Toll-like receptors as diagnostic targets in pellucid marginal degeneration.

The main purpose of this study is to evaluate the diagnostic role of Toll-like receptors 2 (TLR2) and 4 (TLR4) expression in corneal and conjunctival epithelial cells of eyes with pellucid marginal degeneration (PMD) compared to keratoconus patients (KC) and control subjects. A prospective case-control study in 29 PMD eyes, 109 KC eyes and 72 healthy eyes was done. All participants were subjected to a clinical, topographic, aberrometric and tomographic exam with extraction of corneal and conjunctival epithelial cells through scraping. The TLR2 and TLR4 expression was measured with flow cytometry. Receiver operating characteristic (ROC) curve analysis was used to determine the most appropriate cutoff point for predicting the risk of PMD and KC. Correlations between TLR2/TLR4 expression and the severity of PMD/KC were evaluated. A TLRs follow-up review was made 19 ± 4 months after to the first review. As result, mean expression of TLR2 and TLR4 in both corneal and conjunctival epithelial cells was significantly higher in eyes with corneal ectasia (PMD and KC) than in control eyes (all p < 0.05). Conjunctival TLR4 expression showed the highest capacity to diagnose the existence of PMD (odd ratio 42.84; 95% confidence interval:6.20-296.20; p < 0.0001) after adjusting by eye rubbing and steeper corneal meridian. Moreover, we found an association between the TLR2/TLR4 overexpression with the severity of the PMD and KC measured by corneal topographic, aberrometric and tomographic quantitative parameters (all p < 0.05). Differences on TLR2/TLR4 expression between study groups were maintained during the follow-up period. In conclusion, the TLR2/TLR4 overexpression in corneal and conjunctival epithelial cells of PMD and KC patients compared to healthy control subjects have demonstrated their role as diagnostic target in both corneal ectatic disorders.

Immunoresolvent Resolvin D1 Maintains the Health of the Ocular Surface.

The present review focuses on the role of one of the D-series resolvins (Rv) RvD1 in the regulation of conjunctival goblet cell secretion and its role in ocular surface health. RvD1 is the most thoroughly studied of the specialized proresolution mediators in the goblet cells. The anterior surface of the eye consists of the cornea (the transparent central area) and the conjunctiva (opaque tissue that surrounds the cornea and lines the eyelids). The secretory mucin MUC5AC produced by the conjunctival goblet cells is protective of the ocular surface and especially helps to maintain clear vision through the cornea. In health, a complex neural reflex stimulates goblet cell secretion to maintain an optimum amount of mucin in the tear film. The specialized pro-resolution mediator, D-series resolvin (RvD1) is present in human tears and induces goblet cell mucin secretion. RvD1 interacts with its receptors ALX/FPR2 and GPR32, activates phospholipases C, D, and A2, as well as the EGFR. This stimulation increases the intracellular [Ca2+] and activates extracellular regulated kinase (ERK) 1/2 to cause mucin secretion into the tear film. This mucin secretion protects the ocular surface from the challenges in the external milieu thus maintaining a healthy interface between the eye and the environment. RvD1 forms a second important mechanism along with activation of a neural reflex pathway to regulate goblet cell mucin secretion and protect the ocular surface in health.

Use of trypan blue to assess lymphatic function following trabeculectomy.

IMPORTANCE: To illustrate the importance of lymphatic drainage in assessing trabeculectomy bleb function using intracameral trypan blue. BACKGROUND: To study the lymphatic drainage of trabeculectomy blebs using trypan blue, correlate with bleb function and classify them accordingly. DESIGN: Prospective cross-sectional study in a tertiary care centre. PARTICIPANTS: Thirteen glaucoma patients post-trabeculectomy were studied. METHODS: Trypan blue was injected into the anterior chamber. MAIN OUTCOME MEASURES: The duration taken for dye to stain a drainage bleb or lymphatics is recorded. The extent of the lymphatic structures were measured in clock hours. Intraocular pressure (IOP) prior to surgery was recorded. RESULTS: Eight post-trabeculectomy subjects with dye stained lymphatic vessels had lower IOP (12.6 mmHg, P = .013) compared to the five with no lymphatic vessel staining (mean IOP 23.6 mmHg). Lymphatic extent was inversely related to IOP (P = .021). CONCLUSIONS AND RELEVANCE: Eyes with lymphatic connections to drainage blebs had lower IOP and reduced requirement for topical medications. The extent of lymphatic connection to drainage blebs is related to lower IOP.

