Hyperbaric oxygen as a model of lens aging in the bovine lens: The effects on lens biochemistry, physiology and optics.

Age related nuclear (ARN) cataracts in humans take years to form and so experimental models have been developed to mimic the process in animals as a means of better understanding the etiology of nuclear cataracts in humans. A major limitation with these animal models is that many of the biochemical and physiological changes are not typical of that seen in human ARN cataract. In this review, we highlight the work of Frank Giblin and colleagues who established an in vivo animal model that replicates many of the changes observed in human ARN cataract. This model involves exposing aged guinea pigs to hyperbaric oxygen (HBO), which by causing the depletion of the antioxidant glutathione (GSH) specifically in the lens nucleus, produces oxidative changes to nuclear proteins, nuclear light scattering and a myopic shift in lens power that mimics the change that often precedes cataract development in humans. However, this model involves multiple HBO treatments per week, with sometimes up to a total of 100 treatments, spanning up to eight months, which is both costly and time consuming. To address these issues, Giblin developed an in vitro model that used rabbit lenses exposed to HBO for several hours which was subsequently shown to replicate many of the changes observed in human ARN cataract. These experiments suggest that HBO treatment of in vitro animal lenses may serve as a more economical and efficient model to study the development of cataract. Inspired by these experiments, we investigated whether exposure of young bovine lenses to HBO for 15 h could also serve as a suitable acute model of ARN cataract. We found that while this model is able to exhibit some of the biochemical and physiological changes associated with ARN cataract, the decrease in lens power we observed was more characteristic of the hyperopic shift in refraction associated with ageing. Future work will investigate whether HBO treatment to age the bovine lens in combination with an oxidative stressor such as UV light will induce refractive changes more closely associated with human ARN cataract. This will be important as developing an animal model that replicates the changes to lens biochemistry, physiology and optics observed in human ARN cataracts is urgently required to facilitate the identification and testing of anti-cataract therapies that are effective in humans.

Hydrotropic function of ATP in the crystalline lens.

The hypothesis proposed herein is presented to explain the unexpectedly high concentration of ATP and provide evidence to support its hydrotropic function in the crystalline lens determined using 31P NMR. The lens, historically considered to be a metabolically quiescent organ, has the requisite machinery to synthesize ATP, such that the homeostatic level is maintained at about 3 mM. This relatively high concentration of ATP has been found to be consistent among multiple mammalian species including humans. This millimolar quantity is many times greater than the micromolar amounts required for the other known functions of ATP. The recent postulation that ATP at millimolar concentrations functions as a hydrotrope in various cell/tissue homogenates preventing protein aggregation coupled with observations presented herein, provide support for extending the hypothesis that ATP functions as a hydrotrope not only in homogenates but in an intact functioning organ, the crystalline lens. Concentrations of ATP of this magnitude are hypothesized to be required to maintain protein solubility and effectively prevent protein aggregation. This concept is important considering protein aggregation is the etiology for age-related cataractogenesis. ATP is a common ubiquitous intracellular molecule possessing the requisite hydrotropic properties for maintaining intracellular proteins in a fluid, non-aggregated state. It is proposed that the amphiphilic ATP molecule shields the hydrophobic regions on intralenticular fiber cell protein molecules and provides a hydrophilic interfacial surface comprised of the ATP negatively charged triphosphate side chain. Evidence is presented that this side chain is exposed to and has been reported to organize intracellular interstitial water to form an interfacial rheologically dynamic water layer. Such organization of water is substantiated with the effect of deuterium oxide (heavy water) on ATP line widths of the side chain phosphates measured ex vivo by 31P NMR. A novel model is presented to propose how this water layer separates adjacent lens fiber cell proteins, keeping them from aggregating. This hypothesis proposes that ATP can prevent protein aggregation in normal intact lenses, and with declining concentrations can be related to the disease process in age-related cataractogenesis, an affliction that affects every older human being.

The influence of storage method on the transparency of pig crystalline lens / A influência do método de armazenamento na transparência do cristalino de porco

ABSTRACT Purpose: The porcine eye is frequently used as a research model. This paper analyzes the effect of different storage methods on the transparency of pig crystalline lens. Methods: A spectral transmission curve (from 220 to 780 nm) for the crystalline lens was determined experimentally after storage in different conditions: saline solution, formalin, castor oil, and freezing at -80°C. The total transmission in the visible spectrum, which was used as an index of transparency, was calculated from these curves. For comparative purposes, fresh lenses were evaluated and used as controls. Results: Storing the porcine crystalline lens in saline solution or castor oil resulted in a transparency loss of approximately 10% after 24 h and storage in formalin resulted in a loss of nearly 30%. Storage by freezing at -80°C for 4 weeks maintained the transparency of the crystalline lens; the spectral transmission measured immediately after defrosting at room temperature coincided exactly with that of the freshly extracted lens. Conclusions: The transparency of porcine crystalline lens is affected by the storage method. The visible spectrum is the most affected, evidenced by the effect on the transparency and consequently the amount of light transmitted. The results show that freezing at -80°C maintains the transpa rency of the crystalline lens for at least 4 weeks.

