Posterior capsule opacification (PCO) is a major component of secondary cataract, a complication of current cataract surgery practice. This iatrogenic condition occurs in virtually all pediatric cases and to a lesser extent in adults. PCO correlates with the development in the latter half of the 20th Century of extracapsular cataract extraction (ECCE). In these surgeries, the lens capsule is left intact. During ECCE surgery a circular capsulotomy opening is created in the anterior lens capsule, and the cataractous, proteinaceous lens is removed, often via ultrasonic lens liquefaction i.e. phacoemulsification. The posterior, equatorial and remaining anterior portions of the sac-like capsule are left intact, permitting the insertion of an artificial lens into the emptied capsule. However, cells from the monolayer of epithelium on the inner surface of the capsule often begin to proliferate and migrate onto the normally cell-free inner surface of the posterior capsule, and may obscure the central axis of vision. Subsequently, a second surgery is necessary to create a small capsulotomy in the centre of the posterior capsule, usually employing an Nd:YAG laser. However, up to 5% of patients who have capsulotomies may then develop further serious, vision-threatening complications such as macular edema and retinal detachments. This thesis reports the photodynamic therapy (PDT) conditions required to prevent lens epithelial (LE) cell de novo proliferation and migration onto posterior lens capsules in a euthanized rabbit surgical model in order to predict parameters required to prevent PCO in humans. Experiments with primary in vitro cultures of human LE cells have shown rapid delivery of the photosensitizer benzoporphyrin derivative monoacid ring A (BPD-MA) and efficient killing with low light doses of 690 nm red light. Additional studies have shown the efficacy of various viscous agents in protecting the comeal endothelium. During model phacoemulsification ECCE surgeries, the use of hyaluronate viscoelastic carriers addressed the need for containment necessary for localized delivery of photosensitizer in the emptied capsule. Long-term monitoring of PDT-treated rabbit lens capsules in vitro has demonstrated a phototoxic effect including complete cell kill in this surgical model employing the prophylactic use of PDT. / Medicine, Faculty of / Pathology and Laboratory Medicine, Department of / Graduate

Background: Cataracts represent the major cause of blindness worldwide, disproportionately affecting such low-income countries as India. Accordingly, these cases are largely preventable, resulting in avoidable clinical and economic effects. Methods: This analysis consists of three parts: i) an exploratory systematic review of the literature to determine the extent to which the economic impact of blindness in India is known; ii) an epidemiological investigation of ophthalmological diagnosis in school age children in rural India; and iii) an examination of the determinants of cataract incidence and cataract surgical outcomes in a special Indian population through binomial logistic regression. Results: Cost estimates for blindness in India were $4.4 billion in 1997 – further investigation with current figures is needed. Costs also stem from more qualitative effects such as caregivers missing work as a result of depression. Vitamin A deficiency was not evident in school age children, though select ophthalmic diagnoses were associated with age and gender. In the adult population, determinants of cataract incidence included age, gender, occupation and caste, while surgical outcome was mainly dependent on occupation and severity of ophthalmic disease. Conclusions: The issue of blindness in India is multifaceted, and complicated by lack of knowledge of current prevalence. Further investigation into how select factors contribute to ophthalmic health, and into possible preventative strategies, is needed.

No / PURPOSE. Potential vision testing attempts to predict the visual outcome that might be expected as a result of a cataract operation. This report details the clinical utility of critical flicker frequency (CFF) as a potential vision test (PVT). METHODS. CFF thresholds were determined in 31 subjects with age-related idiopathic cataract and no other eye disease, 19 subjects with macular disease (MD) and clear ocular media, and 24 age-matched control subjects. In addition, the CFF technique was administered before cataract surgery in 52 patients and compared with the information provided by presurgical case history and ocular examination alone (ophthalmological judgment [OJ]) and results from two commonly used PVTs (the retroilluminated pinhole and the potential acuity meter). RESULTS. CFF thresholds obtained in the nonsurgical cataract group were unrelated to cataract severity and were similar to those in the control group. In contrast, CFF scores were significantly related to visual acuity (VA) in the MD group. In the pre- and postsurgical studies, OJ predicted postoperative VA very well in patients with moderate cataract and normal fundi and better than all the PVTs. OJ performed less well in patients with comorbid eye disease and dense cataracts, when information from the PVTs would probably have been useful. CFF provided the most accurate predictions of postoperative VA in the small sample of patients with dense cataracts. CONCLUSIONS. CFF was unaffected by cataract, yet sensitive to MD, and provided useful information about the postoperative visual outcome beyond that obtained through history and ocular examination in patients with dense cataracts.

