Vascular tone pathway polymorphisms in relation to primary open-angle glaucoma.

AIMS: Vascular perfusion may be impaired in primary open-angle glaucoma (POAG); thus, we evaluated a panel of markers in vascular tone-regulating genes in relation to POAG. METHODS: We used Illumina 660W-Quad array genotype data and pooled P-values from 3108 POAG cases and 3430 controls from the combined National Eye Institute Glaucoma Human Genetics Collaboration consortium and Glaucoma Genes and Environment studies. Using information from previous literature and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, we compiled single-nucleotide polymorphisms (SNPs) in 186 vascular tone-regulating genes. We used the 'Pathway Analysis by Randomization Incorporating Structure' analysis software, which performed 1000 permutations to compare the overall pathway and selected genes with comparable randomly generated pathways and genes in their association with POAG. RESULTS: The vascular tone pathway was not associated with POAG overall or POAG subtypes, defined by the type of visual field loss (early paracentral loss (n=224 cases) or only peripheral loss (n=993 cases)) (permuted P≥0.20). In gene-based analyses, eight were associated with POAG overall at permuted P<0.001: PRKAA1, CAV1, ITPR3, EDNRB, GNB2, DNM2, HFE, and MYL9. Notably, six of these eight (the first six listed) code for factors involved in the endothelial nitric oxide synthase activity, and three of these six (CAV1, ITPR3, and EDNRB) were also associated with early paracentral loss at P<0.001, whereas none of the six genes reached P<0.001 for peripheral loss only. DISCUSSION: Although the assembled vascular tone SNP set was not associated with POAG, genes that code for local factors involved in setting vascular tone were associated with POAG.

Health literacy and vision-related quality of life.

BACKGROUND: Non-visual factors influence a person's vision-related quality of life (VRQoL). The purpose of this study was to assess the relationship between health literacy and VRQoL in glaucoma patients. METHODS: One hundred and ninety-five subjects with open-angle glaucoma participated in a cross-sectional patient survey and chart review. Subjects were administered a test of health literacy, an assessment of physical and mental well-being, and an assessment of VRQoL, the National Eye Institute 25-Item Visual Function Questionnaire (VFQ-25). Charts were reviewed for visual acuity and visual field results. RESULTS: In univariate analyses, older age (p<0.001), non-White race (p<0.001), worse visual acuity (p<0.001), worse visual field scores (p<0.001), lower level of education (p<0.001), worse health literacy (p<0.001) and worse score on the mental health component of the SF-12 (p = 0.005) were associated with worse VFQ-25 scores. In multivariate analyses, only older age was associated with worse total VFQ-25 scores (p<0.001), although the association between health literacy and the VFQ subscale of dependency remained significant (p = 0.04). CONCLUSIONS: Individuals with a lower health literacy do not appear to have a worse overall VRQoL compared with those with a higher literacy, but worse health literacy is associated with increased dependency.

Myocilin-associated exosomes in human ocular samples.

Mutations in myocilin result in ocular hypertension, likely due to decreased drainage of aqueous humor through the trabecular meshwork. Since less myocilin is found in the aqueous humor of those with disease-causing mutations, understanding myocilin's role in the aqueous humor is of clinical importance. Recently, myocilin was shown to exit cultured trabecular meshwork cells in association with shed vesicles called exosomes. To examine relevance of this finding in a physiological setting, the present study examined three different types of ocular samples for the presence of myocilin-associated exosomes. Using differential centrifugation steps, we found myocilin associated with exosomes isolated from effluent collected from human anterior segments in organ culture and aqueous humor obtained from human cadaveric eyes or from patients undergoing excisional surgery. Similar to results with cultured cells, myocilin associated predominately with exosomes in fresh samples, appeared mostly soluble at later times, and had biochemical properties (density of 1.13-1.19 g/ml in linear sucrose gradient) similar to those characteristics of exosomes. These data indicate that exosomes are present and may facilitate the transport of myocilin into the extracellular space of human ocular cells.

Prevalence of myocilin mutations in adults with primary open-angle glaucoma in Ghana, West Africa.

