Subclinical Tear Film Stability Changes in VDU Operators
Coauthors: Jaroslaw MAREK, MD, PhD

Determining Tear Function and its Correlation with Duration of RA in Known Cases of RA
Coauthors: Mehdi HOSSEINI TEHRANI (1); M.H. MALEKMADANI (3)
(1) Cornea Center, Farabi Eye Hospital (Tehran, Iran); (3) Ophthalmology Clinic, Imam Khomeini General Hospital (Tehran, Iran)

Abstract:
PURPOSE: To evaluate tear film status in RA. Dry Eye is a known complication of Rheumatoid Arthritis. Familiarity with prevalence of dry eye syndrome in RA and its relation to duration of RA, gives the physician the ability to manage this condition as early and effective as possible.
METHODS: Prospective, cross sectional study. A total of 63 eyes from 121 patients (who were reffered from Rheumatology Research center, Imam Khomeini general hospital, Tehran) as known cases of RA according to American College of Rheumatology criteria have participated in this study in Ophthalmology clinic in Imam khoneimi general hospital, Tehran. All eyes have been tested by Tear Break Up Time, Tear Meniscus, Rose Bengal and Schirmer tests (both with and with out anesthesia).
RESULTS: 25 eyes out of 121 total eyes were dry eye (of any grade according to Madrid Triple Classification of Dry Eye Grading system) according to the above tests. These dry eyes stay in the category of Type2 Sjogren Syndrome and all others in the category of Type1 Sjogren Syndrome with involvement of exocrine glands, have been excluded from this study. The prevalence of the dry eye was significantly lower in male population than in females. There was a significant correlation between duration of RA (in years) and Dry Eye (P value less than 0.01) mostly 4 years after diagnosis of RA.
CONCLUSIONS: Patients with RA should be checked and screened for Dry Eye at least in the first 4 - year after diagnosis of RA.

Laser Therapy in Pterygium
Coauthors: Eduardo GOMEZ DEMMEL; José ESPINOSA; Romina FIGUEROA

Abstract:
PURPOSE: To report the experience with the use of diiodo laser photocoagulation for the treatment of small and advanced stationary pterygium.
METHODS: In this study, 57 patients and 62 eyes with small and advanced pterigium were subjected to diiodo laser therapy. Age varied between 27 and 63 years. The most frequent ocular symptoms were: irritation and foreign body sensation. Patients were followed up for a period of 6-24 months.
RESULTS: Thirty-two eyes (52%) had small pterygia and twenty-eight eyes (45%) had advanced pterygia. Two eyes (3.2%) had recurrent pterygia after one surgical excision. The success rate was 98% when laser therapy was employed in small pterygium, while it was 70% in cases with advanced pterygium. Size reduction of pterygia and diminution of symptoms were progressive in the three months following treatment and persisted with the time.
CONCLUSIONS: Diiodo laser treatment is a safe and effective method of therapy for small pterygium, avoids the advancement of pterygia and improves the symptoms. It is an alternative technique to straightforward surgical excision.

Establishment of a Two-dimensional Human Tear Protein Map by Proteomic Analysis
Coauthors: P. MARCELO (2); J. VINH (2); S. DOAN (3); C. CHAUMEIL (1); E. BALLOT (4)
(1) Laboratoire, Centre Hospitalier National d’Ophtalmologie; (2) Laboratoire Neurobiologie (ESPCI, CNRS UMR 7637); (3) Service d’Ophtalmologie Hôpital Bichat; (4) Laboratoire Immunologie Hôpital Saint-Antoine (Paris, France)

Abstract:
PURPOSE: Establish a two-dimensional human normal tear protein map with some identifications by proteomic analysis.
SAMPLES and METHOD: 109 tear sample were collected from 65 normal donors (5 to 72 years old, 42 females and 23 males) using microcapillary tubes. The protein level and the gel agarose electrophoresis patterns of all these samples were normal. Three pools of 3 to 4 mg of protein were used. One mg of tear proteins from each pool was analysed by 2D-electrophoresis, using immobilized pH gradients strips (pH 3-10 and pH 4-7) for the isoelectrophoresis before the separation in function of their molecular weights on 10% and 15% SDS polyacrylamide gels for each pool. Gels were then colloidal Coomassie blue stained. The spots which were common to the different pools were digested with trypsin and their identifications were performed using peptide mass fingerprint analysis obtained by matrix-assisted laser desorption/ ionization-time of flight (MALDI-TOF) mass spectrometry. Identifications were confirmed by aminoacids sequences obtained by nanoscale liquid chromatography electrospray tandem mass spectrometry (LC-ESIMS/MS).
RESULTS: More than 60 spots of molecular weights between 12 and 95 kDa and of iso-electric points between pH 4 and pH 9 were studied: the three major tear protein components, lipocalins, lactoferrin, lysozyme, which are separated by electrophoresis on agarose gel were found under many isoforms for the two first. Many spots revealed isoforms of lipocalins at their theorical molecular weight and at the double of the last one, supposing the existence of dimer of lipocalins. Other proteins were identified. Among them, cystatines SA et SN, Zinc-a2glycoprotein, B2-microglobulin, mammaglobin B, and curiously isoforms of the prolactin-induced protein.
CONCLUSIONS: The understanding of the human protein tear profile should lead to find new objective biomarkers of ocular diseases with diverse applications in the diagnosis of ocular surface pathology.

Masked Study of Corneal Nerves in Dry Eye Patients Using Confocal Microscopy
Coauthors: M. WASFY; J.M. MARTINEZ DE LA CASA; D. MENDEZ; J. GARCIA FEIJOO; J. GARCIA SANCHEZ
Unidad de Superficie e Inflamación Ocular, Hospital Clínico San Carlos (Madrid, Spain)

Abstract:
PURPOSE: The ocular surface, the main lacrimal gland and the interconnecting neural reflex loops comprise a functional unit. An alteration in corneal innervation has been described in dry eye (DE) associated with diabetes mellitus, contact lens use and lasik. The purpose of this study was to evaluate if DE not related with Sjogren’s syndrome (NSDE) and DE related to this syndrome (SSDE) are associated with an alteration of the corneal nerves.
METHODS: Group healthy volunteers less 60 yo (HV<60) (10 females and 1 male) aged 30.7 SD 2.6, group healthy volunteers more 60 yo (HV>60) (8 females and 2 males) aged 68.7 SD 7.1, group SSDE (10 females and 1 male) aged 52.9 SD 8.7 and group NSDE (8 females and 2 males) aged 65.8SD 5.3 yo. Patients were examined with a Confoscan model P4 (Tomey, Germany).
RESULTS: Sub-basal nerve density: HV<60 = 769 SD 88, HV>60 = 624 SD 86, SSDE = 508 SD 128 and NSDE = 593 SD 127 microns/mm2 (p<0.000, ANOVA). There was a statistically significant difference between HV<60 and the other groups (HV>60 p<0.05, SSDE p<0.000 and NSDE p<0.005, Bonferroni). Number of beadings: HV<60 = 198 SD 66, HV>60 = 182 SD 63, SSDE = 387 SD 62 and NSDE = 323 SD 64 /mm (p<0.000, ANOVA). There was a statistically significant difference between healthy volunteers groups and DE groups (p<0.000, Bonferroni).
CONCLUSIONS: SSDE and NSDE patients exhibit corneal innervation abnormalities.

Ophthalmic Formulations Based on Aloe Vera: Characterisation and In Vivo Evaluation in Animal Model
Coauthors: C. SOMMANI; D. MONTI; M. NAJARRO; P. CHETONI;
Dept. of Biorganic Chemistry and Biopharmaceutics, University of Pisa (Pisa, Italy)

Abstract:
PURPOSE: Aloe Vera (Aloe barbadensis Miller) has been employed and utilised by virtue of curative properties for a long time. Aloe Vera gel, the fiber-free mucous exudate of the succulent leaves, is the part of the plant therapeutically used for treating skin ans body tissue wounds (burns, superficial injuries, cutaneous irritation and rush). This gel is largely used today as an important adjuvants in cosmetic formulations due to its moisturizing properties and its ability in increasing wound-healing rate. For these reasons we decided to use Aloe Vera gel as ophthalmic vehicle.
METHODS: Different amounts of Aloe Vera powder were dissolved in different buffer solutions and their rheological properties, pH and tonicity values were determined. The mucoadhesive properties and the ability of the Aloe Vera solutions to produce ferning-like residual were also evaluated. One of the prepared solutions was chosen as polymeric tear substitute and its efficacy in the treatment of Keratoconjunctivitis sicca was evaluated by different in vivo tests (1). Further studies were carried out to evaluate corneal wound-healing and spreading coefficient of this preparation using animal models.
RESULTS and CONCLUSIONS: Tests carried out on formulation containing Aloe Vera highlighted very good characteristics to take advantage of ophthalmic application for treating dry eye diseases.
Reference: (1) Burgalassi S., Panichi L., Chetoni P., Saetone M.F., Boldrini E.: Development of a Simple Dry Eye Model in the Albino Rabbit and Evaluation of Some Tear Substitutes: Ophthalmic Research 1999, 31:229-235.