Cataract disease affects millions of people worldwide. It is characterized by the accumulation of light-scattering bodies within the lens that reduce visual acuity. Cataracts are effectively treated surgically, but at great expense, costing Medicare $3.4 billion in 1997. Development of an alternative therapy for this disease would provide medical and economic benefits. We have investigated a novel antioxidant, the superoxide scavenger Mn(III)-salophen, as a therapeutic agent in the selenite cataract model. Mn(III)-salophen has been shown to protect E. coli colonies against oxidative stress but was untested in a eukaryotic system. A total dose of 300 mmol/kg, given IP in four equal 75 mmol/kg doses spaced four hours apart, protects 75% of neonatal rats from nuclear cataract development five days after selenite injection. Selenite is toxic through its reaction with the endogenous antioxidant glutathione (GSH). The reduction of selenite to selenide through an intermediate, selenodiglutathione (GSSeSG) leads to generation of superoxide radical, one of several toxic oxygen species that can damage the lens. Mn(III)-salophen causes an in vitro preservation of the lifetime of GSSeSG by interrupting the reduction of selenite. We have established that the reduction of GSSeSG to selenide does not use GSH as a reducing agent, but rather depends upon electrons generated in the earlier reduction of selenite to selenodiglutathione. These electrons can be intercepted by known one-electron scavengers, arresting the metabolism of GSSeSG. Extensive proteolysis of lens crystallins and loss of calcium homeostasis occur in cataractous lenses from a rat treated with sodium selenite. The visual protection with Mn(III)-salophen is accompanied by a partial loss of the calcium homeostasis, a net increase in sodium, and calpain-mediated proteolysis of à -crystallins similar to lenses from animals treated with selenite alone. Although preservation of alpha-crystallins may contribute to the greater transparency in the protected lens, generalized à -crystallin proteolysis is insufficient for cataract formation. From these experiments we propose that Mn(III)-salophen minimizes the oxidative stress imposed upon the cell by interfering with the metabolism of selenodiglutathione. This allows the cell to compensate for the loss of cation homeostasis and prevents aggregation of proteolyzed crystallins into cataracts. / Ph. D.

Calcium is an important factor in many cellular activities and in the maintenance of structural integrity of membranes. Calcium accumulation in the rat lens above the normal physiological range (0.11 µmol/g wet weight) has been associated with formation of cataracts. Selenite is known to influence the histological characteristics of the lens and also alter important biochemical functions. In the selenite induced cataract calcium levels increase as much as 5-fold above normal, the increase preceding appearance of mature opacity by 24 hours. Calcium in the lens rapidly exchanges with labelled calcium in the incubation medium and. establishes equilibrium within 30 minutes. Selenite, accumulated in lens in vivo, causes a 5-6 fold increase in lens calcium over controls during a 24 hour incubation in modified Hank's medium at 37°C. In vitro lmM selenite in the culture medium results in a 15-fold increase in lens calcium, a 17-fold increase in sodium/potassium ratio, and a 60% increase in lens hydration. Efflux of calcium from in vivo and in vitro selenite-treated lens is not impaired. The lens retains the capability to transport 3-4 fold greater calcium over controls from lenses which have accumulated excess calcium in the presence of selenite. In vivo selenite treatment results in a 2.5-fold increase only of extracellular water. In in vitro selenite-treated lenses, however, there is a 2.6-fold increase in extracellular water and a 1.8-fold increase of intracellular water. Selenite treatment of the lenses in vivo and in vitro causes a greater influx of calcium into the lens. Calcium accumulation may act in a nonspecific manner altering lens biochemical functions, membranes and structure, causing development of relationships between selenite induced lens fiber opacities. cataract and cell the associated changes in lens calcium content further validates this model for studying the biochemical changes which impact cataract formation. / Master of Science

