High-throughput ultrastructural analysis of macular telangiectasia type 2.

Introduction: Macular Telangiectasia type 2 (MacTel), is an uncommon form of late-onset, slowly-progressive macular degeneration. Associated with regional Müller glial cell loss in the retina and the amino acid serine synthesized by Müller cells, the disease is functionally confined to a central retinal region - the MacTel zone. Methods: We have used high-throughput multi-resolution electron microscopy techniques, optimized for disease analysis, to study the retinas from two women, mother and daughter, aged 79 and 48 years respectively, suffering from MacTel. Results: In both eyes, the principal observations made were changes specific to mitochondrial structure both outside and within the MacTel zone in all retinal cell types, with the exception of those in the retinal pigment epithelium (RPE). The lesion areas, which are a hallmark of MacTel, extend from Bruch's membrane and the choriocapillaris, through all depths of the retina, and include cells from the RPE, retinal vascular elements, and extensive hypertrophic basement membrane material. Where the Müller glial cells are lost, we have identified a significant population of microglial cells, exclusively within the Henle fiber layer, which appear to ensheathe the Henle fibers, similar to that seen normally by Müller cells. Discussion: Since Müller cells synthesize retinal serine, whereas retinal neurons do not, we propose that serine deficiency, required for normal mitochondrial function, may relate to mitochondrial changes that underlie the development of MacTel. With mitochondrial changes occurring retina-wide, the question remains as to why the Müller cells are uniquely susceptible within the MacTel zone.

Ocular Effects of Prenatal Carotenoid Supplementation in the Mother and Her Child: The Lutein and Zeaxanthin in Pregnancy (L-ZIP) Randomized Trial - Report Number 2.

Purpose: Lutein (L) and zeaxanthin (Z) are xanthophyll carotenoids that have been promoted to enhance maternal health and infant visual and neurodevelopment. In this study, we determined the effects of prenatal L and Z supplementation on systemic and ocular carotenoid status in the mother and her newborn infant (NCT03750968). This report focuses on the ocular effects of prenatal carotenoid supplementation. Design: A prospective randomized clinical trial with 47 subjects randomly assigned by 1:1 allocation to receive standard-of-care prenatal vitamins along with 10 mg L and 2 mg Z softgel (Carotenoid Group) or standard-of-care prenatal vitamins with a placebo softgel (Control Group) starting in the first trimester. Subjects: We enrolled low-risk pregnancy subjects aged ≥18 years from the obstetrics and gynecology clinic of the University of Utah Hospital. Methods: Maternal macular, skin, and serum carotenoid concentrations were measured using autofluorescence imaging, resonance Raman spectroscopy, and high-performance liquid chromatography, respectively. Infants' ocular carotenoids and retinal architecture were measured by blue light reflectance imaging and spectral-domain OCT, respectively. Main Outcome Measures: Changes in maternal and infant macular pigment, skin, and serum carotenoid status over the study period. Differences in infants' retinal maturity indicators between the 2 study groups. Results: Following supplementation, there was a statistically significant increase in maternal macular pigment optical volume (P < 0.001) in the Carotenoid Group relative to the Control Group at all study time points, and there was no detectable maternal ocular carotenoid depletion. Infant skin and serum carotenoids increased significantly in the Carotenoid Group compared with the Control Group. As exploratory endpoints, infants in the Carotenoid Group had a 20% increase in macular pigment optical density (P = 0.242) and more mature foveal parameters compared with those in the Control Group. Conclusion: Prenatal carotenoid supplementation significantly increased maternal and infant systemic carotenoids and caused a pattern of increased infant ocular carotenoid status, which may benefit both mothers and their infants' ocular development and function. This study provides important data to design and power a future multicenter study of prenatal carotenoid supplementation in higher-risk pregnancies. Financial Disclosures: The author(s) have no proprietary or commercial interest in any materials discussed in this article.

The Progression of Stargardt Disease as Determined by Spectral-Domain Optical Coherence Tomography over a 24-Month Period (ProgStar Report No. 18).