Development and Long-Term Evaluation of Ocular Surface Conditions Using Ovariectomized Cynomolgus Monkey.

OBJECTIVES: We aim to use the ovariectomized cynomolgus monkey to observe the effect of long-term changes of sex hormone levels on ocular surface. METHODS: Six healthy adolescent cynomolgus monkeys were randomly assigned to control and experimental groups (n=3/group). Bilateral ovariectomy was performed in experimental group, and sham operation was performed on control animals with their ovaries reserved. The concentrations of serum estradiol and testosterone in cynomolgus monkeys' lower-limb venous were detected by radioimmunoassay before surgery, and at 6, 12, 18, and 24 months after the surgery. Phenol red cotton thread test, tear osmolarity, corneal fluorescein staining, and conjunctival impression cytology were performed to evaluate the ocular surface conditions before surgery, and at 6, 12, 18, and 24 months after the surgery. The histological examinations of the bulbar conjunctiva and labial glands tissues were performed at 12 and 24 months after the surgery. RESULTS: There were no significant differences in the serum level of estradiol and testosterone between the experimental and control groups before surgery, at 6, and 12 months after surgery (P>0.05). Serum level of estradiol and testosterone were significantly less in experimental versus control group at 18 and 24 months after surgery (P<0.05). There was no significant differences in phenol red cotton between the experimental and control groups before surgery, at 6, 12, 18, and 24 months after surgery (P>0.05). Tear osmolarity had no significant differences between experimental and control group before surgery (P>0.05), and it was significantly higher in experimental versus control group at all four time-points after surgery (P<0.05). The number of conjunctival goblet cells had no significant differences between experimental and control group before surgery, at 6 months after surgery (P>0.05), and it was significantly lower in experimental versus control group at 12, 18, and 24 months after surgery (P<0.05). There were no significant differences in the corneal fluorescein staining score between the experimental and control groups before surgery, at 6, 12, and 18 months after surgery (P>0.05), and it was higher in experimental than control group at 24 months after surgery (P<0.05). In addition, inflammatory cell infiltration in bulbar conjunctiva and labial glands tissues was observed in experimental group and was unobserved in control group at 24 months after surgery. CONCLUSIONS: Ovariectomy led to decreased sex hormone levels in cynomolgus monkey. In turn, tear osmolarity was increased, the number of conjunctival goblet cells were decreased, the corneal fluorescence staining was increased, and the pathological examination of the bulbar conjunctiva and labial glands tissues were abnormal, phenol red cotton thread test did not change significantly.

Safety of Using Matrix Metalloproteinase Inhibitor in Experimental Glaucoma Filtration Surgery.

We evaluated the safety of matrix metalloproteinase (MMP) inhibitor in experimental glaucoma filtration surgery in an animal model. Fifteen New Zealand white rabbits underwent an experimental trabeculectomy and were randomly allocated into 3 groups according to the adjuvant agent: no treatment group (n = 5), 0.02% mitomycin C (MMC) soaking group (n = 5), and MMP inhibitor (ilomastat) subconjunctival injection group (n = 5). Slit lamp examination with Seidel testing, pachymetry, and specular microscopy was performed preoperatively and postoperatively. The conjunctiva and ciliary body toxicity were evaluated with scores according to the pathologic grading systems. Electron microscopy was used to examine the structural changes in cornea, conjunctiva, and ciliary body. In the ilomastat-treated group, there was no statistically significant change in central corneal thickness preoperatively and at 28 days postoperatively (P = 0.655). There were also no significant changes in specular microscopy findings over the duration of the study in the ilomastat-treated group. The conjunctival toxicity score was 1 in the control group, 1.5 in the ilomastat-treated group, and 2 in the MMC-treated group. When assessing ciliary body toxicity scores, the ilomastat-treated group score was 0.5 and the MMC-treated group score was 1.5. Transmission electron microscopy did not show structural changes in the cornea and ciliary body whereas the structural changes were noticed in MMC group. A single subconjunctival injection of MMP inhibitor during the experimental trabeculectomy showed a less toxic affect in the rabbit cornea, conjunctiva, and ciliary body compared to MMC.

