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[Dependence of lipid peroxidation on pigmentation of the porcine iris].

PURPOSE: Melanin has been shown to act as antioxidant in lipid peroxidation studies. We have now investigated lipid peroxidation in dependence on stromal pigmentation in isolated porcine irises. METHODS: The same number of lightly pigmented and heavily pigmented porcine irises (visual selection) were homogenized in buffer (50 mmol/l Na2HPO4, 50 mmol/l NaH2PO4 and 4 mmol/l sodium azide; 1:20 w/v). 500 microliters homogenate were incubated at 37 degrees C for 5, 10, 20 and 40 min in absence and presence of Fe2+ as inducer of lipid peroxidation. Lipid peroxidation was assayed by the thiobarbituric acid (TBA) test. Results are expressed as nmol of TBA reactive material produced (TBAR) per mg protein. Fe2+ concentration of the supernatant was determined spectrophotometrically with phenanthroline. RESULTS: 70 mumol/l, 180 mumol/l and 360 mumol/l Fe2+ induced lipid peroxidation. A plateau region was reached after 20 min. Lipid peroxidation differed in dependence on stromal pigmentation in porcine irises by a factor of 2.8. 180 mumol/l Fe2+ induced 1.373 +/- 0.138 nmol TBAR/mg protein in lightly pigmented irises compared to 0.491 +/- 0.125 nmol TBAR/mg protein in heavily pigmented irises after 10 min incubation (p < 0.0001, n = 4). On the other hand, the content of Fe2+ in the supernatant was the same within error. CONCLUSIONS: There was a stronger induction of lipid peroxidation in lightly pigmented porcine irises compared to heavily pigmented porcine irises. This effect may be related to the difference in stromal melanin content and its antioxidant activity.

Animals↗

Unoprostone isopropyl ester darkens iris color in pigmented rabbits with sympathetic denervation.

PURPOSE: Unoprostone isopropyl ester (unoprostone) -induced iris color darkening was evaluated in a rabbit model using a cyclooxygenase inhibitor, an alpha(1)-adrenergic antagonist, and sympathetic denervation. MATERIALS AND METHODS: Dutch-belted rabbits were divided into five groups based on type of surgery and eyedrop treatment to both eyes: (1) sham surgery (n = 7); (2) bilateral superior cervical ganglionectomy (SCGx, n = 7); (3) SCGx plus flurbiprofen 0.03% (n = 7); (4) SCGx plus thymoxamine 0.5% (n = 6); and (5) SCGx plus flurbiprofen and thymoxamine (n = 6). All rabbits were treated with unoprostone 0.12% to one eye and its vehicle to the contralateral eye twice daily for 43 weeks after SCGx. Periodic color photographs of paired eyes were scored for difference in eye color. Iris melanin and aqueous humor protein were measured at week 43. RESULTS: Twenty-three of the 26 rabbits with bilateral SCGx and unilateral unoprostone treatment demonstrated a darker iris color on the unoprostone-treated side. The average scores (demonstrating difference in iris color) comparing photographs of treated versus control eyes in the four SCGx groups were higher than those in the sham surgery group (P < 0.03), and higher than at week 0 (P < 0.001). The group pretreated with flurbiprofen and thymoxamine had the highest score of all groups. The aqueous humor protein in unoprostone-treated eyes was higher (P < or = 0.0001) than in vehicle-treated eyes. The melanin content of irides of the denervated groups was higher (P < or = 0.01) in unoprostone-treated than in vehicle-treated eyes. CONCLUSION: Unoprostone produced iris color darkening in pigmented rabbit eyes with sympathetic denervation. Pretreatment with flurbiprofen and thymoxamine appeared to enhance this effect but this was not statistically demonstrated by the study.

Adrenergic alpha-Antagonists↗

Vortex keratopathy of the corneal graft.

We examined the surface epithelium of 30 eyes with color specular microscopy at 3-month intervals 1-18 months following penetrating keratoplasty. At least three examinations were performed on most eyes. We found a vortex keratopathy to be present in 70% of these eyes. In nine eyes, this vortex pattern was present at every examination, and in 12 eyes, it was present at some examinations and not at others. There was no vortex pattern seen at any examination in nine eyes. We also found alterations in the normal epithelial morphology around sutures and at the wound margin, with palisading parallel to the sutures and perpendicular to the wound edge. This altered morphology was also demonstrated by digitization of cells for shape and size, which showed significant differences compared with normals. We conclude that the morphology of the epithelial cells on the surface of the cornea remains abnormal for up to 18 months following penetrating keratoplasty.

