Ophthalmology
Abstract
Dyslipidemia and autophagy have been implicated in the pathogenesis of blinding neovascular age-related macular degeneration (NV-AMD). Very low-density lipoprotein receptor (VLDLR), expressed in photoreceptors with a high metabolic rate, facilitates the uptake of triglyceride-derived fatty acids (FA). Since FA uptake is reduced in Vldlr-/- tissues, more remain in circulation, and the retina is fuel deficient, driving the formation in mice of neovascular lesions reminiscent of retinal angiomatous proliferation (RAP), a subtype of NV-AMD. Nutrient scarcity and energy failure are classically mitigated by increasing autophagy. We find that excess circulating lipids restrain retinal autophagy, which contributes to pathological angiogenesis in the Vldlr-/- RAP model. Triglyceride-derived FA sensed by free fatty acid receptor 1 (FFAR1) restricted autophagy and oxidative metabolism in photoreceptors. FFAR1 suppressed transcription factor EB (TFEB), a master regulator of autophagy and lipid metabolism. Reduced TFEB, in turn, decreased Sirtuin-3 expression and mitochondrial respiration. Metabolomic signatures of mouse RAP-like retinas were consistent with a role in promoting angiogenesis. This signature was also found in human NV-AMD vitreous. Restoring photoreceptor autophagy in Vldlr-/- retinas, either pharmacologically or by deleting Ffar1, enhanced metabolic efficiency and suppressed pathological angiogenesis. Dysregulated autophagy by circulating lipids might therefore contribute to the energy failure of photoreceptors driving neovascular eye diseases, and FFAR1 may be a target for intervention.
Authors
Emilie Heckel, Gael Cagnone, Tapan Agnihotri, Bertan Cakir, Ashim Das, Jin Sung Kim, Nicholas Kim, Geneviève Lavoie, Anu Situ, Sheetal Pundir, Ye Sun, Florian Wünnemann, Kerry A. Pierce, Courtney Dennis, Grant A. Mitchell, Sylvain Chemtob, Flavio A. Rezende, Gregor Andelfinger, Clary B. Clish, Philippe P. Roux, Przemyslaw Sapieha, Lois E.H. Smith, Jean-Sébastien Joyal
Abstract
BACKGROUND Outcome measures sensitive to disease progression are needed for ATP-binding cassette, sub-family A, member 4–associated (ABCA4-associated) retinopathy. We aimed to quantify ellipsoid zone (EZ) loss and photoreceptor degeneration beyond EZ-loss in ABCA4-associated retinopathy and investigate associations between photoreceptor degeneration, genotype, and age.METHODS We analyzed 132 eyes from 66 patients (of 67 enrolled) with molecularly confirmed ABCA4-associated retinopathy from a prospective natural history study with a median [IQR] follow-up of 4.2 years [3.1, 5.1]. Longitudinal spectral-domain optical coherence tomography volume scans (37 B-scans, 30° × 15°) were segmented using a deep learning (DL) approach. For genotype-phenotype analysis, a model of ABCA4 variants was applied with the age of criterion EZ-loss (6.25 mm2) as the dependent variable.RESULTS Patients exhibited an average (square-root-transformed) EZ-loss progression rate of [95% CI] 0.09 mm/y [0.06, 0.11]. Outer nuclear layer (ONL) thinning extended beyond the area of EZ-loss. The average distance from the EZ-loss boundary to normalization of ONL thickness (to ±2 z score units) was 3.20° [2.53, 3.87]. Inner segment (IS) and outer segment (OS) thinning was less pronounced, with an average distance from the EZ-loss boundary to layer thickness normalization of 1.20° [0.91, 1.48] for the IS and 0.60° [0.49, 0.72] for the OS. An additive model of allele severity explained 52.7% of variability in the age of criterion EZ-loss.CONCLUSION Patients with ABCA4-associated retinopathy exhibited significant alterations of photoreceptors outside of EZ-loss. DL-based analysis of photoreceptor laminae may help monitor disease progression and estimate the severity of ABCA4 variants.TRIAL REGISTRATION ClinicalTrials.gov identifier: NCT01736293.FUNDING National Eye Institute Intramural Research Program and German Research Foundation grant PF950/1-1.
