Cone cell dysfunction attenuates retinal neovascularization in oxygen-induced retinopathy mouse model.

Aberrant neovascularization is the most common feature in retinopathy of prematurity (ROP), which leads to the retinal detachment and visual defects in neonates with a low gestational age eventually. Understanding the regulation of inappropriate angiogenic signaling benefits individuals at-risk. Recently, neural activity originating from the specific neural activity has been considered to contribute to retinal angiogenesis. Here, we explored the impact of cone cell dysfunction on oxygen-induced retinopathy (OIR), a mouse model commonly employed to understand retinal diseases associated with abnormal blood vessel growth, using the Gnat2cpfl3 (cone photoreceptor function loss-3) strain of mice (regardless of the sex), which is known for its inherent cone cell dysfunction. We found that the retinal avascular area, hypoxic area, and neovascular area were significantly attenuated in Gnat2cpfl3 OIR mice compared to those in C57BL/6 OIR mice. Moreover, the HIF-1α/VEGF axis was also reduced in Gnat2cpfl3 OIR mice. Collectively, our results indicated that cone cell dysfunction, as observed in Gnat2cpfl3 OIR mice, leads to attenuated retinal neovascularization. This finding suggests that retinal neural activity may precede and potentially influence the onset of pathological neovascularization.

Wnt/ß-Catenin-Signaling Modulates Megakaryopoiesis at the Megakaryocyte-Erythrocyte Progenitor Stage in the Hematopoietic System.

The bone marrow (BM) hematopoietic system (HS) gives rise to blood cells originating from hematopoietic stem cells (HSCs), including megakaryocytes (MKs) and red blood cells (erythrocytes; RBCs). Many steps of the cell-fate decision remain to be elucidated, being important for cancer treatment. To explore the role of Wnt/ß-catenin for MK and RBC differentiation, we activated ß-catenin signaling in platelet-derived growth factor b (Pdgfb)-expressing cells of the HS using a Cre-lox approach (Ctnnb1BM-GOF). FACS analysis revealed that Pdgfb is mainly expressed by megakaryocytic progenitors (MKPs), MKs and platelets. Recombination resulted in a lethal phenotype in mutants (Ctnnb1BM-GOFwt/fl, Ctnnb1BM-GOFfl/fl) 3 weeks after tamoxifen injection, showing an increase in MKs in the BM and spleen, but no pronounced anemia despite reduced erythrocyte counts. BM transplantation (BMT) of Ctnnb1BM-GOF BM into lethally irradiated wildtype recipients (BMT-Ctnnb1BM-GOF) confirmed the megakaryocytic, but not the lethal phenotype. CFU-MK assays in vitro with BM cells of Ctnnb1BM-GOF mice supported MK skewing at the expense of erythroid colonies. Molecularly, the runt-related transcription factor 1 (RUNX1) mRNA, known to suppress erythropoiesis, was upregulated in Ctnnb1BM-GOF BM cells. In conclusion, ß-catenin activation plays a key role in cell-fate decision favoring MK development at the expense of erythroid production.

Untargeted Metabolomic Analysis of Sjögren-Larsson Syndrome Reveals a Distinctive Pattern of Multiple Disrupted Biochemical Pathways.

Sjögren-Larsson syndrome (SLS) is a rare inherited neurocutaneous disease characterized by ichthyosis, spastic diplegia or tetraplegia, intellectual disability and a distinctive retinopathy. SLS is caused by bi-allelic mutations in ALDH3A2, which codes for fatty aldehyde dehydrogenase (FALDH) and results in abnormal lipid metabolism. The biochemical abnormalities in SLS are not completely known, and the pathogenic mechanisms leading to symptoms are still unclear. To search for pathways that are perturbed in SLS, we performed untargeted metabolomic screening in 20 SLS subjects along with age- and sex-matched controls. Of 823 identified metabolites in plasma, 121 (14.7%) quantitatively differed in the overall SLS cohort from controls; 77 metabolites were decreased and 44 increased. Pathway analysis pointed to disrupted metabolism of sphingolipids, sterols, bile acids, glycogen, purines and certain amino acids such as tryptophan, aspartate and phenylalanine. Random forest analysis identified a unique metabolomic profile that had a predictive accuracy of 100% for discriminating SLS from controls. These results provide new insight into the abnormal biochemical pathways that likely contribute to disease in SLS and may constitute a biomarker panel for diagnosis and future therapeutic studies.