INTRODUCTION: The aim of this study was to evaluate the progression of atrophy as determined by spectral-domain optical coherence tomography (SD-OCT) in patients with molecularly confirmed ABCA4-associated Stargardt disease type 1 (STGD1) over a 24-month period in a multicenter prospective cohort study. METHODS: SD-OCT images from 428 eyes of 236 patients were analyzed. Change of mean thickness (MT) and intact area were estimated after semiautomated segmentation for the following individual layers in the central subfield (CS), inner ring (IR), and outer ring (OR) of the ETDRS grid: retinal pigment epithelium (RPE), outer segments (OSs), inner segments (IS), outer nuclear layer (ONL) inner retina (IR), and total retina. RESULTS: Statistically significant decreases of all outer retinal layers (RPE, OS, IS, and ONL) could be observed over a 24-month period both in decline of mean retinal thickness and intact area (p < 0.0001, respectively), whereas the IR showed an increase of retinal thickness in the CS and IR and remained unchanged in the OR. CONCLUSIONS: Significant loss could be detected in outer retinal layers by SD-OCT over a 24-month period in patients with STGD1. Loss of thickness and/or intact area of such layers may serve as potential endpoints for clinical trials that aim to slow down the disease progression of STGD1.

Elovl4b knockout zebrafish as a model for ocular very-long-chain PUFA deficiency.

Very-long-chain PUFAs (VLC-PUFAs) are a group of lipids with chain lengths >24 carbons, and the ELOVL4 (elongation of very-long-chain FA-4) enzyme is responsible for vertebrate VLC-PUFA biosynthesis. Studies on the role of VLC-PUFAs in vision have been hindered because of the need for adequate animal models to capture the global loss of VLC-PUFAs. Since homozygous Elovl4 ablation is lethal in neonatal mice because of catastrophic drying from the loss of their protective skin barrier, we established a zebrafish (Danio rerio) model of Elovl4 ablation. We generated Elovl4b KO zebrafish by creating a 56-bp deletion mutation in exon 2 of the Elovl4b gene using CRISPR-Cas9. We used GC-MS and LC-MS/MS to analyze the VLC-PUFA and lipid profiles from wild-type and Elovl4b KO fish eyes. We also performed histology and visual-behavioral tests. We found that heterozygous and homozygous Elovl4b KO zebrafish eyes had altered lipid profiles and a significantly lower C30 to C36 VLC-PUFA abundance than wild-type fish. Moreover, Elovl4b+/- and Elovl4b-/- KO larvae had significantly lower motor activity in response to light-dark cycles than their age-matched controls. Elovl4b-/- adult fish showed no obvious differences in gross retinal morphology and lamination compared with wild type, except for the presence of lipid droplets within the retinal pigment epithelial cell layer of Elovl4b-/- fish. Our data indicate that the loss of Elovl4b in zebrafish changes ocular lipid profiles and leads to visual abnormalities and subtle retinal changes. These findings highlight the use of zebrafish as a model for VLC-PUFA depletion and ELOVL4-related dysfunction.

It Takes a Village: An Interdisciplinary Approach to Preparing Internal Medicine Residents to Care for Patients at the Intersection of Women's Health, Gender-Affirming Care, and Health Disparities.