Authors
Maximilian Pfau, Catherine A. Cukras, Laryssa A. Huryn, Wadih M. Zein, Ehsan Ullah, Marisa P. Boyle, Amy Turriff, Michelle A. Chen, Aarti S. Hinduja, Hermann E.A. Siebel, Robert B. Hufnagel, Brett G. Jeffrey, Brian P. Brooks
Abstract
Adiponectin receptor 1 (ADIPOR1) is a lipid and glucose metabolism regulator that possesses intrinsic ceramidase activity. Mutations of the ADIPOR1 gene have been associated with non-syndromic and syndromic retinitis pigmentosa. Here we show that the absence of AdipoR1 in mice leads to progressive photoreceptor degeneration, significant reduction of electroretinogram amplitudes, decreased retinoid content in the retina, and reduced cone opsin expression. Single-cell RNA-Seq results indicated that ADIPOR1 encodes the most abundantly expressed ceramidase in mice and one of the two most highly expressed ceramidases in the human retina, next to acid ceramidase ASAH1. We discovered an accumulation of ceramides in the AdipoR1-/- retina, likely due to insufficient ceramidase activity for healthy retina function, resulting in photoreceptor death. Combined treatment with desipramine and L-cycloserine (DC) lowered ceramide levels and exerted a protective effect on photoreceptors in AdipoR1-/- mice. Moreover, we observed improvement in cone-mediated retinal function in the DC-treated animals. Lastly, we found that prolonged DC-treatment corrected the electrical responses of the primary visual cortex to visual stimuli, approaching near-normal levels for some parameters. These results highlight the importance of ADIPOR1 ceramidase in the retina, and show that pharmacological inhibition of ceramide generation can provide a therapeutic strategy for ADIPOR1-related retinopathy.
Authors
Dominik Lewandowski, Andrzej T. Foik, Roman Smidak, Elliot H. Choi, Jianye Zhang, Thanh Hoang, Aleksander Tworak, Susie Suh, Henri Leinonen, Zhiqian Dong, Antonio F.M. Pinto, Emily Tom, Jennings C. Luu, Joan Y. Lee, Xiuli Ma, Erhard Bieberich, Seth Blackshaw, Alan Saghatelian, David C. Lyon, Dorota Skowronska-Krawczyk, Marcin Tabaka, Krzysztof Palczewski
Abstract
BACKGROUND. >1,500 variants in the ABCA4 locus underlie a heterogeneous spectrum of retinal disorders ranging from aggressive childhood-onset chorioretinopathy to milder, late-onset macular disease. Genotype-phenotype correlation studies have been limited in clinical applicability as patient cohorts are typically small and seldom capture the full natural history of individual genotypes. To overcome these limitations, we constructed a genotype-phenotype correlation matrix that provides quantifiable probabilities of long-term disease outcomes associated with specific ABCA4 genotypes from a large, age-restricted patient cohort. METHODS. The study included 112 unrelated patients ≥50 years of age in whom 2 pathogenic variants were identified after sequencing of the ABCA4 locus. Clinical characterization was performed using the results of best-corrected visual acuity, retinal imaging and full-field electroretinogram testing. RESULTS. Four distinct prognostic groups were defined according to the spatial severity of disease features across the fundus. Recurring genotypes were observed in milder prognoses including those associated with a newly defined class of rare hypomorphic alleles. PVS1 (predicted null) variants were enriched in the most severe prognoses; however, missense variants comprised a larger than expected fraction of these patients. Analysis of allele combinations and their respective prognostic severity, showed that certain variants such as p.(Gly1961Glu), and both rare and frequent hypomorphic alleles, are “clinically dominant” with respect to patient phenotypes irrespective of the allele in trans. CONCLUSION. These results provide much needed structure to the complex genetic and clinical landscape of ABCA4 disease and adds a tool to the clinical repertoire to quantitatively assess individual genotype-specific prognoses in patients.
