Crystallization and preliminary X-ray data of human plasma retinol-binding protein.

Crystals of human plasma retinol-binding protein have been obtained from 4.5 M-NaCl buffered at pH 6.8 with 20 mM-cacodylate. The crystals are trigonal with space group R3 and unit cell dimensions, referred to the hexagonal system, a = b = 104.2 A and c = 74.5 A. The crystals diffract to a resolution of 2.0 A.

Associations between plasma levels of vitamins and cataract in the Italian-American Clinical Trial of Nutritional Supplements and Age-Related Cataract (CTNS): CTNS Report #2.

PURPOSE: To investigate the association at baseline between plasma levels of selected vitamins and the presence and type of cataract in the participants in The Italian-American Trial of Nutritional Supplements and Age-related Cataract. METHODS: At baseline, the participants (1020, 710 with "early cataract" and 310 with "no cataract," 55-75 years of age) received an ocular examination, photographic lens grading, and measurement of plasma levels of vitamins A, C, E, beta-carotene, and of red blood cell glutathione reductase activity. RESULTS: In multiple logistic models adjusted for potential confounders, high vitamin C levels were associated with a protective effect on nuclear (N) [OR: 0.54; 95% CI: 0.30, 0.97] and posterior subcapsular (PSC) cataract (OR: 0.37; 95% CI: 0.15, 0.93). High vitamin E levels were associated with increased prevalence of cortical cataract (C) (OR: 1.99; 95% CI: 1.02-3.90), PSC (OR: 3.27; 95% CI: 1.34, 7.96) and of any cataract (OR: 1.86; 95% CI: 1.08, 3.18). CONCLUSIONS: In agreement with some earlier studies, we found higher plasma levels of vitamin C to be associated with reduced prevalence of N and PSC cataracts. The finding of an increased prevalence of some types of cataract with higher levels of vitamin E was unexpected, has not been previously reported, and could be due to unadjusted confounding.

Retroillumination versus reflected-light images in the photographic assessment of posterior capsule opacification.

PURPOSE: To investigate the relative merit of retroillumination and of reflected light slit-lamp-derived photographs in the assessment of the opacification of the posterior lens capsule. METHODS: Retroillumination and slit-lamp-derived reflected-light photographs were taken on 23 consecutive eyes with posterior capsule opacification (PCO) in uncomplicated pseudophakia. Subjective grading was performed on both types of photographs to evaluate the extent and density of posterior capsular opacification. Best-corrected visual acuity (BCVA) before and after YAG laser capsulotomy was used to assess the impact of capsular opacification on visual function. RESULTS: After capsulotomy all patients attained a BCVA > or = 46 letters (> or =20/32) with a mean increase of 25 letters, indicating that PCO was the cause of visual impairment in these patients. The relative capacity of retroillumination and of reflected-light photographs to adequately capture the extent and the severity of posterior capsule opacification varied considerably. Reflected-light images, in addition to frequently producing higher severity scores for the opacity than retroillumination photographs, in 4 of 23 eyes (17.4%) proved to be the only technique able to document the presence of PCO. CONCLUSIONS: Our results indicate that, with respect to retroillumination images, reflected-light photography has an increased ability to adequately capture the presence and the severity of PCO and that the use of only retroillumination images may lead to its underestimation. This may be relevant to clinical studies aiming to evaluate incidence and progression of this condition.

Enzyme activities in human lens epithelium of age-related cataract.

PURPOSE: To investigate associations between enzyme activity of glutathione reductase (GR) with and without added flavin adenine dinucleotide (FAD), glutathione peroxidase (GPX), and 6-phosphogluconic dehydrogenase (6PGDH) in the lens epithelium collected at surgery, and some nutritional and biochemical variables determined in the same individuals during the Italian-American Case-Control Study of age-related cataract. METHODS: One hundred eighty-three epithelium capsule samples were collected from 174 patients undergoing surgery. Data on enzyme activity were obtained from 52 samples for 6-PGDH and from 53 samples for GR and for GPX. The Lens Opacity Classification System II was used to classify and grade cataracts. RESULTS: No correlation was found between enzyme activity in lens epithelium and the same enzymatic activity in erythrocytes (with the exception of a negative correlation between lens and erythrocyte 6PGDH activity), or the type and severity of cataract. No correlation was found between lens GPX activity and plasma selenium and between lens GR activation coefficient (GRAC) and riboflavin intake. Lens GR with added FAD and lens GRAC were significantly correlated to plasma vitamin E level. Lens GRAC was positively correlated to a nutritional vitamin index. CONCLUSIONS: Present data stress the difficulty in verifying the assumption that biochemical indices collected on plasma and on erythrocyte actually reflect the status of these factors in the lens itself.