CHENZHUO, Lucia
7500 Kirby Dr. #1644 - TX 77030 HOUSTON (U.S.A.)
Phone: (+1 918) 5835114 - E-mail: lucychenzhuo@yahoo.com
Topics: 01. Basic science lacrimal research

Structure Remodelling of the Penetrating Corneal Injuries Treated by Human Amniotic Membrane Transplantation
Coauthors: C. REDONDO; J. MURUBE

CHENZHUO, Lucia
7500 Kirby Dr. #1644 - X 77030 HOUSTON (U.S.A.)
Phone: (+1 918) 5835114 – E-mail: lucychenzhuo@yahoo.com
Topics: 01. Basic science lacrimal research

Stroma Niche Controls Human Corneal Epithelial Cells Culture
Coauthors: Surrendra BASTI; De-quan LI; Stephen PFLUGFELDER; Arturo VILLARREAL

Tear Film Changes in Pseudoexfoliation Syndrome: Potential Clinical Implications
Coauthors: Stavroula KOUKOULA; Nikitas FOUNTOULAKIS; Vassilios P. KOZOBOLIS

Abstract:
PURPOSE: Pseudoexfoliation Syndrome (PEX) is characterized by the presence of whitish material at the anterior ocular segment. Previous studies have reported reduced tear secretion and stability in PEX. The present study aims at evaluating potential associations of these findings with clinical parameters, including central corneal thickness (CCT), central corneal mechanical sensitivity (CCMS) and corneal epithelial staining (CES).
METHODS: Patients with PEX in one or both eyes constituted the Study Group (SG). Age and gender-matched patients without PEX in either eye constituted the Control Group (CG). All participants underwent Schirmer test (ST, measured in mm), Break-Up-time test (BUT, measured in sec), evaluation of CCMS (Cochet-Bonnet esthesiometer, Luneau Ophthalmologie, mesured on a scale of 60-5mm of thread length), evaluation of CCT (Pocket ultrasonic corneal pachymeter, Quantel Medical measured in um) and evaluation of CES (individual punctuate epithelial stains measured up to 60, with all numbers above 60 recorded at "60").
RESULTS: The SG included 64 patients (43 males, 67.18%). The CG included 44 patients (29 males, 65.09%). ST scores were significantly lower in the SG, compared with the CG (11.07±1.64mm and 15.11±1.43mm, respectively, p=0.03). BUT scores were also significantly lower in the SG, compared with the CG (6.07±1.99sec and 14.01±3.14sec, respectively, p=0.03). CCMS score was significantly (p=0.02) reduced in the SG compared with the CG (4.51±0.29cm and 5.79±0.58cm, respectively). CES counts were significantly higher in the SG, compared with CG (51.49±4.11 and 42.88±5.28, respectively, p<0.01). Differences in CCT score between the SG and CG were statistically not significant.
CONCLUSIONS: Dry-eye changes in PEX may predispose to an increase in CES and a decrease in CCMS. The reduction in the latter may have implications in the clinical management of PEX patients who may be at risk for corneal abrasions due to the frequent concomitant use of local antiglaucomatous medications.

Endonasal Dacryocystorhinostomy when a Previous Nasolacrimal Duct Surgery Has Failed
Coauthors: A. FERRERAS; R. FERNANDEZ-LIESA; F.M. HONRUBIA
Miguel Servet University Hospital (Zaragoza, Spain)

Abstract:
PURPOSE: To compare the efficacy of endonasal dacryocystorhinostomy (DCR) indicated on a primary basis with the results of this procedure following a failed external or endonasal DCR.
METHODS: 20 eyes of 20 patients were divided in two groups age-matched: the first group included 11 eyes with acquired epiphora without previous nasolacrimal duct surgery, and the second group included 3 eyes with previous endonasal DCR and 6 eyes with previous external DCR. Both groups underwent endonasal DCR with bicanalicular silicone tubing. Nasal mucose and bone were removed with a drill and the lacrimal sac was opened with a sickle knife. Intraoperative mitomycin C (MMC) was applied in the intranasal ostium, in 4 eyes (35.4%) of the first group and in 5 (55.6%) eyes of the second group.
RESULTS: Mean age was 59.82 +/- 12.48 years in the first group and 60.33 +/- 6.92 in the second group. Success rate for the two groups (defined as ostium patency on irrigation and nasal endoscopy) was 90.9% (10/11) and 88.9% (8/9) respectively, at 1 year postoperatively. The success rate in the MMC-treated group was statistically significantly different in the group with previous DCR but not in the group without previous surgery. Complications of endonasal DCR were not greater in frequency or magnitude than those associated with external DCR.
CONCLUSIONS: Endonasal DCR may be indicated as revisional surgery following failed external or endonasal DCR. Success rates may be improved when intraoperative MMC is applied to the intranasal ostium.

Long-Term Efficacy of Endonasal Dacryocystorhinostomy to Correct Nasolacrimal Duct Obstruction
Coauthors: F.J. FERNANDEZ-TIRADO; A.B. PAJARIN; R. FERNANDEZ-LIESA; F.M.HONRUBIA

Abstract:
PURPOSE: To assess the long-term efficacy and safety of endonasal dacryocystorhinostomy (DCR), advantages and limitations of the procedure and intraoperative and postoperative complications.
METHODS: From June 2000 to March 2003, a total of 28 eyes of 28 patients with epiphora were included in the study and followed-up for more than 20 months. All of them underwent endonasal DCR with bicanalicular silicone tubing, performed under general anesthesia. Nasal mucose and bone were removed with a drill and the lacrimal sac was opened with a sickle knife. Intraoperative mitomycin C (MMC) was applied in the intranasal ostium in 15 eyes (55.4%).
RESULTS: Mean age was 56.16 +/- 14.61 years (range: 17 to 79). Success rate was 82.1% (23/28) at 12 and 20 months postoperatively, defined as resolution of epiphora with ostium patency on irrigation and nasal endoscopy. The mean follow-up time was 26.87 +/- 8.58 months (range: 20 to 53). The success rate in the MMC-treated group was not statistically significantly different from MMC-non treated group. The most common intraoperative complication was hemorrhage, and the main cause of failure was progressive cicatricial closure of the ostium, and development of synechiaes between the ostium and the middle turbinate or the septum nasal.
CONCLUSIONS: Endonasal DCR is a good option for the correction of nasolacrimal duct obstruction that avoids cutaneous incision and scar, and is not limited by age. Success rate is similar or higher comparing endonasal DCR with external surgery.

Objective Evaluation of Antiallergic Therapy
Coauthors: Mitchell H. FRIEDLAENDER, MD
Division of Ophthalmology, Scripps Clinic (La Jolla, CA, U.S.A.)

Abstract:
PURPOSE: To objectively evaluate the effect of antiallergic eyedrops on allergic conjunctivitis induced by conjunctival allergen challenge (CAC) using the EES (Erythema, Edema, Sensation) Method.
METHODS: Twenty subjects with a history of allergy to airborne pollens were challenged by topical application of increasing concentrations of either cat dander, grass, or ragweed allergens, until a moderate allergic conjunctival reaction was produced. One week later, eyes were challenged with allergen, and treated 10 minutes later with either one drop of naphazoline hydrochloride 0.025%/pheniramine maleate 0.3% or normal saline. The following week, eyes were treated 10 minutes before CAC with one drop of epinastine hydrochloride 0.05% or olopatadine hydrochloride 0.01%. Erythema was measured 10 minutes before and 10 minutes after instillation of eye drops and allergen with a spectroradiometer. The u’ chromaticity coordinate was recorded. Edema of the lower eye lid margin was measured with a fractional millimeter reticule. Ocular surface sensation was measured with the aesthesiometer of Cochet and Bonnet. Subjective measurements were made by observation (erythema, edema), and questioning (itching). Data were analyzed using a paired sample t-test.
RESULTS: A reduction in conjunctival erythema could be detected objectively and subjectively ten minutes after treatment with a vasoconstrictor/antihistamine eye drop (p = 0.001). There was no significance difference in erythema or edema between the two treatment groups. Subjective itching was significantly less in eyes pretreated with epinastine, compared with eyes pretreated with olopatadine, 10 and 15 minutes after CAC (p = 0.020, p = 0.045).
CONCLUSIONS: Allergic reactions can be measured objectively using the EES Method. A significant reduction in conjunctival erythema could be measured objectively and subjectively after treatment with a vasoconstrictor/antihistamine eye drop. Epinastine and olopatadine were equivalent in their effect on erythema following CAC, but epinastine was superior to olopatadine in the reduction of itching.