Thesis (PhD)--Stellenbosch University, 2001 / ENGLISH ABSTRACT: The eye is that biological instrument which conveys the light of the external world into the inner world of the mind, wherein we receive the miraculous gift of vision. So precious is this gift, that Science must search for ways to keep this portal clear for the flow of light. Indeed, Science is called upon to “make war upon the bloody tyrant, Time.” (Shakespeare W. Sonnet No. 16). For, in the course of ageing, the lens grows cloudy and cataractous. In this battle between Science and Time, we are fortunate to live in an era in which Science is uncovering the molecular basis for the various obstacles to vision. The question arises, whether or not, the ruinous hand of time can be stayed. Due to unrelenting, progressive lens opacification, most of the elderly are destined to be subjected to loss of vision and with passage of time, even blindness. Globally the cataract surgery rate is inadequate to keep pace with the ever growing demand on financial and human resources created by the cataract problem. An immense challenge therefore is directed to primary eye care: “Can cataract be prevented or can its onset at least be postponed?” This laudable ultimate aim can only be achieved once the etiology of cataractogenesis is well understood. This dissertation seeks to examine two previously unrecognized etiological aspects that, if correctly understood and managed, have the potential to achieve preventive ophthalmological goals that may indeed help to stay the ‘ruinous hand of time’. The first aspect deals with the role of lipids and was examined using a study group of dyslipidemic subjects. The first part of the study concluded that dyslipidemic patients develop cortical lens opacities more frequently and at an earlier age than the normal population, and that cortical lens opacities should be regarded as one of the most reliable clinical signs of dyslipidemia. Furthermore, an extremely strong correlation was found to exist between low HDL Cholesterol levels and the development of opacities. Below a HDL-Cholesterol level of 1,5mmol/l, subjects had more than seven-fold higher risk of falling in the lens opacity subgroup than those with HDLCholesterol levels above 1,5mmol/l. An equally strong correlation was demonstrated between high (>5) LDLHDL ratios and the development of lens opacities. Subjects with a LDL:HDL-C ratio below 5 possessed a 2.35 times greater risk of having lenticular opacities than the group with a LDL:HDL-C ratio greater than 5. The prevention or retardation of dyslipidemia associated lens opacities is therefore possible, provided patients with a genetic predisposition are detected early and their blood lipids managed adequately. The second aspect deals with the relationship between age related cataracts and the acetylation status of the individual. This study compellingly submits that the slow acetylator pheno- and genotype may be regarded as a genetic indicator of risk for age related cataract. The ability accurately to classify a patient genotypically and phenotypically, may henceforth be useful in health counseling since, if an individual is identified as being a slow acetylator, additional preventative and precautionary measures may be taken, i.e. the prevention of UVexposure to the eye and caution with the ingestion of xenobiotics like caffeine, commercial dyes, food preservatives, and drugs. Furthermore, such a finding should be taken into account in the long term therapeutic management of glaucoma, with special regard to carbonic anhydrase inhibitors which are sulphonamide-related drugs and totally dependent on the N-acetyltransferase pathway for metabolism. These drugs may accumulate in the slow acetylator over time and exert toxic effects intra-ocularly, conceivably including cataractogenesis. The search for genetic and metabolic mechanisms that may contribute to human cataractogenesis should be pursued with great enthusiasm. This endeavour may help Science to achieve its primary objective, ablate the effects of Time and really aid in preventing cataracts in man. / AFRIKAANSE OPSOMMING: Die wondergawe van visie word