Dietary Nutrient Intake and Progression to Late Age-Related Macular Degeneration in the Age-Related Eye Disease Studies 1 and 2.

PURPOSE: To analyze associations between the dietary intake of multiple nutrients and risk of progression to late age-related macular degeneration (AMD), its subtypes, and large drusen. DESIGN: Post hoc analysis of 2 controlled clinical trial cohorts: Age-Related Eye Disease Study (AREDS) and AREDS2. PARTICIPANTS: Eyes with no late AMD at baseline among AREDS participants (n = 4504) and AREDS2 participants (n = 3738) totaled 14 135 eyes. Mean age was 71.0 years (standard deviation, 6.7 years), and 56.5% of patients were women. METHODS: Fundus photographs were collected at annual study visits and graded centrally for late AMD. Dietary intake of multiple nutrients was calculated from food frequency questionnaires. MAIN OUTCOME MEASURES: Progression to late AMD, geographic atrophy (GA), neovascular AMD, and (separate analyses) large drusen. RESULTS: Over median follow-up of 10.2 years, of the 14 135 eyes, 32.7% progressed to late AMD. For 9 nutrients, intake quintiles 4 or 5 (vs. 1) were associated significantly (P ≤ 0.0005) with decreased risk of late AMD: vitamin A, vitamin B6, vitamin C, folate, ß-carotene, lutein and zeaxanthin, magnesium, copper, and alcohol. For 3 nutrients, quintiles 4 or 5 were associated significantly with increased risk: saturated fatty acid, monounsaturated fatty acid, and oleic acid. Similar results were observed for GA. Regarding neovascular AMD, 9 nutrients were associated nominally with decreased risk-vitamin A, vitamin B6, ß-carotene, lutein and zeaxanthin, magnesium, copper, docosahexaenoic acid, omega-3 fatty acid, and alcohol-and 3 nutrients were associated with increased risk-saturated fatty acid, monounsaturated fatty acid, and oleic acid. In separate analyses (n = 5399 eyes of 3164 AREDS participants), 12 nutrients were associated nominally with decreased risk of large drusen. CONCLUSIONS: Higher dietary intake of multiple nutrients, including minerals, vitamins, and carotenoids, is associated with decreased risk of progression to late AMD. These associations are stronger for GA than for neovascular AMD. The same nutrients also tend to show protective associations against large drusen development. Strong genetic interactions exist for some nutrient-genotype combinations, particularly omega-3 fatty acids and CFH. These data may justify further research into underlying mechanisms and randomized trials of supplementation.

Accelerated Development of Rod Photoreceptors in Retinal Organoids Derived from Human Pluripotent Stem Cells by Supplementation with 9-cis Retinal.

Human pluripotent stem cells (PSCs) can be differentiated into retinal organoids with proper neural layer organization, yet the protocols are technically challenging and time consuming. We have modified a widely used differentiation protocol by switching all-trans retinoic acid with 9-cis retinal to accelerate photoreceptor differentiation and improve morphogenesis. In this report, we provide a detailed and improved protocol to generate retinal organoids from human pluripotent stem cells. For complete details on the use and execution of this protocol, please refer to Kaya et al. (2019).

No CFH or ARMS2 Interaction with Omega-3 Fatty Acids, Low versus High Zinc, or ß-Carotene versus Lutein and Zeaxanthin on Progression of Age-Related Macular Degeneration in the Age-Related Eye Disease Study 2: Age-Related Eye Disease Study 2 Report No. 18.

