Evaluation of the G protein coupled receptor-75 (GPR75) in age related macular degeneration

Journal Articl

Reflection-based imaging of macular pigment distributions in infants and children

pre-printWe have developed a reflection-based capability of the RetCam® platform, an FDA-cleared pediatric retinal-imaging instrument, for the purpose of measuring macular pigment levels as well as their spatial distributions in infants and children. Our modifications include narrow-band blue-wavelength excitation of the macular pigment absorption in combination with spectrally selective blue-wavelength readout of the reflection signals received by the instrument's CCD detector array. Furthermore, an algorithm is developed that allows the computation of optical density maps for the macular pigment relative to peripheral retinal areas. This made it possible for the first time to directly measure macular pigment levels and their spatial features in the developing human retina. In contrast to adults, infants with measurable pigment levels had almost exclusively a narrow, circularly symmetric, pigment distribution. The described methodology holds promise for future investigations into the role of macular pigment in the developing human retina and the effect of dietary interventions in diseases resulting from a lack of normal carotenoid levels

HPLC measurement of ocular carotenoid levels in human donor eyes in the lutein supplementation era.

Journal ArticlePURPOSE: A substantial proportion of the population at risk for visual loss from age-related macular degeneration consumes supplements containing high doses of lutein, but clinical studies to date have shown only modest and variable increases in macular carotenoid pigments in response to supplementation. To determine whether lutein supplementation can indeed alter ocular carotenoid levels, the authors chemically measured levels of lutein, zeaxanthin, and their metabolites in the macula, peripheral retina, and lens of 228 eyes from 147 human donors and correlated these results with retrospective supplement histories from families of selected members of the study population. METHODS: Lenses and circular punches of macula (4-mm diameter) and equatorial peripheral retina (8-mm diameter) were dissected from donor eyes free of ocular disease procured from the local eye bank. The amounts of lutein, zeaxanthin, meso-zeaxanthin, and 3'-oxolutein were determined by HPLC with photodiode array and mass spectral detection. RESULTS: Eighteen percent of eyes from donors age 48 and older had unusually high levels (66.3 +/- 15.1 ng) of macular carotenoids that were three times the rest of the older population's mean level (23.0 + 12.1 ng; P < 0.001). Carotenoid levels in these outliers were also unusually high in the lens and in the peripheral retina. Similar outliers were not present in donors younger than 48. Most of these outliers regularly consumed high-dose lutein supplements before death. Lutein supplementation was uncommon in older donors whose macular carotenoids were in the normal range. CONCLUSIONS: The presence of unusually high levels of macular carotenoids in older donors who were regularly consuming high-dose lutein supplements supports the hypothesis that long-term lutein supplementation can raise levels of macular pigment. Elevated carotenoid levels in the peripheral retina and lens in these same donors could have important implications for understanding why some clinical methods of macular pigment measurement have had difficulty detecting robust and consistent responses in carotenoid supplementation trials

Skin carotenoid status measured by resonance Raman spectroscopy as a biomarker of fruit and vegetable intake in preschool children

pre-printBackground/Objective: Dietary assessment in children is difficult, suggesting a need to develop more objective biomarkers of intake. Resonance Raman spectroscopy (RRS) is a noninvasive, validated method of measuring carotenoid status in skin as a biomarker of fruit/vegetable intake. The purpose of this study was to examine the feasibility of using RRS in preschool children, including describing the inter-individual variability in skin carotenoid status and to identify factors associated with the biomarker in this population. Subjects/Methods: We conducted a cross-sectional study of 381 economically-disadvantaged preschoolers in urban centers in Connecticut (U.S.). 85.5% were black non-Hispanic or Hispanic/Latino, and 14.1% were obese and 16.9% were overweight by age- and sex-specific BMI percentiles. Children had their skin carotenoid status assessed by RRS in the palm of the hand. Fruit/vegetable consumption was assessed by a brief parent/guardian-completed food frequency screener and a liking survey. Results: We observed inter-individual variation in RRS values that was nearly normally distributed. In multiple regression analysis, higher carotenoid status, measured by RRS, was positively associated with fruit/vegetable consumption (p=0.02) and fruit/vegetable preference (p<0.01). Lower carotenoid status was observed among younger children, those participating in the U.S. Supplemental Nutrition Assistance Program (SNAP), and those with greater adiposity (p<0.05 for all). Conclusions: We observed wide variability in skin carotenoid status in a population of young children, as assessed by RRS. Parent-reported fruit/vegetable intake and several demographic factors were significantly associated with RRS-measured skin carotenoid status. We recommend further development of this biomarker in children, including evaluating response to controlled interventions