PURPOSE: Investigators have noted that primary open-angle glaucoma (POAG) in West Africa has an earlier age of onset and appears to be more clinically severe than in the United States and Europe. Primary open-angle glaucoma patients with mutations in myocilin have a similar phenotype. Therefore, we investigated the role of mutations in myocilin in patients with POAG in a West African population. MATERIALS AND METHODS: Patients seen at the Emmanuel Eye Clinic in Accra, Ghana, were recruited for this study. Informed consent was obtained from all study patients. Glaucoma specialists from the sponsoring institution (PC, LWH, or RRA) ascertained all POAG and control patients. Age-matched unaffected controls were obtained in patients with an IOP < 22 mm Hg and normal-appearing optic nerves. PCR amplification of each of the three myocilin exons was performed. Denaturing high-performance liquid chromatography (Transgenomics Corp.) was used to detect allelic differences and samples demonstrating a mobility shift were sequenced in both directions. RESULTS: Ninety unrelated affecteds with POAG and 76 control patients were recruited. Four individuals with severe POAG were found to have novel missense mutations in exon 3. Two exhibit an Asp380Asn mutation and two an Arg342Lys mutation. These changes were not detected in 152 ethnically matched control chromosomes. Fourteen affected individuals and eight controls exhibit a translationally silent polymorphism in codon 325 (Thr325Thr). CONCLUSIONS: A total of 4.4% of patients with POAG have novel disease-associated mutations in myocilin. Mutations in myocilin appear to play a limited role in the pathogenesis of POAG in this region of West Africa.

Hereditary benign intraepithelial dyskeratosis: Report of two cases with prominent oral lesions.

Hereditary benign intraepithelial dyskeratosis is a rare autosomal dominant disorder of the oral and ocular mucosa initially described in the Haliwa-Saponi Native American tribe of North Carolina. We describe 2 sisters with the characteristic oral and ocular findings. This entity should be distinguished from several other diseases that cause white lesions in the mouth including white sponge nevus.

Pseudoexfoliation syndrome in Icelandic families.

AIM: To examine the distribution and clinical ophthalmic characteristics of pseudoexfoliation syndrome (pseudoexfoliation) and glaucoma in Icelandic families. METHODS: Icelandic families containing three or more members aged 70 or older with at least one member with pseudoexfoliation were identified. All family members over age 45 were invited to participate. Visual acuity, Goldmann applanation tonometry, gonioscopy, slit lamp examination before and after dilatation, and dilated fundus examination were performed on all available family members. Pertinent data were obtained from medical records, including ophthalmic history and a medical history of cardiovascular disease, cerebrovascular disease, systemic hypertension, and diabetes mellitus. Participants were classified according to affected status for pseudoexfoliation, glaucoma, and age related macular degeneration. RESULTS: Six families were identified who met the criteria for entry into the study. Of 94 family members who were invited to participate 82 were enrolled (87%). Of these 25 (30%) had pseudoexfoliation syndrome, 51 (62%) were unaffected, and six (7%) were suspects. At least one individual with pseudoexfoliation was identified in the second generation of every family. A parent with pseudoexfoliation was identified in all cases either by examination (4/6) or a review of ophthalmic records (2/6). In all cases the mother was the affected parent. The prevalence of glaucoma was significantly greater in the group with pseudoexfoliation (p <0.0001). Although the presence of age related macular degeneration (ARMD) was highly associated with the presence of pseudoexfoliation, the significance was lost after correction for age (p = 0.69). Although the sample size was small, no association between pseudoexfoliation affected status and cardiovascular disease, cerebrovascular disease, systemic hypertension, or diabetes mellitus was found. CONCLUSIONS: Multiple Icelandic families with pseudoexfoliation in two generations were identified. In all cases where determination was possible, transmission to the second generation was through an affected parent. In each case the affected parent was the mother. Pseudoexfoliation was strongly associated with the presence of glaucoma, but was not associated with either ARMD or systemic disease in this study. These data clearly indicate that pseudoexfoliation is a familial condition and although not conclusive, supports the hypothesis that pseudoexfoliation syndrome is genetically inherited.

Familial occurrence of pigment dispersion syndrome.

