Consecutive dosing of UVB irradiation induces loss of ABCB5 expression and activation of EMT and fibrosis proteins in limbal epithelial cells similar to pterygium epithelium.

Pterygium pathogenesis is often attributed to a population of altered limbal stem cells, which initiate corneal invasion and drive the hyperproliferation and fibrosis associated with the disease. These cells are thought to undergo epithelial to mesenchymal transition (EMT) and to contribute to subepithelial stromal fibrosis. In this study, the presence of the novel limbal stem cell marker ABCB5 in clusters of basal epithelial pterygium cells co-expressing with P63α and P40 is reported. ABCB5-positive pterygium cells also express EMT-associated fibrosis markers including vimentin and α-SMA while their ß-catenin expression is reduced. By using a novel in vitro model of two-dose UV-induced EMT activation on limbal epithelial cells, we could observe the dysregulation of EMT-related proteins including an increase of vimentin and α-SMA as well as downregulation of ß-catenin in epithelial cells correlating to downregulation of ABCB5. The sequential irradiation of limbal fibroblasts also induced an increase in vimentin and α-SMA. Taken together, these data demonstrate for the first time the expression of ABCB5 in pterygium stem cell activity and EMT-related events while the involvement of limbal stem cells in pterygium pathogenesis is exhibited via sequential irradiation of limbal epithelial cells. The later in vitro approach can be used to further study the involvement of limbal epithelium UV-induced EMT in pterygium pathogenesis and help identify novel treatments against pterygium growth and recurrence.

The Role of Limbal Epithelial Stem Cells in Regulating Corneal (Lymph)angiogenic Privilege and the Micromilieu of the Limbal Niche following UV Exposure.

The cornea is a clear structure, void of blood, and lymphatic vessels, functioning as our window to the world. Limbal epithelial stem cells, occupying the area between avascular cornea and vascularized conjunctiva, have been implicated in tissue border maintenance, preventing conjunctivalisation and propagation of blood and lymphatic vessels into the cornea. Defects in limbal epithelial stem cells are linked to corneal neovascularisation, including lymphangiogenesis, chronic inflammation, conjunctivalisation, epithelial abnormalities including the presence of goblet cells, breaks in Bowman's membrane, persistent epithelial defects and ulceration, ocular surface squamous neoplasia, lipid keratopathy, pain, discomfort, and compromised vision. It has been postulated that pterygium is an example of focal limbal deficiency. Previous reports showing changes occurring in limbal epithelium during pterygium pathogenesis suggest that there is a link to stem cell damage. In this light, pterygium can serve as a model disease of UV-induced stem cell damage also characterised by corneal blood and lymphangiogenesis. This review focuses on the role of corneal and limbal epithelial cells and the stem cell niche in maintaining corneal avascularity and corneal immune privilege and how this may be deregulated following UV exposure. We present an overview of the PUBMED literature in the field as well as recent work from our laboratories.

Distribution of conjunctival ultraviolet autofluorescence in a population-based study: the Norfolk Island Eye Study.

OBJECTIVE: The objective of this study was to describe the distribution of conjunctival ultraviolet autofluorescence (UVAF) in an adult population. METHODS: We conducted a cross-sectional, population-based study in the genetic isolate of Norfolk Island, South Pacific Ocean. In all, 641 people, aged 15 to 89 years, were recruited. UVAF and standard (control) photographs were taken of the nasal and temporal interpalpebral regions bilaterally. Differences between the groups for non-normally distributed continuous variables were assessed using the Wilcoxon-Mann-Whitney ranksum test. Trends across categories were assessed using Cuzick's non-parametric test for trend or Kendall's rank correlation τ. RESULTS: Conjunctival UVAF is a non-parametric trait with a positively skewed distribution. Median amount of conjunctival UVAF per person (sum of four measurements; right nasal/temporal and left nasal/temporal) was 28.2 mm(2) (interquartile range 14.5-48.2). There was an inverse, linear relationship between UVAF and advancing age (P<0.001). Males had a higher sum of UVAF compared with females (34.4 mm(2) vs 23.2 mm(2), P<0.0001). There were no statistically significant differences in area of UVAF between right and left eyes or between nasal and temporal regions. CONCLUSION: We have provided the first quantifiable estimates of conjunctival UVAF in an adult population. Further data are required to provide information about the natural history of UVAF and to characterise other potential disease associations with UVAF. UVR protective strategies should be emphasised at an early age to prevent the long-term adverse effects on health associated with excess UVR.