RESUMO Objetivos: Olho de porco é frequentemente usa do como modelos de pesquisa. Este estudo analisa o efeito de di ferentes métodos de armazenamento na preservação da transparência do cristalino de porco. Métodos: Uma curva de transmissão espectral (de 220 até 780 nm) para o cristalino foi experimentalmente determinada após armazenamento em diferentes condições: solução salina, formol, óleo de mamona e congelamento a -80°C. Transmissão total do espectro visível, que foi usada como um índice de transparência foi calculada a partir dessas curvas. Para fins comparativos, lentes frescas foram avaliadas e usadas como controles. Resultados: O armazenamento do cristalino suíno em solução salina ou óleo de mamona resultou uma perda de transparência de aproximadamente 10% após 24 h e o armazenamento em formol resultou uma perda de quase 30%. O armazenamento por congelamento a -80°C durante 4 semanas manteve a transparência do cristalino; a transmissão espectral medida imediatamente após o descongelamen to à temperatura ambiente coincidiu exatamente com a da lente extraída recentemente. Conclusão: A transparência do cristalino suíno é afetada pelo método de armazenamento. O espectro visível é o mais afetado, evidenciado pelo efeito sobre a transparência e consequentemente a quantidade de luz transmitida. Os resultados mostram que o congelamento a -80°C mantém a transparência do cristalino suíno por pelo menos 4 semanas.

The influence of storage method on the transparency of pig crystalline lens.

PURPOSE: The porcine eye is frequently used as a research model. This paper analyzes the effect of different storage methods on the transparency of pig crystalline lens. METHODS: A spectral transmission curve (from 220 to 780 nm) for the crystalline lens was determined experimentally after storage in different conditions: saline solution, formalin, castor oil, and freezing at -80°C. The total transmission in the visible spectrum, which was used as an index of transparency, was calculated from these curves. For comparative purposes, fresh lenses were evaluated and used as controls. RESULTS: Storing the porcine crystalline lens in saline solution or castor oil resulted in a transparency loss of approximately 10% after 24 h and storage in formalin resulted in a loss of nearly 30%. Storage by freezing at -80°C for 4 weeks maintained the transparency of the crystalline lens; the spectral transmission measured immediately after defrosting at room temperature coincided exactly with that of the freshly extracted lens. CONCLUSIONS: The transparency of porcine crystalline lens is affected by the storage method. The visible spectrum is the most affected, evidenced by the effect on the transparency and consequently the amount of light transmitted. The results show that freezing at -80°C maintains the transpa rency of the crystalline lens for at least 4 weeks.

Macular pigment density variation after supplementation of lutein and zeaxanthin using the Visucam® 200 pigment module: Impact of age-related macular degeneration and lens status.

PURPOSE: To assess the evolution of macular pigment optical density (MPOD) following supplementation with various macular formulations obtained with the Visucam® 200, and to study the factors affecting MPOD measurements. MATERIALS AND METHODS: In this prospective, randomized, double-masked multicenter study, patients were divided into 2 groups: group A (patients without retinal pathology who underwent cataract surgery 1 month previously) and group B (patients with neovascular age-related macular degeneration [AMD] in one eye). In each group, half of the patients were randomly assigned to receive a food supplementation either with or without carotenoids (5mg of Lutein and 1mg of Zeaxanthin). Outcome measures included MPOD responses obtained with the Visucam® 200 for one year. RESULTS: In total, 126 subjects (52 men, 74 women) with a mean age of 75.3±7.61 years were enrolled. Mean MPOD values at the time of inclusion were statistically lower in group A (0.088 density unit [DU]) compared to group B (0.163 DU, P<0.05). No statistically significant increase in MPOD was noted in either group, even after discontinuation of the supplementation. By multiple regression analysis, age, female gender, lens status and the presence of AMD seemed to significantly affect MPOD measurements. CONCLUSION: No significant improvement in MPOD seems to be detected with the Visucam® 200 after carotenoid supplementation. The MPOD measurement seems to be highly affected by cataract extraction and the presence of AMD.

Decompression-induced ocular tear film bubbles reflect the process of denitrogenation.