BACKGROUND: Pigment dispersion syndrome affects up to 4% of the white population. It is characterized by the presence of transillumination defects, Krukenberg's spindle and dense trabecular meshwork pigmentation. Open-angle glaucoma will develop in as many as 50% of affected patients. In this study we describe the familial occurrence of pigment dispersion syndrome in six North American pedigrees and the phenotypic characteristics with respect to pigment dispersion syndrome and glaucoma. METHODS: Probands with pigment dispersion syndrome were identified in glaucoma clinics at university eye centres in Ottawa and Durham, NC. Families with two or more affected members were evaluated. All willing members in each family underwent a thorough clinical examination and were classified as affected with pigment dispersion syndrome, suspect or unaffected. The previous medical records were reviewed to obtain the past medical and ocular history, including risk factors for glaucoma. RESULTS: All six families are white. Three families show at least two generations of affected members. Of the 43 subjects examined 58% were women. All 14 affected members showed moderate to heavy trabecular meshwork pigmentation and either Krukenberg's spindle or transillumination defects. The affected members were also considerably more myopic (mean spherical equivalent for the right eye -4.72 dioptres) than the suspect group or the unaffected group (mean spherical equivalent -0.79 D and +1.19 D respectively) (p < or = 0.001), and the intraocular pressure was higher for the affected than the unaffected group (mean for the right eye 20 mm Hg vs. 16 mm Hg) (p = 0.004). Half of those affected also had open-angle glaucoma. INTERPRETATION: We have identified and phenotypically characterized six North American families with autosomal dominant pigment dispersion syndrome. Our ultimate goal is to identify the gene(s) that causes this disorder in order to clarify its molecular etiology and pathophysiology. This may give rise to a molecular classification of the disease as well as provide the foundation for genetic testing and new treatment approaches.

A duplication in chromosome 4q35 is associated with hereditary benign intraepithelial dyskeratosis.

Hereditary benign intraepithelial dyskeratosis (HBID) is an autosomal dominant disorder characterized by elevated epithelial plaques on the ocular and oral mucous membranes. It has been reported primarily, but not exclusively, in individuals of American Indian heritage in North Carolina. We have examined and obtained DNA on two large families affected by HBID. Using genetic linkage analysis we have localized the HBID gene to chromosome 4 (4q35) with a peak LOD score of 8.97. Molecular analysis of these data reveals that all individuals affected with HBID in both families demonstrate the presence of three alleles for two tightly linked markers, D4S1652 and D4S2390, which map to the telomeric region of 4q35. This suggests the presence of a duplication segregating with the disease phenotype that is most likely involved in its causation.

Characteristics and risk factors of infections after glaucoma filtering surgery.

PURPOSE: To define characteristics and potential risk factors of endophthalmitis and blebitis after glaucoma filtering surgery in adults. METHODS: A chart review of all cases of endophthalmitis or blebitis treated at the Duke University Eye Center for 6 years (January 1993 to December 1998) was performed to identify patients with a history of incisional glaucoma surgery. RESULTS: Twenty patients were identified. The filtering bleb was located superiorly in all patients. Blebitis but not endophthalmitis developed in 3 (15%) of 20 patients, and all had visual outcomes of at least 20/25. Endophthalmitis (blebitis and vitritis) occurred in 17 (85%) of 20 patients. Cases of blebitis were treated with topical antibiotics. All cases of endophthalmitis were treated with intravitreal antibiotics, and 3 (18%) of 17 patients also underwent immediate vitrectomy. Initial visual acuity was less than hand motions in 5 (29%) of 17. Final visual acuity was less than 20/200 in only one case of endophthalmitis. In 15 (75%) of 20 patients, the bleb was noted to be thin, avascular, or both. On presentation, 11 (55%) of 20 blebs had Seidel-positive leaks with hypotony. A history of recurrent bleb leaks was documented in 7 (33%) of 20 patients. Pseudophakia was present in 13 (65%) of 20 eyes, and 7 (35%) of 20 had undergone combined cataract and filtering surgery. A prodrome, such as a browache, headache, or external eye inflammation or infection, was documented in previous physician visits in 7 (35%) of 20 patients. No cases occurred in eyes with glaucoma implants. CONCLUSIONS: Patients in whom endophthalmitis develops after trabeculectomy do poorly, even with aggressive medical and surgical intervention. As expected, several patients had thin, avascular, leaking blebs. In addition, hypotony, recurrent bleb leaks, pseudophakia, and more than one filtering surgery may also be associated with blebitis or endophthalmitis after glaucoma filtering surgery. In a surprising number of patients, prodromal signs or symptoms were documented by ophthalmologists days or weeks before the diagnosis of blebitis or endophthalmitis was made.