The effect of mesenchymal stem cell conditioned media on corneal stromal fibroblast wound healing activities.

AIMS: To investigate the effects of conditioned media from mesenchymal stem cells (MSC) on the wound healing activities of corneal stromal fibroblasts. METHODS: Cell cycle analysis and early stage activation of apoptosis, chemotactic chambers and fibroblast-populated type I collagen gels were used to assess corneal stromal fibroblast proliferation, migration and contraction, respectively. Fibroblasts were obtained from human donor corneas and MSC from fresh rat bone marrow. MSC conditioned media and fibroblast culture medium (FCM), with and without calf serum supplementation, were compared. RESULTS: MSC conditioned media and serum-free FCM had an inhibitory effect on the progression of corneal fibroblasts through the cell cycle. There was a significant increase in the number of cells in the G0-G1 phase for MSC conditioned media and serum-free FCM (p=0.001, p=0.97 respectively). Fibroblast migration and relaxed and stressed gel contraction were significantly inhibited by MSC conditioned media and serum-free FCM compared with FCM with serum (all p=0.001). Glucose and lactate analysis confirmed that these factors were not contributing to this effect. CONCLUSION: MSC conditioned media was found to inhibit the wound healing activities of corneal stromal fibroblasts in vitro. Putative factors secreted by MSC could be developed for therapeutic use in corneal repair.

Miyake-Apple study of the rotational stability of the Acrysof Toric intraocular lens after experimental eye trauma.

PURPOSE: To evaluate the rotational stability of the Acrysof Toric intraocular lens (IOL) after experimental eye trauma. METHODS: Two human cadaver eyes were prepared in accordance with the Miyake-Apple technique, with closed-system modification. After phacoemulsification, an Acrysof Toric IOL was implanted in the capsular bag. Experimental traumatisation of the globe was achieved by pressing against the eye wall using a cotton tip or a fingertip, whereas the IOL was observed from the posterior view. Digital photographs of the eye before and after the procedures were overlaid to detect and measure rotation of the IOL. RESULTS: The IOL rotated 5.80 degrees when the traumatic procedures did not cause important leakage from the incision. When the traumatic procedures caused important leakage from the incision and anterior chamber collapse, the IOL rotated 41.00 degrees. CONCLUSION: Ocular trauma can cause rotation of the Acrysof Toric IOL. In the event of an eye trauma with no or insignificant leakage from the incision, the IOL rotates less than when the trauma causes significant leakage from the incision.

Multichannel surface recordings on the visual cortex: implications for a neuroprosthesis.

Using a multi-channel platinum surface electrode array, recordings from cat primary visual cortex were obtained in response to visual stimuli, and electrical stimuli delivered using the elements of the array itself. Neural responses to electrical stimuli were consistent, regardless of stimulus polarity or leading phase (biphasic), although thresholds were lower for monophasic than biphasic pulses. Both visual and electrical stimuli reliably evoked responses with characteristic components, which interacted with each other in a nonlinear summation showing first facilitation then suppression during the window of interaction. The chronaxie for eliciting threshold cortical responses was about 100 mus, and the charge density with a pulse width of 50-100 mus was around 55 muC cm(-2). These data form the basis of understanding the types of cortical responses to stimuli delivered by devices suitable for chronic implantation.

Preventing conjunctival autograft inversion in pterygium surgery.

