CD40 Expressed in Endothelial Cells Promotes Upregulation of ICAM-1 But Not Pro-Inflammatory Cytokines,NOS2 and P2X7 in the Diabetic Retina

PurposeCD40 is an upstream inducer of inflammation in the diabetic retina. CD40 is upregulated in retinal endothelial cells in diabetes. The purpose of this study was to determine whether expression of CD40 in endothelial cells is sufficient to promote inflammatory responses in the retina of diabetic mice.MethodsTransgenic mice with CD40 expression restricted to endothelial cells (Trg-CD40 EC), transgenic control mice (Trg-Ctr), B6, and CD40^−/− mice were made diabetic using streptozotocin. Leukostasis was assessed using FITC-conjugated ConA. Pro-inflammatory molecule expression was examined by real-time PCR, immunohistochemistry, ELISA, or flow cytometry. Release of ATP was assessed by ATP bioluminescence.ResultsDiabetic B6 and Trg-CD40 EC mice exhibited increased retinal mRNA levels of ICAM-1, higher ICAM-1 expression in endothelial cells, and increased leukostasis. These responses were not detected in diabetic mice that lacked CD40 (CD40^−/− and Trg-Ctr). Diabetic B6 but not Trg-CD40 EC mice upregulated TNF-α, IL-1β, and NOS2 mRNA levels. CD40 stimulation in retinal endothelial cells upregulated ICAM-1 but not TNF-α, IL-1β, or NOS2. CD40 ligation did not trigger ATP release by retinal endothelial cells or pro-inflammatory cytokine production in bystander myeloid cells. In contrast to diabetic B6 mice, diabetic Trg-CD40 EC mice did not upregulate P2X₇ mRNA levels in the retina.ConclusionsEndothelial cell CD40 promotes ICAM-1 upregulation and leukostasis. In contrast, endothelial cell CD40 does not lead to pro-inflammatory cytokine and NOS2 upregulation likely because it does not activate purinergic-mediated pro-inflammatory molecule expression by myeloid cells or induce expression of these pro-inflammatory molecules in endothelial cells.     

Retinal OFF-Pathway Overstimulation Leads to Greater Accommodation-Induced Choroidal Thinning

PurposeTo examine the interactions between accommodation and overstimulation of the retinal ON- and OFF-pathways, and their association with changes in choroidal thickness (ChT) and vascularity.MethodsOptical coherence tomography imaging of the choroid of twenty young adults (ages 25 ± 5 years) was performed before and after a series of 30-minute-long viewing tasks, including reading a bright text on dark background (ON-pathway overstimulation) and dark text on bright background (OFF-pathway overstimulation), and a control task of viewing a movie with unbiased ON-/OFF-pathway activation. The viewing tasks were performed with relaxed, and 5 diopter (D) accommodation (induced by soft contact lenses) demands. Both reading texts were matched for the mean luminance (35 cd/m²), luminance contrast (87%), and letter size (approximately 11.8 arc minutes). The change in ChT from baseline associated with contrast polarity and accommodation was examined using linear mixed model analysis.ResultsThe subfoveal ChT decreased significantly by −7 ± 1 µm with 5 D accommodation compared with relaxed accommodation (−3 ± 1 µm; P < 0.001), and by −9 ± 1 µm with OFF-pathway compared with ON-pathway overstimulation (−4 ± 1 µm; P = 0.002) and the control condition (−2 ± 1 µm; P < 0.001). Overstimulation of the OFF-pathway, but not the ON-pathway, resulted in a significantly greater choroidal thinning compared with the control condition, both at relaxed (−7 ± 1 µm; P = 0.003) and 5 D (−11 ± 1 µm; P = 0.005) accommodation levels. Similar changes were also observed for macular total, stromal, and luminal ChT.ConclusionsRetinal OFF-pathway stimulation enhanced the choroidal thinning associated with accommodation, thereby providing a potential mechanism that involves accommodation and the retinal OFF-signaling pathway, linking near work and myopia.     