Antithrombin III, a serpin family protease inhibitor, is a major heparin binding protein in porcine aqueous humor.

Our hypothesis is that the proteins in aqueous humor may be involved in the regulation of outflow facility through the trabecular meshwork and uveoscleral meshwork. In this study, we analyzed the profile of heparin-binding proteins present in porcine aqueous humor to identify and characterize secretory proteins with a binding affinity for heparin. A single step involving heparin-sepharose affinity chromatography of porcine aqueous humor yielded a approximately 60 kDa protein as the major heparin-binding species. This protein was specifically eluted from the column by heparin. The N-terminal sequence and immunological cross reactivity of this protein confirmed its identity as antithrombin III. Aqueous humor from different species, as well as cells from human trabecular meshwork, Schlemm's canal, and lens epithelium, contained detectable amounts of antithrombin III. Based on its known anticoagulative function in endothelial cells and effects on the production of prostacyclin, it is reasonable to speculate that antithrombin III present in aqueous humor might influence the physiology of the trabecular and uveoscleral meshwork and thereby regulate intraocular pressure.

Isolation of primary open-angle glaucomatous trabecular meshwork cells from whole eye tissue.

PURPOSE: Isolation and culture of human trabecular meshwork (TM) cells from primary open-angle glaucomatous (POAG) tissue has proven difficult. The objective of this study was to directly compare the utility of two different isolation methods to obtain viable human TM cells from POAG whole eye tissue. METHODS: Using a blunt dissection technique, human TM tissue was obtained from four pairs of donor eyes (67, 77, 81 and 82 years) with a documented history of POAG. TM tissue from one eye was explanted into tissue culture. TM from the contralateral eye was digested with a collagenase mixture and seeded onto culture plates. RESULTS: Primary cell isolates were obtained from all donors with both techniques. However, only cells obtained using the digestion method (3 of 4 TMs) could be passaged for expansion and freeze-downs (3 x 107 second passage cells/donor). None of the cells obtained from explanted TMs could be passaged. Cells from successful isolations were of uniform size, possessed typical TM morphology and had doubling times < 48 hours. CONCLUSION: These results demonstrate a clear advantage to digesting the extracellular matrix of glaucomatous TM tissue to obtain sufficient numbers of healthy cells for use in experiments. In contrast to cells obtained from explants, cells liberated from POAG TM tissue by digestion appear indistinguishable morphologically and behaviorally from "normal" TM cells.

Genome-wide scan for adult onset primary open angle glaucoma.

Adult onset primary open angle glaucoma is a leading cause of blindness throughout the world. The disease results in an apoptotic death of retinal ganglion cells that is usually associated with an elevation of intraocular pressure. Familial aggregation of the disorder provides evidence for strong genetic influences that are likely to be the result of multiple susceptibility genes. A two-stage genome scan to identify the genomic locations of glaucoma susceptibility genes was performed using an initial pedigree set of 113 affected sibpairs and a second pedigree set of 69 affected sibpairs. Linkage analysis was performed using both model-dependent (lod score) and model-independent affected relative pair and sibpair methods. Twenty-five regions identified by the initial scan were further investigated using the second pedigree set. In the combined data analysis, regions located on chromosomes 2, 6, 9, 11, 14, 17 and 19 continued to produce model-dependent lod scores and/or an MLS >1.0, while five regions (2, 14, 17p, 17q and 19) produced an MLS >2. 0. Multipoint analysis using ASPEX also showed significant results on chromosomes 2, 14, 17p, 17q and 19. These results are an important step towards the identification of genes responsible for the genetic susceptibility to this blinding condition.