BACKGROUND: Surgeon-dependent variables influencing pterygium surgical outcome using the conjunctival autograft technique include conjunctival retraction as a consequence of subepithelial contracting fibrous tissue, and autograft inversion causing necrosis and sloughing of the graft. METHOD: A simple and useful technique of pterygium excision is described, which helps to ensure the correct surface and linear orientations of the conjunctival autograft, and also defines the end point of adequate excision of the subepithelial connective tissue. CONCLUSION: This simple technique of defining the anterior surface and the centrifugal orientation with the letter "G" marked on the graft prevents reverse orientation of the graft.

Mechanosensitivity and the eye: cells coping with the pressure.

The cells of the various organ systems in humans are subject to mechanical forces to which they must respond. Here the authors review what is known of the ways in which the cells of animals, ranging from the prokaryotic to humans, sense and transduce mechanical forces to respond to such stimuli. In what way this pertains to the eye, especially with respect to axial myopia and the pressure related disease of glaucoma, is then surveyed.

Lower eyelid tensometry in younger and older normal subjects.

PURPOSE: This study aimed to establish normative and repeatability data for lower eyelid tensometry (LET) using a newly developed eyelid tensometer. METHODS: In this prospective consecutive observational case series, 32 normal adult subjects, comprising 12 younger (aged 29+/-5 years, 6M:6F) and 20 older subjects (aged 74+/-6 years, 10M:10F), underwent LET. In the younger group, LET was measured for postero-anterior (PA), nasal and temporal displacement. Duplicate measurements were taken and these were repeated on a separate occasion. Associations between repeated measurements and between right and left eyes were determined and the coefficient of repeatability for PA measurements was estimated. Differences in LET between males and females were determined using ANOVA. Only PA tensions, with duplicate measurements, were assessed in the older group, and age effects were determined. RESULTS: Mean temporal tension in the younger group was 7.8+/-2.9 mN (milliNewtons)/mm, while nasal tension was 13.0+/-4.6 mN/mm and PA tension was 11.2+/-5.3 mN/mm. There was a good correlation between interoccasion PA measurements (r=0.82-0.84, P<0.005) and between the right and left eyes (r=0.48, P<0.005). The coefficient of repeatability for PA tension was 1.8 mN/mm. In the younger group, overall eyelid tension (for all directions) for males was significantly higher than for females (ANOVA, P=0.03). For measurements in older subjects on a single occasion, PA tension was 11.1+/-3.5 mN/mm and in younger subjects 12.8+/-4.2 mN/mm (P=0.22). CONCLUSIONS: LET is rapid, repeatable, feasible, and acceptable. PA tension was the most repeatable measurement. Younger males had higher eyelid tension than females, and there was no significant reduction in PA tension with age.

Human ganglion cells express the alpha-2 adrenergic receptor: relevance to neuroprotection.

BACKGROUND/AIM: Alpha-2alpha adrenergic receptor (alpha(2)-AR) agonists are thought to be neuroprotective, preventing retinal ganglion cell death independent of pressure reduction. Previous studies have identified alpha(2)-ARs in rat retina. The authors aimed to demonstrate the presence and localisation of alpha(2)-ARs in human and rat retina and on the rat retinal ganglion cell line, RGC-5. METHODS: Seven postmortem human and three postmortem rat eyes were paraformaldehyde fixed and frozen. RGC-5 cells were also paraformaldehyde fixed. The expression of alpha(2A)-ARs was determined by antibody immunofluorescence. RESULTS: alpha(2A)-AR expression was identified in the human retina, on ganglion cells, and cells in the inner and outer nuclear layers (INL, ONL). Differential alpha(2A)-AR staining patterns in the INL and ONL suggest a further restriction to as yet unidentified neuronal subclasses. The RGC-5 cell line also expressed alpha(2A)-ARs in undifferentiated cells and an increased expression upon fully differentiated cells. CONCLUSION: alpha(2)-AR agonists in addition to their pressure lowering effects in the eye, may act directly upon retinal neurons, including retinal ganglion cells. The presence of alpha(2)-ARs on the RGC-5 cell line allows future investigation of these possible direct effects using in vitro glaucoma model systems.

Case-controlled clinical and histopathological study of conjunctivochalasis.