Location and severity of cortical opacities in different regions of the lens in age-related cataract.

PURPOSE: To examine the distribution of cortical opacities across the lens in the Italian-American Natural History Study of Age-Related Cataract and to study the association between an index of sunlight exposure and the location of cortical cataract within the lens. METHODS: Lens photographs of one eye of 731 persons with cortical opacities (503 with pure and 228 with mixed types of opacity) were included in the analysis. A radial grid superimposed on the photographs was used to assess presence, location, and severity of wedge-shaped cortical opacities. RESULTS: Both the prevalence and the extent of cortical opacities were highest in the inferior-nasal quadrant and lowest in the superior-nasal quadrant of the lens. In polychotomous logistic regression, persons with the greatest excess areal involvement in the inferior half of the lens were more likely to have high exposure to sunlight, as measured by a sunlight index, than persons with excess involvement in the superior half of the lens (odds ratio, 1.73; 95% confidence interval 1.03, 2.93). Excess areal involvement of the inferior lens also was associated with the pure type of cortical cataract and with the total extent of the opacity. CONCLUSIONS: Age-related cortical opacities occur more frequently inferiorly than superiorly and, to a lesser extent, nasally than temporally. Possibly higher exposure of these lens segments to sunlight may explain this preferential location of cortical opacities.

Human lens membrane cation permeability increases with age.

Parallel studies of the ionic balance and membrane permeability characteristics of normal human lenses were carried out in three countries (USA, England and Italy). Similar age-related changes were found in each laboratory. The lens membrane potential and resistance declined markedly with age while internal Na+ and free Ca2+ increased. There was a concomitant stimulation of Na+ and K+ transmembrane fluxes. These data indicate that in the ageing process there is an increasing contribution to membrane ion traffic from a channel, or channels, that permit Na+, K+ and Ca2+ to pass. The increase in permeability coincides exactly with the increase in optical density that occurs in the ageing human lens.

The effect of cataract severity and morphology on the reliability of the Lens Opacities Classification System II (LOCS II).

Data collected from 3646 eyes in the Italian-American Natural History Study of Age-Related Cataract were used to investigate whether the reliability of the Lens Opacities Classification System II (LOCS II) by the severity of the opacity that is being graded or is influenced by the presence and severity of coexisting opacities. Reliability was assessed by comparing the slit-lamp gradings of two clinical examiners (346 eyes) and the gradings performed at the slit lamp with gradings of photographs (3646 eyes). The severity of cortical and nuclear opacities did not affect the reproducibility of slit-lamp gradings, but clinical grading of posterior subcapsular opacities became more reliable as the severity of the posterior subcapsular opacities increased. More advanced coexisting opacities decreased the agreement in the slit-lamp diagnosis of nuclear, but not cortical or posterior subcapsular, opacities. Comparisons of clinical and photographic gradings showed very good to excellent agreement for nuclear and cortical opacities, regardless of the severity of the specific opacity or the severity of the coexisting opacities. Agreement in diagnosing posterior subcapsular opacities was decreased in eyes with milder posterior subcapsular opacities and in eyes with more severe coexisting nuclear and/or cortical opacities. The effect of the severity of the opacity being graded and the severity of coexisting opacities on the reliability of the LOCS II must be considered in studies that use the system to classify and grade cataracts.

Glutathione S-transferase M1 genotype and age-related cataracts. Lack of association in an Italian population.

