea tree oil (TTO) and its derivatives, such as terpinen 4-ol. CONCLUSION: Multiple therapeutic choices were evaluated in this meta-analysis. Pharmacological interventions were superior to non-pharmacological (mechanical, thermal and pulsed light) interventions. It was not possible to establish significant differences between TTO and non-TTO-derived treatments. Adverse reactions were more frequent in TTO-derived treatments, however all were mild. It is necessary to execute studies with longer follow-up times to determine whether re-infestation occurs after the administration of different treatments.

Visual outcomes in hyperopic myopic and emmetropic patients with customized aspheric ablation (Q factor) and micro-monovision.

PURPOSE: To evaluate the visual outcomes, visual performance, and stereoacuity in presbyopic patients following treatment by a change in the corneal asphericity and micro-monovision. METHODS: Presbyopic patients with diverse refractive errors and emmetropes (n = 30 eyes) were treated with a custom Q-ablation profile and micro-monovision in the non-dominant eye. There with a difference of Q - 0.30 in the Q profiles between dominant and non-dominant eyes. Patients were assigned in two groups based on the preoperative spherical equivalent (Group 1 + 4.00 to + 0.50, and group 2 neutral to - 3.00). Binocular uncorrected distance visual acuity (binocular UCVA), best-corrected visual acuity (BCVA), binocular uncorrected near visual acuity (binocular UNVA) preoperative and postoperative, spherical equivalent refraction, contrast sensitivity, and stereopsis were analyzed at 1, 3, and 6 months. RESULTS: The mean age was 52.6 ± 5.1 (SD) years. At six months post-operation, the mean binocular uncorrected distance visual acuity (binocular UDVA) was 0.15 ± 0.04 logMAR (20/25-) in group 1, and 0.11 ± 0.05 logMAR (20/25) in group 2, and binocular uncorrected near vision UNVA was 0.5 ± 0.1 M (20/25 J2) in group 1 and 0.45 ± 0.2 M (20/25 J2) group 2. An increase in stereoacuity was found in both groups. CONCLUSIONS: The correction of refractive defects using customized corneal asphericity was an effective treatment in presbyopic patients. Furthermore, the treatment was well tolerated in this group of patients. Following surgery, the quality of vision was adequate, and the stereovision improved in this cohort of patients.

Topical ivermectin-metronidazole gel therapy in the treatment of blepharitis caused by Demodex spp.: A randomized clinical trial.

PURPOSE: To evaluate the efficacy of topical ivermectin-metronidazole combined therapy in the management of Demodex-associatedblepharitis. METHODS: Sixty patients with a diagnosis of Demodex-associatedblepharitis were recruited in a randomized clinical trial. Thirty receiving topical ivermectin (0.1%)-metronidazole (1%) gel treatment on days 0, 15 and 30. Thirty additional patients were used as a control group receiving vehicle on days 0, 15 and 30. The primary efficacy measure was the number of Demodex spp. mitesin the eyelashes of patients. The secondary outcomes included clinical improvement of signs and adverse events. RESULTS: Complete eradication of Demodex spp. was found in 96.6% of patients in the treatment group. Furthermore, a significant reduction of inflammation signs were found in all treated patients versus controls. None of the patients experienced any adverse effects associated with the treatment. CONCLUSION: Demodex infection was controlled satisfactorily with the ivermectin (0.1%)-metronidazole (1%) gel, and no adverse effects were observed. Application of this gel for the treatment of different parasitic infections of the eyelids could be feasible, and this requires further exploration.

Randomised, prospective clinical trial of platelet-rich plasma injection in the management of severe dry eye.

BACKGROUND: To evaluate the effectiveness of platelet-rich plasma (PRP) injections in the treatment of severe dry eye. METHODS: This prospective, intervention study included patients with severe dry eye who had been diagnosed with Sjogren syndrome. Patients were divided into two groups. The intervention group received PRP (n=15) injections on days 0, 30, 60 and 90, as well as hyaluronic acid five times per day. The comparison group received hyaluronic acid (n=15) five times per day. Subjects were measured at baseline and at 30, 60 and 90 days. The primary outcome measures were changes in corneal staining according to the Oxford classification, results of the Schirmer test and tear break-up time (TBUT). The secondary outcome measures were changes in the Ocular Surface Disease Index (OSDI) and treatment compliance. RESULTS: All subjects completed the study. The intervention group showed improvements in all primary outcome measures when compared with the control group, including a reduction in corneal staining (p<0.001), increase in the mean Schirmer value from 5.6±0.7 to 9.0±1.1 mm, and an increase in TBUT from 4.0±0.4 to 6.4±0.4 s at day 90. An improvement in subjective OSDI values was also found. CONCLUSION: PRP injection is safe and effective in improving tear parameters as well as subjective parameters, and was found to be superior to hyaluronic acid alone in the management of patients with severe dry eye. This represent a novel alternative treatment for severe dry eye. TRIAL REGISTRATION NUMBER: NCT02257957.