Choroidal and Retinal Thickness and Axial Eye Elongation in Chinese Junior Students

PurposeTo explore the associations between macular choroidal and retinal thickness and axial elongation in non-myopic and myopic junior students.MethodsIn this school-based longitudinal observational study, axial length was measured by optical low-coherence reflectometry, and choroidal thickness and retinal thickness were measured by spectral-domain optical coherence tomography. Myopia was defined as non-cycloplegic objective spherical equivalent refraction ≤ −0.50 diopters. Structural equation modeling and multiple linear regression models were used to analyze the associations between baseline choroidal and retinal thickness with axial elongation.ResultsOut of 1307 students examined at baseline in 2017, 1197 (91.58%) returned for follow-up examination in 2018, with a median age of 12.00 years (interquartile range [IQR], 1.00) and included 667 boys (55.72%). Within a 1-year period, the median axial elongation of right eyes was 230 µm (IQR, 180) in boys and 200 µm (IQR, 160) in girls (P = 0.032). The thinner temporal choroidal thickness was associated with greater 1-year axial elongation only in myopic students (β, −0.20; 95% confidence interval [CI], −0.37, −0.03), the thinner temporal retinal thickness was associated with greater 1-year axial elongation in both non-myopic (β, −2.67; 95% CI, −4.52, −0.82) and myopic (β, −0.99; 95% CI, −1.68, −0.30) students, after adjustment for sex, age, and height. Subfoveal and nasal choroidal and retinal thickness were not significantly associated with axial elongation in either non-myopic or myopic students.ConclusionsA thinner temporal choroid at age 12 years may predict greater 1-year axial elongation in myopic students, and a thinner temporal retina may predict greater 1-year axial elongation in both non-myopic and myopic students. This finding may help to identify children at risk and control axial elongation with potential preventive strategies.     

Myeloid Cells in the Mouse Retina and Uveal Tract Respond Differently to Systemic Inflammatory Stimuli

PurposeIn spite of clear differences in tissue function and significance to ocular disease, little is known about how immune responses differ between the retina and uveal tract. To this end we compared the effects of acute systemic inflammation on myeloid cells within the mouse retina, iris-ciliary body, and choroid.MethodsSystemic inflammation was induced in Cx3cr1^gfp/gfp and CD11c-eYFP Crb1^wt/wtmice by intraperitoneal lipopolysaccharide (LPS). In vivo fundus imaging was performed at two, 24, and 48 hours after LPS, and ocular tissue wholemounts were immunostained and studied by confocal microscopy. Flow cytometry was used to investigate the expression of activation markers (MHC class II, CD80, CD86) on myeloid cell populations at 24 hours. For functional studies, retinal microglia were isolated from LPS-exposed mice and cocultured with naïve OT-II CD4+ T-cells and ovalbumin peptide. T-cell proliferation was measured by flow cytometry and cytokine assays.ResultsSystemic LPS altered the density and morphology of retinal microglia; however, retinal microglia did not upregulate antigen presentation markers and failed to stimulate naïve CD4+ T-cell proliferation in vitro. In contrast, uveal tract myeloid cells displayed a phenotype consistent with late-activated antigen-presenting cells at 24 hours. Systemic LPS induced remodeling of myeloid populations within the uveal tract, particularly in the choroid, where dendritic cells were partially displaced by macrophages at 24 hours.ConclusionsThe disparate myeloid cell responses in the retina and uveal tract after systemic LPS highlight differential regulation of innate immunity within these tissue environments, observations that underpin and advance our understanding of ocular immune privilege.     

Evidence of hematopoietic differentiation, vasculogenesis and angiogenesis in the formation of human choroidal blood vessels

Human fetal eyes 8–40 weeks gestation (WG) were examined using markers to hematopoietic stem cells (HSC), vascular precursor cells (VPC), monocytes/macrophages and endothelial cells (EC). Electron microscopy and bromo-deoxyuridene labeling were undertaken to confirm the existence of solid vascular cords and to demonstrate vasculogenesis and angiogenesis in developing choroidal tissue. Our results demonstrated that the earliest incipient choroid consisted of vimentin⁺ mesenchymal precursor cells which downregulated vimentin expression with maturation. Our observations lead us to conclude that these vimentin⁻/CD34⁺/CD44⁺/CD133⁺ HSCs then differentiated into three distinct lineages: single isolated CD34⁻/CD39⁺ VPCs that formed solid vascular cords which lumenized and became lined with CD34⁺ vascular ECs; CD34⁻⁻⁺/CD14⁺/CD68⁺ monocytes that differentiated into tissue macrophages; and CD133⁺/CD34⁻⁻⁺/α-smooth muscle actin⁺ mural precursor cells that matured into smooth muscle cells and pericytes. Blood vessel formation occurred throughout the whole choroid simultaneously, indicative of in situ differentiation. Vasculogenesis, as evidenced by lumenization of solid vascular cords, was responsible for the formation of the entire choroidal area with angiogenesis, in all three layers of the choroid, only adding to vascular density. These results suggest that formation of the human choroid involves three processes: HSC differentiation, vasculogenesis and angiogenesis. Since vasculogenesis takes place independently of VEGF₁₆₅, further insights regarding the molecular mechanisms of vasculogenesis are required to better inform future treatments of choroidal neovascularization.     