Objective: To create an interdisciplinary curriculum to teach key topics at the intersection of women's health, gender-affirming care, and health disparities to internal medicine (IM) residents. Materials and Methods: A core team of faculty from IM, Obstetrics and Gynecology, and Surgery partnered with faculty and fellows from other disciplines and with community experts to design and deliver the curriculum. The resulting curriculum consisted of themed half-day modules, each consisting of three to four inter-related topics, updated and repeated on an ∼3-year cycle. Health equity was a focus of all topics. Module delivery used diverse interactive learning strategies. Modules have been presented to ∼175 residents annually, beginning in 2015. To assess the curriculum, we used formative evaluation methods, using primarily anonymous, electronic surveys, and collected quantitative and qualitative data. Most surveys assessed resident learning by quantifying residents' self-reported comfort with skills taught in the module pre- and postsession. Results: Of 131 residents who completed an evaluation in 2022/23, 121 (90%) "somewhat" or "strongly" agreed with their readiness to perform a range of skills taught in the module. In all previous years where pre- and postsurveys were used to evaluate modules, we observed a consistent meaningful increase in the proportion of residents reporting high levels of comfort with the material. Residents particularly valued interactive teaching methods, and direct learning from community members and peers. Conclusion: Our interdisciplinary curriculum was feasible, valued by trainees, and increased resident learning. The curriculum provides a template to address equity issues across a spectrum of women's and gender-affirming care conditions that can be used by other institutions in implementing similar curricula.

Potassium Iodate Retinotoxicity after an Overdose of Anti-Radiation Pills: A Case of Apocalyptic Retinopathy.

PURPOSE: To report a case of severe retinal toxicity after an attempted suicidal overdose of anti-radiation pills containing a well-known retinal pigment epithelial toxin, potassium iodate. METHODS: Clinical examination and multimodal imaging are provided from a clinic visit eleven years after the suicide attempt. RESULTS: The 42-year-old patient had widespread retinal atrophy in both eyes with near complete absence of the retinal pigment epithelium and severe retinal thinning. A few remaining areas of preserved photoreceptors provided her with 20/60 and 20/200 vision in her right and left eyes, respectively, with very constricted visual fields. CONCLUSION: Despite being first described over 100 years ago, few cases of iodate retinotoxicity have been reported in the published literature, and most are related to accidental or intentional ingestion of iodate salts used to iodize table salt. This unique case of iodate retinotoxicity secondary to anti-radiation tablets highlights the risks of unregulated sale of these agents marketed in the United States as essential components of nuclear apocalypse "go bags" because they have a poor margin-of-safety and no specific warnings of the risk of permanent blindness with overdosage.

Vitreomacular interface abnormalities in the Ghanaian African.

BACKGROUND/OBJECTIVE: Describe vitreomacular interface abnormalities (VMIA) using spectral-domain optical coherence tomography (SD-OCT), and correlations with age-related macular degeneration (AMD) grade in Ghanaian Africans. SUBJECTS/METHODS: Prospective, cross-sectional study of adults aged ≥50 years recruited in Ghana AMD Study. Participant demographics, medical histories, ophthalmic examination, digital colour fundus photography (CFP) were obtained. High-resolution five-line raster OCT, Macular Cube 512 × 128 scans, and additional line scans in areas of clinical abnormality, were acquired. SD-OCT VMI features classified by International Vitreomacular Traction Study Group system and relationships to AMD grade were evaluated. OUTCOMES: VMIA prevalence, posterior vitreous detachment (PVD), vitreomacular adhesions (VMA), vitreomacular traction (VMT), epiretinal membranes (ERM), correlations with AMD grade. RESULTS: The full Ghana AMD cohort included 718 participants; 624 participants (1248 eyes) aged ≥50 years (range = 50-101, mean = 68.8), 68.9% female were included in this analysis. CFP with OCT scans were available for 776 eyes (397 participants); 707 (91.1%) had gradable CFP and OCT scans for both AMD and VMI grading forming the dataset for this report. PVD was absent in 504 (71.3%); partial and complete PVD occurred in 16.7% and 12.0% respectively. PVD did not increase with age (p = 0.720). VMIA without traction and macular holes were observed in 12.2% of eyes; 87.8% had no abnormalities. VMIA was not significantly correlated with AMD grade (p = 0.819). CONCLUSIONS: This provides the first assessment of VMIA in Ghanaian Africans. VMIA are common in Africans; PVD may be less common than in Caucasians. There was no significant association of AMD grade with VMIA.

Dietary fish oil enriched in very-long-chain polyunsaturated fatty acid reduces cardiometabolic risk factors and improves retinal function.