PURPOSE: To investigate possible associations between the gene number and allelic forms of glutathione S-transferase M1 (GSTM1) and the occurrence of nucleic and cortical age-related cataracts. METHODS: Patients with cortical cataract, nuclear cataract, mixed and cortical cataract, and no cataract were sytematically selected from subjects evaluated in the Italian-American Study of the Natural History of Age-Related Cataract. The patients were typed for the A, B, and null alleles of GSTM1 using a variation of the amplification refractory mutation system. RESULTS: Forty-nine percent of patients (50/102) with cortical cataracts, 45% (13/29) with nuclear cataracts, 51% (36/71) with mixed nuclear and cortical cataracts, and 50% of controls (49/98) were homozygous for the null GSTM1 allele. Twenty-five percent of patients (26/102) with cortical cataracts, 24% (7/29) with nuclear cataracts, 31% with mixed nuclear and cortical cataracts, and 27% of controls (26/98) displayed only the A allele for GSTM1. Twenty-four percent of patients (24/102) with cortical cataract, 24% (7/29) with nuclear cataracts, 14% (10/71) with mixed nuclear and cortical cataract, and 18% of controls showed only the B allele for GSTM1. Two percent of patients (2/102) with cortical cataracts, 7% (2/29) with nuclear cataracts, 4% (3/71) with mixed nuclear and cortical cataracts, and 5% of controls (5/98) showed both A and B alleles for GSTM1. CONCLUSIONS: No associations between the GSTM1 alleles, including the null allele, and cataracts were detected in this study.

Influence of type and severity of pure forms of age-related cataract on visual acuity and contrast sensitivity. Italian American Cataract Study Group.

PURPOSE: To compare the relationship between logMAR visual acuity (VA) and cataract severity and between contrast sensitivity (CS) and cataract severity in pure types of age-related lens opacities. METHODS: Analysis included patients followed in the ongoing Italian-American Study of the Natural History of Age-Related Cataract. Lens opacities were classified and graded according to the Lens Opacities Classification System II (LOCS II). Visual acuity was measured with the Early Treatment Diabetic Retinopathy Study Chart. Contrast sensitivity was measured with the Pelli-Robson chart. RESULTS: Data from 1,076 eyes were used for the analysis (366 clear lenses; 550, 124, and 36 eyes with cortical, nuclear and posterior subcapsular cataract, respectively). In age-adjusted analyses, increasing severity of all three cataract types was associated with progressively higher logMAR VA, which translates into poorer acuity, and lower CS scores. For both VA and CS, the effect of increasing severity was greatest for nuclear and least for cortical opacities. After adjusting for age and VA, CS scores were no longer associated with cataract type and severity, with the exception of advanced cortical opacities. CONCLUSIONS: Increased cataract severity, as determined by LOCS II grading, is strongly associated with both VA and CS scores. Contrast sensitivity scores obtained from testing at low spatial frequency do not seem to offer additional information over standard VA testing in early cortical and posterior subcapsular opacities nor in nuclear cataracts.

A dose-response effect between a sunlight index and age-related cataracts. Italian-American Cataract Study Group.

To explore the existence of a dose-response relationship between sunlight exposure and risk of age-related cataracts, we analyzed data collected from 1008 patients with cataracts and 469 control subjects enrolled in the Italian-American Case-Control Study of Age-Related Cataracts. Fourteen variables related to sunlight exposure history were included in the questionnaire administered to the study participants. A sunlight index was constructed and its relationship to the presence of cataracts was modeled by logistic regression. After adjustments for potential confounding variables and for age and sex, a significant dose-response effect (P = 0.01) was detected between the sunlight exposure index and the presence of pure cortical cataracts. With the exception of corticonuclear cataracts, all the other mixed types of opacity also showed a dose-response association with the sunlight index. These data support the hypothesis that sunlight exposure is a risk factor in the development of cortical cataracts, and demonstrate the existence of a dose-response relationship in this association.

Interocular transfer of a visual aftereffect in early-onset esotropia.