vir ons moontlik gemaak deur die oog wat as biologiese instrument die lig van die buitewereld inlaat na die binnewereld van die brein. So kosbaar is hierdie gawe dat die Wetenskap deurgaans moet poog om die poort oop te hou. Inteendeel, die Wetenskap word gemaan deur Shakespeare in sy Sonnet nommer 16 om “oorlog te maak teen die bloeddorstige tiran, Tyd”. Soos ‘n mens ouer word, word die lens dof en ‘n katarak mag vorm. Ten spyte van hierdie stryd tussen ‘Wetenskap’ en ‘Tyd’ leef ons in die gelukkige era waarin die Wetenskap meer en meer leer van die verskeie obstruksies tot visie. Die vraag ontstaan of die rinnewerende hand van ‘Tyd’ gestuit sal kan word. Vanwee ongenaakbare, progressiewe lensvertroebeling is die meeste bejaardes bestem om aan visie verlies, en met verloop van tyd selfs blindheid, te ly. Die wereldwye katarakchirurgie tempo is nie voldoende om by te hou by die immergroeiende finansiele en mannekrag eise wat deur die katarak probleem gestel word nie. Daar word dus ‘n reuse uitdaging aan primere oogsorg gestel naamlik: “Kan katarakte nie eerder voorkom of die aanvang daarvan ten minste uitgestel word nie?” Hierdie prysenswaardige doelwit kan nie bereik word alvorens die etiologie van kataraktogenese goed verstaan word nie. Hierdie tesis ondersoek twee voorheen onerkende etiologiese aspekte wat, indien hulle korrek verstaan en hanteer word, beslis die potensiaal het om die gemelde voorkomende doelwitte te bereik en sekerlik te kan bydrae om die rinnewerende hand van Tyd te stuit. Die eerste aspek spreek die rol van lipiede aan deur te kyk na 'n studiegroep van dislipidemiese persone. Die eerste deel van die studie kom tot die gevolgtrekking dat dislipidemiese pasiente kortikale lens opasiteite meer dikwels en op ‘n vroeer ouderdom ontwikkel as die normale populasie en dat sulke opasiteite beskou moet word as een van die mees betroubare kliniese tekens van dislipidemie. Daar is ook ‘n baie sterk korrelasie gevind tussen lae HDL cholesterol vlakke en die voorkoms van opasiteite. Persone in die studie met ‘n HDL cholesterol vlak laer as 1,5mmol/l het ‘n sewe keer hoer kans gehad om in die lensopasiteit subgroep te val as die met ‘n HDL cholesterol vlak laer as 1,5mmol/l. ‘n Sterk korrelasie tussen ‘n hoe (>5) LDLHDL verhouding en die voorkoms van lens opasiteite is ook gevind. Persone met ‘n LDLHDL verhouding >5 het ‘n 2.35 maal groter risiko gehad om lensopasiteite te he as die met ‘n LDL:HDL verhouding van <5. Die voorkoming of vertraging van dislipiedemie geassosieerde lens opasiteite is dus moontlik, solank persone met ‘n genetiese geneigdheid daartoe vroeg ontdek en hulle bloedlipiede voldoende beheer word. Die tweede deel van die tesis handel oor die verhouding tussen ouderdoms verwante katarakte en die asetilasie status van die individu. Met oortuiging kom hierdie studie tot die gevolgtrekking dat die stadige asetilator fenoen genotipe as 'n genetiese merker vir ouderdoms verwante katarakte beskou moet word. Die vermoe om ‘n individu beide genotipies en fenotipies akkuraat te klassifiseer mag voorts bruikbaar wees in gesondheidsraadgewing. Indien ‘n individu geTdentifiseer is as ‘n stadige asetileerder, kan addisionele voorsorg maatreels getref word soos bv. die voorkoming van blootstelling van die oog aan UV lig sowel as omsigtigheid met die inname van xenobiotika soos kaffei'ene, kleurstowwe, voedsel preserveermiddels en geneesmiddels. Hierdie bevinding moet ook in berekening gebring word in die langtermyn terapeutiese hantering van gloukoom. Die koolsuuranhidrase inhibitore, dikwels gebruik in die behandeling van gloukoom, is sulfonamied-agtige middels en dus totaal afhanklik van die N-asetieltransferase pad vir hulle metabolisme. Hierdie middels kan ophoop in die stadige asetileerder en gegewe genoeg tyd, bes moontlik toksiese intra-okulere effekte tot gevolg he. Die soeke na genetiese en metaboliese meganismes wat mag bydra tot menslike kataraktogenese behoort nagestreef te word met groot entoesiasme. Hierdie strewe mag dalk net vir die 'Wetenskap' bystaan om sy primere mikpunt te bereik, die effek van ‘Tyd’ te neutraliseer en te help om katarakte werklik te voorkom.