PURPOSE: To assess whether genotypes at 2 major loci associated with age-related macular degeneration (AMD), complement factor H (CFH), or age-related maculopathy susceptibility 2 (ARMS2), modify the response to oral nutrients for the treatment of AMD in the Age-Related Eye Disease Study 2 (AREDS2). DESIGN: Post hoc analysis of a randomized trial. PARTICIPANTS: White AREDS2 participants. METHODS: AREDS2 participants (n = 4203) with bilateral large drusen or late AMD in 1 eye were assigned randomly to lutein and zeaxanthin, omega-3 fatty acids, both, or placebo, and most also received the AREDS supplements. A secondary randomization assessed modified AREDS supplements in 4 treatment arms: lower zinc dosage, omission of ß-carotene, both, or no modification. To evaluate the progression to late AMD, fundus photographs were obtained at baseline and annual study visits, and history of treatment for late AMD was obtained at study visits and 6-month interim telephone calls. Participants were genotyped for the single-nucleotide polymorphisms rs1061170 in CFH and rs10490924 in ARMS2. Bivariate frailty models using both eyes were conducted, including a gene-supplement interaction term and adjusting for age, gender, level of education, and smoking status. The main treatment effects, as well as the direct comparison between lutein plus zeaxanthin and ß-carotene, were assessed for genotype interaction. MAIN OUTCOME MEASURES: The interaction between genotype and the response to AREDS2 supplements regarding progression to late AMD, any geographic atrophy (GA), and neovascular AMD. RESULTS: Complete data were available for 2775 eyes without baseline late AMD (1684 participants). The participants (mean age ± standard deviation, 72.1±7.7 years; 58.5% female) were followed up for a median of 5 years. The ARMS2 risk allele was associated significantly with progression to late AMD and neovascular AMD (P = 2.40 × 10-5 and P = 0.002, respectively), but not any GA (P = 0.097). The CFH risk allele was not associated with AMD progression. Genotype did not modify significantly the response to any of the AREDS2 supplements. CONCLUSIONS: CFH and ARMS2 risk alleles do not modify the response to the AREDS2 nutrient supplements with respect to the progression to late AMD (GA and neovascular AMD).

Association of Rare Predicted Loss-of-Function Variants in Cellular Pathways with Sub-Phenotypes in Age-Related Macular Degeneration.

PURPOSE: To investigate the association of rare predicted loss-of-function (pLoF) variants within age-related macular degeneration (AMD) risk loci and AMD sub-phenotypes. DESIGN: Case-control study. PARTICIPANTS: Participants of AREDS, AREDS2, and Michigan Genomics Initiative. METHODS: Whole genome sequencing data were analyzed for rare pLoF variants (frequency <0.1%) in the regions of previously identified 52 independent risk variants known to be associated with AMD. Frequency of the rare pLoF variants in cases with intermediate or advanced AMD was compared with controls. Variants were assigned to the complement, extracellular matrix (ECM), lipid, cell survival, immune system, metabolism, or unknown/other pathway. Associations of rare pLoF variant pathways with AMD sub-phenotypes were analyzed using logistic and linear regression, and Cox proportional hazards models. MAIN OUTCOME MEASURES: Differences in rare pLoF variant pathway burden and association of rare pLoF variant pathways with sub-phenotypes within the population with AMD were evaluated. RESULTS: Rare pLoF variants were found in 298 of 1689 cases (17.6%) and 237 of 1518 controls (15.6%) (odds ratio [OR], 1.11; 95% confidence interval [CI], 0.91-1.36; P = 0.310). An enrichment of rare pLoF variants in the complement pathway in cases versus controls (OR, 2.94; 95% CI, 1.49-5.79; P = 0.002) was observed. Within cases, associations between all rare pLoF variants and choroidal neovascularization (CNV) (OR, 1.34; 95% CI, 1.04-1.73; P = 0.023), calcified drusen (OR, 1.33; 95% CI, 1.04-1.72; P = 0.025), higher scores on the AREDS Extended AMD Severity Scale (Standardized Coefficient Beta (ß)=0.346 [0.086-0.605], P = 0.009), and progression to advanced disease (hazard ratio, 1.25; 95% CI, 1.01-1.55; P = 0.042) were observed. At the pathway level, there were associations between the complement pathway and geographic atrophy (GA) (OR, 2.17; 95% CI, 1.12-4.24; P = 0.023), the complement pathway and calcified drusen (OR, 3.75; 95% CI, 1.79-7.86; P < 0.001), and the ECM pathway and more severe levels in the AREDS Extended AMD Severity Scale (ß = 0.62; 95% CI, 0.04-1.20; P = 0.035). CONCLUSIONS: Rare pLoF variants are associated with disease progression. Variants in the complement pathway modify the clinical course of AMD and increase the risk of developing specific sub-phenotypes.

A Deep Phenotype Association Study Reveals Specific Phenotype Associations with Genetic Variants in Age-related Macular Degeneration: Age-Related Eye Disease Study 2 (AREDS2) Report No. 14.