BACKGROUND/AIMS: Conjunctivochalasis, a secondary cause of the watery eye, is frequently seen in the older age group as an elevation of the bulbar conjunctiva lying along the lateral or central lower lid margin. A prospective, interventional, case-controlled clinical and histopathological study was conducted. The relevant features of 18 patients (29 eyes) who had their conjunctivochalasis resected as part of the surgical management of their watery eye syndrome were examined. In the control group, tissue was obtained from an age matched series of 24 normal subjects undergoing routine cataract surgery. METHODS: 24 controls (24 specimens) and 18 patients (29 specimens) had conjunctival strip biopsies, taken from the usual lid margin level bulbar conjunctiva in line with the inferior limbus (controls), and the clinically apparent conjunctivochalasis (patients). These were submitted for histological study. RESULTS: 23 of 24 control sections demonstrated normal conjunctival variation. Four of 29 patient specimens demonstrated a chronic non-granulomatous conjunctivitis, while three eyes of the patient group (two patients) demonstrated features of elastosis. Of the four patients who had the inflammatory infiltrates, three had functional nasolacrimal duct obstructions (FNLDOs) and one had a primary acquired nasolacrimal duct obstruction (PANDO). Of the two patients who had elastosis, one had an FNLDO and the other had normal lacrimal drainage and was Jones 1 positive. CONCLUSION: Six of 18 patients--that is, seven of 29 specimens of conjunctivochalasis demonstrated signs of elastosis or of chronic non-granulomatous inflammation. Clinically, patients had a spectrum of aetiologies of their watery eye syndrome.

In vivo confocal microscopy of the human cornea.

AIMS: To describe the optics of in vivo confocal microscopy, its advantages over previous methods, and to summarise the literature that arose from its use for the observation of the human cornea. A critical review of the clinical usefulness of this new technology for the corneal examination is undertaken. METHODS: Confocal microscopes obtain increased resolution by limiting the illumination and observation systems to a single point. Rapid scanning is used to reconstruct a full field of view and allows for "real time" viewing. RESULTS: Coronal sections of the in situ epithelium, Bowman's membrane, stroma, and endothelium can be visualised at a resolution of 1-2 micro m. A backscattered light intensity curve allows objective measurements of sublayer thickness and corneal haze to be taken. In vivo confocal microscopy is therefore particularly useful in the areas of infective keratitis, corneal dystrophies, refractive surgery, and contact lens wear, where it aids in differential diagnosis and detection of subtle short and long term changes. Real time endothelial cell assessment can also be performed. CONCLUSION: Because of their ability to visualise living tissue at cellular levels, confocal microscopes have proved useful additions to the current clinical tools.

Australasian orbital and adnexal Wegener's granulomatosis.

OBJECTIVE: To report a retrospective case series of 29 Australian and New Zealand patients with orbital and adnexal Wegener's granulomatosis (WG). DESIGN: Retrospective case series. PARTICIPANTS: Twenty-nine cases of orbital and adnexal WG were identified. METHODS: A number of oculoplastic surgeons and other clinicians in Australia and New Zealand was asked about their experience with orbital and adnexal WG. Clinical data regarding these cases were conveyed by means of a questionnaire. Cases of ophthalmic WG without features of orbital or adnexal disease were excluded. MAIN OUTCOME MEASURES: Data obtained from the questionnaire includes age, gender, limited or generalized disease, antineutrophil cytoplasmic antibody (ANCA) status, symptoms and signs: nasolacrimal obstruction, sinusitis, fistula/orbital bone erosion, orbital mass/proptosis, extraocular muscle/diplopia, visual acuity reduction caused by optic nerve compression, orbital pain, lid edema/erythema, biopsy status, and treatment status. RESULTS: Twenty-nine patients with orbital and adnexal WG were identified and described. Symptoms included awareness of an orbital mass, epiphora, orbital pain and diplopia. Signs included an orbital mass or proptosis (69%), nasolacrimal duct obstruction (52%), limited ocular rotations (52%), lid erythema and edema (31%), bony destruction (21%), and reduced visual acuity (17%). Two patients had a persistent nasolacrimocanthal fistula. Cytoplasmic pattern antineutrophil cytoplasmic antibodies (c-ANCA) were present in 52% of patients, and in 9 of 10 patients with generalized disease. However, c-ANCA was positive in only 32% (6 of 19) of patients with limited WG. Perinuclear pattern antineutrophil cytoplasmic antibodies (p-ANCA) was positive in 10% of cases. CONCLUSIONS: To diagnose and treat ophthalmic WG effectively, the clinician must be aware of its protean orbital and adnexal manifestations. WG may occur with or without systemic involvement, and c-ANCA was negative in approximately half our cases. Our cases also demonstrated two orbital fistulae, an observation previously believed to be rare.

