Animal Lenses Research Articles

NMR pulse relaxation studies on the normal aging and cataractous lens

Proton NMR pulse relaxation experiments have been performed in this and other laboratories mainly on animal lenses and some human lenses. In order to evaluate T1 and T2 analyses as a potentially useful (non-invasive) parameter in delineating lenticular changes in normal aging human rabbit lenses, and in human cataracts, we performed a series of pulse relaxation studies on human eye bank lenses 10 to 93 yr of age, on fresh intact human cataracts and on normal rabbit lenses. Additional spectroscopic studies (fluorescence and 31P-NMR) were also performed on the lenses in order to correlate these parameters with the data relating to 'water phases' in the lens. These experiments demonstrate a potential additional parameter for evaluating 'molecular' changes in the aging and cataractous lens.

Positron Annihilation Studies of Age-Induced Changes in Animal Lenses

Both, the sulfhydryl (SH) concentration and the state of the sulfur in the eye lens have always been of great interest with respect to age-induced changes and alterations associated with cataract formation. In the present experiments the positron annihilation lifetime technique is employed to study changes in the nucleus and the cortex and changes due to aging of lens material from selected species of animals. In all cases where the lens material is separated into the nucleus and the cortex, the intensity of the longest-lived component, I3, is found to be larger in the nucleus than the cortex. This change is opposite to the change in SH concentration, in particular the glutathione (GSH) concentration. La concentration du sulfhydryl (SH) et l'etat due souffre dans le cristallin, dans leurs rapports avec les changements produits par le vieillissement et les alterations associees avec la formation des cataractes, retiennent depuis longtemps l'attention des chercheurs. Dans les experiences dont il est ici question, nous avons essaye d'utiliser la technique d'annihilation des positrons de vie moyenne pour etudier les changements produits dans le nucleus et le cortex et les alterations dues au vieillissement dans le cristallin de certaines especes animales. Dans tous les cas ou le cristallin etait separe en nucleus et cortex, l'intensite de la composante la plus ancienne, I3, s'est averee plus forte dans le nucleus que dans le cortex. Ce qui est exactement a l'oppose de ce qui se passe dans le cas des changements observes dans la concentration du sulfhydryl (SH), et plus particulierement dans celle du glutathione (GSH).

The effect of oxidation on sorbitol pathway kinetics.

The rapid conversion of glucose to sorbitol by aldose reductase and the consequent hyperosmolarity of the cytoplasm has been shown to be the primary cause of the so-called "sugar" or "osmotic" cataract in many animal lenses. It is not as clear, however, that hyperosmolarity is the principal factor in the etiology of cataracts in human diabetic subjects. In fact, the comparatively low activity of aldose reductase in the human lens as compared with several animal lenses, and the osmotically insignificant levels of sorbitol pathway products (sorbitol and fructose), suggest that hyperosmolarity, per se, may not be as important a factor in human cataract formation as it is in animals. We present evidence that the flux of glucose and sorbitol through the rat lens is markedly reduced by oxidative stress (0.1 mM H2O2). Sorbitol accumulation is reduced by 114%, sorbitol turnover is reduced by 78%, sorbitol production is reduced by 90%, fructose accumulation is reduced by 60%, and fructose turnover is reduced by 76% in the presence of 36 mM glucose. H2O2 does not affect glucose turnover, the glucose rate constant, or the ATP level significantly at 36 mM glucose, but at 5.5 mM glucose, 0.2 mM H2O2 leads to a rapid loss of ATP that can be prevented by 0.04 mM sorbinil, an aldose reductase inhibitor. These results suggest that inhibition of aldose reductase by sorbinil renders rat lenses better able to cope with oxidative stress. In the absence of an aldose reductase inhibitor, elevating ambient glucose may render a lens less able to scavenge oxidants by diverting NADPH into sorbitol production.(ABSTRACT TRUNCATED AT 250 WORDS)

Membrane permeability characteristics of perfused human senile cataractous lenses