PURPOSE: Age-related macular degeneration (AMD), a multifactorial disease with variable phenotypic presentation, was associated with 52 single nucleotide polymorphisms (SNPs) at 34 loci in a genome-wide association study (GWAS). These genetic variants could modulate different biological pathways involved in AMD, contributing to phenotypic variability. To better understand the effects of these SNPs, we performed a deep phenotype association study (DeePAS) in the Age-Related Eye Disease Study 2 (AREDS2), followed by replication using AREDS participants, to identify genotype associations with AMD and non-AMD ocular and systemic phenotypes. DESIGN: Cohort study. PARTICIPANTS: AREDS and AREDS2 participants. METHODS: AREDS2 participants (discovery cohort) had detailed phenotyping for AMD; other eye conditions; cardiovascular, neurologic, gastrointestinal, and endocrine disease; cognitive function; serum nutrient levels; and others (total of 139 AMD and non-AMD phenotypes). Genotypes of the 52 GWAS SNPs were obtained. The DeePAS was performed by correlating the 52 SNPs to all phenotypes using logistic and linear regression models. Associations that reached Bonferroni-corrected statistical significance were replicated in AREDS. MAIN OUTCOME MEASURES: Genotype-phenotype associations. RESULTS: A total of 1776 AREDS2 participants had 5 years follow-up; 1435 AREDS participants had 10 years. The DeePAS revealed a significant association of the rs3750846 SNP at the ARMS2/HTRA1 locus with subretinal/sub-retinal pigment epithelial (RPE) hemorrhage related to neovascular AMD (odds ratio 1.55 [95% confidence interval 1.31-1.84], P = 2.67 × 10-7). This novel association remained significant after conditioning on participants with neovascular AMD (P = 2.42 × 10-4). Carriers of rs3750846 had poorer visual acuity during follow-up (P = 6.82 × 10-7) and were more likely to have a first-degree relative with AMD (P = 5.38 × 10-6). Two SNPs at the CFH locus, rs10922109 and rs570618, were associated with the drusen area in the Early Treatment Diabetic Retinopathy Study Report (ETDRS) grid (P = 2.29 × 10-11 and P = 3.20 × 10-9, respectively) and the center subfield (P = 1.24 × 10-9 and P = 6.68 × 10-8, respectively). SNP rs570618 was additionally associated with the presence of calcified drusen (P = 5.38 × 10-6). Except for positive family history of AMD with rs3750846, all genotype-phenotype associations were significantly replicated in AREDS. No pleiotropic associations were identified. CONCLUSIONS: The association of the SNP at the ARMS2/HTRA1 locus with subretinal/sub-RPE hemorrhage and poorer visual acuity and of SNPs at the CFH locus with drusen area may provide new insights in pathophysiological pathways underlying different stages of AMD.

A small plant with big benefits: Fenugreek (Trigonella foenum-graecum Linn.) for disease prevention and health promotion.

Plant-derived natural products have long-standing utility toward treating degenerative diseases. It is estimated that about two-thirds of world population depend on traditional medicine for primary medical needs. Fenugreek (Trigonella foenum-graecum Linn.), a short-living annual medicinal plant belonging to Fabaceae family, is used extensively in various parts of the world as herb, food, spice, and traditional medicine. Fenugreek is considered as one of the oldest medicinal plants and its health-promoting effects have been cited in Ayurveda and traditional Chinese medicine. The investigations into the chemical composition and pharmacological actions have seen a renaissance in recent years. Extensive preclinical and clinical research have outlined the pharmaceutical uses of fenugreek as antidiabetic, antihyperlipidemic, antiobesity, anticancer, anti-inflammatory, antioxidant, antifungal, antibacterial, galactogogue and for miscellaneous pharmacological effects, including improving women's health. The pharmacological actions of fenugreek are attributed to diverse array of phytoconstituents. The phytochemical analysis reveals the presence of steroids, alkaloids, saponins, polyphenols, flavonoids, lipids, carbohydrates, amino acids, and hydrocarbons. This review aims to summarize and critically analyze the current available literature to understand the potential of fenugreek for disease prevention and health improvement with special emphasis on cellular and molecular mechanisms. Current challenges and new directions of research on fenugreek are also discussed.

Efficacy of FurosapTM, a novel Trigonella foenum-graecum seed extract, in Enhancing Testosterone Level and Improving Sperm Profile in Male Volunteers.