Endothelin-1, Endothelin-Converting Enzyme-1 and Lactoferrin in Chalazia
Coauthors: L. MASSAI; P. CARBOTTI; M. MENCARELLI; C. ALESSANDRINI; G. GRASSO
Dipartimento di Scienze Oftalmologiche e Neurochirurgiche; Dipartimento di Scienze Anatomiche e Biomediche, Universitá di Siena (Siena, Italy)

Abstract:
PURPOSE: Chalazion is a chronic granulomatous inflammatory process of the eyelid. It develops for bacterial cause around a meibomian gland as a foreign body reaction to lipid produced by the gland and released into the surrounding tissue because of retention of meibomian gland secretions. The lipids of the meibomian gland are important for the stability of tear film and reduce evaporation of the fluid. The tear concentration of the lactoferrin, a iron-binding molecule appears to be closely related to tear production, and has been shown to be a good predictor of tear film stability in Sjögren’s syndrome. The aim of the present study was to investigate possible local endothelin-1 (ET-1) production by inflammatory cells, (ET-converting enzyme-1 (ECE-1)) and lactoferrin expression in chalazion sessions.
METHODS: Forty chalazia removed surgically by excision from suffering patients were studied. Chalazia sections were stained with hematoxylin and eosin for histological examination and incubated with specific antiserum for ET-1, (ECE-1) and lactoferrin for the singular and double immunohistochemical technique. Only the cells of the meibomian glands behind in the sebaceous transformation and the deeper cells of meibomian central duct immunostain for ET-1. Furthermore, ET-1 positive cell, specially neutrophils and macrophages, were located in the stroma, around the alveoli, and within the lipogranuloma.
RESULTS: The ECE-1 was present strongly in conjunctival epithelial cells, in tarsal glandular cells and their ducts, scarcely in the secreting portion of the meibomian glands, suggesting a local ET-1 production by inflammatory cells. Lactoferrin positive cells were in meibomian glands and in the stroma, specially neutrophils in the chalazia sections.
CONCLUSIONS: The presence of ET-1 (and ECE-1) in the chalazia could demonstrate a local production of ET-1 which might behave as pro-inflammatory mediator and regulate the lactoferrin production by neutrophils and meibomian gland cells acting in autocrine-paracrine role.

Bilateral Programmable Infusion Pumps Implanted in CREST Syndrome
Coauthors: J. DE ANDRES; P. UDAONDO; M. SERRANO; M. DIAZ-LLOPIS; D. SALOM, M. GARCIA-POUS, L. AMSELEM, A. PEREZ, M. HERNANDEZ
Hospital General Universitario de Valencia; Instituto Oftalmológico de Valencia (IOVA); Fundación Oftalmológica del Mediterráneo (FOM)

Abstract:
PURPOSE: We present one patient with severe dry eye secondary to a CREST syndrome treated with a bilateral programmable artificial tear pump, to achieve a continuous artificial tear infusion with daily and nightly tear flow fluctuations.
METHODS: Using Murube’s technique we implanted two pumps in the abdominal region, conected to the conjunctival superior sac of both eyes by a silicone tube.
RESULTS: No intraoperative nor postoperative complications were observed. The reservoir and the silicone tube was well tolerated and the patient experimented a dramatical improve of dry eye signs and symptoms after the surgery.
CONCLUSIONS: To our knowledge this is the first time that a bilateral programmable infusion pumps has been implanted to treat a dry eye secondary to a CREST syndrome. It has been a safe and effective procedure for our patient.

Wash of Lachrymal Sac with Triamcinolone and Ciprofloxacin in Acute Dacriocystitis
Coauthors: Dr. Diana Virginia FUENTES RIVERA; Dr. Alberto ESPEJO YEP; Dr. Bertha Yesenia IZARRA BEJARANO; Dr. Erico Iván CONCEPCION ALVARADO
Alberto Sabogal Hospital (Lima, Peru)

Abstract:
OBJECTIVE: Describe a local treatment for acute bacterial infectious dacriocystitis.
METHODS: Between January of 2003 and December of 2004, at the Ophthalmology Service of the Alberto Sabogal Sologuren Hospital,Essalud; Callao-Peru; 50 cases of Acute Dacriocystitis were selected. Females were predominant. The common features were acute inflammation of the lachrymal sac with tumefaction, congestion, purulent secretion and, in some cases, fistulization with external drainage. The wash of the lachrymal sac was with triamcinolone (10 mg./ml) and ciprofloxacin (0.3%), with previous anaesthesia of the sac with lidocaine 2%. In some cases we repeated the procedure after 7 days.
RESULTS: Eighty percent of cases required a single procedure, 20% repeated the procedure on the 7th day; in all cases remission of the process, and closure of fistula when it was presented, occurred before 14th day.
CONCLUSIONS: The present procedure allows the remission of acute bacterial infection of the lachrymal sac, is useful in diminishing the oral use of antibiotics, and is advisable because it allows a clean lachrymal sac, adapted for one dacriocystorhinostomy when indicated.

GONZALEZ, J.R.
Laboratorios Sophia - TX 77030 GUADALAJARA (MEXICO)

Cyclosporine-A Level in Serum after Topical Application of Eye Drops from Modusik-A Ofteno™
L.M. BAIZA-DURAN; J.A. RODRIGUEZ-TREVILLA; M.M. GONZALEZ-LOMELI; R. TORNERO-MONTAÑO
Laboratorios Sophia (Guadalajara, Mexico)

Abstract:
BACKGROUND:  Cyclosporine-A (CsA) is a peptide with potent immunosuppressive activity. Several studies indicate that CsA affects mainly proliferating T-cells. CsA has been used in ophthalmology, to improve some conditions related to dry eye syndrome. Using CsA can promote natural tears production.
AIMS: To evaluate the systemic absorption of CsA after the topical installation of Modusik-A Ofteno™ eye drops used in the treatment of dry eye syndrome.
METHODS: Modusik-A Ofteno™ consists of 0.1% CsA associated to Sophisen™ (US patent 6,071,958). 1 ml blood samples were obtained from 26 patients who received Modusik-A Ofteno™ eye drops each 12 hours during a period from 3 to 6 months. Sera from blood samples were processed for CsA with (FPIA) Fluorescence Polarization Immunoassay with antibodies against Cyclosporine-A.
RESULTS: Twenty-six patients were included in this study, all of them signed an informed consent and were receiving CsA solution eye drops through 3 to 6 months.   The mean value of CSA level in sera detected was of 1.75 ng/mL with a Standard deviation +/- 2.58 in a range 0f 0 to 9, distributed as follow 73%  (19 patients) sera level was between 0 and 2 ng/mL, 11.5% (3 patients) Shown between 3 and 4ng/mL, 7.7% (2 patients) between 5 and 6 ng/mL, 3.8% (1 patient) between 7 and 8 ng/mL and 3.8% (1 patient) between 9 and 10 ng/mL.
CONCLUSIONS: The results suggest that the systemic absorption of CsA when is applied as eye drops is minimum if we compare it with the blood level of CsA during treatment to avoid organ transplant reject or some others immunologic diseases management. This indicates that topical application of Modusik-A Ofteno™ does not reach risky CsA sera levels.

GONZALEZ, J.R.
Laboratorios Sophia - TX 77030 GUADALAJARA (MEXICO)

Creatinine and Urea Sera Levels Determination after 0.1% Cyclosporine Eyedrops Instillation
L.M. BAIZA-DURAN; J.A. RODRIGUEZ-TREVILLA; M.M. GONZALEZ-LOMELI; R. TORNERO-MONTAÑO
Laboratorios Sophia (Guadalajara, Mexico)

Abstract:
PURPOSE: To determine if kidney function is affected by instillation of 0.1% Cyclosporine-A (CsA) eye drops, serum creatinine and urea levels were determined.
METHODS: The study included 40 healthy volunteers with no eye problems. All read and signed an informed consent letter. Each volunteer received one drop of 0.1% cyclosporine (w/v) on each eye, twice a day for 60 days. Blood samples were analyzed for creatinine and urea levels before and after the 60 days of treatement.
RESULTS: The average baseline serum creatinine level was 0.94 mg/dl, and urea was 32.5 mg/dl. After 60 days of CsA treatment, the data for creatinine and urea were 0.89 mg/dl and 29.1 mg/dl respectively. The results showed no evidence of changes.
CONCLUSIONS: After 60 days of 0.1% (w/v) CsA eye drops instillation, creatinine and urea sera levels do not indicate a risky condition for kidney health.