Active matrilysin (MMP-7) in human pterygia: potential role in angiogenesis.

PURPOSE: Pterygia are invasive, proliferative fibrovascular growths, with the matrix metalloproteinase (MMP) family of enzymes strongly implicated in the pathogenesis of these lesions. The purpose of this study was to determine the cellular distribution and activation status of matrilysin (MMP-7) in pterygia. METHODS: Resected pterygia (n = 8) and normal conjunctiva (n = 8) were sectioned and analyzed immunohistochemically with two different epitope-specific anti-MMP-7 monoclonal antibodies (Abs) which differentiate pro- and active MMP-7. The specificity of each Ab was confirmed by Western blot analysis of p-aminophenylmercuric acetate (APMA)-activated and latent recombinant MMP-7. Pterygia (n = 4) and autologous normal conjunctiva (n = 4) were placed in organ culture to determine the activation status of secreted MMP-7. RESULTS: Precursor and active forms of MMP-7 were detected in epithelial cells from both pterygia and normal conjunctiva. Intense immunoreactivity for pro- and active MMP-7 was also observed in the pterygium vasculature, but was essentially absent from conjunctival vessels. Pro-MMP-7 was also identified in the epithelial basement membrane and associated with matrix components in pterygia. The 141-7B2 Ab reacted with the 30-kDa latent MMP-7, and the IM47L Ab precipitated a 19-kDa active enzyme, thus confirming the differential specificity of each Ab. Pro- and active MMP-7 were increased 1.4- and 2.7-fold, respectively, in the supernatants from organ-cultured pterygia compared with conjunctiva. CONCLUSIONS: This study is the first to specifically localize an active MMP species in pterygia and strengthens the hypothesis that these enzymes are involved in the pathogenesis of this disease. The data also suggest that MMP-7 may play a significant role in the angiogenesis that characterizes this lesion. Future studies will be directed at determining whether targeting MMP activity may be useful for treatment of pterygia.

Lower nasal distribution of cortical cataract: the Blue Mountains Eye Study.

Previous reports have indicated that cortical cataract commences in the lower nasal lens, possibly due to sunlight exposure because of the shallow brow temporally. The present study aimed to assess the lens sector distribution of cortcal cataract in a population. The Blue Mountains Eye Study assessed 3654 residents aged 49-97 years; 3270 phakic subjects had retroillumination lens photographs graded using the Wisconsin method, which divides the lens into eight radial sectors with a grid overlay. Graders estimated percentage of cortical opacity in each sector. The lower nasal area consistently had the greatest opacity, a pattern present at each age and similar in both sexes, despite the 20% greater cortical cataract in women. The mean area of lens cortex involved by opacities in the lower nasal hemisphere was four fold greater than in the upper temporal hemisphere at each age. The lower nasal distributon was highly symmetrical when both eyes were affected. When > 20% of the lower nasal lens quadrant had cortical opacity, 88% of bilaterally affected subjects had cortical cataract n the same quadrant of the fellow eye. The lower nasal distribution may indicate a role for sunlight in the aetiology of cortcal cataract, which could be considered when examining other cortical cataract risk factors, such as diabetes, vascular disease and hormonal factors in women.

Clinical evaluation of twice-daily emedastine 0.05% eye drops (Emadine eye drops) versus levocabastine 0.05% eye drops in patients with allergic conjunctivitis.