Ocular Surface Temperature During Scleral Lens Wearing in Patients With Keratoconus.

OBJECTIVE: To evaluate the ocular surface temperature using an infrared thermography camera before and after wearing scleral lens in patients with keratoconus and correlate these results with the tear production and stability. METHODS: A pilot, experimental, short-term study has been performed. Twenty-six patients with keratoconus (36.95±8.95 years) participated voluntarily in the study. The sample was divided into two groups: patients with intrastromal corneal ring (KC-ICRS group) and patients without ICRS (KC group). Schirmer test, tear breakup time (TBUT), and ocular surface temperature in the conjunctiva, limbus, and cornea were evaluated before and after wearing a scleral lens. RESULTS: The patients wore the scleral lenses from 6 to 9 hours with average of 7.59±0.73 hours. No significant changes in Schirmer test and TBUT were found for both groups. No temperature differences were found between the KC-ICRS and the KC groups for all zones evaluated. There was a slight, but statistically significant, increase in the inferior cornea, temporal limbus, and nasal conjunctival temperature for KC-ICRS group and temporal limbus temperature decreasing for the KC group after wearing scleral lens (P<0.05). The conjunctiva and limbus temperature was statistically higher than the central cornea for both groups before and after scleral lenses wearing (P<0.05), but no difference in the peripheral cornea was found. No statistically significant differences in the central corneal temperature were found between the groups after scleral lens wearing (P>0.05). CONCLUSION: Scleral contact lens seems not to modify the ocular surface temperature despite the presence of the tear film stagnation under the lens.

Prosthetic Replacement of the Ocular Surface Ecosystem Scleral Lens Therapy for Exposure Keratopathy.

OBJECTIVES: To investigate the utility of Prosthetic Replacement of the Ocular Surface Ecosystem (PROSE) scleral lenses in patients with exposure keratopathy, with outcomes based on changes in visual acuity, visual function, and corneal staining. METHODS: A retrospective interventional case series of patients evaluated and treated from September 2009 through June 2014 at a single multi-specialty institutional practice. Eighteen of 29 patients with exposure keratoconjunctivitis, lagophthalmos, ectropion, or lid retraction, referred to USC Eye Institute after failing conventional therapies completed PROSE scleral lens fitting and were included in the study. Visual function was assessed before and after PROSE fitting with the Ocular Surface Disease Index (OSDI) survey. Visual acuity (VA) and corneal staining changes were also evaluated before and after treatment. RESULTS: Visual acuity improved from 0.60±0.68 logMAR pre-PROSE to 0.25±0.34 logMAR (Z=-3.81, P=0.00014) post-PROSE, which corresponds to an improvement of about 20/80 to 20/35 on Snellen VA. Ocular Surface Disease Index scores improved from 56.54±29.75 pre-PROSE to 24.98±21.23 post-PROSE (Z=-2.98, P=0.0029), and corneal staining values decreased from 2.17±0.84 pre-PROSE to 0.64±0.70 post-PROSE (Z=-3.27, P=0.011). CONCLUSIONS: The results suggest that PROSE scleral lens therapy is effective in patients with exposure keratopathy who had failed conventional therapies and can serve as an alternative to lid surgery.

Lacritin and other autophagy associated proteins in ocular surface health.