Background: Dietary fiber rich fenugreek (Trigonella foenum-graecum) seeds have exhibited cardioprotective, hypolipidemic and other health benefits. Furosap (FS), an innovative, patented, 20% protodioscin-enriched extract was developed in our laboratory from fenugreek seeds. This study examined the free and total testosterone levels, sperm profile and morphology, sexual health, mood and mental alertness, and broad spectrum safety parameters of FS in 50 male volunteers following supplementation over a period of 12 weeks. Methods: Institutional Review Board (IRB) and other regulatory approvals were obtained for our study. This one-arm, open-labelled, multi-center study was conducted in 50 male volunteers (age: 35 to 65 years) over a period of 12 weeks to determine the efficacy of FS (500 mg/day/subject) on free and total testosterone levels, sperm profile, sperm morphology, libido and sexual health, mood and mental alertness, and broad spectrum safety parameters. Results: Free testosterone levels were improved up to 46% in 90% of the study population. 85.4% of the study population showed improvements in sperm counts. Sperm morphology improved in 14.6% of volunteers. Majority of the subjects enrolled in the study demonstrated improvements in mental alertness and mood. Furthermore, cardiovascular health and libido were significantly improved. Extensive safety parameters were evaluated which included blood chemistry data. No significant changes were observed in serum lipid function, cholesterol, triglyceride, HDL and LDL levels, hemogram (CBC), hepatotoxicity and nephrotoxicity. Conclusion: Overall, the results demonstrate that FS, enriched in 20% protodioscin, is safe and effective in attenuating testosterone levels, healthy sperm profile, mental alertness, cardiovascular health and overall performance in human subjects.

Efficacy of a Novel Fenugreek Seed Extract (Trigonella foenum-graecum, Furocyst) in Polycystic Ovary Syndrome (PCOS).

Polycystic ovary syndrome (PCOS) is one of the most prevalent hormonal disorders among women of reproductive age causing irregular menstrual cycles, excessive body or facial hair, miscarriage and infertility. The latter being a most common PCOS symptoms. Because the symptoms are seemingly unrelated to one another, PCOS is often overlooked and undiagnosed. The present study is an open label, one-arm, non-randomized, post-marketing surveillance study in 50 premenopausal women (18-45 years, BMI<42) diagnosed with PCOS using a novel Trigonella foenum-graecum seed extract (fenugreek seed extract, Furocyst, 2 capsules of 500 mg each/day) extract, enriched in approximately 40% furostanolic saponins, over a period of 90 consecutive days. The study was conducted to determine its efficacy on the reduction of ovarian volume and the number of ovarian cysts. Ethical committee approval was obtained for this study. Furocyst treatment caused significant reduction in ovary volume. Approximately 46% of study population showed reduction in cyst size, while 36% of subjects showed complete dissolution of cyst. It is important to mention that 71% of subjects reported the return of regular menstrual cycle on completion of the treatment and 12% of subjects subsequently became pregnant. Overall, 94% of patients benefitted from the regimen. Significant increases in luteinizing hormone (LH) and follicular stimulating hormone (FSH) levels were observed compared to the baseline values. Extensive blood chemistry, hematological and biochemical assays demonstrated the broad-spectrum safety. Furocyst caused significant decrease in both ovarian volume and the number of ovarian cysts. Serum ALT, BUN and CK were assessed to demonstrate the broad-spectrum safety of Furocyst. No significant adverse effects were observed. In summary, Furocyst was efficacious in ameliorating the symptoms of PCOS.

Safety and efficacy of a novel Prunus domestica extract (Sitoprin, CR002) on testosterone-induced benign prostatic hyperplasia (BPH) in male Wistar rats.