Human cataractous lenses were removed by the cryoprobe technique and were maintained for up to 24 hr in a solution of similar ionic composition to human aqueous humour. The bimodal distribution of internal sodium concentrations was similar to that previously reported for unincubated human lenses. Lenses with lower total and free sodium contents had relatively higher membrane potentials and they lost 86Rb at a slower rate than lenses with high internal sodium. The 86Rb efflux in these lenses was stimulated four-fold by removing external calcium. The efflux was reduced by inereasing external calcium, but was increased during a small (60 mosmol) hyperosmotic shock. A similar hyperosmotic shock also surprisingly increased 86Rb efflux. Lenses with increasing internal sodium (and calcium) levels showed an inereasing rate of loss of 86Rb and the stimulation by calcium removal was progressively diminished. The efflux from lenses with disturbed ion levels was also relatively to changes in external osmolarity and to increasing external potassium. Lenses with raised sodium concentrations also had an increased inulin space. Frog, rat and rabbit lenses were also exposed to the same range of stimuli and only frog lenses responded to the low calcium solution with more than a four-fold increase in efflux rate. Although only a two- to four-fold increase in efflux rate was obtained from rabbit lenses exposed to Ca-free conditions, this was the only type of animal lens so far tested that, like the human lens, responded to both hyperosmotic and isosmotic shocks with an increase in efflux rate. All three species of mammalian lenses responded with an increase in efflux rate when exposed to a hyperosmotic test solution while in the frog, the efflux rate from the lens decreased. The glucose efflux from human cataractous lenses was inhibited by cytochalasin B in a similar manner to the efflux from rat and frog lenses. It was concluded, therefore, that the cryoprobed human lens can be kept for a limited period in a relative simple artificial aqueous humour solution. The potassium permeability characteristics of low sodium cataracts remained relatively intact and showed a unique response (relative to lenses from other animals) when exposed to various stimuli that are known to be potentially cataractogenic.

An Experimental and Clinical Evaluation of Lens Transparency and Aging

The normal human lens is a precisely formed structure containing about 65% water and 35% organic matter, the latter being mainly (greater than 90%) structural proteins. The lens becomes encapsulated at the 13 mm embryonic stage and is consequently divorced from a direct blood supply for life. It provides the investigator with an organ that exists in an environment analogous to a single-cell type of tissue culture system and can be reproduced in vitro. Since the cornea and lens are transparent one can directly monitor morphologic and biochemical aging changes in vivo. Recent developments now permit us to monitor certain molecular changes as well. Thus this organ can serve as a unique model for studying in vivo aging parameters. In vivo UV slit lamp densitography to monitor lens fluorescence provides us with a useful diagnostic and prognostic screening method to identify lens photodamage due to excess UV exposure (occupational or accidental), photosensitizers (e.g. psoralens) as well as the normal age related increase in lenticular fluorescence. Cortical opacities are also a common finding in older individuals. In vitro NMR (human and animal lenses) and in vivo (animal experiments) currently in progress are rapidly elucidating the physicochemical basis for the development of light scattering areas in the aging lens cortex. This review demonstrates the feasibility of utilizing biophysical methods, in particular, optical spectroscopy and NMR analyses to delineate and monitor specific age related parameters in the ocular lens, in vivo as well as in vitro.

Lens metabolism and aging: enzyme activities and enzyme alterations in lenses of different species during the process of aging.

Extensive investigations on the metabolism of animal lenses over the life span indicate various changes that, regarded collectively, suggest a decrease in metabolic energy. A factor analytic interpretation and discussion of these results indicate that enzyme modifications might be one of the fundamental causes of the age-related changes in lens metabolism. Detailed investigations on the kinetic and physicochemical properties of certain enzymes of the carbohydrate breakdown in lenses of different ages verify this assumption. Since the changes observed are found mostly in the lens nucleus protein neosynthesis no longer takes place, posttranslational modifications appear to be responsible.

Lens plasma membrane: Isolation and biochemical characterization

Lens plasma membrane from different animal lenses has been prepared by the acylation procedure. Using three different criteria: heat aggregation of intrinsic membrane polypeptides, immunochemistry and solubilization of intrinsic proteins at low (40 mM) LIS concentration, it has been shown that these preparations are essentially free of cytoplasmic contaminants. Using the results obtained with acylated membrane as the reference of purity, it has been shown that both sucrose gradient centrifugation of bovine WI protein and urea washing of old human lenses give impure membrane preparations. The main intrinsic polypeptides (mol. wt 26 000 and 22 000) of human lens membrane have been purified and characterized. It has been shown by enzymatic digestion and amino terminal analysis of the residual membrane-bound fragments that the amino terminal halves of the polypeptides are embedded in the lipid bilayer and are probably blocked at their amino terminal sites. Lipid analyses of human and bovine lens membranes suggest that the protein to total lipid ratio is 1:1. Carbohydrate analyses of chromatographically separated intrinsic membrane polypeptides indicate the presence of 1 mol glucose/1 mol protein.

