Fusun Kilic

Date of Award


Degree Type


Degree Name

Doctor of Philosophy


Cataract formation is a complex, multifactorial and multistep process: it is associated with cellular and biochemical changes in the lens. In the present work, steps in the pathway to cataract development were followed in vitro using two different reagents which induce cataract formation by two different mechanisms: elevated glucose (55.6 mM), as a model for in vivo diabetic cataractogenesis and an actin monomer-stabilizer, cytochalasin D (10{dollar}\sp{lcub}-5{rcub}{dollar} M, CD). Findings were utilized to classify and arrange the events in a common pathway and to focus on the mechanism of cataract production.;The development of cortical subcapsular opacity in cultured lenses was elucidated as a time-dependent phenomenon by measuring different parameters: (i) distribution of the living and dead cells in the lens through a confocal microscope following their fluorescent staining, (ii) appearance of the lens observed daily by a dissection microscope, and (iii) changes in and/or leakage of intracellular components ({dollar}\gamma{dollar}-crystallin, LDH, and magnesium ion). The findings suggested possible mechanisms for the action of the cataract-causative agents, glucose and CD, which initiate membrane damage in the lens cells in both model systems and help locate these factors at common steps in their pathways to cataract.;Using this reproducible lens incubation system, various potential anti-cataract agents e.g., antioxidants, sorbitol-lowering agent, and other agent were tested in an in vitro diabetic model systems. All these agents were relatively effective in preventing cataract formation in the in vitro diabetic model system. These findings support the hypothesis that the damage processes in the presence of glucose involve glycation and oxidative stress. The effect of an antioxidant, Vitamin C (VC), was also tested to explore the factors involved in the mechanism of CD-cataract formation. Prevention of damage by VC is consistent with the hypothesis that oxidative stress is associated with the disorganization of actin and cytoskeletal network during cataract formation.;The timing and localization of calcium uptake in the lens was studied in both cataractous model systems by radioactive and fluorescent labelling techniques. The lens calcium uptake increased parallel to the incubation time in cataract causative agents. The effect of CD was earlier and more severe than the glucose effect. The results of lens glucose-treatment, as well as inhibitors showed that elevated calcium could be one of the factors leading to cataract development. Finally, the role of calpain-proteolysis of spectrin/fodrin was studied during cataract formation process in both model systems. The results suggest that the proteolysis of spectrin/fodrin does not precede cataract formation but it is a consequence of precataractous changes.



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