Identification of lutein and zeaxanthin oxidation products in human and monkey retinas

Journal ArticlePURPOSE: To characterize fully all the major and minor carotenoids and their metabolites in human retina and probe for the presence of the oxidative metabolites of lutein and zeaxanthin. METHODS: Carotenoids of a composite of 58 pairs of human retinas and a monkey retina were elucidated by comparing their high-performance liquid chromatography (HPLC)-ultraviolet/visible absorption spectrophotometry (UV/Vis)-mass spectrometry (MS) profile with those of authentic standards prepared by organic synthesis. RESULTS: In addition to lutein and zeaxanthin, several oxidation products of these compounds were present in the extracts from human retina. A major carotenoid resulting from direct oxidation of lutein was identified as 3-hydroxy-beta, epsilon-caroten-3'-one. Minor carotenoids were identified as: 3'-epilutein, epsilon,epsilon-carotene-3,3'-diol, epsilon,epsilon-carotene-3,3'-dione, 3'-hydroxy-epsilon,epsilon-caroten-3-one, and 2,6-cyclolycopene-1,5-diol. Several of the geometric isomers of lutein and zeaxanthin were also detected at low concentrations. These were as follows: 9-cis-lutein, 9'-cislutein, 13-cis-lutein, 13'-cis-lutein, 9-cis-zeaxanthin, and 13-cis-zeaxanthin. Similar results were also obtained from HPLC analysis of a freshly dissected monkey retina. CONCLUSIONS: Lutein, zeaxanthin, 3'-epilutein, and 3-hydroxy-beta,epsilon-caroten-3'-one in human retina may be interconverted through a series of oxidation-reduction reactions similar to our earlier proposed metabolic transformation of these compounds in humans. The presence of the direct oxidation product of lutein and 3'-epilutein (metabolite of lutein and zeaxanthin) in human retina suggests that lutein and zeaxanthin may act as antioxidants to protect the macula against short-wavelength visible light. The proposed oxidative-reductive pathways for lutein and zeaxanthin in human retina, may therefore play an important role in prevention of age-related macular degeneration and cataracts

Macular pigment Raman detector for clinical applications

Journal ArticleClinical studies of carotenoid macular pigments (MP) have been limited by the lack of noninvasive, objective instruments. We introduce a novel noninvasive optical instrument, an MP Raman detector, for assessment of the carotenoid status of the human retina in vivo. The instrument uses resonant excitation of carotenoid molecules in the visible wavelength range, and quantitatively measures the highly specific Raman signals that originate from the single- and double-bond stretch vibrations of the pi-conjugated carotenoid molecule's carbon backbone. The instrument is a robust, compact device and suitable for routine measurements of MP concentrations in a clinical setting. We characterized and tested the instrument in clinical studies of human subjects to validate its function and to begin to establish its role as a possible screening test for macular pathologies. We also show that the MP Raman spectroscopy technology has potential as a novel, highly specific method for rapid screening of carotenoid antioxidant levels in large populations at risk for vision loss from age-related macular degeneration, the leading cause of blindness of the elderly in the developed world

Transformations of selected carotenoids in plasma, liver, and ocular tissues of humans and in nonprimate animal models

Journal ArticlePURPOSE: To determine the stereochemistry of carotenoids in human ocular tissues in comparison with plasma and liver and to elucidate the possible transformations of dietary (3R,3'R,6'R)-lutein and (3R,3'R)-zeaxanthin in the eye. Similarly, to characterize the carotenoid profiles in the eye tissues, plasma, and liver of quails and frogs to determine whether these can serve as appropriate nonprimate animal models for metabolic studies. METHODS: Configurational isomers of carotenoids and their nondietary by-products from pooled human plasma, liver, retinal pigment epithelium (RPE-choroid), ciliary body, iris, and lens were characterized and quantified by high-performance liquid chromatography (HPLC) on a chiral column. Carotenoids and their nondietary by-products in pooled extracts from quail and frog plasma, liver, retina, RPE-choroid, iris, and lens were similarly characterized and quantified. RESULTS: (3R,3'R,6'R)-lutein, (3R,3'R)-zeaxanthin, (3R,3'S; meso)-zeaxanthin, (3R,3'S,6'R)-lutein (3'-epilutein), 3-hydroxy-beta, epsilon -carotene-3'-one, and 5Z- and all-E-lycopene were detected in nearly all human ocular tissues examined. (3R,3'S; meso)-zeaxanthin was not detected in the human plasma and liver but was present in human macula, retina, and RPE-choroid. (3S,3'S)-zeaxanthin was detected in human macula in minute quantities. The carotenoid profiles in quail and frog ocular tissues were somewhat similar to those in humans, with the exception that lycopene was absent. Frog retina, plasma, and liver revealed the presence of (3S,3'S)-zeaxanthin. CONCLUSIONS: The most likely transformations of carotenoids in the human eye involve a series of oxidation-reduction and double-bond isomerization reactions. Quail and frog appear to possess the appropriate enzymes for conversion of dietary (3R,3'R,6'R)-lutein and (3R,3'R)-zeaxanthin to the same nondietary by-products observed in humans and thus may serve as excellent nonprimate animal models for metabolic studies