Lens membranes XV. Comparative study of membrane and cytoplasmic proteins from human, monkey and other animal lenses

Various characteristics of membrane and cytoplasmic proteins from human, monkey and other animal lenses have been studied. The polypeptide patterns of lens fiber membranes of man and monkey are very similar. They contain two major polypeptides: a main intrinsic polypeptide of 27 dalton (MIP) and its 22000–23000 dalton conversion product, which can both be extracted by chloroform-methanol. Monkey MIP is immunologically identical to human MIP. Both show a reaction of partial identity with the MIPs of calf, sheep and pig. A weak cross-reactivity is found between primate MIP and chicken MIP. EDTA-extractable proteins (EEP) of 3200 and 35000 dalton can be isolated from monkey lens membrans with low recovery, while EDTA fails to extract any EEP from human lens membranes. The individual crystallins of man and monkey are immunologically closely related. They show reaction of partial identity with the crystallins of calf, sheep and pig. Chicken α - and β -crystallin are partially identical to α - and β -crystallin from mammalian and primate lenses.

Ultraviolet action spectrum for fluorogen production in the ocular lens.

Abstract— …Previous work has demonstrated that fluorescent material (360nm excitation, 440nm emission), whose concentration normally increases with age in human lenses, can be generated artificially by exposing cultured human or animal lenses to UV radiation. In the present paper we report measurements of the rate of production of this fluorescent material in rat lenses in vitro as a function of UV irradiation wavelength. A plot of the observed rate of fluorogen production normalized to constant photon flux vs irradiation wavelength shows little action at 360 or 320nm, increases sharply at 300nm, remains relatively constant in the range 300–280nm, and then exhibits a further gradual rise from 270–250nm. The results on rat lenses are compared with results reported elsewhere for tryptophan in aqueous solution. The action spectrum for photochemical destruction of tryptophan in solution closely parallels that for fluorogen production in rat lenses. This result and other evidence suggest that photochemical destruction of tryptophan might be the initial event in UV‐induced fluorogen production in the ocular lens.

Cataracts: Chemistry, Mechanisms, Therapy.

Dr. Lerman's most recent book, Cataracts: Chemistry, Mechanisms, Therapy, examines closely and evaluates thoroughly the knowledge available about the production of cataracts. This richly illustrated monograph consists of a detailed consideration of the embryology and morphology of the lens and adjacent structures, including many electron micrographs of the human and animal lens. The carbohydrate, protein, and nucleic acid metabolism of the lens is discussed with a thoroughness that is more than sufficient for the average reader. The process of lens aging and regeneration and the mechanisms of experimental cataracts and their relationship to the cataractous process in man are unraveled in a clear, factual manner. The author presents a simplified classification of cataracts, discusses the lens-induced inflammations, and deals briefly with the therapy of each abnormality. This monograph is particularly well written and presents an organized, comprehensible discussion of the lens from an embryologic, physiologic, biochemical, and therapeutic aspect. The

STUDIES ON THE EXCITED STATES OF PROTEINS

During the 1950s, Szent-Gyorgyi began exploring the physical properties of biological substances (a field he called bioenergetics, electronic biology, or sub-molecular biology). In this article, he and Richard Steele reported some of their studies which tested the optical properties (e.g., fluorescence and phosphorescence) of animal lens protein, several amino acids, and proteins such as albumin, trypsin, and myosin.

ON EXCITATION OF BIOLOGICAL SUBSTANCES

During the 1950s, Szent-Gyorgyi began exploring the physical properties of biological substances (a field he called bioenergetics, electronic biology, or sub-molecular biology). In this study, he and Richard Steele tested the optical properties (e.g., fluorescence) of the structural proteins in animal lenses, six different amino acids, in adenine and several of its derivatives, including adenosine triphosphate (ATP), DNA and RNA, and several other substances.

Paper electrophoresis of hydrosoluble proteins of the crystalline lens

By using paper electrophoresis, the authors succeeded in demonstrating different protein fractions in human and animal lenses. Qualitative and quantitative differences were noted in different animals, between the cortical and the nuclear areas of the lens, and also according to age.

Investigation on the crystalline lens

Although a large number of papers have been written on the nature and possible causes of cataract, it is only recently that any attempt has been made to attack the problem from a biochemical standpoint. It was a thought, therefore, that experiments on the biochemistry of the normal animal lens might be of some value in the solving of this problem.