Effects of genipin corneal crosslinking in rabbit corneas.

PURPOSE: To evaluate the effect of genipin, a natural crosslinking agent, in rabbit eyes. SETTING: Department of Ophthalmology, Universidad Nacional de Colombia Centro de Tecnologia Oftalmica, Bogotá, Colombia. DESIGN: Experimental study. METHODS: Ex vivo rabbit eyes (16; 8 rabbits) were treated with genipin 1.00%, 0.50%, and 0.25% for 5 minutes with a vacuum device to increase corneal permeability. Penetration was evaluated using Scheimpflug pachymetry (Pentacam). In the in vivo model (20 rabbits; 1 eye treated, 1 eye with vehicle), corneas were crosslinked with genipin as described. Corneal curvature, corneal pachymetry, and intraocular pressure (IOP) assessments as well as slitlamp examinations were performed 0, 7, 30, and 60 days after treatment. RESULTS: In the ex vivo model, Scheimpflug pachymetry showed deep penetration in the rabbit corneas with an increase in corneal density and a dose-dependent relationship. Corneal flattening was observed in treated eyes (mean 4.4 diopters ± 0.5 [SD]) compared with the control eyes. Pachymetry and IOP were stable in all evaluations. No eye showed toxicity in the anterior chamber or in the lens. CONCLUSIONS: Corneal crosslinking induced by genipin produced significant flattening of the cornea with no toxicity in rabbit eyes. This crosslinking could be useful in the treatment of corneal ectasia and in the modification of corneal curvature. FINANCIAL DISCLOSURE: None of the authors has a financial or proprietary interest in any material or method mentioned.

Restoration of human lacrimal function following platelet-rich plasma injection.

PURPOSE: The aim was to evaluate the effect of autologous platelet-rich plasma on lacrimal function in patients with severe dry eye. METHODS: A prospective interventional case series design was adopted. Four patients with severe lacrimal dysfunction and severe dry eye were treated. Platelet rich-activated plasma (1 mL) was injected adjacent to the lacrimal gland on day 0 and at 4, 8, and 12 weeks. The objective parameters included a Schirmer test, ocular surface staining, and tear break-up time (TBUT). The patients were followed up for 12 weeks after the first injection. RESULTS: All cases showed a significant improvement in lacrimal volume (from 3.3 ± 0.8 mm to 11.1 ± 2.3 mm). In all the patients, an increase in tear break-up time values and a decrease in ocular staining (basal 8.0 ± 0.61-2.8 ± 0.5) with subjective improvement occurred. None of the patients presented any adverse effect, and none reported pain or discomfort. Additionally, no complications were observed. CONCLUSIONS: Injected platelet-enriched plasma was found to be safe and effective in increasing lacrimal production and in improving ocular staining secondary to severe dry eye. This approach could be an alternative for the management of these patients, although additional studies are required to perfect the technique.

Corneal crosslinking with genipin, comparison with UV-riboflavin in ex-vivo model.