GONZALEZ, J.R.
Laboratorios Sophia - TX 77030 GUADALAJARA (MEXICO)

Safety Efficacy and Tolerance Clinical Evaluation of a 0.1% Cyclosporine-A Eyedrops Compared to those in 0.05% Cyclosporine-A and Placebo as Treatment for Moderated to Severe Dry Eye Syndrome
L.M. BAIZA-DURAN; J.A. RODRIGUEZ-TREVILLA; M.M. GONZALEZ-LOMELI; R. TORNERO-MONTAÑO
Laboratorios Sophia (Guadalajara, Mexico)

Abstract:
INTRODUCTION: Cyclosporine-A (CsA) can improve tears production in moderated to severe dry eye patients even without Sjögren syndrome, which turn dry eye treatment in an alternative to treat them instead of lubricants eyedrops which are limited to improve symptoms, CsA promotes tears production with all its characteristics, properties and advantages.
PURPOSE: To determine and compare safety, efficacy and tolerance of two different concentrations of CsA aqueous solution and placebo eyedrops for treatment of patients with dry eye syndrome.
METHODS: We compared the effects of 0.1% CsA (group A), 0.05% CsA (group B), and vehicle or placebo (group C), were evaluated symptoms and  signs in patients affected by moderated to severe dry eye syndrome. This was a 120 days multicenter, randomized, double-blind clinical trial was preceded by two weeks wash-out period.
RESULTS: Patients receiving Modusik-A Ofteno^Ò (0.1% Cyclosporine) eye drops improved  72% in Schirmer I test compared to its own baseline, while 0.05% Cyclosporine improved 60.09% and placebo improved 32.2%. Also with Modusik-A Ofteno^Ò corneal surface conditions improved in 100% of the patients with moderated to severe dry eye syndrome. Compared to Placebo, patients with Modusik-A Ofteno^Ò improved 85% more in foreign body sensation. Also, burn sensation diminished 55% more when Modusik-A Ofteno^Ò compared to placebo. Dryness sensation diminished 40% in patients who used Modusik-A Ofteno^Ò compared to placebo. Weeping diminished 40% in patients with dry eye syndrome.
CONCLUSIONS: CsA reduced complaints and improved major ocular signs in patients suffering from moderate to severe dry eye syndrome. The group treated with 0.1% CsA aqueous solution outperformed the other 2 study groups.
Key words: Dry eye, Cyclosporine A, randomized clinical trial.

GONZALEZ, J.R.
Laboratorios Sophia - TX 77030 GUADALAJARA (MEXICO)

In Vitro Study of Corneal Retention of Cyclosporine-A from Different Formulations
J.D. QUINTANA-HAU; L.M. BAIZA-DURAN; J.A. RODRIGUEZ-TREVILLA; M.M. GONZALEZ-LOMELI; R. TORNERO-MONTAÑO
Laboratorios Sophia (Guadalajara, Mexico)

Abstract:
INTRODUCTION: Cyclosporine-A (CsA) is valuable in the treatment of diseases, such as dry eye, uveitis and corneal transplantation. Its acts by selective inhibition of interleukin-2 release during the activation of T-cells and causes suppression of the cell-mediated immune response. Several attempts were made to improve ocular CsA penetration. Modusik-A Ofteno^® is a new product developed by Laboratorios Sophia, which contains Cyclosporine-A.
PURPOSE: Compare the amount of CsA retained in cornea using an in vitro model with different formulations.
METHODS: It was compared Modusik-A Ofteno^® against Restasis™ (0.05% CsA in castor oil; Allergan, Irvine CA) and Modusik-A Ofteno^® against 2.0% CsA in olive oil as vehicle, by In Vitro passive diffusion.
RESULTS: Modusik-A Ofteno^® showed to be a better source when the total amount of CsA in the cornea was compared to 0.10% o CsA in olive oil. When the concentration of CsA was increased to 2.0% in olive oil the concentration of the drug detected in cornea was similar compared to Modusik-A. Using the same method two different concentrations of CsA (0.10% and 0.05% Modusik-A Ofteno^®) were compared to Restasis. A slight increase of CsA in cornea was observed when the donor was 0.10% Modusik-A Ofteno^® as donor.
CONCLUSIONS: Challenging 2.0% CsA in olive oil and Modusik-A Ofteno^® by passive diffusion we did not find an improvement in amount of CsA in cornea when an oil formulation was used as a donor. It appears that cornea has restricted passive diffusion capability for CsA. These results suggest that Modusik-A Ofteno^® is a proper formulation to deliver CsA to the eye.
Key words: Dry eye, cyclosporine A, passive diffusion.

E.A.G.L.E.S., Large Scale Survey about Ocular Allergy and Quality of Life in Europe
Coauthors: Sophie PAGNY (2)
Fondation Rothdchild (Paris, France); (2) Allergan France

Abstract:
OBJECTIVE: To assess ophthalmologists’ appreciation of ocular allergy symptoms, treatment and quality-of-life (QOL) perceptions.
DESIGN: EAGLES was designed as an observational study, using questionnaires for both the patients and their ophthalmologist. The survey enrolled a total of 4115 patients suffering from ocular allergy in France, Italy and Spain.
RESULTS: Majority of patients were women (61%), with a median age of 36 years. 70% had already experienced eye allergy (74% seasonal, 26% perennial). The high percentage of patients without history of ocular allergy (30%) was surprising. These could be patients having their first ocular allergy manifestation or patients with dry eye syndrome mimicking ocular allergy, since it is well known that both conditions share common clinical signs and symptoms. More than half the patients (52%) had been prescribed medications before the consultation (cromoglycate sodium: 56%; antihistaminics: 40%; tears, antiseptics solution: 31%). Most common symptoms were itching (81%), hyperaemia (64%), tearing (52%) and burning (38%). Burning sensation is not specific for ocular allergy but is often present since ocular allergy is always more or less associated with dry eye syndrome. From the QOL data, the activities causing most concerns were reading/writing or using a computer(50%), travelling or outdoor activities (31%). 48% reported feeling very tired and 44% experienced irritability and about 30% of patients declared feeling socially embarrassed. Patient’s first expectation is a treatment for a fast and effective relief (80%). Most prescribed treatment category was the new generation of anti-allergic eye drops, antihistamines with mast cell stabilizing properties, sometimes associated with steroids and artificial tears.
CONCLUSIONS: Ocular allergy affects patients in their quality of life and everyday activities, including office working and social relationships. Expectations from the patients for a fast and effective relief match with the new generation of anti-aller.

Flow Cytometric Analysis of Conjunctival HLA-DR Expression in Patients with Superior Limbic Keratoconjunctivitis
Coauthors: Anna M. THEISL; Hans J. HEINTZ *; Eva-Maria HALLER-SCHOBER; Otto SCHMUT; Reingard M. AIGNER *
Dept. of Ophthalm., Medical University (Graz, Austria); *Dept. of Radiology, Division of Nuclear Medicine, Medical Univ. (Graz, Austria)

Abstract:
PURPOSE: Superior limbic keratoconjunctivitis (SLK) is a rare disease of the upper bulbar and tarsal conjunctiva and cornea. The most commonly recognized theory concerning the pathogenesis is based on increased friction between tarsal and bulbar conjunctiva. Another theory postulates an autoimmune process. Therefore, the expression of the inflammatory marker HLA-DR on conjunctival epithelial cells was determined.
PATIENTS and METHODS: 12 Patients with SLK were compared to 15 healthy persons, to 15 patients with Sjögren syndrome and to 15 dry eye patients without Sjögren syndrome. HLA-DR expression on the bulbar conjunctiva was determined after brush cytology using quantitative flow cytometric analysis. Tear breakup time and Schirmer test without local anesthesia were performed and the ocular surface was assessed by fluorescein and lissamin green staining, as well as impression cytology.
RESULTS: Patients with SLK had a significant higher expression of HLA-DR on the conjunctival epithelial cells compared to the healthy control group. Highest values were observed in patients with Sjögren syndrome. There was no significant difference of HLA-DR expression between SLK patients and dry eye patients without Sjögren syndrome. SLK patients showed a significant decrease in tear breakup time and slightly reduced Schirmer test values. The morphology of the conjunctiva observed by impression cytology showed extreme pathological changes.
CONCLUSIONS: Increased HLA-DR expression in patients with SLK suggests an inflammatory component in the pathogenesis of this disease. Anti inflammatory therapy might be an additional benefit for these patients.

Structural and Functional Aspects of Collectins SP-A and SP-D in the Lacrimal System
Coauthors: Kristin RECKER (1); Dirk EHRICH (2); Saddettin SEL(2); Friedrich PAULSEN (1)
(1) Depts. Of Anatomy&Cell Biology and (2) Dept. of Ophthalmology, Martin-Luther-Univ. Halle (Halle, Germany)

Abstract:
Surfactant proteins (SP)-A and D are collectins, a family of collagenous carbohydrate binding proteins (collagenous C-type lectins). They are importants components of innate immune response and participate in aspects of immune and inflammatory regulation. SP-A and AP-D can facilitate phagocytosis by opsonizing bacteria, fungi and viruses, stimulate oxidative burst by phagocytes and modulate proinflammatory cytokine production. Both collectins can also provide a link between innate and adaptive immune response by promoting differentiation and chemotaxis of dendritic cells. Our study examined the expression, production and regulation of SP-A and -D in lacrimal gland, conjunctiva, cornea and nasolacrimal ducts. Espresion of SP-A and -D was amplified by RT-PCR to total mRNA in all tissues. Proteins were detected by Western blot analysis in all samples. Immunohistochemical analysis revealed both collectins in lacrimal gland acinus cells as well as conjunctival epithelial cells whereas corneal epithelial cells did not stained positive. Furthermore, we compared the relative expression of SP-A and -D in healthy nasolacrimal ducts with cases of dacryostenosis by real-time PCR. The expression level was much higher in pathological tissues. Our studies suggest that SP-A and -D play an important role in the immune response of the ocular surface and lacrimal system.
Supported by Wilhelm-Roux-program FKZ 09/18 and DFG PA738/1-5

Abstract:
PURPOSE: To study the changes of tear production before and after IEMF influence in case of dry kerato-conjunctivitis.
METHODS: 92 patients (182 eyes) with dry kerato conjunctivitis (DKC) were observed. The treatment with IEMF was recommended to all patients.
RESULTS: At early stages of dry kerato-conjunctivitis Schirmer probe was (9.0 + 0.25) mm, but after IEMF influence it was (13.0 + 0.38) mm (p < 0.001). In patients with DR indications Schirmer probe was (55.6 + 0.31) mm before and after influence of impulse electromagnetic field. Break-up-time of lacrimal membrane was 9,7 +0.21 /sec/ at the mild stage of disease, then it was increased to 13,6 + 0.32 /sec/ after IEMF influence. At moderate stage of disease, break-up-time was 9,1 + 0.26 /sec/, after IEMF influence it was increased to 11,2 + 0.44 /sec/. At severe stage of disease break-up-time was 7,5 + 0.28 /sec/, after IEMF influence it was increased to 9,2 + 0.48 /sec/.
CONCLUSION: The reliable increasing of break-up-time in all patients with DKC points out the stability of lacrimal membrane has been improved under IEMF influence from the components, responsible for tear production.