The efficacy of a novel Prunus domestica bark extract (Sitoprin, CR002) was investigated on testosterone propionate (TP)-induced benign prostatic hyperplasia (BPH) in male Wistar rats. BPH was induced by daily subcutaneous administration of TP (3.0 mg/kg) over a period of 15 days (interim sacrifice group) and for an additional 21 days (terminal sacrifice group). We evaluated the dose-dependent efficacy (0, 50, 100 and 200 mg/kg body weight/day) of CR002 and a control group against BPH, and compared with a reference standard Prunus africana extract (CR001). Extensive clinical examinations were carried out on days 1, 7, 14, 21, 28 and 35 of treatment period to determine the onset, duration and severity of clinical signs. Clinical pathology, hematology, biochemistry and histopathology were performed on days 15 and 35, prior to necropsy. Animals were fasted overnight prior to blood collection. Prostate glands and tissues were examined. On day 36, histopathology of ventral prostrate of control rats demonstrates single layer of columnar mucin secreting epithelial cells along with a lumen occupied with eosinophilic secretion. In contrast, CR002 and CR001 groups (100 and 200 mg/kg/day) exhibited no hyperplasia and proliferation of epithelial cells. Prostate histopathology of these treated groups was comparable with control rats. The hyperplasia and hypertrophy of prostrate was reduced to single-layered cell indicating the efficacy of CR002 and CR001. Overall, results demonstrate that CR002 exhibits therapeutic efficacy/activity in TP-induced BPH in rats, which is comparable to CR001.

Safety, efficacy and toxicological evaluation of a novel, patented anti-diabetic extract of Trigonella Foenum-Graecum seed extract (Fenfuro).

Safety and anti-diabetic efficacy of a novel, proprietary Trigonella foenum-graecum seed extract [novel fenugreek extract (FE), Fenfuro™, CR0010810) enriched in furostanolic saponins (>60% w/w, HPLC) were assessed. Concerning safety, we undertook studies dealing with acute oral toxicity, 28-d sub-chronic toxicity and Ames' bacterial reverse mutation assay that revealed no toxicity. Concerning efficacy, we examined beneficial effects of the extract on rats with type 2 diabetes (T2D). Male Sprague-Dawley rats received a high-fat diet for 2 weeks followed by streptozotocin (STZ, 35 mg/kg i.p.) to produce T2D. Seven days post-STZ, rats showing ≥300 mg/dl fasting plasma glucose level (PGL) were included in the study. FE (150- or 450- mg/kg p.o.) and glipizide (5 mg/kg p.o.) were administered once daily for 20 d and then twice daily for another 10 d (total 30 d). Blood samples were collected at 0, 10, 20 and 30 d of treatment and estimated for fasting plasma triglyceride (PTG), total cholesterol and insulin levels. After 30 d, FE and glipizide-treated diabetic animals were treated in combination with or without metformin (100 mg/kg) twice daily for another 10 d. FE did not influence body weight, feed and water intake. FE (150 mg/kg p.o.) reduced PTG levels in T2D rats by 22%, 24.6% and 29% at 10, 20 and 30 d of treatment, respectively, while glipizide (5 mg/kg p.o.) reduced the PTG levels by 57.4%, 46.2% and 39.4% at these time points. FE (450 mg/kg) treatment in STZ-induced diabetic rats produced significant hypoglycemic activity (approximately 31.5%) as compared to insulin (48.2% with 1 U/kg i.p.). FE (150 mg/kg p.o.) and metformin (100 mg/kg p.o.) combined produced significant reduction (20.7%) of PGL in T2D rats. No adverse effects were observed. We conclude after extensive in vitro and in vivo safety and efficacy studies that FE is safe and effective in treating T2D.

No clinically significant association between CFH and ARMS2 genotypes and response to nutritional supplements: AREDS report number 38.