Authors
Winston Lee, Jana Zernant, Pei-Yin Su, Takayuki Nagasaki, Stephen H. Tsang, Rando Allikmets
Abstract
Ischemic retinopathies including diabetic retinopathy are major causes of blindness. While neurons and Müller glia are recognized as important regulators of reparative and pathologic angiogenesis, the role of mononuclear phagocytes (MPs), such as microglia/macrophages, is unclear, particularly microglia, the resident retinal immune cells. Here we found microglial/macrophage activation in human diabetic retinopathy, especially in neovessels from human neovascular membranes in proliferative retinopathy, including TNF-α expression. There was similar activation in the mouse oxygen-induced retinopathy (OIR) model of ischemia-induced neovascularization. Glucagon-like peptide-1 receptor (GLP-1R) agonists are in clinical use for glycemic control in diabetes and are also known to modulate microglia. We investigated the effect of a long-acting GLP-1R agonist, NLY01. Following intravitreal administration, NLY01 selectively localized to MPs in OIR retina. NLY01 modulated MP but not retinal endothelial cell viability, apoptosis, and tube formation in vitro. In OIR, NLY01 treatment inhibited MP infiltration and activation, including microglia/macrophage expression of cytokines in vivo. NLY01 significantly suppressed global induction of retinal inflammatory cytokines, promoted reparative angiogenesis, and suppressed pathologic retinal neovascularization. Collectively, these findings indicate the important role of microglia/macrophages in regulation of retinal vascularization in ischemia and suggest modulation of MPs as a new treatment strategy for ischemic retinopathies.
Authors
Lingli Zhou, Zhenhua Xu, Yumin Oh, Rico Gamuyao, Grace Lee, Yangyiran Xie, Hongkwan Cho, Seulki Lee, Elia J. Duh
Abstract
Fibrotic posterior capsular opacification (PCO), a major complication of cataract surgery, is driven by transforming growth factor β (TGFβ). Previously, αV integrins were found to be critical for the onset of TGFβ-mediated PCO in vivo, however, the functional heterodimer was unknown. Here, β8 integrin conditional knockout (β8ITGcKO) lens cells (LCs) were observed to attenuate their fibrotic responses, while both β5 and β6 integrin null LCs underwent fibrotic changes similar to WT at 5 days PCS. RNAseq revealed that β8ITGcKO LCs attenuated their upregulation of integrins and their ligands, as well as known targets of TGFβ induced signaling at 24 hours PCS. Treatment of β8ITGcKO eyes with active TGFβ1 at the time of surgery rescued the fibrotic response. Treatment of wild type mice with an anti- αVβ8 integrin function blocking antibody at the time of surgery ameliorated both canonical TGFβ signaling and LC fibrotic response PCS, and treatment at 5 days PCS, after surgically induced fibrotic responses are established, largely reversed this fibrotic response. These data suggest that αVβ8 integrin is a major regulator of TGFβ activation by LCs PCS and that therapeutics targeting αVβ8 integrin could be effective for fibrotic PCO prevention and treatment.
Authors
Mahbubul H. Shihan, Samuel G. Novo, Yan Wang, Dean Sheppard, Amha Atakilit, Thomas D. Arnold, Nicole M. Rossi, Adam P. Faranda, Melinda K. Duncan
Abstract
Mutations in the cilium-associated protein CEP290 cause retinal degeneration as part of multi-organ ciliopathies or as retina-specific diseases. The precise location and the functional roles of CEP290 within cilia and, specifically, the connecting cilia (CC) of photoreceptors, remain unclear. We used superresolution fluorescence microscopy and electron microscopy (TEM) to localize CEP290 in the CC and in primary cilia of cultured cells with sub-diffraction resolution, and to determine effects of CEP290 deficiency in three mutant models. Radially, CEP290 localizes in close proximity to the microtubule doublets in the region between the doublets and the ciliary membrane. Longitudinally, it is distributed throughout the length of the CC whereas it is confined to the very base of primary cilia in hRPE-1 cells. We found Y-shaped links, ciliary sub-structures between microtubules and membrane, throughout the length of the CC. Severe CEP290 deficiencies in mouse models did not prevent assembly of cilia or cause obvious mislocalization of ciliary components in early stages of degeneration. There were fewer cilia and no normal outer segments in the mutants, but the Y-shaped links were clearly present. These results point to photoreceptor-specific functions of CEP290 essential for CC maturation and stability following the earliest stages of ciliogenesis.