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.

Spatial distribution of metabolites in the retina and its relevance to studies of metabolic retinal disorders.

INTRODUCTION: The primate retina has evolved regional specialisations for specific visual functions. The macula is specialised towards high acuity vision and is an area that contains an increased density of cone photoreceptors and signal processing neurons. Different regions in the retina display unique susceptibility to pathology, with many retinal diseases primarily affecting the macula. OBJECTIVES: To better understand the properties of different retinal areas we studied the differential distribution of metabolites across the retina. METHODS: We conducted an untargeted metabolomics analysis on full-thickness punches from three different regions (macula, temporal peri-macula and periphery) of healthy primate retina. RESULTS: Nearly half of all metabolites identified showed differential abundance in at least one comparison between the three regions. Furthermore, mapping metabolomics results from macula-specific eye diseases onto our region-specific metabolite distributions revealed differential abundance defining systemic metabolic dysregulations that were region specific. CONCLUSIONS: The unique metabolic phenotype of different retinal regions is likely due to the differential distribution of different cell types in these regions reflecting the specific metabolic requirements of each cell type. Our results may help to better understand the pathobiology of retinal diseases with region specificity.

Intraretinal pigmented cells in retinal degenerative disease.

PURPOSE: Invasion of pigmented cells into the retina occurs in retinal degenerative diseases, such as macular telangiectasia type 2 (MacTel) and retinitis pigmentosa (RP). These intraretinal pigmented cells may be derived from the retinal pigment epithelium (RPE), but differences and similarities between intraretinal pigmented cells and RPE have so far not been well characterised.Clinicopathologic case report. METHOD: Here, we compared intraretinal pigment cells with RPE cells by immunohistochemistry. Immunohistological stains for classic RPE markers (RPE65, CRALBP and KRT18) and blood vessel markers (lectin and collagen 4) were done on sections from postmortem eye tissue from two MacTel donors, an RP donor and a control donor. MAIN OUTCOME MEASURES: Presence of specific immunohistochemistry markers on intraretinal pigmented and RPE cells. RESULTS: We found that intraretinal pigmented cells did not express RPE65 and CRALBP, with a small subset expressing them weakly. However, they all expressed KRT18, which was also present in normal RPE cells. Interestingly, we also found clusters of KRT18-positive cells in the retina that were not pigmented. CONCLUSIONS: Our findings suggest that RPE cells invading the retina dedifferentiate (losing classic RPE markers) and can be pigmented or unpigmented. Therefore, the number of RPE cells invading the retina in retinal degenerative disease may be underappreciated by funduscopy.

Description of a patient cohort with Hereditary Sensory Neuropathy type 1 without retinal disease Macular Telangiectasia type 2 - implications for retinal screening in HSN1.