Corneal endothelial cell density and microvascular changes of retina and optic disc in autosomal dominant polycystic kidney disease

Purpose:Vascular endothelial dysfunction in autosomal dominant polycystic kidney disease (ADPKD) may affect the retinal vascular parameters due to structural similarities of kidney and retina. We aimed to evaluate the microvascular changes of retina and optic disc and also corneal endothelial cell density in patients with ADPKD.Methods:Forty-six eyes of 23 patients with ADPKD (Group 1), and 46 eyes of 23 sex- and age-matched healthy controls (Group 2) were included in this cross-sectional study. Demographic and ophthalmic findings of participants were collected. Corneal endothelial cell density (CECD) measurements were obtained by noncontact specular microscopy. Foveal retinal thickness, peripapillary retinal nerve fiber layer (RNFL) thickness, vessel density in different sections of the retina and optic nerve head were analyzed by optical coherence tomography angiography.Results:The mean ages were 41 ± 11 years for Group 1 and 39 ± 10 years for Group 2 (P = 0.313). CECD values were significantly lower in group 1 when compared to group 2 (2653 ± 306 cells/mm² and 2864 ± 244 cells/mm², respectively, P < 0.001). The foveal retinal thickness and RNFL thickness were similar, but superior quadrant thickness of RNFL was significantly lower in Group 1 than Group 2 (126 ± 14 μm vs. 135 ± 15 μm, P = 0.003). In Group 1, whole image of optic disc radial peripapillary capillary densities were significantly lower compared to Group 2 (49.4 ± 2.04%, and 50.0 ± 2.2%, respectively, P = 0.043). There was no significant difference regarding superficial, deep retinal vessel densities, foveal avascular zone and flow areas between the groups (P > 0.05 for all).Conclusion:Lower CECD values and decreased superior quadrant RNFL thickness, and microvascular densities of optic disc were revealed in patients with ADPKD. Evaluation of CECD and retinal microvasculature may be helpful in the management of these patients.     

Choroidal Changes in Eyes With Polypoidal Choroidal Vasculopathy After Anti-VEGF Therapy Imaged With Swept-Source OCT Angiography

PurposeSwept-source optical coherence tomography angiography was used to investigate choroidal changes and their association with pigment epithelial detachments (PEDs) in eyes with polypoidal choroidal vasculopathy (PCV) after treatment with vascular endothelial growth factor (VEGF) inhibitors.MethodsPatients with treatment-naïve PCV were included and underwent anti-VEGF therapy. Mean choroidal thickness (MCT), choroidal vascularity index (CVI), and PED volume measurements were obtained before and after treatment.ResultsThirty-four treatment-naïve PCV eyes from 33 patients were included. The PED volume decreased after treatment (P < 0.05). The MCT decreased from 223.0 ± 79.6 µm at baseline to 210.9 ± 76.2 µm after treatment (P < 0.001). The CVI at baseline was 0.599 ± 0.024, and the CVI after treatment was 0.602 ± 0.023 (P = 0.16). There was a correlation between the decreased PED volumes and the decreased MCT measurements (r = 0.47; P = 0.006). Also, there was a correlation between the decreased PED volumes and the increased CVI measurements (r = −0.63; P < 0.001).ConclusionsIn treatment-naïve eyes with PCV, the decreases in PED volumes were correlated with the decrease in MCT and the increase in CVI measurements. We propose that, at baseline, the PCV lesions serve as high-volume arteriovenous shunts between choroidal arterial and venous circulation, causing transudation into the choroidal stroma. We propose that, after treatment, the blood flow through the vascular shunt is reduced, the excess stromal transudation is resorbed, and the exudation from the neovascular lesion is reduced, resulting in thinning of the choroid, resolution of the PEDs, and an increase in the CVI due to the resorption of excess choroidal transudation.     