Very-long-chain polyunsaturated fatty acids (VLCPUFAs; C24-38) constitute a unique class of PUFA that have important biological roles, but the lack of a suitable dietary source has limited research in this field. We produced an n-3 C24-28-rich VLCPUFA-oil concentrated from fish oil to study its bioavailability and physiological functions in C57BL/6J mice. The serum and retinal C24:5 levels increased significantly compared to control after a single-dose gavage, and VLCPUFAs were incorporated into the liver, brain, and eyes after 8-week supplementation. Dietary VLCPUFAs resulted in favorable cardiometabolic changes, and improved electroretinography responses and visual performance. VLCPUFA supplementation changed the expression of genes involved in PPAR signaling pathways. Further in vitro studies demonstrated that the VLCPUFA-oil and chemically synthesized C24:5 are potent agonists for PPARs. The multiple potential beneficial effects of fish oil-derived VLCPUFAs on cardiometabolic risk and eye health in mice support future efforts to develop VLCPUFA-oil into a supplemental therapy.

Eye-mounting goggles to bridge the gap between benchtop experiments and in vivo robotic eye surgery.

A variety of robot-assisted surgical systems have been proposed to improve the precision of eye surgery. Evaluation of these systems has typically relied on benchtop experiments with artificial or enucleated eyes. However, this does not properly account for the types of head motion that are common among patients undergoing eye surgery, which a clinical robotic system will encounter. In vivo experiments are clinically realistic, but they are risky and thus require the robotic system to be at a sufficiently mature state of development. In this paper, we describe a low-cost device that enables an artificial or enucleated eye to be mounted to standard swim goggles worn by a human volunteer to enable more realistic evaluation of eye-surgery robots after benchtop studies and prior to in vivo studies. The mounted eye can rotate about its center, with a rotational stiffness matching that of an anesthetized patient's eye. We describe surgeon feedback and technical analyses to verify that various aspects of the design are sufficient for simulating a patient's eye during surgery.

Macular Pigment Carotenoids and Bisretinoid A2E.

Lutein (L), zeaxanthin (Z), and meso-zeaxanthin (MZ) are the three macular pigments (MP) carotenoids that uniquely accumulate in the macula lutea region of the human retina. L and Z are obtained by humans through dietary intake. The third MP, MZ, is rarely present in diet, and its abundance in the human fovea is due to the metabolic conversion of dietary L by the retinal pigment epithelium's RPE65 enzyme. The major functions of MP in ocular health are to filter high-intensity, phototoxic blue light and to act as effective antioxidants for scavenging free radicals. The pyridinium bisretinoid, N-retinylidene-N-retinylethanolamine (A2E), contributes to drusen formation in dry age-related macular degeneration (AMD) and to the autofluorescent flecks in autosomal recessive Stargardt disease (STGD1). Retinal carotenoids attenuate A2E formation and can directly and indirectly alleviate A2E-mediated oxidative damage. In this chapter, we review these more recently recognized interconnections between MP carotenoids and A2E bisretinoids.

Role of VLC-PUFAs in Retinal and Macular Degeneration.

Very-long-chain polyunsaturated fatty acids (VLC-PUFAs) are a special class of fatty acids that are present in the retina and a few other human tissues. They cannot be synthesized de novo and are rarely present in dietary sources. Structurally, these lipids are composed of a proximal end with a typical saturated fatty acid character and a distal end more characteristic of common PUFAs. They have not been studied in detail until recently due to their low abundance in these tissues and technical difficulties in assaying these lipids by conventional chromatography. This unique class of lipids has chain lengths greater than 24 carbons, with the longest typically 38 carbons long. There is increasing interest in understanding their roles in the maintenance of retinal membrane integrity and the prevention of macular degeneration and inherited retinal diseases.

Understanding the Roles of Very-Long-Chain Polyunsaturated Fatty Acids (VLC-PUFAs) in Eye Health.