Pathogenic variants in the genes encoding serine palmitoyl transferase (SPTLC1 or SPTLC2) are the most common causes of the rare peripheral nerve disorder Hereditary Sensory Neuropathy Type 1 (HSN1). Macular telangiectasia type 2 (MacTel), a retinal disorder associated with disordered serine-glycine metabolism, has been described in some patients with HSN1. This study aims to further investigate this association in a cohort of people with HSN1. Fourteen patients with a clinically and genetically confirmed diagnosis of HSN1 from the National Hospital for Neurology and Neurosurgery (NHNN, University College London Hospitals NHS Foundation Trust, London, United Kingdom) were recruited to the MacTel Registry, between July 2018 and April 2019. Two additional patients were identified from the dataset of the international clinical registry study (www.lmri.net). Ocular examination included fundus autofluorescence, blue light and infrared reflectance, macular pigment optical density mapping and optical coherence tomography. Twelve patients had a pathogenic variant in the SPTLC1 gene, with p.Cys133Trp in 11 cases (92%) and p.Cys133Tyr in one case (8%). Four patients had a variant in the SPTLC2 gene. None of the patients showed clinical evidence of MacTel. The link between HSN1 and MacTel seems more complex than can solely be explained by the genetic variants. An extension of the spectrum of SPTLC1/2-related disease with phenotypic pleiotropy is proposed. HSN1 patients should be screened for visual symptoms and referred for specialist retinal screening, but the association of the two diseases is likely to be variable and remains unexplained.

VEGFR1 signaling in retinal angiogenesis and microinflammation.

Five vascular endothelial growth factor receptor (VEGFR) ligands (VEGF-A, -B, -C, -D, and placental growth factor [PlGF]) constitute the VEGF family. VEGF-A binds VEGF receptors 1 and 2 (VEGFR1/2), whereas VEGF-B and PlGF only bind VEGFR1. Although much research has been conducted on VEGFR2 to elucidate its key role in retinal diseases, recent efforts have shown the importance and involvement of VEGFR1 and its family of ligands in angiogenesis, vascular permeability, and microinflammatory cascades within the retina. Expression of VEGFR1 depends on the microenvironment, is differentially regulated under hypoxic and inflammatory conditions, and it has been detected in retinal and choroidal endothelial cells, pericytes, retinal and choroidal mononuclear phagocytes (including microglia), Müller cells, photoreceptor cells, and the retinal pigment epithelium. Whilst the VEGF-A decoy function of VEGFR1 is well established, consequences of its direct signaling are less clear. VEGFR1 activation can affect vascular permeability and induce macrophage and microglia production of proinflammatory and proangiogenic mediators. However the ability of the VEGFR1 ligands (VEGF-A, PlGF, and VEGF-B) to compete against each other for receptor binding and to heterodimerize complicates our understanding of the relative contribution of VEGFR1 signaling alone toward the pathologic processes seen in diabetic retinopathy, retinal vascular occlusions, retinopathy of prematurity, and age-related macular degeneration. Clinically, anti-VEGF drugs have proven transformational in these pathologies and their impact on modulation of VEGFR1 signaling is still an opportunity-rich field for further research.

Synergistic effect of vascular endothelial growth factor gene inactivation in endothelial cells and skeletal myofibres on muscle enzyme activity, capillary supply and endurance exercise in mice.

NEW FINDINGS: What is the central question of this study? Does vascular endothelial growth factor (VEGF) expressed by both endothelial cells and skeletal myofibres maintain the number of skeletal muscle capillaries and regulate endurance exercise? What is the main finding and its importance? VEGF expressed by both endothelial cells and skeletal myofibres is not essential for maintaining capillary number but does contribute to exercise performance. ABSTRACT: Many chronic diseases lead to exercise intolerance, with loss of skeletal muscle capillaries. While many muscle cell types (myofibres, satellite cells, endothelial cells, macrophages and fibroblasts) express vascular endothelial growth factor (VEGF), most muscle VEGF is stored in myofibre vesicles which can release VEGF to signal VEGF receptor-expressing cells. VEGF gene ablation in myofibres or endothelial cells alone does not cause capillary regression. We hypothesized that simultaneously deleting the endothelial cell (EC) and skeletal myofibre (Skm) VEGF gene would cause capillary regression and impair exercise performance. This was tested in adult mice by simultaneous conditional deletion of the VEGF gene (Skm/EC-VEGF-/- mice) through the use of VEGFLoxP, HSA-Cre-ERT2 and PDGFb-iCre-ERT2 transgenes. These double-deletion mice were compared to three control groups - WT, EC VEGF gene deletion alone and myofibre VEGF gene deletion alone. Three weeks after initiating gene deletion, Skm/EC-VEGF-/- mice, but not SkmVEGF-/- or EC-VEGF-/- mice, reached exhaustion 40 min sooner than WT mice in treadmill tests (P = 0.002). WT, SkmVEGF-/- and EC-VEGF-/- , but not Skm/EC-VEGF-/- , mice gained weight over the 3 weeks. Capillary density, fibre area and capillary: fibre ratio in soleus, plantaris, gastrocnemius and cardiac papillary muscle were similar across the groups. Phosphofructokinase and pyruvate dehydrogenase activities increased only in Skm/EC-VEGF-/- mice. These data suggest that deletion of the VEGF gene simultaneously in endothelial cells and myofibres, while reducing treadmill endurance and despite compensatory augmentation of glycolysis, is not required for muscle capillary maintenance. Reduced endurance remains unexplained, but may possibly be related to a role for VEGF in controlling perfusion of contracting muscle.