Punctal Plug Occlusion for the Treatment of the Epithelial Keratitis after Keratoplasty
Coauthors: Hitoshi WATANABE; Kohji NISHIDA; Naoyuki MAEDA; Hitoshi HAYASHI; Yasuo TANO
Dept. of Ophthalm., Osaka University Medical School (Osaka, Japan)

Abstract:
PURPOSE: To investigate whether the punctal plug occlusion is effective for the treatment of intractable superficial keratitis after penetrating keratoplasty.
METHODS: Eight eyes of eight patients with intractable keratitis after penetrating keratoplasty were studied. A silicone punctal plug was inserted in both the upper and lower punctum of the operated eye. The ocular surface was evaluated based on the fluorescein staining score of the punctate epithelial erosions on the grafted cornea before and after the implantation of the plugs. Schirmer’s test was also performed and the efficacy of the punctal plug occlusion was evaluated.
RESULTS: Punctal plug insertion was of benefit to the superficial keratitis of postkeratoplasty eyes. There appeared to be a progressive improvement and there was a significant improvement between before and after the treatment (p<0.001, one way repeated measures ANOVA). Improvement of the epithelial keratitis on the grafted cornea was relatively rapid in the first 2 weeks after plug insertion, in accordance with the increase in tear volume and better distribution of the tear film on the grafted cornea. The healing rate of the keratitis on the corneal graft became slow after 2 weeks as the tear volume remained steady value.
CONCLUSIONS: Punctal plug occlusion is an effective treatment for the intractable superficial keratitis in patients who underwent penetrating keratoplasty. Placement of punctal plugs improves the inadequate distribution and enables the tear film to cover the entire corneal graft by increasing the tear volume in a relatively short period of time.

Efficacy of Artificial Tears Solution, Plunctual Plug Therapy and Oral Pilocarpine for the Treatment of Dry Eye in Patients with Sjögren Syndrome: a Randomised 24 Week Controlled Study
Coauthors: V.T. DIAZ ALEMAN; G. FERNANDEZ-BACA VACA; D. PERERA SANZ; J. RODRIGUEZ MARTIN

Abstract:
PURPOSE: To evaluate artificial tears solution, punctual plug therapy and oral pilocarpine for the treatment of dry eye in patients with Sjögren syndrome.
METHODS: A 24 week, randomised controlled study was performed. 30 patients with dry eye were included, 10 of the patients were randomly assigned to receive artificial tears, 10 received superior puncta occlusion and 10 received oral pilocarpine, 5 mg four times a day. Patients were evaluated at baseline and throughout the study every 4 weeks for their subjective global assessment of dry eyes and for their objective assessment of dry eyes, with Schirmer’s I, II tests, fluorescein test and rose Bengal test.
RESULTS: Most of the patients had significant improvement in subjective assessment of dry eyes, but patients treated with superior puncta occlusion showed greater objective improvement as measured by fluorescein test and patients receiving oral pilocarpine showed the greatest objective improvement, measured by the rose Bengal test, while Schirmer’s I, II tests showed no differences between the treated groups. Although in patients who had been taking oral pilocarpine, several adverse effects were observed, diaphoresis was the most commonly reported.
CONCLUSIONS: The use of 5 mg of oral pilocarpine four times a day had a beneficial effect on subjective eye symptoms, in addition an improvement of rose Bengal staining was noted, but adverse effects limited its use. While punctual plug therapy showed similar results and was well tolerated.

Endoscopy Study of the Intranasal Ostium in External Dacryocystorhinostomy Postoperative and Influence of the Saline Solution and the 5 Fluorouracyl
Coauthors: Ana Maria MARCONDES, PhD; José NEWTON KARA, PhD
Depts. Of Otorhinolaringology&Ophthalmology, Universidad Estadual de Campinas (Campinas, Brasil)

Abstract:
PURPOSE: The present study aims at studying, through the endoscopy, the structural changes of the intranasal ostium that take place in the external dacryocystorhinostomy postoperative, and to evaluate the influence of the use of the saline solution and the 5 FU in the evolution of the ostium, seeking a complementary alternative for the treatment of bad prognosis dacryocystitis.
METHODS: There were 50 patients distributed in following groups: Group I - saline solution - 13 patients underwent dacryocystorhinostomy with the use of saline solution during the surgery; Group II - 5 fluorouracil - 17 patients underwent dacryocystorhinostomy and an injection of 5 FU during the surgery; Group III - control - 11 patients underwent only surgical procedures; Group IV - 5 fluorouracil - 9 patients underwent dacryocystorhinostomy associated with three injections of 5 FU.
RESULTS: The evaluation of each one of the groups through the Wilcoxon test revealed there was a significant reduction of the ostium area only in the control group during the 30th and 60th days’ postoperative (p<0,05). The comparative study between the four groups through the Kruskal-Wallis test did not show significant changes in the first and second month after the surgery. The difference in area during the two postoperative months in the groups was also not significant.
DISCUSSION: This study makes clear that the size of the final surgical ostium is always smaller than the initial surgical orifice. The option to use the 5 FU was based on the fact that this anti-metabolite have had positive results in ocular surgeries, when the main objective is to interfere in the healing processe but there was not significant changes in the first and second month. The difference in area during the two postoperative months in the groups was also not significant.
CONCLUSIONS: The result of this study suggest that the use of 5 FU in external DCR does not influence, in a significant way, the final size of the surgical fistula during the two months evaluation.

Use of Hyaluronidase in the Idiopatic Functional Blockage of the Lacrimal Drainage System
Coauthors: Ana Carolina FAVA SALATA, MD; Felipe DO CARMO CARVALHO, MD; Ana Maria MARCONDES, PhD
Depts. Of Otorhinolaringology&Ophthalmology, Universidad Estadual de Campinas (Campinas, Brasil)

Abstract:
PURPOSE: To study the use of irrigations of hyaluronidase for the treatment of idiopatic blockage of the lacrimal drainage system.
METHODS: There were made applications in 13 lacrimal systems (10 patients) with functional blockage. The diagnose was based on probing and on the dacryocystography. The radiological exams were taken before and after the applications, and the patients were asked about the improvement of the symptoms after them. Each lacrimal system was irrigated with 2 ml of hyaluronidase solution, in four sessions, with an interval of two days.
RESULTS: There were clinical and radiological improvement in 75% lacrimal systems.
CONCLUSIONS: Treatment of the idiopatic functional blockage with hyaluronidase can be a new therapeutic option.

Use of Hyaluronidase in the Functional Blockage of the Lacrimal Drainage System
Coauthors: Alessandra KOBAIYASHI, MD; Marcos DANTAS, MD; Ana Maria MARCONDES, PhD
Depts. Of Otorhinolaringology&Ophthalmology, Universidad Estadual de Campinas (Campinas, Brasil)

Abstract:
Epiphora in the absence of anatomically obstructive lesion is considered as a functional blockage of the lacrimal drainage system.
PURPOSE: The objective of this study is to assess the effect of hyaluronidase irrigations in the treatment of idiopatic functional blockage.
METHODS: The diagnosis was based on a clinic exam (biomicroscopy, probing and lacrimal irrigation), dacryocystography (DCG) and dacryoscintigraphy (Dci). Thirty one patients underwent irrigations comparing the effect of hyaluronidase, destilled water and saline solution.
RESULTS: Subjective improvement was referred by 10 (83,3%) of hyaluronidase group, 4 (44,4%) of destilled group and 5 (50%) of saline solution. Objective improvement occurred in 50% in hyaluronidase group, 11,1% in destilled group and 10% in saline solution.
DISCUSSION: No statistically significant difference for hyaluronidase (p=0,1956) was observed when comparing subjective improvement among the groups. There was not also statistically difference comparing the objective improvement among the groups (p=0,182).