OBJECTIVE: To determine whether genotypes at 2 major loci associated with late age-related macular degeneration (AMD), complement factor H (CFH) and age-related maculopathy susceptibility 2 (ARMS2), influence the relative benefits of Age-Related Eye Disease Study (AREDS) supplements. DESIGN: Unplanned retrospective evaluation of a prospective, randomized, placebo-controlled clinical trial of vitamins and minerals for the treatment of AMD. SUBJECTS: AREDS participants (mean age, 69 years) who were at risk of developing late AMD and who were randomized to the 4 arms of AREDS supplement treatment. METHODS: Analyses were performed using the Cox proportional hazards model to predict progression to late AMD (neovascular or central geographic atrophy). Statistical models, adjusted for age, gender, smoking status, and baseline AMD severity, were used to examine the influence of genotypes on the response to therapy with 4 randomly assigned arms of AREDS supplement components: placebo, antioxidants (vitamin C, vitamin E, ß-carotene), zinc, or a combination. MAIN OUTCOME MEASURES: The influence of the genotype on the relative treatment response to the randomized components of the AREDS supplement, measured as progression to late AMD. RESULTS: Of the 1237 genotyped AREDS participants of white ethnicity, late AMD developed in 385 (31.1%) during the mean follow-up of 6.6 years. As previously demonstrated, CFH genotype (P = 0.005), ARMS2 (P< 0.0001), and supplement were associated individually with progression to late AMD. An interaction analysis found no evidence that the relative benefits of AREDS supplementation varied by genotype. Analysis of (1) CFH rs1061170 and rs1410996 combined with ARMS2 rs10490924 with the 4 randomly assigned arms of AREDS supplement and (2) analysis of the combination of CFH rs412852 and rs3766405 with ARMS2 c.372_815del443ins54 with the AREDS components resulted in no interaction (P = 0.06 and P = 0.45, respectively, before multiplicity adjustment). CONCLUSIONS: The AREDS supplements reduced the rate of AMD progression across all genotype groups. Furthermore, the genotypes at the CFH and ARMS2 loci did not statistically significantly alter the benefits of AREDS supplements. Genetic testing remains a valuable research tool, but these analyses suggest it provides no benefits in managing nutritional supplementation for patients at risk of late AMD.

Free radical scavenging, antioxidant and cancer chemoprevention by grape seed proanthocyanidin: an overview.

A large number of investigations have demonstrated a broad spectrum of pharmacological and therapeutic benefits of grape seed proanthocyanidins (GSP) against oxidative stress and degenerative diseases including cardiovascular dysfunctions, acute and chronic stress, gastrointestinal distress, neurological disorders, pancreatitis, various stages of neoplastic processes and carcinogenesis including detoxification of carcinogenic metabolites. GSP exhibited potent free radical scavenging abilities in both in vitro and in vivo models. GSP exerted significant in vivo protection against structurally diverse drug and chemical-induced hepatotoxicity, cardiotoxicity, neurotoxicity, nephrotoxicity and spleentoxicity. GSP also protected against idarubicin and 4-hydroxyperoxy-cyclophosphamide-induced cytotoxicity toward human normal liver cells. GSP exhibited selective cytotoxicity toward selected human cancer cells, while enhancing the growth and viability of normal cells. GSP exhibited potent modulatory effects of pro-apoptotic and apoptotic regulatory bcl-XL, p53, c-myc, c-JUN, JNK-1 and CD36 genes. Long-term exposure to GSP may serve as a novel chemoprotectant against three stages of DMN-induced liver carcinogenesis and tumorigenesis including initiation, promotion and progression. GSP may selectively protect against oxidative stress, genomic integrity and cell death patterns in vivo. These results demonstrate that GSP may serve as a novel therapeutic intervention against carcinogenesis.

Nrl-knockout mice deficient in Rpe65 fail to synthesize 11-cis retinal and cone outer segments.

PURPOSE: To define rod and cone function further in terms of visual cycle mechanism, the retinal phenotype resulting from Rpe65 (retinoid isomerase I) deficiency in Nrl(-)(/)(-) mice having a single class of photoreceptors resembling wild-type cones was characterized and outcomes of retinoid supplementation evaluated. METHODS: Rpe65(-)(/)(-)/Nrl(-)(/)(-) mice were generated by breeding Rpe65(-)(/)(-) and Nrl(-)(/)(-) strains. Retinal histology, protein expression, retinoid content, and electroretinographic (ERG) responses were evaluated before and after treatment with 11-cis retinal by intraperitoneal injection. Results Retinas of young Rpe65(-)(/-)/Nrl(-)(/-) mice exhibited normal lamination, but lacked intact photoreceptor outer segments at all ages examined. Rpe65, Nrl, and rhodopsin were not detected, and S-opsin and M/L-opsin levels were reduced. Retinyl esters were the only retinoids present. In contrast, Nrl(-)(/)(-) mice exhibited decreased levels of retinaldehydes and retinyl esters, and elevated levels of retinols. ERG responses were elicited from Rpe65(-)(/-)/Nrl(-)(/-) mice only at the two highest intensities over a 4-log-unit range. Significant retinal thinning and outer nuclear layer loss occurred in Rpe65(-)(/-)/Nrl(-)(/-) mice with aging. Administration of exogenous 11-cis retinal did not rescue retinal morphology or markedly improve ERG responses. CONCLUSIONS: The findings provide clarification of reported cone loss of function in Rpe65(-)(/-)/Nrl(-)(/-) mice, now showing that chromophore absence results in destabilized cone outer segments and rapid retinal degeneration. The data support the view that rod-dominant retinas do not have a cone-specific mechanism for 11-cis retinal synthesis and have potential significance for therapeutic strategies for rescue of cone-rich retinal regions affected by disease in the aging human population.

