Retinal Blood Oxygen Saturation and Angiogenic and Inflammatory Biomarkers in Type 2 Diabetes

Introduction: Diabetic retinopathy (DR) is a major source of visual loss in the world, including North America. A number of hyperglycemia-related pathways have been associated with DR onset and progression. Disturbances in the retinal vasculature appear to play a vital role in DR, resulting in biochemical and functional vascular changes. Therefore, this study investigated retinal blood oxygen saturation and angiogenic and inflammatory biomarkers in DR. Methods: Chapter 3 and 5: FD-OCT Doppler blood flow was non-invasively measured using a prototype system based on the RTVue (Optovue Inc., USA). A minimum of six separate FD-OCT Doppler measurements was acquired. Chapter 4, 5, 7: Non-invasive hyperspectral retinal (HR) imaging was acquired in participants with mild-to-moderate NPDR and age-matched controls. For each subject, six repeated HRC images were acquired at wavelengths of 586 and 605nm. Results: Chapter3: The individual COV medians for retinal blood flow were 7.5% and 9.2% for young and elderly subjects, respectively. The group mean CORs for retinal blood flow for young participants were 6.4µl/min and for elderly subjects were 10.5µl/min. Chapter 4: Retinal blood oxygen saturation in the arterioles of healthy controls was 92.97±1.6%, and in the venules was 55.90±4.8%. Retinal blood oxygen saturation for diabetic subjects with NPDR was significantly higher at 94.65±2.2% (p=0.015) in the arterioles and 64.13±4.3% (p<0.001) in the venules. Chapter 5: Total retinal blood flow was significantly lower in NPDR when compared to controls (42.66 vs 32.97; p=0.004). There was no relationship between total retinal blood flow and venular oxygen saturation (r=0.2). Chapter 6: Angiopoietin 2, IL-8, HGF was significantly higher in NPDR patients than in control patients (p=0.005, p=0.034, p=0.018, respectively) and EGF was significantly lower in NPDR patients when compared to controls (p=0.025). Chapter 7: The study demonstrated a correlation between retinal blood oxygen saturation and Ang 2, HGF and EGF but did not find any correlation for IL-8, TGF-β even though these biomarkers were significantly higher in the diabetic group. Conclusions: Chapter 3: Doppler OCT gave consistent and repeatable blood flow measurements within retinal venules in normal subjects. Chapter 4: A higher blood oxygen saturation could be the result of less oxygen consumtion due to cell death. Chapter 5: There is no correlation between retinal blood flow and retinal blood oxygen saturation. Chapter 6: Further investigation of Ang 2, HGF, IL-8, EGF, TGF-β could be used to better understand the pathophysiology of DR. Chapter 7: The result of this study revealed a relationship between the biomarkers that might result in cell death and higher retinal blood oxygen saturation.1 yea