Distribution of Somatostatin and Somatostatin Receptors in the Lacrimal System
Coauthors: Rolf MENTLEIN (2); Kristin RECKER (1); Christian KINDLER (1); Sadettin SEL (3)
(1) Dept. of Anatomy and Cell Biology, Martin-Luther-Univ. of Halle-Wittenberg (Halle-Germany); (2) Dept. of Anatomy, Christian Albrecht Univ. Of Kiel(Kiel, Germany); (3) Dept. of Ophthalmology, Martin-Luther-Univ. of Halle-Wittenberg (Halle-Germany)

Abstract:
Recent investigations revealed the presence of somatostatin (SS) in the excretory system of the lacrimal gland and in tear fluid. To get deeper insights into a possible role of SS at the ocular surface and in the lacrimal system the distribution pattern of SS and its receptors 1-5 (SSTR1-5) were investigated by means of RT-PCR, Western-blot analysis and immunohistochemistry in lacrimal gland, tear fluid, conjunctiva, cornea, nasolacrimal duct epithelium as well as conjunctival and corneal epithelial cell lines. Cell culture experiments with corneal epithelial cell lines were undertaken to analyze a possible impact of SS on the activation of MAP-kinases ERK1/2. The results confirmed the presence of SS in lacrimal gland and tear fluid. Expression of SSTRs1-5 was visible in lacrimal gland, whereas immunohistochemistry detected only SSTRs 1, 2 and 5. These latter receptors were also detected in conjunctiva, cornea, nasolacrimal ducts as well as corneal and conjunctival epithelial cell lines by RT-PCR and immunohistochemistry. Western blot analysis confirmed the presence of SSTR2 in all investigated tissues. Cell culture experiments with corneal epithelial cells revealed that SS alone had nearly no effect on ERK1/2 phosphorylation but a strongly increasing activation was seen when SS was used in combination with VEGF or EGF. This effect was further enhanced if SS was replaced by SS analoga like octreotid or L054.522. These primary data support an autocrine and paracrine role of SS in the lacrimal system and at the ocular surface and implicate a role of SS in corneal angiogenesis.
Supported by DFG program grant PA 738/6-1 and Wilhelm-Roux-program FKZ 09/17

VEGF and its Receptors in Pterygium
Coauthors: Matthias GEBHARDT(1); Rolf MENTLEIN (1); Ulrich SCHAUDIG (2); Thomas PUFE (1); Kristin RECKER (3), Bernhard NÖLLE(4), Kais AL-SAMIR (2), Gerd GEERLING (5)
(1) Dept. Of Anatomy and (4) Dept. of Ophthalm., Christian Albrecht Univ. Of Kiel; (2) Dept. of Ophthalm., Univ. Hosp. Hamburg-Eppendorf; (3) Dept. of Anatomy&Cell Biology, Martin-Luther Univ. Of Halle-Wittenberg; (5) Dept. of Ophthalm. Univ. Of Lübeck (Germany)

Abstract:
The study was undertaken to evaluate the role of vascular endothelial growth factor (VEGF) and its receptors VEGFR1 and VEGFR2 in pterygia and compare it with healthy conjunctiva. Tissue specimens from patients treated for primary pterygia were analyzed using immunohistochemistry and molecular biological methods. Healthy conjunctiva from specimens treated for cataracts, as well as specimens from the conjunctiva, limbus and lens of both eyes of body donors served as controls. Surgical specimens of pterygiac and normal conjunctiva specimens were examined by histology using antibodies against VEGF and its receptors, and. The other part of both groups of specimens as well as specimens of body donors were prepared and analyzed by means of RT-PCR, real-time-PCR, ELISA and Western-blot. VEGF and VEGFRs were analyzed to identify the splice variants of VEGF as well as the distribution and amount of VEGF and both receptors in pterygia and the control tissues. In analysis of specimens from pterygia patients as well as normal conjunctiva VEGF121 and VEGF165 were identified as the only VEGF splice forms expressed. In addition to VEGF, VEGFR1 and VEGFR2 were detected in pterygium and conjunctiva and immunostained within the epithelium of pterygia and conjunctiva and on intrapterygial and intraconjunctival endothelial cells. Levels of VEGFR-1 and -2 mRNA were lower in pterygia than in conjunctiva but similar in limbal and pterygium samples. VEGF levels were higher in pterygia than in conjunctiva, but were similar in the limbus and pterygia. The results strongly support the presumption that pterygia arise from limbal epithelial cells and that human conjunctiva is not a suitable control for the analysis of pterygia. Moreover, the results suggest that VEGF might play anactive role in the physiology of conjunctival epithelial cells.
Supported by DFG program grant PA 738/6-1 and Wilhelm-Roux-program FKZ 09/17

Mucins and TFF-Peptides of Human Lacrimal Gland
Coauthors: Gesa SCHÄFER (2); Werner HOFFMANN (2); Monica BERRY (3)
Dept. of Anatomy and Cell Biology, Martin-Luther-Univ. of Halle-Wittenberg (Halle-Germany); (2) Inst. of Molecular Biology&Medical Chemistry, Otto von Guericke Univ. (Magdeburg, Germany); (3) Division of Ophthalm., Univ. Of Bristol (Bristol, UK)

Abstract:
The Objective of this study was to determine whether the lacrimal gland synthesizes mucins and TFF (trefoil)-peptides and whether mucins are changed with age or in cases of dry eye. Expression and distribution of mucins and TFF-peptides in human lacrimal glands was monitored by reverse transcription-polymerase chain reaction analysis, dot blot and immunohistochemistry. Normal tissues from cadavers were investigated plus four with dry eye condition treated with artificial tears. Expression studies detected mRNAs for mucins MUC1, -4, -5AC, -5B, -6 and -7. The message for MUC4 was present in cases of dry eye, but only in some normal glands from individuals. MUC6 mRNA was detected only in about half of the investigated samples. Immunoblot analysis detected MUC1, -4, -5AC, -5B, -7. immunohistochemistry revealed membrane-bound MUC1 at the apical surface of acinal cells, MUC5AC associated with goblet cells of excretory ducts, MUC5B and -7 in the cytoplasm of acinar cells, and MUC7 also in epithelial cells of excretory ducts. MUC4 mucin was detected only in those individuals in which message was identified. In dry eyes, MUC5AC and -5B were localized in the same acinar cells. Dot-blot analysis clearly revealed increased amounts of MUC4, -5AC, and -5B in the glands of persons who received treatment for dry eyes. Expression studies detected mRNAs for mucins TFF1 and -3. immunohistochemistry revealed for TFF3 in some lacrimal acinar cells. The results confirm that the human lacrimal gland synthesizes a spectrum of mucins , part of them might be correlated with age. The production of TFF3 by the lacrimal gland is unclear at present. Changes in the mucin production in cases of dry eye support the assumption that lacrimal gland mucins are involved in the pathophysiological events that occur at the ocular surface.
Supported by DFG grant PA 738/1-4, National Eye Research Center (Grant 2000/13) and Sicca-Forschungsförderung

NGF Effect on Epithelial Cell Cultures
Coauthors: V. BARBARO; D. MERLO; M. DE LUCA; S. BONINI; A. LAMBIASE
Epithelial Regional Research Center, Veneto Eye Bank Foundation (Venice, Italy); Dept. of Ophthalm., University "Campus Biomedico" (Rome, Italy)

Abstract:
The aim of this study is to evaluate the effects of NGF on primary culture of human corneal and conjunctival epithelia.
NGF concentration was evaluated by ELISA in culture medium of corneal and conjunctival epithelial cells at different time. NGF levels were related to proliferation and differentiation markers of the cells culture. In order to investigate the effect of NGF on proliferative compartment, exogenous NGF at concentrations between 0,01 to 250 ng/ml was added to the culture medium. The total colony forming efficiency and the percentage of aborted colonies were evaluated at different NGF concentrations. The inhibition of endogenous NGF was evaluated in parallel cultures by adding neutralizing anti-NGF antibody in concentration ranging between 100 to 250 ng/ml. Pathways of NGF involved were analysed by culturing conjunctival epithelium in presence of increasing concentration of NGF and antibodies against the TrkA and the p75 receptors. Effects on differentiation was investigated by RT-PCR analysis, immunocitological stainings and western blot analysis on cell pellets and culture medium.
Primary culture of human corneal and conjunctival epithelial cells produce and release endogenous NGF in the culture medium in a time-dependent fashion during clonal growth. The concentration of NGF in the medium raise a 3-5 fold increase before confluence of both conjunctival and corneal cultures with a corresponding increase in the total number of daughter colonies. Addition of exogenous NGF did not show an increase in p63 highly positive cells, but showed an increase in 14-3-3 sigma expressing cells and MUC 5AC expression levels, suggesting a effect on differentiation of conjunctival cells. The biological effect was reversed by the antibody reacting with endogenous NGF. On going study suggests that different NGF receptors may differentially modulate conjunctival cell differentiation.
This study confirms the role of NGF in the pathophysiology of the ocular surface. Moreover the demonstration of NGF role in the differentiation of conjunctival epithelium highlights potential involvement of this neurotrophin in a wide spectrum of ocular surface diseases.