Evaluation of the Effect of a Novel β3-Adrenergic Agonist on Choroidal Vascularity

PurposeTo determine the effect of the new β₃-agonist (mirabegron), which is used for overactive bladder (OAB) treatment, on central retinal thickness (CRT) and choroidal vascularity.Material and MethodsThe 26 eyes of 26 cases using 50 mg tablet mirabegron once per day for OAB were included in this prospective case control study. The CRT, choroidal thickness (ChT), and choroidal vascularity were measured at baseline, week 1 (W1), month 1 (M1), month 2 (M2), and month 3 (M3). Subfoveal ChT measurement included the total subfoveal choroidal thickness (SFCT), and the small and large choroidal vessel layer (SCVL and LCVL) thickness. The total choroidal area (TCA), lumen area (LA), stromal area (SA), stroma/lumen ratio, and choroidal vascularity index (CVI) were measured with the Image-J software.ResultsThe largest SFCT increase compared to baseline was at M1 (26.8 ± 40.8 µm, P = 0.001). The subfoveal SCVL thickness showed a significant decrease at M2 and M3 (−6.0 ± 8.9 µm, P = 0.002; −7.8 ± 13.4 µm, P = 0.046, respectively). LCVL thickness showed a significant increase at W1, M1, and M2, with the largest at M1. CVI showed a significant increase at M1, M2, and M3 (P < 0.05 for all). The TCA, LA, and SA showed a significant increasing trend at all follow-up periods. LA/SA decreased at W1 because of stromal expansion but increased at M3 with more prominent vascular dilatation. CRT values showed no significant change.ConclusionsMirabegron had a significant effect on choroidal thickness. Choroidal vascular response is in the form of narrowing in the choriocapillaris and enlargement in the Haller''s layer.     

Differences in Retinal and Choroidal Vasculature and Perfusion Related to Axial Length in Pediatric Anisomyopes

PurposeThe purpose of this study was to evaluate the interocular differences in choroidal vasculature, choriocapillaris perfusion, and retinal microvascular network, and to explore their associations with interocular asymmetry in axial lengths (ALs) in children with anisomyopia.MethodsRefractive error, AL, and other biometric parameters were measured in 70 children with anisomyopia. Using optical coherence tomography (OCT) and OCT-angiography, we measured the submacular choroidal thickness (ChT), total choroidal area (TCA), luminal area (LA), stromal area (SA), choroidal vascularity index (CVI), choriocapillaris flow deficit (CcFD), retinal vessel density (VD), and foveal avascular zone (FAZ) area.ResultsThe mean interocular differences in spherical equivalent refraction and AL were −2.26 ± 0.94 diopters and 0.95 ± 0.46 mm, respectively. Submacular ChT, TCA, LA, SA, and CVI were all significantly lower in the more myopic (longer AL) eyes than in the less myopic (shorter AL) fellow eyes. In eyes with longer ALs, both the CcFD and FAZ areas were significantly greater, whereas the superficial and deep retinal VDs were significantly less. After adjusting for corneal power and intraocular pressure, interocular differences in LA (β = −0.774), SA (β = −0.991), and CcFD (β = 0.040) were significantly associated with interocular asymmetry in AL (all P < 0.05).ConclusionsIn pediatric anisomyopes, eyes with longer ALs tended to have lower choroidal vascularity and choriocapillaris perfusion than the contralateral eyes with shorter ALs. Longitudinal investigations would be useful follow-ups to test for a causal role of choroidal circulation in human myopia.     

Choroidal Thickness and Its Association With Age,Axial Length,and Refractive Error in Chinese Adults

PurposeTo identify the association between the choroidal thickness (ChT) with age and axial length (AL) under different refractive errors (REs) in Chinese adults.MethodsSwept-source optical coherence tomography was used to measure ChT in 2126 right eyes of 2126 participants. The participants were classified as having pathologic myopia (PM), high myopia without PM (HM), low myopia (LM), and nonmyopia (non-M) according to their REs and META-PM (the Meta-Analysis of Pathologic Myopia) classification criteria.ResultsThe mean age was 52.49 ± 20.39 years (range, 18−93 years), and the mean RE was −5.27 ± 5.37 diopters (D; range, −25.5 to +7.75 D). The mean average ChT was 159.25 ± 80.75 µm and decreased in a linear relationship from non-M to PM (190.04 ± 72.64 µm to 60.99 ± 37.58 µm, P < 0.001). A significant decline in ChT was noted between 50 and 70 years (r = −0.302, P < 0.001) and less rapidly after the age of 70 years (r = −0.105, P = 0.024). No correlation was noted between age and ChT under 50 years (P = 0.260). A significantly higher association with AL was noted in the central fovea (β^HM = −23.92, β^LM = −23.88, β^Non-M = −18.80, all P < 0.001) and parafoveal ChT (β^HM = −22.87, β^LM = −22.31, β^Non-M = −18.61, all P < 0.001) when compared with the perifoveal region (β^HM = −19.80, β^LM = −18.29, β^Non-M = −13.95, all P < 0.001). Within each group of PM, HM, LM, and non-M, regression analysis showed that the coefficients of age and AL with different macular regions of ChT varied significantly.ConclusionsChT was negatively correlated with age after 50 years. The thinning of the choroid was more prominent in the center and parafoveal regions as AL increased. Varied distributions of ChT decrease associated with AL and age were noted among different refractive groups.     