Genetic susceptibility to age-related macular degeneration: a paradigm for dissecting complex disease traits.

Age-related macular degeneration (AMD) is a progressive neurodegenerative disease, which affects quality of life for millions of elderly individuals worldwide. AMD is associated with a diverse spectrum of clinical phenotypes, all of which include the death of photoreceptors in the central part of the human retina (called the macula). Tremendous progress has been made in identifying genetic susceptibility variants for AMD. Variants at chromosome 1q32 (in the region of CFH) and 10q26 (LOC387715/ARMS2) account for a large part of the genetic risk to AMD and have been validated in numerous studies. In addition, susceptibility variants at other loci, several as yet unidentified, make substantial cumulative contribution to genetic risk for AMD; among these, multiple studies support the role of variants in APOE and C2/BF genes. Genome-wide association and re-sequencing projects, together with gene-environment interaction studies, are expected to further define the causal relationships that connect genetic variants to AMD pathogenesis and should assist in better design of prevention and intervention.

Radioprotective and antioxidant activity of fractionated extracts of berries of Hippophae rhamnoides.

Plants are an abundant source of medicinal compounds, some of which are useful in combating free radical-mediated oxidative stress. In the present study, initially two fractions designated REC-1001 (flavonoid-rich fraction) and REC-1002 (flavonoid-poor fraction) of Hippophae rhamnoides were screened on the basis of their reducing power in the aqueous phase. REC-1001 was selected for further study, since it exhibited 27.38 times higher antioxidant activity than REC-1002. REC-1001 also showed significant (P < .05) membrane protection potential at 50 microg/mL, which was attributed to its ability to scavenge peroxyl radicals (64.82 +/- 1.25% scavenging within 1,440 min). A significant (P < .05) difference of 67.02% in free radical scavenging activity at 1,000 ng/mL between REC-1001 and vitamin E demonstrated the extract fraction's worth in radiation protection. Such activities were attributed to the presence of quercetin, isorhamnetin, and kaempferol in this fraction. Further, REC-1001 was found to be nontoxic up to 200 mg/kg of body weight. This research suggests that the REC-1001 fraction of H. rhamnoides extract is a safe and effective antioxidant nutraceutical product.

Interaction of retinal bZIP transcription factor NRL with Flt3-interacting zinc-finger protein Fiz1: possible role of Fiz1 as a transcriptional repressor.

NRL (neural retina leucine zipper) is a basic motif leucine zipper transcription factor of the Maf-subfamily. Multiple phosphorylated isoforms of NRL are detected specifically in rod photoreceptors. NRL regulates the expression of several rod-specific genes, including rhodopsin and cGMP phosphodiesterase beta-subunit, in synergy with other transcription factors (e.g. the homeodomain protein CRX). Missense mutations in the human NRL gene are associated with autosomal dominant retinitis pigmentosa, whereas the loss of its function leads to rodless retina in Nrl-knockout mice that exhibit enhanced S-cone function. To further elucidate the molecular mechanism(s) underlying NRL-mediated transcriptional regulation, we used yeast two-hybrid screening to isolate NRL-interacting proteins in the retina and report the identification of Flt3-interacting zinc-finger protein, Fiz1. Interaction of Fiz1 and NRL-leucine zipper was validated by GST pulldown assays and co-immunoprecipitation from bovine retinal nuclear extracts. Fiz1 suppressed NRL- but not CRX-mediated transactivation of rhodopsin promoter activity in transiently transfected CV1 cells. The mRNA and the protein for both Fiz1 and its only other known interacting protein Flt3, a receptor tyrosine kinase, are expressed in the retina. Our results indicate potential cross-talk among signaling pathways in the retina and suggest that the function of NRL is modulated by its interaction with specific repressor proteins.