The interocular transfer of the tilt aftereffect was studied in strabismic patients with early-onset esotropia and in controls. Squinting subjects were divided into a microtropia subgroup with anomalous binocular vision and a large-angle esotropia subgroup with total suppression of the deviated eye. Patients with alternating microtropia and large-angle esotropia showed normal or moderately reduced interocular transfer of the aftereffect. Patients with monocular microtropia with medium- or low-degree amblyopia showed a reduced monocular visual aftereffect in the dominant eye and practically no transfer to the nondominant eye. Adaptation of the nondominant eye and transfer of the aftereffect to the dominant eye was normal. The interocular transfer of the tilt aftereffect is a poor indicator of the state of binocularity of a squinting subject, and the clinical usefulness of this test is limited.

Preliminary evaluation of a simple photographic cataract classification for epidemiological surveys.

A preliminary evaluation of the agreement between clinical and photographic cataract classification and its reproducibility by utilizing a very simple classification system is presented. Photographic classification was based on color Zeiss 75-SL transparencies. Results indicate that photo-derived cataract classification based on slitlamp photographs has good validity for nuclear opacities but tends to underestimate posterior subcapsular cataracts.

The isolation by self forming gradients of Percoll of plasma membrane enriched fractions from bovine retinal pigment epithelium.

A method is described for the preparation of plasma membrane enriched fractions from bovine retinal pigment epithelium (RPE) by means of differential centrifugation followed by the use of self forming gradients of Percoll. A detailed analysis of the distribution of organelle specific markers (nuclei, mitochondria, lysosomes, endoplasmic reticulum, cytosol) in the different fractions is presented. Comparison of 125I-wheat germ agglutinin (WGA) binding with more conventional plasma membrane enzyme markers demonstrates that also in RPE radiolabeled lectin is a specific and extremely sensitive marker to follow quantitatively the distribution of outer cell membranes. Results of 125I-WGA displacement experiments indicate that plasma membranes are mostly (90%) composed of right side out vesicles or sheets. On the basis of 125I-WGA radioactivity the overall recovery of plasma membranes was about 10% and purification over 15 fold. NADH cytochrome c reductase activity, which is shown to be a specific marker for endoplasmic reticulum in retinal pigment epithelium, has been utilized to evaluate microsomal contamination of the plasma membrane preparation.

Amino-acid transport and protein synthesis in the rabbit lens: absence of cryoprobe effect.

Recently developed organ-culture techniques have been used to investigate the effects of cryoprobe treatment on rabbit lenses. Uptake of 14C-tyrosine into cryoprobe treated and control lenses was followed for 96 h. Lens proteins were separated by gel filtration and incorporation of label measured in the individual crystallins. The cryoprobe treatment had no measurable effect on lens water, Na+, K+ or Ca++ content, tyrosine transport or the incorporation of tyrosine into the crystallins, during the period of the experiment.

Cytoskeleton abnormalities in human senile cataract.

The cytoskeletal pattern of the most superficial layers (cortex and epithelium) of senile cataractous lenses has been analyzed by PAGE-SDS. While the nuclear type of cataract and age-matched transparent human lenses have superimposable protein patterns, lenses with cortical cataract demonstrate appreciable modifications of their cytoskeletal composition. The most evident change is the decrease of fodrin and the marked reduction or even the absence of the 98 Kd band. Fodrin may be completely removed from the water insoluble fraction (WIF) of cortical cataract by extraction in low ionic strength buffer, a treatment which only partially solubilizes this protein in transparent control lenses.

86-Rb efflux in normal and cataractous human lenses.

86Rb efflux has been studied in normal lenses and in human senile cataracts. The rate constant (Ki) of the efflux gradually increases in cataractous lenses with progression of lens damage. Efflux experiments run in the presence of BaC12 suggest that a progressive activation of BaC12 inhibitable efflux routes occurs in cataractous lenses. In the final stages of opacification the ineffectiveness of BaC12 enriched or Ca++ free media on the efflux suggests that a direct disruption of the lens membranes has occurred.

Oxidative cross-linking of fodrin parallels a membrane conductance increase in the mammalian lens.

An oxidative cross-linking of the lens spectrin-like protein fodrin was induced by incubating WKY-rat lenses in the presence of the SH-reagent diamide. The oxidation of fodrin was paralleled by an increase in lens membrane conductance. The time relationship between these two events as well as the reversibility of both, achieved by incubating the lens in the presence of dithiothreitol, indicate that normal permeability characteristics of the lens membranes require the integrity of the membrane attached cytoskeleton.