The Relationship between Retinal Vascular Reactivity and Arteriolar Diameter

ABSTRACT Purpose: The primary aim of the study (i.e. Chapter 3) was to compare the magnitude of retinal vascular reactivity in arterioles of varying diameter in healthy, young subjects. The secondary aims were to determine: a) if there are any order effects in terms of provoking vasoconstriction or vasodilation first; and b) the repeatability of the vascular reactivity measurements. An additional aim (i.e. Chapter 4) was to determine the effect of healthy aging on the relationship between retinal vascular reactivity and vessel diameter. Method: The sample comprised 10 healthy, young subjects (mean age 26.5 years, SD 4.04) and 7 healthy, older subjects (mean age 55.43 years, SD 5.41). Each subject from the young age group attended for three sessions. The first session was used to determine eligibility and select hemodynamic measurement sites. At sessions 2 and 3, O2 and CO2 were sequentially administered to the subjects using a face mask and sequential re-breathing circuit (to maintain standardized hyperoxia and hypercapnia). The order of vasoconstriction and vasodilation was varied across sessions 2 and 3. The design of the protocol was simplified for the subjects from the older age group. Each subject from the older group attended for one visit. O2 and CO2 were administered to the subjects using a face mask and sequential re-breathing circuit. The order of gas provocation was varied among the subjects (i.e. hyperoxia or hypercapnia first). For both groups, measurements of vessel diameter, centerline blood velocity and derived blood flow were acquired at each condition (i.e. baseline, during stabilized vasoconstriction, vasodilation, and recovery) at two discrete measurement sites along the supero-temporal arteriole. Results: The results of the repeated measures ANOVA showed a significant difference between the narrow and wide measurement sites for the younger group for flow (p≤ 0.0003) and a significant influence of inspired gas provocation on flow for both protocols (p<0.0001). In addition, the interaction of measurement site and inspired gas provocation was significant (p<0.0001). The magnitude of retinal vascular reactivity showed a significantly greater blood flow response for the wide measurement site (p<0.0001). O2 provocation resulted in vasoconstriction that was still present up to 10 minutes after cessation of the stimulus (order effect of O2; p≤0.046). No such order effect was apparent for CO2 provocation (order effect of CO2; p=0.352). The group mean blood flow Coefficient of Repeatability (COR) for the narrow measurement site was 0.74 µl/min (relative to group mean flow of 4.85 µl/min ± SD 1.31) and for the wide measurement site was 1.49 µl/min (relative to group mean flow of 11.29 µl/min ± SD 3.55). There was no difference between the young and the older age groups in retinal vascular reactivity for both the narrow (two-tailed Student t-test, p=0.8692) and wide (two-tailed Student t-test, p=0.2795) measurement sites. Conclusion: This study demonstrated that the magnitude of retinal vascular reactivity was greater for arteriolar measurement sites with wider baseline vessel diameters. In addition, it demonstrated that hyperoxic provocation resulted in a persistent vasoconstriction up to 10 minutes after cessation of the stimulus. The study demonstrated that the repeatability of retinal blood flow measurements in absolute terms is lower for smaller diameter vessels. Finally, this study also suggests that age does not affect the relationship between retinal vascular reactivity and vessel diameter

Variability and repeatability of quantitative, fourier-domain optical coherence tomography doppler blood flow in young and elderly healthy subjects

Purpose. The purpose of this study was to determine the within-session variability and between-session repeatability of spectral Fourier-domain optical coherence tomography (Doppler FD-OCT) Doppler retinal blood flow measurements in young and elderly subjects. Methods. Doppler FD-OCT blood flow was measured using the RTVue system. One eye of each of 20 healthy young (24.7 ± 2.7 years) and 16 healthy elderly (64.6 ±5.1 years) subjects was randomly selected, and the pupil was dilated. The double circular scanning pattern of the RTVue was employed. Six Doppler FD-OCT measurements (i.e., each separate measurement comprising an upper and a lower nasal pupil scan) were acquired at each session. Measurements were repeated approximately 2 weeks later. Total retinal blood flow was calculated by summing flow from all detectable venules surrounding the optic nerve head. The coefficient of variation (COV) and coefficient of repeatability (COR) were calculated for each individual. Results. The individual COVs for retinal blood flow for young subjects ranged from 0.4% to 20.4% (median 7.5%) and for the elderly subjects ranged from 0.6% to 34.6% (median 9.2%). The group mean CORs for retinal blood flow for young participants were 6.4 μL/min (median 5.91 μL/min, relative to a mean effect 39.8 μL/min) and for elderly subjects were 10.5 μL/min (median 9.2 μL/min, relative to a mean effect 46.4 μL/min). Conclusions. Doppler FD-OCT gave consistent and repeatable blood flow measurements within retinal venules in normal subjects. Considering the individual variation in blood flow measurements, confidence limits for retinal hemodynamics need to be determined on an individual basis

The relationship between retinal nerve fiber layer thickness and total retinal blood flow in primary open angle glaucoma.