Excisional bleb revision to correct overfiltration or leakage.

OBJECTIVES: To evaluate the effectiveness of excisional revision of filtering blebs for hypotony or leakage when more conservative measures have failed. PATIENTS AND METHODS: Retrospective review of all patients who underwent excisional revision of a filtering bleb for hypotony (intraocular pressure [IOP] < 5 mm Hg) or leakage during a 3 year period. The revision consisted of excision of the avascular bleb, mobilization of the surrounding conjunctiva, and suturing of the conjunctiva at the limbus. RESULTS: Sixteen patients were included in the study. The average age was 66.3 +/- 14.8 years (range, 39-83). Revision followed trabeculectomy in 11 cases, combined phacoemulsification-trabeculectomy in three cases, and inadvertent blebs in two cases. Five cases had bleb leaks without hypotony, four cases had hypotony alone, and seven cases had both hypotony and a bleb leak. Average follow-up after bleb revision was 25 +/- 11 months (range, 9-43). Average IOP increased from 3.8 +/- 5.6 mm Hg (range, 0-22) to 11.9 +/- 4.1 mm Hg (range, 3-18), with an average of 1.1 +/- 1.1 medication (range, 0-3). The IOP at the last visit was < 15 mm Hg in all but two patients, with 10 of the 16 patients requiring medications. At the last follow-up examination, visual acuity had improved > or = two lines in nine patients and was reduced two lines in one patient. Five patients had early postoperative limbal wound leaks; resuturing was required in one case. CONCLUSIONS: Excisional bleb revision is an effective technique to correct hypotony or leakage after filtering surgery when other methods have failed. Intraocular pressure control is often maintained with the use of medications.

Excimer laser effects on outflow facility and outflow pathway morphology.

PURPOSE: To determine the relative contributions to aqueous outflow resistance of the tissues distal to the inner wall of Schlemm's canal. METHODS: While performing constant pressure perfusion at 10 mm Hg, a 193-nm excimer laser (Questek) was used to precisely remove portions of sclera, unroofing Schlemm's canal while leaving the inner wall intact. The laser beam was masked to produce a beam 2 mm by 1 mm. The laser output was constant at a fluency of 75 mJ/cm2 and 20 Hz. The excimer laser at a frequency of 1 Hz was used as the aiming beam. Photoablation was performed on human cadaver eyes at the limbus at an angle of 0 degrees to 45 degrees from the optical axis. As the excimer photoablations progressed, Schlemm's canal was visualized by the fluorescence of the Barany's solution containing fluorescein dye. After perfusion fixation the eyes were immersion-fixed overnight. The facility of outflow before (Co) and after (Ce) the excimer ablation was measured in 7 eyes. RESULTS: The facility of outflow increased in all eyes after the excimer sinusotomy, from a mean of 0.29+/-0.02 before the sinusotomy to 0.37+/-0.03 microl/min per mm Hg after (P < 0.05). The mean ratio of outflow facility after and before ablation (Ce/Co) was 1.27+/-0.08 (range, 1.20-1.39), a reduction of outflow resistance of 21.3%. Using the formula of Ellingsen and Grant (1972), percentage of resistance to outflow eliminated = 100 [1 - alphaCo/Ce - (1 - alpha)Co], where alpha = fraction of the circumference dissected. Assuming that because of circumferential flow approximately 50% of Schlemm's canal is drained by the single opening made in the outer wall ablation studies, this results in resistance to outflow eliminated of 35%, which is consistent with the calculated eliminated resistance derived from the data of Rosenquist et al., 1989. Light and scanning electron microscopy confirmed the integrity of the inner wall Schlemm's canal underlying the area of ablation. CONCLUSIONS: The results provide direct evidence indicating that approximately one third of resistance to outflow in the human eye lies distal to the inner wall Schlemm's canal in an enucleated perfused human eye.

Decrease in intraocular pressure after orbital decompression for thyroid orbitopathy.