Lipids serve many roles in the neural system, from synaptic stabilization and signaling to DNA regulation and neuroprotection. They also regulate inflammatory responses, maintain cellular membrane structure, and regulate the homeostatic balance of ions and signaling molecules. An imbalance of lipid subgroups is implicated in the progression of many retinal diseases, such as age-related macular degeneration (AMD), retinitis pigmentosa, and diabetic retinopathy, and diet can play a key role in influencing these diseases' onset, progression, and severity. A special class of lipids termed very-long-chain polyunsaturated fatty acids (VLC-PUFAs) is found exclusively in mammalian vertebrate retinas and a few other tissues. They comprise <2% of fatty acids in the retina and are depleted in the retinas of patients with diseases like diabetic retinopathy and AMD. However, the implications of the reduction in VLC-PUFA levels are poorly understood. Dietary supplementation studies and ELOVL4 transgene studies have had positive outcomes. However, much remains to be understood about their role in retinal health and the potential for targeted therapies against retinal disease.

FLUORESCENCE LIFETIME IMAGING OPHTHALMOSCOPY IN PATIENTS WITH MACULAR TELANGIECTASIA TYPE 2 WITH AND WITHOUT DIABETES.

PURPOSE: Macular telangiectasia type 2 (MacTel) is a vision-altering retinal disease with a high prevalence of diabetes. Differences between patients with MacTel with and without diabetes were investigated using fluorescence lifetime imaging ophthalmoscopy (FLIO). METHODS: Eighty-six patients with MacTel (59 ± 12 years) were included. 40 patients (46%) did not have diabetes, 16 patients (19%) were prediabetic, and 30 patients (35%) were diabetic. Of these, seven had diabetic retinopathy. 18 diabetic patients without MacTel and 42 age-matched healthy controls were included. FLIO lifetimes (FLTs) were obtained in short (SSC, 498-560 nm) and long (LSC, 560-720 nm) spectral channels from different areas of interest using a Heidelberg Engineering FLIO. RESULTS: Fundus autofluorescece lifetimes did not show significant differences when comparing diabetic with nondiabetic MacTel eyes (MacTel zone, SSC, diabetic: 243 ± 65 ps; nondiabetic: 232 ± 51 ps; P = 1.0; LSC, diabetic: 327 ± 66 ps; nondiabetic: 309 ± 54 ps; P = 0.582). Longitudinal changes were similarly unrelated to diabetes status. A nonsignificant trend of increased FLT progression with higher body mass index was found. Fundus autofluorescece lifetimes in diabetic patients without MacTel were significantly shorter within the MacTel zone and longer in the periphery compared with diabetic patients with MacTel. CONCLUSION: Although MacTel has a high prevalence of diabetes, FLTs from the MacTel zone are unrelated to diabetes. Fluorescence lifetime imaging ophthalmoscopy retains diagnostic abilities in patients with MacTel even in the presence of prediabetes, diabetes, and advanced diabetic retinopathy. The lack of diabetic FLT changes in the periphery of diabetic patients with MacTel is an interesting finding that needs further investigation.

The value of pre-symptomatic genetic risk assessment for age-related macular degeneration: the Moran AMD Genetic Testing Assessment (MAGENTA) study-a study protocol for a randomized controlled trial.