PURPOSE: The efficacy and safety of emedastine 0.05% eye drops (Emadine; Alcon Laboratories, Inc, Fort Worth, Texas), a new H(1) antagonist, were studied in comparison to levocabastine 0.05% eye drops (Livostin; Janssen-Cilag N V, Berchem, Belgium) during a twice-daily treatment schedule for 6 weeks in adult and pediatric patients with seasonal allergic conjunctivitis. METHODS: In a prospective, multicenter, randomized, double-masked, parallel group study, 222 patients with allergic conjunctivitis were randomized (221 received treatment) to either emedastine or levocabastine, instilled twice daily for 6 weeks. Patient diaries were completed four times daily (before the morning and evening instillations, at noon, and in the afternoon), and clinical examinations were conducted at regular intervals. Primary efficacy variables of ocular redness and itching and secondary efficacy variables of chemosis, eyelid swelling, patient diary data, and physician's global assessment were analyzed. RESULTS: Both emedastine and levocabastine produced a statistically significant (P =.0001) reduction in itching and redness within 5 minutes of the first instillation. All signs and symptoms improved progressively over the 6-week treatment period. After 7 days of use, and throughout the remainder of the study, emedastine was statistically superior to levocabastine (P <.006) in preventing and alleviating the signs and symptoms (itching, redness, chemosis, and eyelid swelling) of allergic conjunctivitis. CONCLUSIONS: Emedastine 0.05% eye drops administered twice daily are more efficacious than levocabastine 0.05% eye drops in the prevention and treatment of the signs and symptoms of allergic conjunctivitis in adults and children of 4 years and above. Both emedastine 0.05% eye drops and levocabastine 0.05% eye drops were well tolerated.

Transcaruncular medial orbitotomy for stabilization of the posterior limb of the medial canthal tendon.

PURPOSE: A method to stabilize the posterior limb of the medial canthal tendon (MCT), using a transcaruncular medial orbitotomy (TMO) approach, is described in a stepwise fashion. The technique described is a modified version of procedures published by Ritleng, Crawford and Collin, and Fante and Elner METHODS: A prospective clinical evaluation of MCT stabilization via the TMO approach was undertaken in I I consecutive patients who presented with MCT laxity as one of the features of their ectropion. These cases are initially described in detail in two representative case reports, and summarized in 11 cases. The stepwise surgical approach is outlined. RESULTS: All patients had improved symptomatology in terms of epiphora and comfort. Furthermore, in all cases the lid position was improved or normalized. In four of the I cases (36%) the lower punctum did not ultimately reside in the lacrimal ake, but the punctal position was nevertheless improved and the MCT was stabilized. CONCLUSION: The TMO procedure provides both excellent MCT stabilization and adequate placement of the lower lacrimal punctum onto the globe. It does not require canalicular resection, and avoids continued anterior displacement of the medial lower lid which may occur when only the anterior limb of the MCT is addressed surgically.

Kinetic observational exophthalmometry: a simple clinical method of assessing the relative axial positions of the eyes.

A simple clinical method for detecting the presence of unilateral proptosis or enophthalmos is described. It is important that in a patient with orbital disease, a detailed ophthalmic and orbital history be taken and a careful clinical evaluation be performed. In the clinical assessment of such a patient, it is also essential to determine the presence of any asymmetry of eye position after excluding any asymmetry in eye size or in the skeletal structure of the face. Observation of the patient's head from above and noting the relative positions of the two eyes has been the traditional method in the clinical assessment of proptosis. We believe that this method of assessment of proptosis has not been adequately described or fully exploited in the past. This paper describes an extension of the same technique. It involves the examiner making the observation from above while slowly moving the patient's head passively from a position of neck flexion to one of neck extension, and then noting which eye in a patient comes into view first. We believe that this kinetic technique provides an accurate method of noting any asymmetry in eye position in relation to the supraciliary ridge. It should therefore prove to be a clinically useful method in the initial assessment of a patient with suspected ocular displacement.