Advantage may be taken of macroautophagy ('autophagy') to promote ocular health. Autophagy continually captures aged or damaged cellular material for lysosomal degradation and recyling. When autophagic flux is chronically elevated, or alternatively deficient, health suffers. Chronic elevation of flux and stress are the consequence of inflammatory cytokines or of dry eye tears but not normal tears invitro. Exogenous tear protein lacritin transiently accelerates flux to restore homeostasis invitro and corneal health invivo, and yet the monomeric active form of lacritin appears to be selectively deficient in dry eye. Tissue transglutaminase-dependent cross-linking of monomer decreases monomer quantity and monomer affinity for coreceptor syndecan-1 thereby abrogating activity. Tissue transglutaminase is elevated in dry eye. Mutation of arylsulfatase A, arylsulfatase B, ceroid-lipofuscinosis neuronal 3, mucolipin, or Niemann-Pick disease type C1 respectively underlie several diseases of apparently insufficient autophagic flux that affect the eye, including: metachromatic leukodystrophy, mucopolysaccharidosis type VI, juvenile-onset Batten disease, mucolipidosis IV, and Niemann-Pick type C associated with myelin sheath destruction of corneal sensory and ciliary nerves and of the optic nerve; corneal clouding, ocular hypertension, glaucoma and optic nerve atrophy; accumulation of 'ceroid-lipofuscin' in surface conjunctival cells, and in ganglion and neuronal cells; decreased visual acuity and retinal dystrophy; and neurodegeneration. For some, enzyme or gene replacement, or substrate reduction, therapy is proving to be successful. Here we discuss examples of restoring ocular surface homeostasis through alteration of autophagy, with particular attention to lacritin.

Evaluation of filtering blebs exhibiting transconjunctival oozing using anterior segment optical coherence tomography.

PURPOSE: To explore the features of filtering blebs exhibiting transconjunctival oozing via three-dimensional anterior segment optical coherence tomography (3D AS-OCT). METHODS: In this cross-sectional study, 131 eyes of 131 patients exhibiting filtering blebs were examined. Of those, 20 eyes were excluded as flat-shaped, non-functioning bleb. Transconjunctival oozing was defined as transconjunctival aqueous egress evident on the bleb surface, in the absence of any point leak observable using a slit-lamp, as confirmed by application of digital pressure. Total bleb height, the height of the fluid-filled cavity, and bleb wall thickness and density were measured using 3D AS-OCT. Patient age, the etiology of glaucoma, postoperative follow-up period, number of glaucoma medication classes prescribed, intraocular pressure (IOP), grade of bleb vascularity, and bleb parameters were compared in eyes with and without bleb oozing. RESULTS: Sixty (54.0 %) of 111 eyes excluding non-functioning flat blebs exhibited oozing; mean IOP value (11.7 ± 4.5 vs. 14.8 ± 4.0 mmHg) and bleb vascularity grade (1.5 ± 0.7 vs. 2.4 ± 1.0) were lower than those of eyes without oozing. Total bleb height (1.1 ± 0.4 vs. 0.9 ± 0.4 mm), bleb wall thickness (0.7 ± 0.4 vs. 0.5 ± 0.3 mm), and bleb wall density (131.3 ± 45.7 vs. 180.9 ± 39.8 optical density units) differed significantly between the two groups (oozing vs. non-oozing). CONCLUSION: Transconjunctival oozing after trabeculectomy with MMC was associated with a low IOP, low-level bleb vascularity, an elevated total bleb height, a thicker bleb wall, and low bleb wall density.

Transplantation of PEDF-transfected pigment epithelial cells inhibits corneal neovascularization in a rabbit model.