Cell Movement↗

Neuronal activity in macaque SEF and ACC during performance of tasks involving conflict.

It has been suggested on the basis of previous studies involving functional MRI (fMRI) and single-neuron recording that neurons of the supplementary eye field (SEF) and anterior cingulate cortex (ACC) monitor conflict. To test this idea, we carried out microelectrode recording in monkeys performing a color-conditional eye movement task in which red and green cues instructed leftward and rightward saccades, respectively. In a variant inducing conflict by spatial incompatibility, the cue was presented either at the location of the target (no conflict) or opposite the location of the target (conflict). In a variant inducing conflict by reversal, the foveal cue either remained one color (no conflict) or reversed color after 100 ms (conflict), with the monkey required to follow the instruction conveyed by the second color. In both tasks, conflict was evident in behavioral measures (reduced percent correct and slowed reaction time) and in physiological measures (reduced strength of directional activity among direction-selective neurons). In the SEF, there was a tendency for neurons to fire more strongly on trials involving conflict, but this effect took the form of modulation of task-related activity among direction-selective neurons, not of a pure conflict-monitoring signal. In the ACC, there was no conflict-related enhancement. These results are incompatible with the idea that the SEF and ACC contain populations of neurons specialized for monitoring conflict.

Action Potentials↗

Analysis of diode, argon, and Nd: YAG peripheral iridectomy in cadaver eyes.

We compared semiconductor diode, neodymium (Nd): YAG and argon laser iridectomies performed in 14 cadaver eyes using Miyake videographic, histologic and scanning electron analysis. We found that the diode laser created an iridectomy in < 10 applications in blue, hazel and brown irides using 1000-1200 mW over 0.5 seconds. In hazel and blue irides the argon laser created iridectomies at parameters similar to the diode, but required multiple low energy burns in brown irides. Scanning electron microscopy showed pigment epithelial loss with the diode and Nd:YAG laser, and heaped up epithelium at the iridectomy site with the argon laser. Nd:YAG laser had the greatest iridectomy size. This study suggests a possible clinical advantage of diode versus argon laser iridectomies in reduced applications in brown irides and in preventing postoperative pigment occlusion. However, diode laser demonstrated no definite advantage over Nd:YAG laser.

Cadaver↗

Zinc uptake and storage: the role of fundus pigmentation.

BACKGROUND: Age-related macular degeneration (AMD) is associated with lower melanin pigmentation and is more prevalent among the elderly Caucasian population than among Africans. A correlation between light iris colour, fundus pigmentation and the incidence of AMD is reported. Moreover, melanin represents the main storage of zinc in the eye. Zinc enhances antioxidant capacity through its function as a cofactor of important enzymes or by influencing gene expression of regulatory elements in the eye. In this study, we investigated the uptake and storage of zinc in the human choroid/retinal pigment epithelium (RPE) complexes in dependence on the fundus pigmentation as judged by the iris colour. MATERIAL AND METHODS: Choroid/RPE complexes of blue and brown human eyes were used. Tissues without any substitution served as controls. Specimens from choroid/RPE complexes were incubated with 100 microM zinc chloride for 24 h. After incubation, pieces of the complexes were stored to investigate the uptake of zinc. The rest of the tissues were kept for 3 and 7 days in culture medium (DMEM) for storage examination. The concentration of zinc was measured by inductively coupled plasma mass spectrometry. RESULTS: After 24 h of zinc treatment the concentration of zinc in the choroid/RPE complexes of blue eyes was not significantly increased. The concentration of zinc in highly pigmented tissues (brown eyes) was increased by the factor 5.1 after 24 h and remained at high levels after 3 days (factor 4.4) and 7 days (factor 2.8). CONCLUSIONS: Zinc uptake in the choroid/RPE complex correlates to the iris colour. Alterations of the degree of iris pigmentation result in differences of zinc uptake and storage in the choroids. A potential protective role of zinc may be more prominent in dark- than in light-coloured eyes.

Adult↗

Ku86 deficiency leads to reduced intrachromosomal homologous recombination in vivo in mice.