Longitudinal Assessment of Remnant Foveal Cone Structure in a Case Series of Early Macular Telangiectasia Type 2.

Purpose: To determine the extent of remnant cone structure within early foveal ellipsoid zone (EZ) lesions in macular telangiectasia type 2 longitudinally using both confocal and split detector adaptive optics scanning light ophthalmoscopy (AOSLO). Methods: Spectral domain optical coherence tomography (SDOCT), confocal and split detector AOSLO were acquired from seven patients (10 eyes) with small (early) EZ lesions on SDOCT secondary to macular telangiectasia type 2 at baseline, 6 months, and 12 months. The presence of cone structure on AOSLO in areas of EZ loss as well as cones at 1° eccentricity, and their change over time were quantified. Results: By split detector AOSLO, remnant cone structure was identified within and on the borders of all foveal EZ lesions. Within the extent of these lesions, cone spacing ranged from 4.97 to 9.95 µm at baseline, 5.30 to 6.10 µm at 6 months, and 4.99 to 7.12 µm at 12 months. Four eyes with significantly smaller EZ lesions showed evidence of recovery of EZ reflectivity on SDOCT B-scans. Remnant cone structure was identified in some areas where EZ reflectivity recovered at the following time point. Eyes that showed recovery of EZ reflectivity had a continuous external limiting membrane. Conclusions: Remnant cone structure can persist within small SDOCT-defined EZ lesions, which can wax and wane in appearance over time. AOSLO can help to inform the interpretation of SDOCT imaging. Translational Relevance: The absence of EZ in early macular telangiectasia type 2 and other retinal conditions needs careful interpretation because it does not always indicate an absence of underlying cone structure. The integrity of the external limiting membrane may better predict the presence of remnant cone structure and recovery of EZ reflectivity.

Systemic lipid dysregulation is a risk factor for macular neurodegenerative disease.

Macular Telangiectasia type 2 (MacTel) is an uncommon bilateral retinal disease, in which glial cell and photoreceptor degeneration leads to central vision loss. The causative disease mechanism is largely unknown, and no treatment is currently available. A previous study found variants in genes associated with glycine-serine metabolism (PSPH, PHGDH and CPS1) to be associated with MacTel, and showed low levels of glycine and serine in the serum of MacTel patients. Recently, a causative role of deoxysphingolipids in MacTel disease has been established. However, little is known about possible other metabolic dysregulation. Here we used a global metabolomics platform in a case-control study to comprehensively profile serum from 60 MacTel patients and 58 controls. Analysis of the data, using innovative computational approaches, revealed a detailed, disease-associated metabolic profile with broad changes in multiple metabolic pathways. This included alterations in the levels of several metabolites that are directly or indirectly linked to glycine-serine metabolism, further validating our previous genetic findings. We also found changes unrelated to PSPH, PHGDH and CPS1 activity. Most pronounced, levels of several lipid groups were altered, with increased phosphatidylethanolamines being the most affected lipid group. Assessing correlations between different metabolites across our samples revealed putative functional connections. Correlations between phosphatidylethanolamines and sphingomyelin, and glycine-serine and sphingomyelin, observed in controls, were reduced in MacTel patients, suggesting metabolic re-wiring of sphingomyelin metabolism in MacTel patients. Our findings provide novel insights into metabolic changes associated with MacTel and implicate altered lipid metabolism as a contributor to this retinal neurodegenerative disease.