BACKGROUND: The effect of thyroid orbitopathy on intraocular pressure (IOP) remains controversial. We carried out a study to determine the effect of orbital decompression surgery on the IOP in patients with advanced thyroid orbitopathy. METHODS: The records of 12 consecutive patients (22 eyes) who underwent decompression surgery for severe thyroid orbitopathy between 1985 and 1996 were reviewed. All patients were maintained on essentially the same medications before and after surgery. The IOP readings, obtained by means of applanation tonometry in primary gaze, from the pre- and postoperative visits were recorded, and the net change was calculated. RESULTS: The mean preoperative and postoperative IOP values were 19.8 mm Hg and 16.8 mm Hg respectively, a significant difference (p = 0.008). Seven of eight eyes with an IOP of 21 mm Hg or greater preoperatively had a postoperative IOP less than 21 mm Hg; these eyes showed a mean decrease in IOP of 5.6 mm Hg. The degree of preoperative IOP elevation was found to be a strong predictor of the amount of IOP lowering after surgery (p = 0.014). INTERPRETATION: Our results support the concept that orbital congestion associated with thyroid orbitopathy produces an increase in IOP by elevation of episcleral venous pressure (EVP) and that orbital decompression may reduce the IOP by decreasing EVP. Decompression surgery may obviate the need for more aggressive management of glaucoma in patients with severe thyroid orbitopathy.

A clinical comparison of transscleral cyclophotocoagulation with neodymium: YAG and semiconductor diode lasers.

PURPOSE: To compare the efficacy of transscleral cyclophotocoagulation using a neodymium: YAG (Nd:YAG) or semiconductor diode laser in controlling intraocular pressure in patients with refractory glaucoma. METHODS: In a prospective study, 95 eyes of 91 patients with refractory glaucoma randomly received Nd:YAG or diode cyclophotocoagulation. Patients were followed for a mean of 10.4 months (10.42 +/- 3.16, mean +/- SD). We compared available data preoperatively and at 1 week, 1 month, 6 months, and 12 months postoperatively. Data analyzed were corrected visual acuity, intraocular pressure, and the type of glaucoma. RESULTS: There was a statistically significant decrease in intraocular pressure after both Nd:YAG and diode cyclophotocoagulation at each time period. However, there were no significant differences in postoperative intraocular pressure or visual acuity change between Nd:YAG and diode procedures. CONCLUSIONS: Compared with the Nd:YAG laser for transscleral cyclophotocoagulation, the diode laser has technological advantages including portability, durability, and smaller size, while providing equivalent postoperative intraocular pressure and visual acuity change.

Gln368STOP myocilin mutation in families with late-onset primary open-angle glaucoma.

PURPOSE: To examine families ascertained for late-onset primary open-angle glaucoma (POAG) to determine mutations in the gene coding for myocilin. METHODS: The diagnosis of late-onset POAG was defined as age at diagnosis more than 35 years, intraocular pressure (IOP) 22 mm Hg or more in both eyes or 19 mm Hg or more while the patient was taking two glaucoma medications, glaucomatous optic neuropathy in both eyes, and visual field loss consistent with optic nerve damage in at least one eye of the proband. Two of three criteria were required in other family members. DNA from all families was screened for polymorphisms in myocilin using single-strand conformation polymorphism analysis. All polymorphisms were sequenced for mutations. RESULTS: Eighty-three affected people in 29 families with late-onset POAG were screened for mutations. Three mutations, two novel missense (Thr377Met and Glu352Lys) and one nonsense (Gln368STOP), were identified. The missense mutations did not segregate with the disease phenotype in these families. The nonsense mutation was found in 3 of 29 unrelated families with POAG. All affected family members and 8 of 12 in whom glaucoma was suspected had the Gln368STOP mutation. All people with this mutation had elevated IOP, and 78% had POAG by age 70. CONCLUSIONS: Three mutations were identified in the gene coding for myocilin in families with late-onset POAG. Of these, the Gln368STOP mutation was highly associated with the development of glaucoma. All people with this mutation had glaucoma or elevated IOP by age 70. In the United States, the Gln368STOP mutation in myocilin is strongly associated with the development of late-onset POAG. However, factors in addition to the presence of this mutation seem to play a role in the development of ocular hypertension and glaucoma in these families.