Ku70 and Ku86 together with DNA-PKcs form the DNA-dependent protein kinase (DNA-PK) complex that is involved in DNA double-strand break repair by nonhomologous end joining. We investigated the effect of Ku86 mutation on intrachromosomal homologous recombination (HR) resulting in deletions in vivo in mice. We quantified such deletion events using a phenotypic pigmentation assay. Deletion of one copy of a 70 kb DNA duplication in the pink-eyed unstable (pun) allele results in reversion to the wildtype pink-eyed dilution (p) gene, allowing black pigment accumulation in cells of the retinal pigment epithelium (RPE). We found that the frequency of homologous recombination was significantly reduced in Ku86 deficient mice. Furthermore, the proliferation of cells in which recombination events occurred was reduced and developmentally delayed in the Ku86 deficient mice. These data indicate a role for Ku86 directly or indirectly in homologous recombination in vivo.

Animals↗

Protection from light damage by ocular pigmentation: analysis using experimental chimeras and translocation mice.

Experimental mouse chimeras and Cattanach's translocation mice, both with alternating pigmented and non-pigmented cells of the retinal pigment epithelium (RPE), have been exposed to various periods of constant fluorescent light. Photoreceptor degeneration was always more severe in the central retina than in the peripheral retina, and it was independent of the pigmentation phenotype of the immediately overlying RPE. The findings suggest that although RPE melanosomes lower total retinal irradiance by absorption of light, they do not provide direct protection from light damage for the immediately underlying photoreceptor cells by a biochemical mechanism as previously suggested.

Animals↗

Mosaic expression of a tyrosinase fusion gene in albino mice yields a heritable striped coat color pattern in transgenic homozygotes.

Genetically albino mouse eggs were injected with an inducible transgene comprising the wild-type tyrosinase (monophenol, L-dopa: oxygen oxidoreductase, EC 1.14.18.1) cDNA and the metallothionein gene promoter in the expectation that the transgene would be expressed to different extents in the various developing pigment cell clones of at least some individuals, thereby leading to patterned coats. This proved to be the case. Five transgenic mice had lightly pigmented patterns of transverse stripes visualizing melanoblast proliferation and migration dorsoventrally on each side. Similar patterns have been seen in genetically mosaic mouse models produced from conjoined blastomeres of different color genotypes and in many naturally patterned genotypes of mice. Four of the transgenics had heritable patterns and autosomal transgene integration. Their homozygous descendants were darker than hemizygotes and transmitted the basic pattern through many generations. Eyes were also pigmented, with clonal patches of melanized cells in the retinal pigment epithelium. The skin was dark due to many pigmented dermal melanocytes, whereas relatively few were in the hair follicles. This "inversion" is attributable to precocious maturation and migratory arrest of many melanoblasts during passage through the dermis en route to the hair bulbs. Patterning in these mice is considered in light of the view, previously proposed, that phenotypically different clones, or phenoclones, may exist in virtually all mammalian cell types and that many genes may be associated with cis-acting control regions causing variations in their expression that are mitotically perpetuated. We point out that mosaic expression has many implications for development as well as neoplasia. In the latter case, the potential for tumor susceptibility may be affected by clonal variation without further gene mutations or deletions. Thus, mice with variegating transgenes can provide molecular access to gene control mechanisms and to their consequences in development and disease.

Albinism↗

Effect of the barring gene on eye pigmentation in the fowl.

Pigment cells of the iris, pecten, retinal pigment epithelium, and choroid of the wild-type jungle fowl (JF) and the barred Plymouth rock (BPR) breeds of adult chickens were studied at both light and electron microscopic levels. BPR choroidal tissues had 2.8 times fewer melanophores than the JF choroid, and BPR melanophores also contained 2.4 times fewer melanosomes, which tended to clump together in variously sized clusters. The melanosomes were often irregular in shape, smaller in diameter, and less mature (stage III) than those granules in the JF. The retinal pigment epithelium of both JF and BPR breeds contained a single epithelial layer of columnar cells. Rod-shaped melanosomes were present in the more apical regions of this cell type in both breeds. Both JF and BPR irides contained a multilayered posterior pigmented epithelium of columnar shaped cells that were densely filled with large spherical granules. Intercellular spaces with interdigitating cytoplasmic projections were present between pigment cells of both breeds. The pecten melanophores of both breeds were dendritic with melanosomes that were larger and fewer in numbers than those pigment cells of the iris and choroid. Intercellular spaces were present between cells in both breeds, with numerous villous-like pigment cell extensions. Choroid melanophores contained very little, if any, acid phosphatase activity. Approximately one-half of the retinal pigment epithelial cells observed contained small amounts of diffuse acid phosphatase activity in both breeds. The iris and pecten melanophores of both breeds contained profuse acid phosphatase activity scattered throughout their cytoplasms. Sparse tyrosinase activity was seen in iris and pecten pigment cells, whereas no tyrosine activity was observed in choroid melanophores or in retinal pigment epithelial cells in the two breeds, indicating that little new melanogenesis occurs in adult pigmented eye tissues. The results show that the barring gene reduces the number and melanin content of the choroidal melanophores in homozygous male BPR chickens as compared to the wild-type JF chickens. Whether this gene prevents the initial migration of embryonic neural crest cells (future melanophores) to the choroid or whether some of the choroidal melanophores prematurely degenerate in the embryo of young birds is yet to be determined. If the latter is the case, this choroid system may serve as a model for a genetic hypomelanotic disease such as vitiligo.