PURPOSE: To investigate the efficacy and safety of Genipin and UV-riboflavin crosslinking (UV-CLX) in corneal crosslinking. METHODS: Porcine eyes were separated in groups for each crosslinker, genipin 0.25% UV-CLX (clinical crosslinking procedure), glutaraldehyde 0.1% (gold standard crosslinker), and control eyes. Intraocular pressure (IOP) was continuously monitored by a pressure sensor cannulated to the anterior chamber and the volume was changed. The changes in ocular pressure as a function of change of the ocular volume were evaluated. Ocular rigidity was calculated as the exponential of polynomial quadratic fit. Endothelial damage was evaluated in a viability assay with alizarin red staining as the changes in cell counts. RESULTS: Significant changes in IOP were observed in the globes were the cornea was stiffened with genipin and UV-CLX treatment (volume 200 µl: Genipin 19.4 mmHg, UVCRX 18.8 mmHg, glutaraldehide 23.9 mmHg, versus control 14.7 mmHg, and 400 µl genipin 31.5 mmHg, UV-CLX 26.0 mmHg, glutaraldehide 37.3 mmHg versus control 18.7 mmHg). The mean ocular ridigity coefficient was genipin 0.0078 mmHg/µl, UV-CLX 0.0065 mmHg/µl, glutaraldehide 0.0092 mmHg/µl, and 0.0046 mmHg/µl for control eyes. Endothelial cell damage was 5.9±1.8% (control), 10.3±1.7% (UV-CLX), 9.4±1.5% (Genipin 0.25%), and 40.1±6.2% (glutaraldehide). Some granules were observed in the UV-CLX group. Reduction of keratocites was observed in the UV CRX crosslinking. CONCLUSIONS: Corneal crosslinking was similar between UV-CLX and genipin with minimal toxicity to endothelial cells. Stiffened corneas by any method induced substancially higher IOP elevation when ocular volume is increased.

Effect of genipin collagen crosslinking on porcine corneas.

PURPOSE: To evaluate the effect of genipin, a natural crosslinker, on porcine corneas. SETTING: Department of Ophthalmology, Universidad Nacional de Colombia, Bogota, Colombia. METHODS: Corneal strips (12.0 mm x 2.3 mm) were harvested from porcine eyes and treated by incubation with genipin at concentrations of 1.00%, 0.25%, and 0.10%. Parallel corneal strips from the same eye were used as untreated controls. After treatment at 20 degrees C for 40 minutes, tensile strain measurements were performed in a biomaterial tester. Porcine button corneas were treated with genipin 0.25% for 15 minutes and then digested by bacterial collagenase. Treated and untreated corneas were evaluated by light microscopy. RESULTS: Young modulus and stiffness in treated corneas increased in a concentration-dependent manner. Genipin increased resistance to corneal collagenase 5-fold in comparison with the controls. A decrease in the interlamellar space in treated corneas was also observed. CONCLUSIONS: Corneal collagen crosslinking induced with genipin produced a significant increase in biomechanical strength and resistance to bacterial collagenase. This crosslinker could be useful in treating corneal ectasia and corneal infectious and noninfectious diseases involving corneal melting.

Nucleoside-derived antagonists to A3 adenosine receptors lower mouse intraocular pressure and act across species.

The purpose of the study was to determine whether novel, selective antagonists of human A3 adenosine receptors (ARs) derived from the A3-selective agonist Cl-IB-MECA lower intraocular pressure (IOP) and act across species. IOP was measured invasively with a micropipette by the Servo-Null Micropipette System (SNMS) and by non-invasive pneumotonometry during topical drug application. Antagonist efficacy was also assayed by measuring inhibition of adenosine-triggered shrinkage of native bovine nonpigmented ciliary epithelial (NPE) cells. Five agonist-based A3AR antagonists lowered mouse IOP measured with SNMS tonometry by 3-5 mm Hg within minutes of topical application. Of the five agonist derivatives, LJ 1251 was the only antagonist to lower IOP measured by pneumotonometry. No effect was detected pneumotonometrically over 30 min following application of the other four compounds, consonant with slower, smaller responses previously measured non-invasively following topical application of A3AR agonists and the dihydropyridine A3AR antagonist MRS 1191. Latanoprost similarly lowered SNMS-measured IOP, but not IOP measured non-invasively over 30 min. Like MRS 1191, agonist-based A3AR antagonists applied to native bovine NPE cells inhibited adenosine-triggered shrinkage. In summary, the results indicate that antagonists of human A3ARs derived from the potent, selective A3 agonist Cl-IB-MECA display efficacy in mouse and bovine cells, as well. When intraocular delivery was enhanced by measuring mouse IOP invasively, five derivatives of the A3AR agonist Cl-IB-MECA lowered IOP but only one rapidly reduced IOP measured non-invasively after topical application. We conclude that derivatives of the highly-selective A3AR agonist Cl-IB-MECA can reduce IOP upon reaching their intraocular target, and that nucleoside-based derivatives are promising A3 antagonists for study in multiple animal models.