BACKGROUND: The purpose of this study was to investigate the effect of recombinant pigment epithelium-derived factor (rPEDF), secreted by ARPE-19 cells transfected with the human PEDF gene and transplanted subconjunctivally in normal and in rabbits in which corneal neovascularization was elicited by a chemical burn. METHODS: Twenty grey Chinchilla Bastard rabbits were randomly assigned to four groups; neovascularization was induced in groups A, B, and C by alkali cauterization. Seven days later, group A received no cell implantation, non-transfected ARPE-19 cells were implanted subconjunctivally in group B, and PEDF-transfected ARPE-19 cells were implanted subconjunctivally in groups C and D (non-cauterized). In-vivo rPEDF secretion was analyzed by immunoblotting, and ELISA of extracts of conjunctival tissue samples taken at different time points. Digital photographs acquired on days 7, 14, and 21 after cauterization were evaluated for lead vessel length, vascular invasion area, and overall neovascularization rate. RESULTS: At days 14 and 21 after cauterization, significant differences were observed between groups A, B, and C in lead vessel length (day 21: 5.91 ± 0.45, 5.11 ± 1.22, 3.79 ± 0.59 mm, repectively), vascular invasion area (day 21: 35.5 ± 8.65, 34.86 ± 4.92, 19.2 ± 5.03 mm(2) respectively), and rate of corneal neovascularization. Compared to controls, neovascularization was reduced by 37.5 % on day 14 and 47 % on day 21. Analysis of conjunctival tissue extracts showed that rPEDF was secreted by the transplanted PEDF-transfected cells. CONCLUSION: Subconjunctivally transplanted, PEDF-transfected ARPE-19 cells secrete rPEDF, which inhibits the corneal neovascularization elicited by alkali cauterization.

Optical edge effects create conjunctival indentation thickness artefacts.

PURPOSE: Conjunctival compression observed in ultrahigh resolution optical coherence tomography (UHR-OCT) images of contact lens edges could be actual tissue alteration, may be an optical artefact arising from the difference between the refractive indexes of the lens material and the conjunctival tissue, or could be a combination of the two. The purpose of this study is to image the artefact with contact lenses on a non-biological (non-indentable) medium and to determine the origins of the observed conjunctival compression. METHODS: Two-dimensional cross-sectional images of the edges of a selection of marketed silicone hydrogel and hydrogel lenses (refractive index ranging from 1.40 to 1.43) were acquired with a research grade UHR-OCT system. The lenses were placed on three continuous surfaces, a glass sphere (refractive index n = 1.52), a rigid contact lens (n = 1.376) and the cornea of a healthy human subject (average n = 1.376). The displacement observed was analysed using ImageJ. RESULTS: The observed optical displacement ranged between 5.39(0.06) µm with Acuvue Advance and 11.99(0.18) µm with Air Optix Night & Day when the lens was imaged on the glass reference sphere. Similarly, on a rigid contact lens displacement ranged between 5.51(0.03) and 9.72(0.12) µm. Displacement was also observed when the lenses were imaged on the human conjunctiva and ranged from 6.49(0.80) µm for the 1-day Acuvue Moist to 17.4(0.22) µm for the Pure Vision contact lens. CONCLUSIONS: An optical displacement artefact was observed when imaging a contact lens on two rigid continuous surfaces with UHR-OCT where compression or indentation of the surface could not have been a factor. Contact lenses imaged in situ also exhibited displacement at the intersection of the contact lens edge and the conjunctiva, likely a manifestation of both the artefact and compression of the conjunctiva.

Healing indicators after pterygium excision by optical coherence tomography.

PURPOSE: To establish the sequence of tomographic changes in the tissue recovery process after pterygium excision and to propose healing indicators. METHODS: Optical coherence tomography (OCT) images were taken at 1 week, 1, 3 and 6 months after lesion excision in 73 eyes of 73 patients (33 male, 40 female; mean age 50, S.D. 5.0, range 40-70 years) with primary nasal pterygium. Biomicroscopy was performed at each visit and at 12 months, to diagnose clinical healing or lesion recurrence. The presence of well demarcated corneal epithelium, conjunctival epithelium, limbal demarcation area, and graft thickening were analysed. Comparisons between tomographic data of both clinical situations were made at each time point using contingency tables. RESULTS: Eleven eyes displayed lesion recurrence (R group) and 62 eyes showed no recurrence (NR group). Normal anatomical structures, corneal and conjunctival epithelium and limbal demarcation area, were identified by OCT images in a higher percentage of NR cases over time, compared to the R group where most of the cases presented without these markers of tissue recovery. In contrast, the variable graft thickening, which analysed a pathological event, revealed similar results in both groups (p > 0.05; Fisher's exact statistic), with a clear decrease of cases which showed graft thickening over time. Differences between groups started at 1 month, when no eye had yet presented clinical recurrence, with greater identification of corneal epithelium in the NR group (p = 0.04; Fisher's exact statistic). At 3 months, corneal and conjunctival epithelium identification tended to be more frequent in the NR than in the R group (in both cases, p = 0.0001; Fisher's exact statistic). Finally, at 6 months these different patterns consolidated, with a significantly higher number of limbal demarcation areas being identified in the NR group (p = 0.001; Fisher's exact statistic). In fact, this landmark of a normally structured limbus was never found in the R group. CONCLUSIONS: The sequence of tissue restoration, according to OCT images, seems to start in the cornea and end in the limbal area, similar to the process of pterygium injury in reverse. Although the visualisation of corneal epithelium could be an early indicator of successful surgery, identification of the limbal demarcation area, as a normal limbal pattern in OCT images, seems to be a better positive predictive value in diagnosing healing.