published_or_final_version / Biochemistry / Master / Master of Philosophy

Background Cataract surgery is the most common surgical procedure carried out in the developed world and surgery volumes have increased considerably during the last decades. Various aspects of the surgical procedure, including surgical incision size and intraocular lens materials, have changed substantially, improving the safety and the quality of the outcome. Previous research has primarily focused on the visual function results with a short follow-up time. Long-term population-based studies, exceeding a few years, presenting visual functional results postoperatively, have not been published. Aims To determine the effects of cataract surgery on subjectively experienced visual function and visual acuity in a defined population, and compare the results between sub-groups, on a long-term basis. Methods In this prospective, population-based investigation, all patients with presenile and senile cataract (n=810), operated on during a one-year period (1997-98), at Umeå University Hospital were included. The frequency of cataract surgery at that time, was 5.2 per 1000 population studied. Visual acuity was tested and an eye examination was performed before surgery, 4-8 weeks postoperatively, and five and ten years after surgery. Subjective visual function was assessed using self-administered questionnaires (VF-14) at all occasions. Statistical evaluations comprised analyses of variance, Mann-Whitney U-test, chi-square test, multiple linear regression, a life-table calculation, and Cox’s proportional hazard model. Results Five years after cataract surgery, subjective and objective visual function remained stable in most patients. The most frequent cause of deterioration of visual acuity and decrease in VF-14 scores was agerelated macular degeneration (ARMD). Two thirds of the patients in the cohort were women. They were significantly older than the men and more often operated on both eyes. After adjustment for age and visual acuity, women cataract surgery patients assessed their visual function worse than men both before surgery and 4 months postoperatively. Five years after surgery these differences were no longer significant. At baseline, 13% of the patients were diabetics. At the five-year followup, subjective and objective visual function remained stable in most surviving diabetics, and the longitudinal visual function was not significantly worse compared with the non-diabetics. Ten years after surgery, 28% had received treatment for posterior capsular opacification (PCO). A significantly larger proportion of patients less than 65 years at surgery (37%) compared with those 65 years or older (20%) had been treated. Conclusions Most patients sustain their level of visual acuity and visual function also five and ten years after cataract surgery. Ocular co-morbidity, such as ARMD, is the major cause of longitudinally reduced visual function. Patients suffering from diabetes did not have a significantly worse visual function after five years. A surprisingly large proportion of patients had received treatment for PCO after ten years.

Congenital cataract is a leading cause of visual disability among children worldwide. It has a heterogeneous genetic basis; the cellular and molecular mechanisms for cataractogenesis remain elusive. A spontaneously occurred autosomal dominant mouse mutant named Secc, which displays small eye, cataract and closed eyelid, has been obtained in our laboratory. By gene mapping and DNA sequencing, we identified a single nucleotide deletion at position 273 of the Cryga gene, leading to a frame-shift from the 3rd Greek Key motif of the A-crystallin (Cryga). The aim of this study is to investigate the pathogenic mechanisms underlying the development of cataract in the Secc mutant, as a disease model for understanding human congenital cataract. Initial phenotype analysis showed that cataract was initiated in E14.5 CrygaSecc mutant embryos, the nuclei of the primary lens fibres were scattered and failed to align in the equatorial region. By E16.5, the secondary lens fibre cells were abnormally arranged with poor lens suture formation. Apoptotic cells were found in the centre of the lens as shown by TUNEL assay, cytoskeleton and cell adhesion in the lens centre were disturbed as shown in immunohistochemistry analysis. Previously by western blotting it was found that mutant -crystallins were enriched in the insoluble fraction. I hypothesized that mutant A-crystallins might be misfolded and protein aggregates were then formed. In this study, aggregation was observed in semi-thin sections stained with toluidine blue. By co-staining using custom-made anti-Secc antibody, CrygaSecc protein was found to be ubiquitinated and was wrapped around by vimentin. Clearly, in the Secc mutant lens, aggresomes were formed for the disposal of the misfolded proteins and to maintain cell survival. However, ultimately cell death would occur in the mutant lens and contributed to cataract formation. It is known that misfolded proteins would trigger unfolded protein response (UPR) and heat shock protein (HSP) responses to facilitate folding and to prevent misfolded proteins from intoxicating the cell. In order to determine which stress response pathway was triggered, gene expression analysis by qRT-PCR was performed. The expression of genes involved in the UPR pathways including BiP, CHOP and spliced variant of XBP-1 were all up-regulated significantly in E14.5 and 16.5 mutant lenses. In addition, among different ER stress related genes, cytosolic chaperones and autophagy related genes, Hsp70 and BiP were upregulated, while Hsp40 and Hsp90aa were downregulated in the homozygotes. The results suggested that both UPR and HSP response pathways were triggered during cataractogenesis in the Secc mutant. In conclusion, mutant A-crystallin appeared to trigger UPR, HSPs and cell death in the fibre cells, while autophagy was not triggered. In the lens fibre cells, the ubiquitin-proteasomal pathway was utilized for the removal of misfolded CrygaSecc proteins. However, the stress perpetuated as the lens grew and produced more mutant proteins. The mutant cells lost their normal cell adhesion, failed to maintain the proper lens architecture, leading to cataract formation. Similar cellular mechanisms could be implicated in human congenital cataract or age-related cataract development. / published_or_final_version / Biochemistry / Doctoral / Doctor of Philosophy

Mode of access: World Wide Web. / Thesis (Ph. D.)--University of Hawaii at Manoa, 2004. / Includes bibliographical references (leaves 86-97). / Electronic reproduction. / Also available by subscription via World Wide Web / xv, 97 leaves, bound ill. 29 cm