Use of D.E.Q. to Evaluate Symptoms of Ocular Dryness in a University Population
Coauthors: Gonzallo CARRACEDO; Inmaculada FERNANDEZ; Jesús PINTOR

Abstract:
PURPOSE: The present study tries to check up on the presence of dry eye problems and also to characterize the typical ocular symptoms associated to this pathology in a random university population.
METHODS: Two different questionnaires were provided, one for non-contact lens wearers (dry eye questionnaire) and one for contact lens wearers (contact lens dry eye questionnaire). These questionnaires were filled by the individuals at the School of Optics and Optometry. Both forms included scales to measure the prevalence and frequency of different ocular surface symptoms.
The sample of the study comprised 115 individuals. Eighty two of them completed the dry eye questionnaire and 33 filled the contact lens dry eye questionnaire. We consider symptomatic patients those who reported any symptom even if it was infrequent.
RESULTS: The most common ocular symptoms reported from the dry eye questionnaire were tired eyes (90%); light sensitivity and how uncomfortable were different air qualities (85% and 80% respectively); the necesity of closing the eyes (68%) and eye discomfort (62%). Some of these symptoms, tired eyes and eye discomfort, experimented an increase in intensity in the evening.
For the contact lens dry eye questionnaire the most frequent symptoms were eye dryness (90%); eye discomfort (87%); blurry vision with a percentage of appearance of 78%, and irritation (72%). In this case, all the symptoms were more obvious at the end of the day.
CONCLUSIONS: Our results show that symptoms of tired eyes and air qualities as well as eye dryness and blurry vision are relatively frequent in this random university population. The intensity of these symptoms appear to increase at the end of the day.

Peribulbar Complex Choristoma in Association with Ipsilateral Nasal Hypoplasia and Lacrimal Obstruction
Coauthors: Z. TABATABAI, MD; A. SADEGHI, MD; A. KASAI, MD; A. RAHIMI, MD

Abstract:
We encountered a 3-year-old boy with epiphora, a striking mass on the nasal bridge and ipsilateral nasal hypoplasia. CT-Scan imaging showed multiple calcified areas within the tumor in addition to linear defect in frontal bone, hypoplastic left ethmoidal sinus and left nasal cavity, and absence of left nasal concha. The patient had no history of seizure and neurologic deficit. Ocular developmental exams were normal. After performing excisional biopsy of the tumor, histopathologic analysis revealed complex choristoma composed of cartilage and bone. The most appropriate name for this malformation, which we have not found described in the literature, seems to be "nasal hypoplasia with complex choristoma".

Is the y+-Transporter Involves in the Pathogeneses of Pterygia?
Coauthors: Christian KINDLER; Friedrich PAULSEN
Dept. Of Anatomy&Cell Biology, Martin-Luther-Univ. Halle (Halle, Germany)

Abstract:
The junction at the eye between the cornea and conjunctiva/sclera is called the limbus. It contains epithelial stem cells and is a barrier that prevents overgrowth of the conjunctiva onto the corneal surface. A pterygium is an invasive and highly vascularized growth, thought to arise from activated and proliferating limbal epithelial stem cells. The pathogenesis of pterygia is uncertain at present. Epidemiologic studies have implicated environmental factors such as ultraviolet light, chronic irritation, and inflammation as causative factors. Other popular hypotheses postulate modifications in apoptosis pathways as well as involvement of matrix metalloproteinases and their inhibitors. Recent investigations have shown an activation of different inflammatory gene products like for example trefoil-faktor-peptides (TFF-peptides) or inducible nitricoxide-synthase (iNOS) in pterygia. By means of RT-PCR, Western blot and immunohistochemistry our examinations reveal the relevance of the important cationic amino acid transporter in mammalia cells, the y+-system which transports the substrate of iNOS, l-arginine. The y+-system was analyzed in different tissues of the lacrimal system and ocular surface (lacrimal gland, conjunctiva, cornea, nasolacrimal ducts), in cell lines of corneal and conjunctival origin and in pterygia. We could show that the proinflammatory cytokinesIL-lb, TNFa and INFg that are upregulated in pterygium pathology upregulate the y+-transporter-expression in epithelial conjunctival cells suggesting an involvement of the y+-system in pterygium pathology. Quantification of mRNA was done by real-time-RT-PCR. The investigations shall open new prospects of preexisting therapeutical concepts in the treatment of pterygia.

Impression Citology Study of Corneal Epithelium Reconstructed by Limbal Transplantation, Preserved Human Amniotic Membrane, and Cultivated Limbal Cell Transplantation in Patients with Limbal Stem Cell Deficiency
Coauthors: S. LOPEZ GARCIA; E. CORREDERA

Abstract:
OBJECTIVE: To determine by impression cytology the morphological changes of corneal cells reconstructed by limbal allo-transplantation (LAT), amniotic membrane transplantation (AMT) and cultivated limbal cell transplantation (CLT) in patients with limbal stem cell deficiency.
METHODS: Impression cytology was performed in 14 eyes from 14 patients with LAT, in 17 eyes from 14 patients with AMT and in 6 eyes from 6 patients with CLT. These consisted of 7 eyes with Stevens-Johnson syndrome, 10 with chemical injuries, 6 with ocular cicatricial pemphigoid and 11 with aniridia. All of these eyes had total stem cell deficiencies. Corneal impression cytology was collected: prior to transplantation, after 3, 6, 9, 12, 24 and 36 months. The grade of squamous metaplasia was evaluated in corneal and limbal cells. Limbal deficiency was diagnosed if conjunctival goblet cells were found on the corneal surface.
RESULTS: There were not significant differences in impression cytology between any disease’s groups. Eyes with LAT showed a continuous decrease in squamous metaplasia and no limbal deficiency after surgery. In eyes with AMT there were a continuous decrease in squamous metaplasia without limbal deficiency, but 6 months after surgery appeared a significant increase of squamous metaplasia and limbal deficiency. In CLT eyes there were similar findings that than LAT.
CONCLUSIONS: Impression cytology can be used to diagnose and monitor corneal diseases with limbal deficiency and indicates that additional limbal stem cell transplantation is needed for effective corneal surface reconstruction in patients with limbal deficiency. AMT only can be used for severe stem cell deficiencies during 6 months.

Lacrimal Obstruction: Technical Options and Results
Coauthors: J. MAESO RIERA; J. FALCO FAGES; E. CAPELLA PEREZ; C. PERMANYER FUSTER

Abstract:
PURPOSE: Try to find more easy and harmless techniques to achieve a good long-term result in the recovery of a normal lachrymal function.
METHODS: We present our results on three treatment groups: Group 1 (G1): 350 endonasal dacryocystorhinostomies (EN-DCR); Group 2 (G2): 75 external dacryocystorhinostomies (EX-DCR); and, Group 3 (G3): 87 lachrymal stents. All procedures have been performed between April 1995 and December 2001. Patients have been followed up by clinical criteria (G1, 2 and 3), nasal endoscopy (G1), lachrymal irrigation (G2 and 3) dacryocystography (G3). The mean follow up period has been 16 months.
RESULTS: After one year follow-up our success rate has been: 96,57% (338/350) for EN-DCR, 88,67% (65/75) for external DCR and 62,06% (54/87) for lachrymal stents. The most frequent failure causes have been formation of a nasal mucosal bridges in surgical cases, and incasion of the prosthesis by fibrous tissue in stents.
CONCLUSIONS: Nowadays we have enough alternatives to try to achieve a functional solution for each case of lachrymal obstruction. No technique can assure long term success in all the cases, but there are some techniques with a comparable effectiveness. The choice ought to depend on the surgeon’s experience.

Comparison of Cell-suspension and Explant Culture of Rabbit Limbal Epithelial Cells
Coauthors: Xiaomin ZHANGA; Xin TANGA; Jian JIA; Xiaorong LIA; Jing SUNA, Zhiwei MA, Jiaqin YUAN, Zhong Chao HAN
Eye Centre of Tianjin Medical College (Tianjin, People’s Republic of China)

Abstract:
Currently, most investigators directly use limbal explants to culture corneal epithelial cells. However, it has not been identified that limbal stem cells do readily migrate from the limbal explants onto culture plate or amniotic membrane carrier. In this study a cell-suspension culture system for rabbit limbal stem cells was developed and compared with the direct explant method in the aspect of stem cells content in the culture system. Rabbit limbal epithelial cells were dissociated from rabbit eyes by dispase and single cell suspension was made for cell-suspension culture. ?Np63 expression of cultured rabbit limbal epithelial cells by cell-suspension technique and explant explant technique was detected. In cell-suspension culture, isolated cell-suspension was evaluated by flow cytometric analysis for vimentin expression and residual limbal tissue after dispase treatment was examined by scanning electron microscopy. In limbal epithelial cells suspension less than 5% cells were vimentin positive. Examination of residual limbal tissue confirmed that all the limbal epithelial cells had been removed. Histological examination revealed that with cell-suspension culture the cultured epithelial cells could differentiate better than with explant technique. In cells cultured with cell-suspension, there were much more cells expressing ?Np63 than in explant cultured cells. In cells cultured with explants, most of ?Np63 labelling cells mustered around the explants, and peripheral cells on the slides were ?Np63 negative. These results suggested that with pure limbal epithelial cells suspension including basal cells, which could directly enter into culture system, cell-suspension culture technique was significantly superior to explant culture technique in terms of stem cells content.
Keywords: limbal; stem cells; cornea; rabbit; ?Np63; cell suspension

Evaporation Rate from an Artificial Tear Meniscus
Coauthors: B.J. WALLACE
Nuffield Laboratory of Ophthalmology, University of Oxford (Oxford, U.K.)