Acid Phosphatase↗

[Vascular architectonics of eye and orbital space in color reflection of doppler spectrum energy].

Super-sharp gray scale image and highly sensitive digital wide-band Doppler, three-dimensional visualization of vessels in the pulsed energy Doppler mode were used in examination of the posterior segment of the eye, optic nerve, and orbital space. Twelve healthy volunteers (24 eyes) aged 25-45 years were examined. The architectonics of the optic disk and the blood filling of the arterial Zinn-Haller circle, reticular platelet, choroid, posterior long and posterior short ciliary arteries were studied. Vascular architectonics of the retrobulbar space was studied by the three-dimensional reconstruction in the energy Doppler mode and the borders of the vascular plexus round the optic nerve were defined.

Adult↗

A standardized approach for iris color determination.

Latanoprost, the phenyl-substituted prostaglandin F2alpha, has been found to be an effective agent for glaucoma therapy. This prostaglandin derivative exerts ocular hypotensive activity but is also associated with an untoward side effect, namely iris color changes. Latanoprost provoked iris color changes in cynomolgus monkeys and in multicenter clinical trials. Until now photographs were taken and compared with color plates to document these changes. The disadvantage of this method is obvious, i.e., the color luminance varies between measurements due to changes in the developer. Furthermore, subjective comparison of color changes relative to color plates rendered judgment subject to impression and opinion rather than to objective data. Therefore, a computerized method using a 3-CCD video camera attached to a slit lamp was developed. The signals were transferred to a computer and a single frame, which was "frozen" by means of a "grabber card." Camera and the computer had previously been calibrated and color plates were measured to check the standard conditions. They were evaluated by a software program displaying average color (as red, green, and blue values) of the selected area. This method provides a fast and accurate way to quantify color changes in the iris of both experimental animals and clinical trials.

Animals↗

Choroidal neovascularization in black patients.

OBJECTIVE: To characterize choroidal neovascularization (CNV) in black patients examined at a retinal disease referral center. DESIGN: Retrospective review of the medical records of all patients diagnosed as having CNV to identify black patients with CNV. SETTING: Single tertiary retinal referral center that included four ophthalmologists. PATIENTS: All patients diagnosed as having CNV between April 1990 and October 1992. MAIN OUTCOME MEASURES: Prevalence, demographic information, fundus photographic and fluorescein angiographic characteristics, natural history, and response to laser photocoagulation of CNV in black patients. RESULTS: Black patients comprise 15% of all patients seen at this center. Of 1725 patients identified as having CNV who were seen at the center during a 2.5-year period, only 25 were black (1.4%). In these patients, CNV was associated with a variety of retinal diseases, the most frequent being age-related macular degeneration. The average age of the study group was 54 years, women outnumbered men 2:1, and 13 of the patients developed bilateral lesions. Twelve of the 38 lesions were extrafoveal on presentation, and five of these were peripapillary. In the laser-treated eyes, recurrence of CNV was frequent and associated with visual loss. CONCLUSIONS: Choroidal neovascularization seems to be rare in blacks among a retinal disease referral center population. The overall presentation, natural history, and response to laser treatment seems to be similar to that of white patients. No feature of CNV in black patients was identified that would suggest that results of randomized clinical trials of laser photocoagulation for CNV are not valid for these patients.

Baltimore↗