Low nitric oxide synthases (NOSs) in eyes with age-related macular degeneration (AMD)

Nitric oxide (NO) production by vascular endothelium is important in regulation of blood flow. Reduced production of NO can adversely affect blood flow and other vascular functions. We investigated the expression of three nitric oxide synthase (NOS) isoforms in retina and choroid of aged human eyes and eyes with AMD. Alkaline phosphatase immunohistochemistry was performed using antibodies against inducible (iNOS), neuronal (nNOS), and endothelial (eNOS) NOSs on cryopreserved sections from aged control donor eyes (n = 13) and eyes with AMD (n = 22). CD34 antibody was used as an endothelial cell (EC) marker. Three independent masked observers scored the intensity of the immunohistochemical reaction product. Mean scores from the aged control and AMD eyes were statistically compared. In aged control retinas, nNOS was in ganglion cells (RGCs) and neurons of both nuclear layers. In choroid, perivascular nerve fibers and retinal pigment epithelial (RPE) cells were nNOS⁺. eNOS and iNOS were confined to the retinal and choroidal vascular ECs. Some cells presumably melanocytes or dendritic cells in choroid were also eNOS⁺. In AMD eyes, nNOS was significantly lower in RGCs, neurons, retinal vessels and RPE (p ≤ 0.05) compared to the aged control eyes. iNOS and eNOS showed no significant differences between aged control and AMD eyes except that there was significantly less eNOS in choroidal arteries (p = 0.006) and choroidal cells (p = 0.03) of AMD eyes. Although NO was not measured directly, these findings suggest that there is less NO produced in AMD eyes. The decrease in retinal nNOS in AMD eyes is probably related to neuronal degeneration. The decrease in nNOS and eNOS in AMD choroid could be associated with vasoconstriction and hemodynamic changes.     

Semaphorin 3fa Controls Ocular Vascularization From the Embryo Through to the Adult

PurposePathological blood vessel growth in the eye is implicated in several diseases that result in vision loss, including age-related macular degeneration and diabetic retinopathy. The limits of current disease therapies have created the need to identify and characterize new antiangiogenic drugs. Here, we identify the secreted chemorepellent semaphorin-3fa (Sema3fa) as an endogenous anti-angiogenic in the eye.MethodsWe generated a CRISPR/Cas9 sema3fa zebrafish mutant line, sema3fa^ca304/304. We assessed the retinal and choroidal vasculature in both larval and adult wild-type and sema3fa mutant zebrafish.ResultsWe find sema3fa mRNA is expressed by the ciliary marginal zone, neural retina, and retinal pigment epithelium of zebrafish larvae as choroidal vascularization emerges and the hyaloid/retinal vasculature is remodeled. The hyaloid vessels of sema3fa mutants develop appropriately but fail to remodel during the larval period, with adult mutants exhibiting a denser network of capillaries in the retinal periphery than seen in wild-type. The choroid vasculature is also defective in that it develops precociously, and aberrant, leaky sprouts are present in the normally avascular outer retina of both sema3fa^ca304/304 larvae and adult fish.ConclusionsSema3fa is a key endogenous signal for maintaining an avascular retina and preventing pathologic vascularization. Furthermore, we provide a new experimentally accessible model for studying choroid neovascularization (CNV) resulting from primary changes in the retinal environment that lead to downstream vessel infiltration.     

Choroidal and Retinal Changes After Systemic Adrenaline and Photodynamic Therapy in Non-Human Primates