BACKGROUND: Age-related macular degeneration (AMD) is an irreversible blinding eye condition with complex genetic and environmental etiologies. Genetic testing for AMD for previously identified multiple-risk single nucleotide polymorphisms can help determine an individual's future susceptibility. However, such testing has been discouraged until evidence shows that providing such information to symptomatic or pre-symptomatic individuals will alter their disease course. Therefore, we designed this study to investigate whether knowledge of AMD risk could stimulate the adoption of a healthier lifestyle that could lower the incidence of AMD later in life. We hypothesize that pre-symptomatic individuals informed of a high genetic risk of AMD are more likely to make quantifiable, positive lifestyle changes relative to participants informed of lower genetic risk or randomized to deferred disclosure of genetic testing results. METHODS: The Moran AMD Genetic Testing Assessment (MAGENTA) study is a phase 2, single-center, prospective, double-masked, randomized controlled trial conducted at the John A. Moran Eye Center, University of Utah, Salt Lake City, Utah, USA. Participants are randomized by a 3:1 allocation ratio to immediate and deferred disclosure groups and followed for 12 months. Skin, ocular, and serum carotenoid status, as well as nutritional and social surveys, are assessed at study visits. Skin carotenoid assessment is by resonance Raman spectroscopy and reflectance spectroscopy, ocular carotenoids are measured with Heidelberg Spectralis autofluorescence imaging and fluorescence lifetime imaging ophthalmoscopy (FLIO), and serum carotenoids are quantified using high-performance liquid chromatography. The primary outcome evaluates changes in skin carotenoid status in response to genetic risk disclosure. The secondary outcomes examine changes in ocular and serum carotenoid status in response to genetic risk disclosure. Also, we will correlate AMD genetic risk with baseline ocular and systemic carotenoid status and FLIO. DISCUSSION: MAGENTA will provide much-needed evidence on whether pre-symptomatic testing for AMD risk can lead to quantifiable long-term changes in behavior and lifestyle associated with a lower incidence of AMD later in life. Findings from the MAGENTA trial will facilitate the design of a future larger, longer-term, multicenter phase 3 trial that could feature subgroup analysis, expanded measures of lifestyle modification, and potential active nutritional interventions. TRIAL REGISTRATION: ClinicalTrials.gov NCT05265624 . Registered on March 3, 2022.

Systemic Effects of Prenatal Carotenoid Supplementation in the Mother and her Child: The Lutein and Zeaxanthin in Pregnancy (L-ZIP) Randomized Trial -Report Number 1.

BACKGROUND: Adding carotenoids, particularly lutein (L) and zeaxanthin (Z), to prenatal micronutrient formulations has been promoted to enhance infant visual and neural development and to maintain maternal health. Although these claims are biologically plausible, they are not yet supported by a compelling prospective trial. OBJECTIVE: We investigated the effect of prenatal carotenoid supplementation on biomarkers of maternal and infant systemic carotenoid status. METHODS: We randomly assigned 47 first trimester pregnant subjects by 1:1 allocation to receive standard-of-care prenatal vitamins plus a 10 mg L and 2 mg Z softgel (the Carotenoid group) or standard-of-care prenatal vitamins with a placebo softgel (the Control group) for 6-8 mo. Maternal carotenoid concentrations in the serum and skin at the end of each trimester and postpartum were measured with HPLC and resonance Raman spectroscopy, respectively. Infants' systemic carotenoid status was assessed using similar techniques but optimized for infants. Repeated measures and paired t-tests were determined, and a P value < 0.05 was considered statistically significant. RESULTS: After supplementation, there was a statistically significant increase in maternal serum L + Z concentrations, serum total carotenoid concentrations, and skin carotenoid status (P < 0.001 for all) in the Carotenoid group relative to the Control group at all study time points. Similarly, infants whose mothers were in the Carotenoid group had a significant 5-fold increase in cord blood L + Z concentrations, over a 3-fold increase in cord blood total carotenoids, and a 38% increase in skin carotenoids compared with the Control group (P < 0.0001 for all). In addition, there was a strong positive, statistically significant correlation between postpartum maternal and infant systemic carotenoid status (P < 0.0001). CONCLUSION: Prenatal carotenoid supplementation significantly increased maternal and infant systemic (skin and serum) carotenoid status, which may benefit pregnant women and their infants' health. This trial was registered at clinicaltrials.gov as NCT03750968.

Prevalence and severity of diabetic retinopathy at first presentation to vitreoretinal services in Bhutan: a 3-year national study.