Ocular surface disease in pseudoexfoliation syndrome.

Aim of the study is to determine connection between pseudoexfoliation (PEX) syndrome and symptoms and signs of ocular surface disease. Tear film break-up time test, Schirmer II test and assessment of lid parallel conjunctival folds were performed in 40 PEX syndrome patients and 40 controls. All data was statistically analyzed. Results show statistically significant difference in every component between groups, most prominent in tear film break up time test. We have concluded that patients with PEX syndrome have higher predisposition of tear function disorders and that both components of dry eye syndrome are present in PEX syndrome.

Scleral fixation underneath an intact conjunctiva.

PURPOSE: To describe a technique for the fixation of intraocular lenses or capsular tension rings through the ciliary sulcus or pars plana. METHODS: This alternative technique allows burial of the knot and the free suture ends within the sclera, without using a scleral flap or pocket or even a conjunctival opening. RESULTS: No suture exposure has occurred with the use of the suture burial technique in approximately 25 eyes in 4 years. Additional benefits include less patient discomfort and an improved anatomic and cosmetic outcome starting from the first postoperative day. CONCLUSIONS: This technique causes minimal tissue damage and scarring with almost no postoperative discomfort.

Understanding the stimulus of an air-jet aesthesiometer: computerised modelling and subjective interpretation.

PURPOSE: Assessment of corneal sensory function is more frequently being carried out using novel non-contact methods, which promise better stimulus control and a greater intensity range than more traditional techniques. An examination of the characteristics of the air-jet stimulus of these 'non-contact' aesthesiometers is important to enable an understanding of how these instruments stimulate the cornea. The purpose of this study was to model the air-jet stimulus of the CRCERT-Belmonte Aesthesiometer (CBA) and its interaction with the cornea. METHOD: A computerised simulation was used to model the CBA stimulus and this model was then examined in the context of verbal descriptions of the supra-threshold stimulus provided by 27 human subjects with normal corneas. The computational fluid dynamics model and the subjective descriptions considered CBA airflow values between 70 and 400 mL min(-1) . RESULTS: The computational fluid dynamics results showed that the CBA air-jet stimulus behaves as expected in terms of force exerted and stabilises and subsides quickly during the 1 s duration for which it is turned on/off. The computer modelling demonstrated that the stimulus of the CBA consists of a central core of air flow, similar in diameter to the CBA nozzle, which exerts maximum pressure at the corneal apex. This core is surrounded by a less prominent zone of lower pressure, extending out to the corneal periphery at higher flow rates. The computer modelling results were confirmed by the subjective descriptions of intensity and corneal area stimulated, whereas relation to duration of sensation was more equivocal. CONCLUSIONS: The CBA probably stimulates sensory receptors over a greater area of the ocular surface compared to the traditional nylon filament aesthesiometer and may recruit neurons sensitive to additional stimulus modalities. Future computational models should also incorporate temperature as an important aspect of the corneal response. This study has effectively demonstrated that it is possible to create a virtual model of an air-jet aesthesiometer stimulus using computational methods and that this model is corroborated by subjective descriptions provided by subjects.

Fluids of the ocular surface: concepts, functions and physics.