PurposeTo determine the tomographic, angiographic, and histologic changes in the choroid and retina of cynomolgus monkeys after systemic adrenaline and verteporfin photodynamic therapy (vPDT).MethodsSix cynomolgus monkeys (12 eyes) were treated with vPDT only (n = 2), adrenaline only for eight weeks (n = 2), adrenaline for eight weeks with vPDT at week 4 (n = 4), and adrenaline for 12 weeks and vPDT at week 8 (n = 4). Spectral-domain optical coherence tomography, angiography, and autofluorescence were performed at baseline and every 14 days thereafter until 28 days after adrenaline therapy or vPDT. Choroid parameters included choroidal thickness (CT) changes and structural changes using semiautomated image binarization. Histology with light and electron microscopy was performed.ResultsAdrenaline resulted in subfoveal CT increase at week 4 compared with baseline (3.4%, P = 0.010), with further increase at week 8 (3.9%, P = 0.007). This correlated with choroidal luminal area increase (16.0% at week 8 compared with baseline, P = 0.030). Outer retinal changes included subretinal fluid, ellipsoid zone (EZ) disruption, photoreceptor elongation, and sub/intraretinal bright dots. Hypocyanescent spots surrounded by leakage was observed. Histology showed dilated choroidal vessels, intracytoplasmic vacuoles, and retinal pigment epithelium (RPE) enlarged basal infoldings. The vPDT decreased subfoveal CT at four weeks after vPDT (−7.5%, P = 0.007). This correlated with choroidal stromal area decrease (−18.0%, P < 0.010). Within the treatment spot, there was outer retinal atrophy, EZ disruption, irregular RPE thickening, intense hypoautofluorescence, hyperfluorescence, and hypocyanescence. On histology, there were outer retina, RPE, and choroid changes.ConclusionsAdrenaline induces choroidal vessel dilation and CT increase. The vPDT decreases CT because of a reduction in choroidal stromal component.     

Retinal VEGF-A Overexpression Is Not Sufficient to Induce Lymphangiogenesis Regardless of VEGF-C Upregulation and Lyve1+ Macrophage Infiltration

PurposeNo lymphatic vessels have been identified in the retina. This study investigated whether pathological VEGF-A–overexpressing diabetic retina causes lymphangiogenesis.MethodsThree genetic mouse models of diabetic retinopathy (DR) (Akita [Ins2^+/−], Kimba [vegfa^+/+], and Akimba [Akita × Kimba] mice) were used. Retinas were examined by fundus photography, fluorescence angiography (FA), and immunostaining to detect lymphangiogenesis or angiogenesis. Lyve1-GFP (Lyve1^EGFP/Cre) mice were used to examine Lyve1-expressing cells by immunostaining. Lymphatic-related factors were investigated in mouse retina and vitreous fluid from proliferative diabetic retinopathy (PDR) patients by RT-PCR and ELISA, respectively. Aged Kimba and Akimba mice were used to examine the retinal phenotype at the late phase of VEGF overexpression.ResultsFA and immunostaining showed retinal neovascularization in Kimba and Akimba mice but not wild-type and Akita mice. Immunohistochemistry showed that lymphangiogenesis was not present in the retinas of Akita, Kimba, or Akimba mice despite the significant upregulation of lymphatic-related factors (Lyve1, podoplanin, VEGF-A, VEGF-C, VEGF-D, VEGFR2, and VEGFR3) in the retinas of Kimba and Akimba mice by RT-PCR (P < 0.005). Furthermore, lymphangiogenesis was not present in aged Kimba or Akimba mice. Significantly increased numbers of Lyve1-positive cells present in the retinas of Kimba and Akimba mice, especially in the peripheral areas, were CD11b positive, indicating a macrophage population (P < 0.005). VEGF-C in PDR vitreous with vitreous hemorrhage (VH) was higher than in PDR without VH or a macular hole.ConclusionsRetinal VEGF-A overexpression did not cause typical lymphangiogenesis despite upregulated lymphatic-related factors and significant Lyve1-positive macrophage infiltration.     

Subcellular Comparison of Visible-Light Optical Coherence Tomography and Electron Microscopy in the Mouse Outer Retina

PurposeWe employed in vivo, 1.0-µm axial resolution visible-light optical coherence tomography (OCT) and ex vivo electron microscopy (EM) to investigate three subcellular features in the mouse outer retina: reflectivity oscillations inner to band 1 (study 1); hyperreflective band 2, attributed to the ellipsoid zone or inner segment/outer segment (IS/OS) junction (study 2); and the hyperreflective retinal pigment epithelium (RPE) within band 4 (study 3).MethodsPigmented (C57BL/6J, n = 10) and albino (BALB/cJ, n = 3) mice were imaged in vivo. Enucleated eyes were processed for light and electron microscopy. Using well-accepted reference surfaces, we compared micrometer-scale axial reflectivity of visible-light OCT with subcellular organization, as revealed by 9449 annotated EM organelles and features across four pigmented eyes.ResultsIn study 1, outer nuclear layer reflectivity peaks coincided with valleys in heterochromatin clump density (−0.34 ± 2.27 µm limits of agreement [LoA]). In study 2, band 2 depth on OCT and IS/OS junction depth on EM agreed (−0.57 ± 0.76 µm LoA), with both having similar distributions. In study 3, RPE electron dense organelle distribution did not agree with reflectivity in C57BL/6J mice, with OCT measures of RPE thickness exceeding those of EM (2.09 ± 0.89 µm LoA). Finally, RPE thickness increased with age in pigmented mice (slope = 0.056 µm/mo; P = 6.8 × 10⁻⁷).ConclusionsVisible-light OCT bands arise from subcellular organization, enabling new measurements in mice. Quantitative OCT–EM comparisons may be confounded by hydration level, particularly in the OS and RPE. Caution is warranted in generalizing results to other species.     