Macular Telangiectasia Type 2: Visual Acuity, Disease End Stage, and the MacTel Area: MacTel Project Report Number 8.

PURPOSE: To report the visual acuity measures from the macular telangiectasia type 2 (MacTel) registry and to investigate and describe phenotypic findings in eyes with substantial vision loss resulting from MacTel. DESIGN: Cross-sectional multicenter study. PARTICIPANTS: Participants in the MacTel Natural History Observation Registration Study. METHODS: Best-corrected visual acuity (BCVA) data, retinal imaging data, and clinical data were accessed from the MacTel Study databases in May 2019. MAIN OUTCOME MEASURES: Frequency distribution of BCVA and its relationship to age; morphologic changes in eyes with very late disease stages, defined by a BCVA of 20/200 or worse; average retinal thickness of macular subfields on OCT; and dimensions of the area affected by MacTel (i.e., the MacTel area). RESULTS: Best-corrected visual acuity was 20/50 or worse in 37.3% and 20/200 or worse in 3.8% of 4449 eyes of 2248 patients; 18.4% and 0.7% of all patients showed bilateral BCVA of 20/50 or worse and 20/200 or worse, respectively. Asymmetry between right and left eyes was present (median BCVA, 71 letters vs. 74 letters), a finding supported by more advanced morphologic changes in right eyes. Participant age correlated with BCVA, but the effect size was small. If a neovascularization or macular hole were present, bilateral occurrence was frequent (33% or 17%, respectively), and BCVA was better than 20/200 (79% or 78%, respectively) or 20/50 or better (26% or 13%, respectively). Eyes with advanced disease (BCVA, ≤20/200) showed the following characteristics: (1) atrophy of the foveal photoreceptor layer with or without associated subretinal fibrosis; (2) an affected area, termed MacTel area, limited to a horizontal diameter not exceeding the distance between the temporal optic disc margin and foveal center, and the vertical diameter not exceeding approximately 0.8 times this distance (exceptions were eyes with large active or inactive neovascular membranes); (3) reduced retinal thickness measures within the MacTel area; and (4) less frequent retinal greying and more frequent hyperpigmentations compared with eyes that have better BCVA. CONCLUSIONS: Severe vision loss is rare in MacTel and is related to photoreceptor atrophy in most people. Results indicate disease asymmetry with slightly worse vision and more advanced disease manifestation in right eyes. MacTel-related neurodegeneration does not spread beyond the limits of the MacTel area.

RhoJ integrates attractive and repulsive cues in directional migration of endothelial cells.

During angiogenesis, VEGF acts as an attractive cue for endothelial cells (ECs), while Sema3E mediates repulsive cues. Here, we show that the small GTPase RhoJ integrates these opposing signals in directional EC migration. In the GTP-bound state, RhoJ interacts with the cytoplasmic domain of PlexinD1. Upon Sema3E stimulation, RhoJ released from PlexinD1 induces cell contraction. PlexinD1-bound RhoJ further facilitates Sema3E-induced PlexinD1-VEGFR2 association, VEGFR2 transphosphorylation at Y1214, and p38 MAPK activation, leading to reverse EC migration. Upon VEGF stimulation, RhoJ is required for the formation of the holoreceptor complex comprising VEGFR2, PlexinD1, and neuropilin-1, thereby preventing degradation of internalized VEGFR2, prolonging downstream signal transductions via PLCγ, Erk, and Akt, and promoting forward EC migration. After conversion to the GDP-bound state, RhoJ shifts from PlexinD1 to VEGFR2, which then terminates the VEGFR2 signals. RhoJ deficiency in ECs efficiently suppressed aberrant angiogenesis in ischemic retina. These findings suggest that distinct Rho GTPases may act as context-dependent integrators of chemotactic cues in directional cell migration and may serve as candidate therapeutic targets to manipulate cell motility in disease or tissue regeneration.