General adoption of the ocular surface (OS) concept has advanced the therapy of the external eye. Fresh physical findings have prompted new concepts; examples taken from each section of the text are: (i) ever-present lipid sealant bridges the palpebral fissure capping the three-dimensional 'OS' sac. The muco-aqueous pool (MAP) is thus enclosed, secluded from atmosphere, evaporation mitigated. Hence, the OS is conceptually, a compartment. The term 'dacruon' (otherwise 'tear film') has been coined for the combined fluids of the OS, viz. lipid film and MAP. (ii) Investigative techniques of physics yield data on (say) surface tension and viscosity, and on functions such as anchorage of dacruon base to the varied mucosae of the OS, lubrication, renovation of intermarginal fluid layers as the eye opens after each blink, and refinement of optics and vision by the fluids attached to the cornea. (iii) Physical events in the opening eye produce the unique 'black line' phenomenon in which capillary force induces subsurface flows into thirsty menisci, bringing about parameniscal dark grooves, pupil-ward of each meniscus. Attenuation of fluorescein in the shallowed fluid gaps behind each groove makes the dye appear unilluminated ('black lines') relative to adjacent full-thickness MAP fluid glowing under cobalt-blue illumination. Isolated from cornea by grooves and gaps, the meniscal fluid cannot pass freely over the cornea. It therefore streams through the menisci to nasolacrimal outflow.

Assessing the sensory function of the ocular surface: implications of use of a non-contact air jet aesthesiometer versus the Cochet-Bonnet aesthesiometer.

Assessment of corneal sensory function is commonly carried out using the Cochet-Bonnet aesthesiometer. The limitations of this instrument have lead to development of newer instruments, such as the CRCERT-Belmonte aesthesiometer, which utilise a jet of air as their stimulus. Recent work, however, has demonstrated contradictory effects on ocular surface sensitivity when measured with different types of aesthesiometer. The purpose of the present study was to compare clinical measurements of corneal sensitivity obtained with the CRCERT-Belmonte and the Cochet-Bonnet aesthesiometers and to examine their stimulus characteristics in terms of force exerted. No association was found between central corneal sensitivity measured with the two aesthesiometers in a group of normal subjects. Sensitivity was measured to be lower with the Cochet-Bonnet aesthesiometer than with the CRCERT-Belmonte. Over half of the subjects could not be assessed with the standard Cochet-Bonnet filament and 11% could not be assessed with either filament, as their corneal sensitivity was outside of the range of the instrument. In contrast, all subjects were within the stimulus range of the CRCERT-Belmonte aesthesiometer. Corneal sensitivity measurements made with the non-contact CRCERT-Belmonte aesthesiometer and the Cochet-Bonnet aesthesiometer are not comparable. Due to dissimilarities in the composition of their stimuli, and thus mode of stimulation, it is possible that the two instruments measure different aspects of the neural response. The underestimation of corneal sensitivity by the Cochet-Bonnet aesthesiometer and its inability to measure sensitivity of some subjects at all are important considerations in the assessment of sensitivity loss. Subtle changes are unlikely to be detected with this instrument, particularly at higher sensitivity levels. The Cochet-Bonnet should therefore be used with caution and the 0.08 mm diameter used as the filament of choice. Adoption of a non-contact aesthesiometer as standard for ocular sensitivity measurement should be considered. The non-contact instrument allows superior stimulus reproducibility and better control over stimulus characteristics, in addition to the ability for exploration of the response of all three types of neuro-receptors on the ocular surface.

Effects of osmolarity on human epithelial conjunctival cells using an electrical technique.

BACKGROUND: The purpose of this study is to report the effect of different media osmolarity on a cell line monolayer of normal human conjunctival epithelia (IOBA-NHC) using Electric Cell-substrate Impedance Sensing (ECIS). METHODS: We built our own ECIS system. We fabricated biocompatible microelectrodes. We used a monolayer of IOBA-NHC cells with media at different osmolarities (315, 360, 446, and 617 mOsm/l). RESULTS: When there is an increase in hyperosmolarity, there is a slight decrease in the measured resistance of the naked microelectrode (without cells), whereas its capacitance remained practically unchanged. The evaluation of resistance and capacitance of a microelectrode covered by a monolayer of IOBA-NHC in relation to a naked microelectrode showed no difference in the standard media (315 mOsm/l), a small difference with 360 mOsm/l, and significant differences with hyperosmolarities of 446 mOsm/l and 610 mOsm/l. The resistance with a confluent cell monolayer is up to three times greater compared to the value of the resistance of the naked electrode with standard media. CONCLUSIONS: Both resistance and capacitance measurements for the cell monolayer were sensitive to changes in osmolarity.