Abstract:
PURPOSE: All previous studies on evaporation consider loss of water from the thin but large pre-ocular and the resultant hyperosmolarity. The compartment hypothesis (Bron et al. Adv. Exp Med Biol 2002;506:1087-95) also considers the marginal meniscus, which has a significant area but a smaller area/volume ratio, so that evaporative loss has less influence on hyperosmolarity. We devised an artificial meniscus to study aspects of this problem.
METHODS: A thread of liquid was placed in the angle between two glass microsope slides, cemented together at right angles so that 2mm of the lower slide projected (representing the lid margin) while the upper slide represented the near-vertical ocular surface. Meniscometry images were obtained at the slit-lamp biomicroscope using the Tearscope with an added striped target, and the radius of curvature of the meniscus was calculated relative to images of standard glass tubes. A series of photographs was taken as evaporation was allowed to take place over 14-15 minutes at room temperature.
RESULTS: The dependence of meniscus radius on fluid volume was found to deviate little from a circular-quadrant model over the range of radii found in the normal human eye, so volume and surface area could be calculated from a known radius during drying. The mean rate of evaporation was aproximately 8.9 nl/mm2/min, or 148 x 10-7 g/cm2/sec in the units usually used for the whole eye.
CONCLUSIONS: The measured rate is for bare water without a surface layer of lipid. Data for human eyes without lipid is lacking, but it is estimated that the meibomian lipid layer reduces the rate by between four- and twenty-fold, which agrees well with reported whole-eye values. For the whole eye, the total lenght of meniscus is about 50mm or roughly 10 mm2, compared to about 260 mm2 for the pre-ocular film. Our result shows that the artificial meniscus can give useful information about evaporation from the tear meniscus, and will be modified to allow measurement of the effect of surface lipid layers.

TORRES SUAREZ, E.
Hopital Universitario Ramón y Cajal – MADRID (SPAIN)

Massive Lacrimal Sac and Ductus Dilatation
C. LOPEZ CABALLERO; M. SALES SANZ; E. MATEOS SANCHEZ
Hospital Universitario Ramón y Cajal (Madrid, Spain)

Abstract:
INTRODUCTION: Obstructive epiphora is usually a clinical sign of chronic dacryocystitis. In most of cases, diagnosis of obstructive epiphora is possible with a proper anamnesis, slit lamp examination and lacrimal pathway syringing, although sometimes we are obliged to perform imaging tests such as dacryocystography and CT examination to rule out nasosinusal disease.
CASE REPORT: We present the case of a 70-year-old woman with periocular pain and progressive left inferior eyelid swelling of two weeks´ duration. She also complained of chronic tearing of her left eye. A CT scan of the orbits  revealed a big mass in the lacrimal area with erosion of the floor and medial orbital wall.
DISCUSSION: CT allows to differentiate between dacryocystocele and maxillar mucocele. Correct interpretation of the radiologic anatomy is essential to achieve the right diagnosis.

Validation of Two Dry Eye Experimental Models in Rabbits
Coauthors: José M. BENITEZ DEL CASTILLO (1); Rocío HERRERO (2); Irene MOLINA (2); Julián GARCIA-SANCHEZ (1);
(1) Hospital Clínico San Carlos; (2) Facultad de Farmacia UCM (Madrid, Spain)

Abstract:
PURPOSE: To evaluate two dry eye experimental models in rabbits: hyposecretory and evaporative.
METHODS: 7 females NZ albino rabbits weighting 3-4 Kg were used to develop both experimental models. The hyposecretory model was developed by instillation of one drop 1% atropine sulphate t.i.d in one eye for one month. To develop the evaporative model the rabbits were anaesthetized with 2 ml of im ketamine HCI and 1 ml of im xilocine and one eye was kept open using a lid speculum (temperature 21ºC and humidity 48% were constant). Dry eye was assessed by Schirmer’s test (in the first model) and by examination of corneal staining using fluorescein (in both models).
RESULTS: After one month the hyposecretory model induction was not achieved. Neither Schirmer’s test reduction nor fluorescein staining was obtained. The evaporative model was achieved in all the animals. The keratitis duration increased with the repetition of the evaporative model induction.
CONCLUSIONS: In our hands, the atropine hyposecretory dry eye model could not be inducted, nevertheless, we could develop an evaporative dry eye model useful to investigate new therapeutic alternatives.

Evolution of the Lacrimonasal Duct in Vertebrates
Coauthors: Juan Pablo ALDECOA; Guillermo FRIDRICH

Abstract:
PURPOSE: To show the difference of the lacrimonasal bone duct, between five kinds of vertebrates, in different branches of his evolution.
METHODS: We presented the head of the case, his skull, showing the lacrimonasal duct introducing a wire made of steel, in throug the duct. In a case of an Alligator, Weasel, Sheep, Beaver, Monkey. We take X-rays, in a frontal and a lateral view, and in the case of Man were we use a Dacriocistography.
RESULTS: We can see the different leanings of the lacrimonasal duct, in the less developed are more horizontal, changing the angle, to a complete horizontal in man, opposed as we see with the cribosa plate which shows an inverse mechanism.
CONCLUSIONS: We believe is interesting for specialist in lacrimal studies, to observe the variation of the lacrimal evacuation.

Decreased Tear Lipocalin Concentration in Patients with Meibomian Gland Dysfunction
Coauthors: Hiroshi MOCHIZUKI (1, 2); Masataka KAWAI (1); Kazuo TSUBOTA (1); Thomas J. BRYCE (3)
(1) Dept. of Ophthalm., Keio Univ. School of Medicine (Tokyo, Japan); (2) Div. For Vision Research, National Inst. of Sensory Organs, National Tokyo Medical Center (Tokyo, Japan); (3) Dept. of Health Sociology, Tokyo Univ. Graduate School of Medicine (Tokyo, Japan)

Abstract:
PURPOSE: Recent studies have demonstrated that tear lipocalin (TL) ans phospholipids play a crucial role in maintaining tear film stability. We examined the level of TL in patients with meibomian glans dysfunction (MGD), and correlated these data with the severity of their clinical disorder.
METHODS: Twelve patients with obstructive MGD, 12 patients with seborrheic MGD and 12 age-matched normal control subjects participated in this study. Three microliters of unstimulated tears were collected with a micropipette from the inferior tear meniscus in the right eye of all subjects. Tear samples were fractionated by high-performance liquid chromatography, and TL concentrations were assayed with a bicinchoninic acid technique.
RESULTS: The mean concentration ± S.D. of TL in patients with obstructive MGD and seborrheic MGD was 0.89 ± 0.19 mg/mL, and 1.05 ± 0.22 mg/mL, respectively. In both groups, mean TL concentration was significantly lower than that of normal controls (p = 0.00021 and p = 0.00042, respectively, Mann-Whitney U-test). There were no significant differences in total protein concentration among the three groups. TL concentrations were not significantly correlated with the results of the Schirmer or cotton thread tests, or with the rose bengal staining score, but did correlate positively with tear film BUT (r = 0.67) and fluorescein staining scores (r = -0.74).
CONCLUSIONS: Our results suggest that TL deficiency may be a predisposing factor for the manifestation of symptoms in MGD.

The Evaluation of Corneal Changes Wearing Soft Contact Lens by Confocal Microscope
Coauthors: Xiaorong LI; Hui-min SUN; Jiaqing YUAN
Tianjin Medical University Eye Center (Tianjin, People’s Republic of China)

Abstract:
OBJECTIVE: To investigate the corneal tissue changes induced by soft contact lens with confocal microscope.
METHODS: In this study, 36 patients who came from myopia clinic of Tianjin Medical University Eye Center with a history of long term contact lens wearing (average 5.60±3.38 years) were investigated. The mean age was 28.54±12.4 yrs (range 18~45). The mean wearing time was over 8 hours every day. The mean manifest spherical equivalence was -5.57±0.67D (-1.0~-9.0D), and a control group of 41 (82 eyes) myopia subjects without a history of contact lens wearing were investigated. The average age was 28.54±12.4 yrs (range 18~52). The average manifest spherical equivalence was -5.94±4.06D (-1.25~-13D). All of the patients were examined by Confocal Microscope.
RESULTS: The thickness of the epithelium of the contact lens group was thinner (42.59±6.13um) than that of control group and the density of anterior stroma keratocytes was decreased (1243±116/um2). There was a significant statistical difference. In the corneal tissue, there were morphological alterations. The density of the endothelial cell, the thickness of cornea and stroma showed no changes. There were many microdots deposit in stroma. The polymorphology and pleomorphism of endotheliun showed alterations.
CONCLUSION: Long term contact lens wearing could induce corneal tissue changes.
Key words: contact lens, cornea, confocal microscope