Authors
Valencia L. Potter, Abigail R. Moye, Michael A. Robichaux, Theodore G. Wensel
Abstract
Dry eye disease affects over 16 million adults in the U.S. and the majority of cases are due to Meibomian gland dysfunction. Unfortunately, the identity of the stem cells involved in Meibomian gland development and homeostasis are not well-elucidated. Here, we report that loss of Krox20, a zinc-finger transcription factor involved in development of ectoderm-derived tissues, or deletion of KROX20-expressing epithelial cells disrupts Meibomian gland formation and homeostasis, leading to dry eye disease secondary to Meibomian gland dysfunction. Ablation of Krox20-lineage cells in adult mice also resulted in dry eye disease, implicating Krox20 in homeostasis of the mature Meibomian gland. Lineage tracing and expression analyses revealed a restricted KROX20 expression pattern in the ductal areas of the Meibomian gland, although Krox20-lineage cells generate the full, mature Meibomian gland. This suggests that KROX20 marks a stem/progenitor cell population that differentiates to generate the entire Meibomian gland. Our Krox20 mouse models provide a powerful system that delineated the identity of stem cells required for Meibomian gland development and homeostasis, and can be used to investigate the factors underlying these processes. They are also robust models of Meibomian gland dysfunction-related dry eye disease with a potential for use in pre-clinical therapeutic screening.
Authors
Edem Tchegnon, Chung-Ping Liao, Elnaz Ghotbi, Tracey Shipman, Yong Wang, Renee M. McKay, Lu Q. Le
Abstract
BACKGROUND. This study systematically investigated circulating and retinal tissue lipid determinants of human diabetic retinopathy (DR) to identify underlying lipid alterations associated with severity of DR. METHODS. Retinal tissues were retrieved from postmortem human eyes including 19 individuals without diabetes, 20 with diabetes but without DR, and 20 with diabetes and DR for lipidomic study. In a parallel study, serum samples from 28 American Indians with type 2 diabetes from the Glia River Indian Community including 12 without DR, 7 with mild non-proliferative DR (NPDR), and 9 with moderate NPDR were selected. A mass-spectrometry-based lipidomic platform was used to measure serum and tissue lipids. RESULTS. In the postmortem retinas, we found a graded decrease of long chain acylcarnitines, longer chain fatty acid ester of hydroxyl fatty acids, diacylglycerols, triacylglycerols, phosphatidylcholines, and Ceramide[NS] in central retina from no diabetes to diabetes with DR. The American Indians’ sera also exhibited a graded decrease in circulating long-chain acylcarnitines and a graded increase in the intermediate length saturated and monounsaturated triacylglycerols from no DR to moderate NPDR. CONCLUSION. These findings suggest diminished synthesis of complex lipids and impaired mitochondrial β-oxidation of fatty acids in retinal DR with parallel changes in circulating lipids. TRIALREGISTRION. ClinicalTrials.gov NCT00340678 FUNDING. Supported by NIH grants K08DK106523, R03DK121941, P30DK089503, P30DK081943, P30DK020572, P30 EY007003, The Thomas Beatson Foundation, and JDRF Center for Excellence (5-COE-2019-861-S-B).
Authors
Patrice E. Fort, Thekkelnaycke M. Rajendiran, Tanu Soni, Jaeman Byun, Yang Shan, Helen C. Looker, Robert G. Nelson, Matthias Kretzler, George Michailidis, Jerome E. Roger, Thomas W. Gardner, Steven F. Abcouwer, Subramaniam Pennathur, Farsad Afshinnia
Abstract
The metabolic environment is important for neuronal cells, such as photoreceptors. When photoreceptors undergo degeneration, as occurs during retinitis pigmentosa (RP), patients have progressive loss of vision that proceeds to full blindness. Currently, there are no available treatments for the majority of RP diseases. We performed metabolic profiling of the neural retina in a preclinical model of RP and found that tricarboxylic acid (TCA) cycle intermediates were reduced during disease. We then determined that, 1) promoting citrate production within the TCA cycle in retinal neurons during disease progression protects the photoreceptors from cell death and prolongs visual function, 2) that supplementation with single metabolites within the TCA cycle can provide this therapeutic effect in vivo over time, and, 3) that this therapeutic effect is not specific to a particular genetic mutation but has broad applicability for patients with RP and other retinal degenerative diseases. Overall, targeting TCA cycle activity in the neural retina promotes photoreceptor survival and visual function during neurodegenerative disease.
Authors
Ashley A. Rowe, Pinkal D. Patel, Ruth Gordillo, Katherine J. Wert
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