PURPOSE: To determine the prevalence and severity of diabetic retinopathy (DR) at first presentation among diabetic patients attending national vitreoretinal (VR) services in Bhutan STUDY DESIGN: Retrospective cross-sectional study METHODS: We included all diabetic patients in Bhutan who presented for retinal evaluation for the first time over a 3-year period (2013-2016). Data including demography, clinical details, diagnostic tests, and clinical staging of DR were analyzed. RESULTS: A total of 843 diabetic patients, aged 57.2 ± 12.0 (range 18-86) years, were enrolled. The majority were male (452, 53.6%; cumulative frequency [cf] 391, 46.4%; P = .14) and from urban settings (570, 67.6%; cf 273; 32.4%) and did not have modern schooling (555, 65.8%). Hypertension was the most common systemic comorbidity (501, 59.4%). The prevalence of DR was 42.7%, with mild nonproliferative DR (NPDR) being the most common type (187, 51.9%), followed by moderate NPDR (88, 24.4%) and proliferative DR (45, 12.5%). In addition, 120 patients had clinically significant macular edema (CSME), with a prevalence of 14.2%. Best-corrected visual acuity (BCVA) of 6/60 or worse occurred in 231 eyes (13.7%), and 41 patients (4.86%) had BCVA of 6/60 or worse bilaterally due to DR/CSME. A logistic regression model indicated that the major determinant of DR was the duration of diabetes, the odds rising by 1.27× with each year of disease (P < .0001). CONCLUSION: The prevalence of DR, including CSME, was high. Although a national DR screening program is established in Bhutan, there is a need to accelerate health education, community screening, and referral systems to reduce the prevalence of DR and CSME.

Prenatal Carotenoid Supplementation With Lutein or Zeaxanthin Ameliorates Oxygen-Induced Retinopathy (OIR) in Bco2-/- Macular Pigment Mice.

Purpose: Premature infants at risk of retinopathy of prematurity (ROP) miss placental transfer of the carotenoids lutein (L) and zeaxanthin (Z) during the third trimester. We previously demonstrated that prenatal L and Z supplementation raised carotenoid levels in infants at birth in the Lutein and Zeaxanthin in Pregnancy (L-ZIP) study (NCT03750968). Based on their antioxidant effects and bioavailability, we hypothesized that prenatal maternal supplementation with macular carotenoids would reduce the risk of ROP. To test this hypothesis, we utilized "macular pigment mice" genetically engineered to take up L and Z into the retina in a model of oxygen-induced retinopathy (OIR). Methods: Pregnant Bco2-/- mice were divided into nine experimental subgroups based on the type of supplementation (L, Z, or placebo) and on the maternal supplementation start date corresponding to the three trimesters of human fetal development (E0, E11, and P1). Pups and nursing mothers were exposed to 75% O2 for 5 days (P7-P12) and returned to room air for 5 days (P12-P17). Pups were killed at P12 and P17, and their retinas were analyzed for vaso-obliteration and intravitreal neovascularization. Results: Pups of pregnant mice supplemented with L or Z had significant reductions in areas of vaso-obliteration and intravitreal neovascularization compared to placebo. Prenatal carotenoid supplementation starting at E0 or E11 was significantly more protective against OIR than postnatal supplementation starting at P1. Conclusions: Prenatal supplementation with L and Z was beneficial in a mouse OIR model. We recommend testing prenatal L and Z supplementation in future human clinical trials to prevent ROP.

iPSC-derived retinal pigmented epithelial cells from patients with macular telangiectasia show decreased mitochondrial function.

