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.

No description available.

The commission on radiological protection through publication 118 decided to recommend a change to the eye dose limit in 2011. ICRP recommendations made in publications, especially ‘publication 60’ and its subsequent update ‘publication 103’ has served as standards for regulatory authorities worldwide in limiting ionizing radiation exposure both to workers and members of the public. For example in Canada, the Canadian Nuclear Safety Commission (CNSC) generally directly adopts recommendations from ICRP. The previous dose limit for the lens of the eye was 150 mSv year-1, based on Publication 60 and 103. Regulatory agencies worldwide have been using this value and subsequently nuclear facilities, hospitals and universities have designed their radiation protection program based on this dose limit for several decades. The new revised eye dose limit now being equivalent to the whole body dose limit will pose significant challenges for sectors where the eye exposure was not characterized as the limit was previously five times over the whole body exposure. A two-step approach was used in conducting this study, firstly a through literature search was conducted on the effects of ionizing radiation to the eye, its radiobiology, fundamentals in established both dose limits was analyzed. Secondly, the authors spent time researching institutions that use ionizing radiation and interviewed engineers, medical physicists, radiation safety officers and regulators from a wide array of fields and industries. Based on the ICRP publications, the review of the literature and the interviews conducted with the nuclear industry, there is consensus in Canada and among IAEA member states that the dose limit for the lens of the eye should be reduced from the original proposed limit of 150 mSv per year. However not to the recommendations suggested by ICRP 118, but, to a standard reasonable and an achievable limit that is 50 mSv per year. / Thesis / Master of Science (MSc) / The International Commission on Radiological Protection (“ICRP”), the independent governing body responsible for radiation protection, since the early 1950s has been issuing recommendations that are widely used as radiological protection standards by regulatory agencies worldwide, primarily UN member states. Since its inception in 1928, the ICRP has served as the basis for radiation protection and value based judgements in protecting both human and non-human biota. In 2011, the commission published (ICRP Pub. 118) its review of epidemiological studies and decided to recommend a change to the previously established eye dose limit. Based on the review of the literature and the research conducted within the academic, veterinary, nuclear and medical industry, there is general consensus in Canada and among IAEA members states that the dose limit for the lens of the eye should be reduced from the original proposed limit, but not to the recommendations suggested by ICRP 118.

The absolute clarity of the lens of the eye is vital in the visual system. The unique structural and physiological properties of the lens are tightly integrated with highly ordered protein content to allow the lens to remain transparent. Consequently, any alteration or disturbance of these highly ordered proteins can affect the optical properties of the lens. In humans, cataracts are the major cause of blindness, yet the exact aetiology of cataract formation (cataractogenesis) is not fully understood. The purpose of the current research was to investigate whether deregulation of the Ca²⁺-dependent enzyme, calpains, following changes in lens Ca²⁺ homeostasis, is a key mechanism leading to undesired cleavage of a number of proteins that are linked with maintaining lens transparency and contributing to cataractogenesis. An ovine lens culture (in vitro) system and the heritable ovine cataract (in vivo) model were used to test the research hypothesis. The Ca²⁺ ionophore, ionomycin, was used to induce a Ca²⁺ overload and in vitro opacification during lens culture. Opacity in the lens was graded by a computer image analysis program. Protein profile (SDS-PAGE, 2-DE and Western detection), calpain activity (casein zymography), lens structure (microscopic view) and cytotoxicity level (LDH leakage assay) were analysed in Ca²⁺-induced opaque lenses. The involvement of calpain during opacification was further examined by applying synthetic exogenous calpain inhibitors to the in vitro system. Two novel exogenous calpain inhibitors were also assessed for their therapeutic potential in preventing the progression of cataracts in the in vivo cataract model by topical administration of the inhibitor direct to the sheep's eye over a 11 week period. HPLC was used to detect the penetration of these compounds into ocular tissues. Sustained Ca²⁺ influx into cultured lenses caused dense opacification. The opacity was characterised by formation of a turbid fraction and cell death in the outer cortex of the ovine lens. There was increased calpain autolysis associated with the progress of opacification, indicating increased calpain activity. Major degradation of the cytoskeletal proteins, spectrin and vimentin, was observed whilst there was limited degradation of the lens structural soluble proteins, crystallins, in response to a Ca²⁺ flux. Lens proteins were protected from degradation by adding synthetic calpain inhibitors to the culture medium. Topical administration of novel anti-calpain molecules failed to retard the progression of cataractogenesis in the ovine inherited cataract model. Further investigation of drug penetration showed that efficacy of inhibitory compounds was limited by permeability of these molecules across the cornea and the ability of the molecules to reach and penetrate into the lens. The ovine lens Ca²⁺-induced opacification (OLCO) model in this thesis has provided a model to understand the role of Ca²⁺ homeostasis in lens transparency. With sustained intracellular Ca²⁺ level, the degradation of cytoskeletal elements is highly correlated with calpain activity. Cataractogenesis is the pathological response to the loss of lens Ca²⁺ homeostasis in this model. The current results support the hypothesis that the deregulation of calpain activity is a trigger for a series of cascading events, leading to death of the cells in the lens.