Intravenous indocyanine green dye is insufficient for robust immune cell labelling in the human retina.

It is not currently possible to reliably visualise and track immune cells in the human central nervous system or eye. Previous work demonstrated that indocyanine green (ICG) dye could label immune cells and be imaged after a delay during disease in the mouse retina. We report a pilot study investigating if ICG can similarly label immune cells within the human retina. Twelve adult participants receiving ICG angiography as part of routine standard of care were recruited. Baseline retinal images were obtained prior to ICG administration then repeated over a period ranging from 2 hours to 9 days. Matched peripheral blood samples were obtained to examine systemic immune cell labelling and activation from ICG by flow cytometry with human macrophage cultures as positive controls. Differences between the delayed near infrared ICG imaging and 488 nm autofluorescence was observed across pathologies, likely arising from the retinal pigment epithelium (RPE). Only one subject demonstrated ICG signal on peripheral blood myeloid cells and only three distinct cell-sized signals appeared over time within the retina of three participants. No significant increase in immune cell activation markers were detected after ICG administration. ICG accumulated in the endosomes of macrophage cultures and was detectable above a minimum concentration, suggesting cell labelling is possible. ICG can label RPE and may be used as an additional biomarker for RPE health across a range of retinal disorders. Standard clinical doses of intravenous ICG do not lead to robust immune cell labelling in human blood or retina and further optimisation in dose and route are required.

Binocular Inhibition of Reading in Macular Telangiectasia Type 2.

Purpose: To assess the presence of binocular gain in macular telangiectasia type 2 (MacTel) and its correlation to paracentral scotomas. Methods: Sixty-eight patients with MacTel were consecutively recruited for a cross-sectional analysis. Best-corrected visual acuity (BCVA), reading acuity, and reading speed were tested monocularly and binocularly. Macular retinal sensitivity was examined with fundus-controlled perimetry (microperimetry). Scotomas were quantified by their size, their depth, and their proximity to the fovea. Results: Binocular reading speed and acuity were lower than monocular reading speed and acuity in the functionally better eye (142 vs. 159 words per minute and 0.43 vs. 0.28 log reading acuity determination, P < 0.001). Magnitude of binocular inhibition of reading speed was correlated to the degree of interocular functional difference (R2 = 0.61, P < 0.001). This correlation was not found for reading acuity or BCVA (R2 < 0.03). Binocular reading speed was negatively correlated to size of right and left eye scotomas, with bigger effect size for left eye scotomas. The magnitude of binocular inhibition was correlated to size of left eye scotomas, but not of right eye scotomas. When both eyes had similar scotoma characteristics, the right eye was more frequently the better reading eye. Conclusions: We provide evidence for the presence of binocular inhibition of reading performance in MacTel, likely due to binocular rivalry. This may result from the characteristic paracentral scotomas in noncorresponding retinal fields and, in particular, a disruptive projection of scotomas in reading direction arising from the left eyes. Patients may benefit from occluding one eye while reading.

Serine and Lipid Metabolism in Macular Disease and Peripheral Neuropathy.