Retinal and choroidal vascular changes in newly diagnosed celiac disease: An optical coherence tomography angiography study

Purpose:To investigate the retinal and choroidal microcirculation changes in celiac disease (CD) patients via optical coherence tomography angiography (OCT-A).Methods:This cross-sectional study included 44 pediatric patients with newly diagnosed CD and 44 healthy pediatric subjects. The vascular densities (VD) of the superficial, deep, and choriocapillar plexuses (VDs, VDd, and VDcc, respectively) (%), the superficial and deep foveal avascular zones (FAZs and FAZd) (%), the central macular thickness (CMT) (mm), and the subfoveal choroidal thickness (SFCT) (mm) were measured with swept-source OCT-A in addition to a complete ophthalmological examination.Results:Mean ages of the CD patients and the healthy participants were 12.02 ± 2.9 and 13.6 ± 2.3 years, respectively. The central sectors of the VDs and VDd measurements were found to be significantly higher in the study group compared to the control group (p = 0.006; P = 0.001, respectively), and the temporal and nasal values of the VDcc measurements were significantly lower in the study group than in the control group (p < 0.05 for both values). CMT and FAZ metrics did not differ between the groups (p > 0.05). SFCT was significantly reduced (p = 0.001), and choroidal thinning was more considerable in female CD patients (p = 0.045).Conclusion:CD seems to affect macular and choroidal microcirculation. The reduced choriocapillaris plexus parameters and choroidal thickness may provide disease activity information.     

Diabetic Macular Ischemia: Influence of Optical Coherence Tomography Angiography Parameters on Changes in Functional Outcomes Over One Year

PurposeTo prospectively evaluate whether diabetic macular ischemia detected with coherence tomography angiography (OCTA) is associated with change in functional outcomes over a period of one year.MethodsThis is a one-year prospective, observational study that included 56 eyes with varying levels of diabetic retinopathy. All participants underwent best corrected visual acuity evaluation, swept-source OCTA and microperimetry at baseline and repeated at one year. Parafoveal vessel densities (VD) and foveal avascular zone (FAZ) areas were generated from OCTA in the superficial and deep vascular plexuses. The influence of baseline and change in OCTA parameters on change in visual acuity and retinal sensitivity over one year was evaluated.ResultsOver the one-year follow-up period, 16% (9) of eyes had at least one line worsening in BCVA and 7% (4) of eyes had at least 5% decrease in retinal sensitivity compared to baseline. Diabetic retinopathy progressed in 12.5%. Mean superficial vascular plexus (SVP) FAZ area increased (0.32 ± 0.15 to 0.39 ± 0.18 mm², P = 0.003) and parafoveal VD in deep vascular plexus (DVP) decreased (49.8 ± 3.7% to 48.8 ± 2.9%, P = 0.040) at one year compared to baseline. In the multivariate regression analysis, larger baseline DVP FAZ area was associated with worsening of BCVA over one year (β = 0.16 logMAR per mm², 95% CI 0.02 to 0.31, P = 0.032). In addition, larger decreases in SVP VD (β = −4.18 db per 10% decrease, 95% CI −6.55 to −1.80, P = 0.002) was associated with worsening of retinal sensitivity over one year.ConclusionsProgression of parafoveal microvasculature changes over one year can be detected using OCTA. Larger baseline DVP FAZ area on OCTA is predictive of worsening in visual outcomes, and larger decreases in SVP VD were associated with worsening of retinal sensitivity over a course of one year in diabetic individuals.     