MMP expression in leaking filtering blebs and tears after glaucoma filtering surgery.

OBJECTIVE: Glaucoma filtering surgery may be compromised by cystic blebs which develop more frequently when anti-metabolites are used to arrest wound healing. Matrix metalloproteinases (MMPs) and the naturally occurring tissue inhibitors of metalloproteinases (TIMPs) are essential in connective tissue remodeling and wound healing. This study aimed to determine whether filtering blebs display increased expression of MMP-2, MMP-9, TIMP-1 and TIMP-2, and whether it is reflected in tear fluid. METHODS: Tissue samples from leaking blebs (n = 5) and control conjunctiva (n = 5) were evaluated by immunohistochemistry for MMP-2, MMP-9, TIMP-1 and TIMP-2. Tear fluid was collected from 12 patients (12 eyes) with cystic blebs and ten patients (ten eyes) with flat blebs following trabeculectomy with Mitomycin C applied and 16 controls. MMP levels were evaluated by zymography and TIMP levels by Western blot analysis. RESULTS: Conjunctival tissue was obtained from five eyes with cystic leaking blebs and five control eyes undergoing cataract surgery. More extensive MMP-2 and MMP-9 expression was found in the epithelial and stromal layers of blebs than in control conjunctiva. TIMP-1and TIMP-2 were expressed in all layers of the blebs, but only in the epithelium of control conjunctiva. MMP-2 and proMMP-2 activity in tears from eyes with flat blebs was significantly higher than that of controls, while activity in tears of eyes with cystic blebs was significantly higher than in those with flat blebs. There was no difference in MMP-9 activity between tears of control and post-filtering surgery eyes. CONCLUSIONS: Increased MMPs and TIMPs expression is associated with the formation of filtering blebs, suggesting involvement of MMPs in bleb remodeling. MMP-2 and ProMMP-2 levels in tear fluid may be markers for bleb configuration.

A solute gradient in the tear meniscus. I. A hypothesis to explain Marx's line.

Marx's line is a line of mucosal staining behind the mucocutaneous junction. It can be demonstrated throughout life in all normal lids by staining with lissamine green and related dyes. Of all the body orifices, only the mucosae of the eye and mouth are directly exposed to the atmosphere. In this paper, we suggest that for the eye, this exposure leads to the formation of Marx's line. The tear meniscus thins progressively toward its apex, where it is pinned at the mucocutaneous junction of the lid. It also thins toward the black line, which segregates the meniscus from the tear film after the blink. We predict that, because of the geometry of the tear meniscus, evaporation generates a solute gradient across the meniscus profile in the anteroposterior plane, which peaks at the meniscus apices at the end of the interblink. One outcome would be to amplify the level of tear molarity at these sites so that they reach hyperosmolar proportions. Preliminary mathematical modeling suggests that dilution of this effect by advection and diffusion of solute away from the meniscus apex at the mucocutaneous junction will be restricted by spatial constraints, the presence of tear and surface mucins at this site, and limited fluid flow. We conclude that evaporative water loss from the tear meniscus may result in a physiological zone of hyperosmolar and related stresses to the occlusal conjunctiva, directly behind the mucocutaneous junction. We hypothesize that this stimulates a high epithelial cell turnover at this site, incomplete epithelial maturation, and a failure to express key molecules such as MUC 16 and galectin-3, which, with the tight junctions between surface epithelial cells, are necessary to seal the ocular surface and prevent penetration of dyes and other molecules into the epithelium. This is proposed as the basis for Marx's line. In Part II of this paper (also published in this issue of The Ocular Surface), we address additional pathophysiological consequences of this mechanism, affecting lid margins.