BACKGROUND: Identifying mechanisms of diseases with complex inheritance patterns, such as macular telangiectasia type 2, is challenging. A link between macular telangiectasia type 2 and altered serine metabolism has been established previously. METHODS: Through exome sequence analysis of a patient with macular telangiectasia type 2 and his family members, we identified a variant in SPTLC1 encoding a subunit of serine palmitoyltransferase (SPT). Because mutations affecting SPT are known to cause hereditary sensory and autonomic neuropathy type 1 (HSAN1), we examined 10 additional persons with HSAN1 for ophthalmologic disease. We assayed serum amino acid and sphingoid base levels, including levels of deoxysphingolipids, in patients who had macular telangiectasia type 2 but did not have HSAN1 or pathogenic variants affecting SPT. We characterized mice with low serine levels and tested the effects of deoxysphingolipids on human retinal organoids. RESULTS: Two variants known to cause HSAN1 were identified as causal for macular telangiectasia type 2: of 11 patients with HSAN1, 9 also had macular telangiectasia type 2. Circulating deoxysphingolipid levels were 84.2% higher among 125 patients with macular telangiectasia type 2 who did not have pathogenic variants affecting SPT than among 94 unaffected controls. Deoxysphingolipid levels were negatively correlated with serine levels, which were 20.6% lower than among controls. Reduction of serine levels in mice led to increases in levels of retinal deoxysphingolipids and compromised visual function. Deoxysphingolipids caused photoreceptor-cell death in retinal organoids, but not in the presence of regulators of lipid metabolism. CONCLUSIONS: Elevated levels of atypical deoxysphingolipids, caused by variant SPTLC1 or SPTLC2 or by low serine levels, were risk factors for macular telangiectasia type 2, as well as for peripheral neuropathy. (Funded by the Lowy Medical Research Institute and others.).

High-Resolution In Vivo Fundus Angiography using a Nonadaptive Optics Imaging System.

PURPOSE: We provide a proof of concept for the detailed characterization of retinal capillary features and surrounding photoreceptor mosaic using a customized nonadaptive optics angiography imaging system. METHODS: High-resolution fluorescein angiography (FFA) and/or indocyanine green angiography (ICGA) images were obtained using a modified Heidelberg retina angiograph (HRA2) device with a reduced scan angle enabling 3° field of view. Colocalized images of the photoreceptor mosaic also were captured in vivo using the same instrument. Visibility of vascular subbranches were compared between high-resolution images and conventional fundus angiography (FA) with a 30° field of view. RESULTS: High-resolution angiographic and infrared images (3° × 3° field of view, a 10-fold magnification) were obtained in 10 participants. These included seven patients with various retinal diseases, including myopic degeneration, diabetic retinopathy, macular telangiectasia, and central serous chorioretinopathy, as well as three healthy controls. Images of the retinal vasculature down to the capillary level were obtained on angiography with the ability to visualize a mean 1.2 levels more subbranches compared to conventional FA. In addition, imaging of the photoreceptor cone mosaic, to a sufficient resolution to calculate cone density, was possible. Movement of blood cells within the vasculature also was discernible on infrared videography. CONCLUSIONS: This exploratory study demonstrates that fast high-resolution angiography and cone visualization is feasible using a commercially available imaging system. TRANSLATIONAL RELEVANCE: This offers potential to better understand the relationship between the retinal neurovascular system in health and disease and the timing of therapeutic interventions in disease states.

Dark-Adapted Two-Color Fundus-Controlled Perimetry in Macular Telangiectasia Type 2.

Purpose: Macular telangiectasia type 2 (MacTel) is a bilateral neurodegenerative disorder of the central macula. Previous findings indicated more functional impairment in low light conditions. We sought to further characterize retinal dysfunction using dark-adapted two-color fundus-controlled perimetry ("scotopic microperimetry"). Methods: Participants of the MacTel Natural History Observation Registry study and age-matched healthy controls underwent retinal imaging including dual wavelength autofluorescence macular pigment optical density (MPOD) measurement. Retinal sensitivity was assessed with scotopic microperimetry using cyan (505 nm) and red (627 nm). Disease was graded into classes of MPOD loss (0 to 3). For perimetry analysis, the differences of the mean sensitivities (MacTel minus controls) were compared at each test location and the results were aggregated to global indices. Results: Thirty-four eyes (19 patients, mean age 62.2 years) were compared with 25 eyes (25 controls, mean age 61.5 years). Both cyan and red sensitivity were lower in MacTel. This was more pronounced at one- and three-degree eccentricity. Eyes with MPOD class 0 did not exhibit a functional deficit. Class 1 had impaired cyan, but normal red sensitivity. Class 2 and 3 behaved similarly and had impaired cyan and red sensitivity with a relatively higher cyan impairment. Conclusions: Rods might be compromised to a greater extent than cones. Linking to previous studies, our results might also hint toward (postreceptoral) dysfunction of the cone system in very early disease stages. Macular pigment loss and global perimetry indices seemed to reflect functional impairment and might be useful as adjunct measures for disease progression.