Diverse macular dystrophy phenotype caused by a novel complex mutation in the ELOVL4 gene

Journal ArticlePURPOSE: A 5-bp deletion in ELOVL4, a photoreceptor-specific gene, has been associated with autosomal dominant (ad) macular dystrophy phenotypes in five related families, in which phenotypes range from Stargardt-like macular dystrophy (STGD3; Mendelian Inheritance in Man 600110) to pattern dystrophy. This has been the only mutation identified in ELOVL4 to date, which is associated with macular dystrophy phenotypes. In the current study, the potential involvement was investigated of an ELOVL4 gene variation in adSTGD-like and other macular dystrophy phenotypes segregating in a large unrelated pedigree from Utah (K4175). METHODS: The entire open reading frame of the ELOVL4 gene was analyzed by direct sequencing in a proband from the K4175 family. The combination of denaturing high-performance liquid chromatography (DHPLC) analysis and direct sequencing of all available family members was used to further assess segregation of identified ELOVL4 variants in the pedigree. RESULTS: A complex mutation, two 1-bp deletions separated by four nucleotides, was detected in all affected members of the family. The mutation results in a frameshift and the truncation of the ELOVL4 protein, similar to the effect of the previously described 5-bp deletion. CONCLUSIONS: The discovery of a second mutation in the ELOVL4 gene segregating with macular dystrophy phenotypes confirms the role of this gene in a subset of dominant macular dystrophies with a wide range of clinical expressions and suggests a role for modifying genes and/or environmental factors in the disease process

Genotype-phenotype analysis of ABCR variants in macular degeneration probands and siblings

Journal ArticlePURPOSE: Single-copy variants of the autosomal recessive Stargardt disease (STGD1) gene ABCR (ABCA4) have been shown to confer enhanced susceptibility to age-related macular degeneration (AMD). To investigate the role of ABCR alleles in AMD further, genotype-phenotype analysis was performed on siblings of patients with AMD who had known ABCR variants. This genetically related population provides a cohort of subjects with similar age and ethnic background for genotype-phenotype comparison to the original probands. METHODS: All available siblings of 26 probands carrying probable disease-associated ABCR variants were examined clinically. Blood samples were collected from these siblings for genotype analysis to search for the ABCR variant alleles corresponding to the isofamilial proband. RESULTS: Nineteen of 33 siblings from 15 families carried the respective proband's variant ABCR allele. Some families exhibited concordance of ABCR alleles with macular degeneration phenotype, but others did not. Exudative AMD was uncommon among both probands and siblings. CONCLUSIONS: Although population studies have indicated that some ABCR variant alleles may enhance susceptibility to AMD, investigation of the extent of ABCR involvement by kindred analysis is complicated by a plethora of environmental and other hereditary factors not investigated in the current study that may also play important roles

Nonmydriatic fluorescence-based quantitative imaging of human macular pigment distributions.

Journal ArticleWe have developed a CCD-camera-based nonmydriatic instrument that detects fluorescence from retinal lipofuscin chromophores ("autofluorescence") as a means to indirectly quantify and spatially image the distribution of macular pigment (MP). The lipofuscin fluorescence intensity is reduced at all retinal locations containing MP, since MP has a competing absorption in the blue-green wavelength region. Projecting a large diameter, 488 nm excitation spot onto the retina, centered on the fovea, but extending into the macular periphery, and comparing lipofuscin fluorescence intensities outside and inside the foveal area, it is possible to spatially map out the distribution of MP. Spectrally selective detection of the lipofuscin fluorescence reveals an important wavelength dependence of the obtainable image contrast and deduced MP optical density levels, showing that it is important to block out interfering fluorescence contributions in the detection setup originating from ocular media such as the lens. Measuring 70 healthy human volunteer subjects with no ocular pathologies, we find widely varying spatial extent of MP, distinctly differing distribution patterns of MP, and strongly differing absolute MP levels among individuals. Our population study suggests that MP imaging based on lipofuscin fluorescence is useful as a relatively simple, objective, and quantitative noninvasive optical technique suitable to rapidly screen MP levels and distributions in healthy humans with undilated pupils