PURPOSE: To confirm that the tear film bubbles observed after decompression from hyperbaric exposure are due to denitrogenation and to assess the time course of denitrogenation based on the number of ocular tear film bubbles. METHODS: The study comprised two parts. In the first, subjects (n = 8) were compressed to a pressure of 2.0 ATA (atmospheres absolute; depth of 10 meters of sea water [msw]) for 60 minutes in a hyperbaric chamber on two separate occasions. On one occasion they breathed air, whereas on the second occasion they inspired pure oxygen. Before and within 30 minutes after each dive, the subjects' tear film was examined with a slit lamp microscope and the average number of bubbles recorded. Ultrasound reflectivity of the lens-vitreous humor compartments was also examined. In the second part of the study, subjects (n = 8) participated in two simulated dives in the hyperbaric chamber: 4.0 ATA (30 msw) for 15 minutes and 2.5 ATA (15 msw) for 180 minutes. The former was a no-stop decompression dive, whereas the latter required a 43-minute decompression stop at 3 msw. Ocular tear film examinations were conducted before the dive, as well as 30 minutes and 1 day, 2 days, and 3 days after the dives. RESULTS: The number of tear film bubbles increased significantly (P < 0.05) after the air dives to 2.0 ATA for 60 minutes, whereas there was no significant postdecompression increase in tear film when oxygen was inspired by the subjects during the dive. Posterior lens-vitreous humor reflectivity increased significantly after decompression from 2 ATA, when air was the breathing mixture, whereas no change in reflectivity was observed when oxygen was inspired during the dive. In the second part of the study, there was a significant elevation in tear film bubbles for 2 days after the two dives (P < 0.05). There was no significant difference in the number of ocular tear film bubbles between the two dives. CONCLUSIONS: After a hyperbaric air exposure, tear film bubbles reflect the process of denitrogenation, which may persist for up to 2 days after the decompression.

Ocular bubble formation as a method of assessing decompression stress.

Tear film bubble formation and ultrasound reflectivity of the lens-vitreous humor compartments were monitored following simulated dives in a hyperbaric chamber. the sensitivity of these methods in determining decompression stress was compared with the results of precordial Doppler ultrasound. In addition, the utility of these diagnostic techniques in testing decompression dive profiles was evaluated. Eleven divers completed two series of chamber dives according to the decompression schedule of the Professional Association of Diving Instructors. The first dive series comprised dives to 70 feet of seawater (fsw) for 15, 29, and 40 min. The second series comprised maximum duration no-stop decompression dives to 40 fsw for 140 min, 70 fsw for 40 min, 90 fsw for 25 min, and 120 fsw for 13 min. Before and immediately after each dive, the following measurements were obtained from each subject: eye surface tear film bubble counts with a slit-lamp microscope, lens and vitreous humor reflectivity using A- and B-mode ophthalmic ultrasonic scan, and precordial Doppler ultrasonic detection of venous gas bubbles. Tear film bubble assessment and ocular scanning ultrasound were observed to be more sensitive in detecting decompression stress than the conventional Doppler ultrasonic surveillance of the precordial region. In contrast to precordial Doppler ultrasonic surveillance, which failed to detect any significant changes in circulating bubbles, tear film bubble formation displayed a dose-response relationship with increasing duration of the 70-fsw dives. Reflectivity changes of the lens-vitreous humor interface were not significant until the no-stop decompression limit was reached. In addition, for each of the no-stop decompression limit dives, increases in the average tear film bubble formation and lens-vitreous humor interface reflectivity were similar. Ocular bubble observations may provide a practical and objective ocular bubble index for analyzing existing decompression schedules and predicting individual susceptibility to decompression sickness.

Selenium-induced cataract--a correlation of dry mass content and light scattering.

Selenium-induced cataracts in young rats were used to compare light scattering and dry mass concentration along the lens axis. Selenium-treated rats and control rats were examined 1, 2, 4 and 6 days after subcutaneous injection of selenium or 0.9% NaCl. The development of nuclear cataract was followed with a Scheimpflug slit-lamp camera. Light-scattering was determined by densitometry of the negatives taken by the Scheimpflug camera. Dry mass concentration was determined by quantitative microradiography of the extracted lenses. Increasing light scatter was seen from day 2 to day 6. The dry mass concentration, however, was not changed until day 6. Two distinct zones were found surrounding the center of the nucleus, one with increased and one with decreased dry mass. The rapid changes in dry mass concentration were probably caused by shifts in water distribution within the lens. Similar zones of hydration have also been found in human senile nuclear cataract. Selenium-induced cataract in the rat is discussed in relation to human nuclear cataract.