p53 plays a crucial role in endothelial dysfunction associated with hyperglycemia and ischemia.

p53 is a guardian of the genome that protects against carcinogenesis. There is accumulating evidence that p53 is activated with aging. Such activation has been reported to contribute to various age-associated pathologies, but its role in vascular dysfunction is largely unknown. The aim of this study was to investigate whether activation of endothelial p53 has a pathological effect in relation to endothelial function. We established endothelial p53 loss-of-function and gain-of-function models by breeding endothelial-cell specific Cre mice with floxed Trp53 or floxed Mdm2/Mdm4 mice, respectively. Then we induced diabetes by injection of streptozotocin. In the diabetic state, endothelial p53 expression was markedly up-regulated and endothelium-dependent vasodilatation was significantly impaired. Impairment of vasodilatation was significantly ameliorated in endothelial p53 knockout (EC-p53 KO) mice, and deletion of endothelial p53 also significantly enhanced the induction of angiogenesis by ischemia. Conversely, activation of endothelial p53 by deleting Mdm2/Mdm4 reduced both endothelium-dependent vasodilatation and ischemia-induced angiogenesis. Introduction of p53 into human endothelial cells up-regulated the expression of phosphatase and tensin homolog (PTEN), thereby reducing phospho-eNOS levels. Consistent with these results, the beneficial impact of endothelial p53 deletion on endothelial function was attenuated in EC-p53 KO mice with an eNOS-deficient background. These results show that endothelial p53 negatively regulates endothelium-dependent vasodilatation and ischemia-induced angiogenesis, suggesting that inhibition of endothelial p53 could be a novel therapeutic target in patients with metabolic disorders.

Contrast sensitivity and visual acuity under low light conditions in macular telangiectasia type 2.

BACKGROUND/AIM: Macular pigment optical density (MPOD) is centrally depleted early on in macular telangiectasia type 2 (MacTel). Contrast sensitivity (CS) might be related to MPOD, and thus impaired in early MacTel. The effect of low luminance was assessed on both CS and best corrected visual acuity (BCVA). METHODS: This is a cross-sectional study. Pelli-Robson charts were used for CS testing at 1 m in photopic (110 lux) and mesopic (1 lux) conditions. BCVA was tested with ETDRS charts and low luminance visual acuity (LLVA) with a 2.0 log unit neutral density filter. MPOD was obtained with dual-wavelength autofluorescence. RESULTS: One hundred and three eyes of 52 patients with MacTel (mean±SD age 62.9±10.2, range 35-77) were compared with 34 healthy eyes of 17 controls (mean±SD age 65.2±7.4, range 53-78). CS was significantly lower in the eyes with MacTel. This impairment was higher in low light conditions (low light contrast sensitivity (LL-CS)). Eyes at the early stages of MacTel had significantly lower LL-CS than controls, but normal (photopic) CS. The results were similar but less pronounced for BCVA/LLVA. Decrease in CS was correlated with loss of MPOD. CONCLUSIONS: Low light conditions have a detrimental effect on visual performance in MacTel. Impaired CS might correlate with MPOD depletion as a pathognomonic finding in MacTel. Functional impairment might precede structural disintegration, indicating dysfunction at the cellular level. The applied tests might be useful as additional functional assessments in clinical routine and as outcome measures in future interventional clinical trials.