Mammalian lens inter-fiber resistance is modulated by calcium and calmodulin.

The relationship between Ca2+ and lens fiber cell communication was investigated in the isolated intact rat lens by using radiotracer and electrophysiological techniques. The lens internal calcium was increased by adding the SH oxidant diamide (1 mM), by incubating in a sodium-free (n-methylglucamine) solution or by increasing external calcium from 1 to 10 mM. A 12 hours incubation in diamide produced a ten-fold increase in 45Ca uptake into the lens which was accompanied by a ten-fold increase in internal resistance. Incubation in Na-free solution or in 10 mM Ca2+ both produced a 5-fold increase in 45Ca content, while the increase in internal resistance was five and six fold respectively. This uncoupling was prevented in the diamide and Na-free treated lenses by omitting Ca2+ from the incubation medium. Fiber cell uncoupling was noticed in each of these experimental conditions after approximately 5 hours incubation, and good recovery was obtained in the high calcium solution if the stress was removed. The calmodulin antagonists calmidazolium (3 microM) and W7 (100 microM) both prevented uncoupling in the high calcium solution, provided there was a 2 hours preincubation period in calcium-free solution containing antagonist before the stress was applied. These data indicate that lens fiber cell communication is required by Ca2+ and calmodulin.

Self-assessment of the quality of vision: association of questionnaire score with objective clinical tests.

PURPOSE: To investigate the association of a quality of life-visual function questionnaire with an objective clinical test of visual function. METHODS: We have developed a questionnaire to assess self-reported visual satisfaction in ophthalmic patients suffering from chronic eye conditions causing visual impairment. The questionnaire was administered to 120 patients suffering from age-related cataract, chronic open angle glaucoma, age-related macular degeneration, branch retinal vein occlusion, and presbyopia or minor refractive defects. All the participants also underwent determination of visual acuity, contrast sensitivity, glare, and visual field. RESULTS: The questionnaire has a good reproducibility, a high internal consistency, and is able to discriminate between the different groups of patients. The total questionnaire score is significantly associated with the results of all visual function tests with the exception of glare. When entered into a multiple linear regression model, near visual acuity and contrast sensitivity are still considerably associated with the total questionnaire score. The psychological attitude of the patient towards his/her health problem is also associated with the total average score. CONCLUSIONS: Overall, the model explains 49% of the variance in the average questionnaire score.

An independent evaluation of the Age-Related Eye Disease Study (AREDS) cataract grading system.

PURPOSE: To assess intra- and interobserver reproducibility of the Age-Related Eye Disease Study (AREDS) system for grading lens opacities and to provide data on its capacity to reliably detect changes in lens status. METHODS: Independent and replicate grading of 40 sets of lens photographs (one slit-lamp and two retroillumination photographs) were performed by three experienced observers. Patients were participants in the Collaborative Italian-American Clinical Trial of Nutritional Supplements which is testing the effect of a mineral-multivitamin supplement on age-related cataract (CTNS). Scatterplots and intraclass correlation were used to assess measurement error. RESULTS: Analysis revealed good intra- and interobserver reproducibility of the system. Greatest intraobserver measurement error showed 100% of pairs within 10% areal difference for cortical cataract, 97.5% within 15% areal difference for posterior subcapsular cataract, and 100% within 1 density unit difference for nuclear opacity. Greatest interobserver measurement error showed 95% of pairs within 10% areal difference for cortical cataract, 97.5% within 15% areal difference for posterior subcapsular cataract, and 97.5% within 1.5 density unit difference for nuclear opacity. CONCLUSIONS: The AREDS lens opacities grading system appears to be sufficiently reliable to detect changes of at least 10% areal involvement for cortical, 15% areal involvement for posterior subcapsular, and 1.0 units for nuclear opacities. It therefore seems sufficiently sensitive to adequately monitor progression of lens opacities in a longitudinal study of patients with early cataract. Its applicability in a population with advanced or complex mixed opacities must await further testing.