Correlation between retinal sensitivity and cystoid space characteristics in diabetic macular edema

Purpose:To evaluate the correlation between retinal sensitivity and cystoid space characteristics in eyes with diabetic macular edema (DME).Results:Subject''s mean age was 57 ± 9 years. The mean logarithm of minimum angle of resolution BCVA was 0.4 ± 0.2. The intraclass correlation coefficient for inter- and intra-grader assessment of cystoid space volume by manual delineation was 0.99 and 0.99, respectively. Mean total ICS volume was 0.4 ± 0.4 mm³ and for the foveal center, subfield was 0.1 ± 0.1 mm³. Mean retinal sensitivity was 12.89 ± 10 dB; however, foveal retinal sensitivity was 12.3 ± 11.1 dB. We found no significant correlation between BCVA and total cystoid space volume (r = 0.33, P = 0.06). Correlation between total retinal sensitivity and total ICS was negative and nonsignificant (r = −0.17, P = 0.36). Correlation between foveal retinal sensitivity and foveal cystoid space intensity was moderate and marginally significant (r = −0.43, P = 0.05).Conclusion:Total cystoid space volume was not significantly correlated with BCVA or total retinal sensitivity in subjects with DME. Foveal cystoid space optical intensity was negatively correlated with foveal retinal sensitivity. These findings suggest further investigation of cystoid space characteristics in the setting of DME may be of value.     

Para-inflammation in the aging retina

Para-inflammation is a tissue adaptive response to noxious stress or malfunction and has characteristics that are intermediate between basal and inflammatory states (Medzhitov, 2008). The physiological purpose of para-inflammation is to restore tissue functionality and homeostasis. Para-inflammation may become chronic or turn into inflammation if tissue stress or malfunction persists for a sustained period. Chronic para-inflammation contributes to the initiation and progression of many human diseases including obesity, type 2 diabetes, atherosclerosis, and age-related neurodegenerative diseases. Evidence from our studies and the studies of some others suggests that para-inflammation also exists in the aging retina in physiological conditions and might contribute to age-related retinal pathologies. The purpose of this review is to introduce the notion of “para-inflammation” as a state between frank, overt destructive inflammation and the non-inflammatory removal of dead or dying cells by apoptosis, to the retinal community.In diabetes and atherosclerosis, leukocytes particularly monocytes and vascular endothelial cells are constantly under noxious stress due to glycaemic and/or lipidaemic dysregulation. These blood-borne stresses trigger para-inflammatory responses in leukocytes and endothelial cells by up-regulating the expression of adhesion molecules or releasing cytokines/chemokines, which in turn cause abnormal leukocyte–endothelial interactions and ultimately vascular damage. In the aging retina, on the other hand, oxidized lipoproteins and free radicals are considered to be major causes of tissue stress and serve as local triggers for retinal para-inflammation. Microarray analysis has revealed the up-regulation of a large number of inflammatory genes, including genes involved in complement activation and inflammatory cytokine/chemokine production, in the aging retina. Para-inflammatory responses in the neuroretina of aged mice are characterized by microglial activation and subretinal migration, and breakdown of blood–retinal barrier. At the retinal/choroidal interface para-inflammation is manifested by complement activation in Bruch's membrane and RPE cells, and microglia accumulation in subretinal space. With age, para-inflammatory changes have also been observed in the choroidal tissue, evidenced by 1) increased thickness of choroid; 2) increased number of CD45⁺CRIg⁺ macrophages; 3) morphological abnormalities in choroidal melanocytes; and 4) fibrosis in choroidal tissue. An increased knowledge of contribution of retinal para-inflammation to various pathological conditions is essential for the better understanding of the pathogenesis of various age-related retinal diseases including diabetic retinopathy, glaucoma and age-related macular degeneration.     

Choroidal vascularity index as an indicator of vascular status of choroid,in eyes with nanophthalmos

PurposeTo investigate choroidal vascular index (CVI) in eyes with nanophthalmos (NO) with the use of optical coherence tomography (OCT).MethodsMacular enhanced depth imaging OCT scans of 25 eyes of 25 patients with NO and age–gender-matched 25 eyes of 25 control subjects were analysed. Images were binarized using the ImageJ software, and total choroid area (TCA), luminal area (LA) and stromal area (SA) were acquired. The main outcome measure was CVI, defined as the ratio of LA to TCA.ResultsTwenty-five eyes of 25 patients with NO and age–gender-matched control subjects were enrolled. The mean TCA, SA and LA were found to be significantly higher in patients with NO (2.51 ± 0.44 vs. 1.91 ± 0.35 mm², P < 0.001; 0.86 ± 0.17 vs. 0.63 ± 0.13 mm², P < 0.001; and 1.65 ± 0.29 vs. 1.27 ± 0.23 mm², P < 0.000, respectively). On the contrary, CVI did not significantly differ between the two groups (65.72, 67.68, P = 0.099).ConclusionAs a novel OCT-based marker, CVI could be used to assess vascular status of the choroid in eyes with NO and can provide better understanding of the pathogenesis of this disease.Subject terms: Eye abnormalities, Scleral diseases, Predictive markers, Hereditary eye disease, Uveal diseases