Patient-derived induced pluripotent stem cells (iPSCs) provide a powerful tool for identifying cellular and molecular mechanisms of disease. Macular telangiectasia type 2 (MacTel) is a rare, late-onset degenerative retinal disease with an extremely heterogeneous genetic architecture, lending itself to the use of iPSCs. Whole-exome sequencing screens and pedigree analyses have identified rare causative mutations that account for less than 5% of cases. Metabolomic surveys of patient populations and GWAS have linked MacTel to decreased circulating levels of serine and elevated levels of neurotoxic 1-deoxysphingolipids (1-dSLs). However, retina-specific, disease-contributing factors have yet to be identified. Here, we used iPSC-differentiated retinal pigmented epithelial (iRPE) cells derived from donors with or without MacTel to screen for novel cell-intrinsic pathological mechanisms. We show that MacTel iRPE cells mimicked the low serine levels observed in serum from patients with MacTel. Through RNA-Seq and gene set enrichment pathway analysis, we determined that MacTel iRPE cells are enriched in cellular stress pathways and dysregulation of central carbon metabolism. Using respirometry and mitochondrial stress testing, we functionally validated that MacTel iRPE cells had a reduction in mitochondrial function that was independent of defects in serine biosynthesis and 1-dSL accumulation. Thus, we identified phenotypes that may constitute alternative disease mechanisms beyond the known serine/sphingolipid pathway.

Mechanism for the selective uptake of macular carotenoids mediated by the HDL cholesterol receptor SR-BI.

The macular carotenoids lutein and zeaxanthin are taken up from the bloodstream into the human retina through a selective process, for which the HDL cholesterol receptor scavenger receptor BI (SR-BI) in the cells of retinal pigment epithelium (RPE) is thought to be a key mediator. However, the mechanism of SR-BI-mediated selective uptake of macular carotenoids is still not fully understood. Here, we investigate possible mechanisms using biological assays and cultured HEK293 cells, a cell line without endogenous SR-BI expression. Binding affinities between SR-BI and various carotenoids were measured by surface plasmon resonance (SPR) spectroscopy, which shows that SR-BI cannot bind lutein or zeaxanthin specifically. Overexpression of SR-BI in HEK293 cells results in more lutein and zeaxanthin taken up than ß-carotene, and this effect can be eliminated by an SR-BI mutant (C384Y) whose cholesterol uptake tunnel is blocked. Next, we determined the effects of HDL and hepatic lipase (LIPC), SR-BI's partners in HDL cholesterol transport, on SR-BI-mediated carotenoid uptake. HDL addition dramatically reduced lutein, zeaxanthin, and ß-carotene in HEK293 cells expressing SR-BI, but the cellular lutein and zeaxanthin are higher than ß-carotene. LIPC addition increases the uptake of all three carotenoids in HDL-treated cells, and promotes the transport of lutein and zeaxanthin better than ß-carotene. Our results suggest that SR-BI and its HDL cholesterol partner HDL and LIPC may be involved in the selective uptake of macular carotenoids.

Divergent amino acid and sphingolipid metabolism in patients with inherited neuro-retinal disease.

OBJECTIVES: The non-essential amino acids serine, glycine, and alanine, as well as diverse sphingolipid species, are implicated in inherited neuro-retinal disorders and are metabolically linked by serine palmitoyltransferase (SPT), a key enzyme in membrane lipid biogenesis. To gain insight into the pathophysiological mechanisms linking these pathways to neuro-retinal diseases we compared patients diagnosed with two metabolically intertwined diseases: macular telangiectasia type II (MacTel), hereditary sensory autonomic neuropathy type 1 (HSAN1), or both. METHODS: We performed targeted metabolomic analyses of amino acids and broad sphingolipids in sera from a cohort of MacTel (205), HSAN1 (25) and Control (151) participants. RESULTS: MacTel patients exhibited broad alterations of amino acids, including changes in serine, glycine, alanine, glutamate, and branched-chain amino acids reminiscent of diabetes. MacTel patients had elevated 1-deoxysphingolipids but reduced levels of complex sphingolipids in circulation. A mouse model of retinopathy indicates dietary serine and glycine restriction can drive this depletion in complex sphingolipids. HSAN1 patients exhibited elevated serine, lower alanine, and a reduction in canonical ceramides and sphingomyelins compared to controls. Those patients diagnosed with both HSAN1 and MacTel showed the most significant decrease in circulating sphingomyelins. CONCLUSIONS: These results highlight metabolic distinctions between MacTel and HSAN1, emphasize the importance of membrane lipids in the progression of MacTel, and suggest distinct therapeutic approaches for these two neurodegenerative diseases.