Purpose: To investigate the relationship between Doppler SD-OCT derived retinal blood flow (RBF) and the retinal nerve fiber layer (RNFL) thickness in patients with primary open angle glaucoma (POAG) and healthy age-matched controls. Methods: Thirty three POAG patients (age, 65±8 years; Humphrey Field Analyzer, HFA, mean deviation, MD, -2.28±3.85) and 33 healthy controls (age, 63±5 years; HFA MD -0.48±1.56) were recruited. The Doppler SD-OCT retinal blood flow measurement was taken using the circum-papillary double circular scan protocol in the RTVue system (Optovue Inc., Freemont, CA, USA). A minimum of six RBF scans were acquired. Flow parameters were calculated only from the valid scans. Peri-papillary RNFL thickness was measured using RTVue’s RNFL scan protocol. Results: The total RBF in the POAG group was significantly lower than in the control group (group mean POAG RBF = 30.61±9.29mL/min; group mean control RBF = 40.68±11.32mL/min; p=<0.01). Superior and inferior RBF were also significantly lower in the POAG group (p=<0.01 for both superior & inferior RBF). The average RNFL thickness, superior and inferior RNFL thickness were significantly lower in the POAG group (p=<0.01). Linear regression analysis showed a significant positive correlation between the total RBF and average RNFL thickness (r=0.38, p=0.03), and the superior RBF and superior RNFL thickness (r=0.37, p=0.01). There was no correlation between the inferior RBF and RNFL thickness (r=0.30,p=0.09). Conclusions: There was an association between thinning of the RNFL and reduced inner retinal perfusion in the eyes of patients with POAG

LXRs regulate features of age-related macular degeneration and may be a potential therapeutic target

Effective treatments and animal models for the most prevalent neurodegenerative form of blindness in elderly people, called age-related macular degeneration (AMD), are lacking. Genome-wide association studies have identified lipid metabolism and inflammation as AMD-associated pathogenic pathways. Given liver X receptors (LXRs), encoded by the nuclear receptor subfamily 1 group H members 2 and 3 (NR1H3 and NR1H2), are master regulators of these pathways, herein we investigated the role of LXR in human and mouse eyes as a function of age and disease and tested the therapeutic potential of targeting LXR. We identified immunopositive LXR fragments in human extracellular early dry AMD lesions and a decrease in LXR expression within the retinal pigment epithelium (RPE) as a function of age. Aged mice lacking LXR presented with isoform-dependent ocular pathologies. Specifically, loss of the Nr1h3 isoform resulted in pathobiologies aligned with AMD, supported by compromised visual function, accumulation of native and oxidized lipids in the outer retina, and upregulation of ocular inflammatory cytokines, while absence of Nr1h2 was associated with ocular lipoidal degeneration. LXR activation not only ameliorated lipid accumulation and oxidant-induced injury in RPE cells but also decreased ocular inflammatory markers and lipid deposition in a mouse model, thereby providing translational support for pursuing LXR-active pharmaceuticals as potential therapies for dry AMD

A recommended “minimum data set” framework for SD-OCT retinal image acquisition and analysis from the Atlas of Retinal Imaging in Alzheimer’s Study (ARIAS)

Introduction: We propose a minimum data set framework for the acquisition and analysis of retinal images for the development of retinal Alzheimer\u27s disease (AD) biomarkers. Our goal is to describe methodology that will increase concordance across laboratories, so that the broader research community is able to cross‐validate findings in parallel, accumulate large databases with normative data across the cognitive aging spectrum, and progress the application of this technology from the discovery stage to the validation stage in the search for sensitive and specific retinal biomarkers in AD. Methods: The proposed minimum data set framework is based on the Atlas of Retinal Imaging Study (ARIAS), an ongoing, longitudinal, multi‐site observational cohort study. However, the ARIAS protocol has been edited and refined with the expertise of all co‐authors, representing 16 institutions, and research groups from three countries, as a first step to address a pressing need identified by experts in neuroscience, neurology, optometry, and ophthalmology at the Retinal Imaging in Alzheimer\u27s Disease (RIAD) conference, convened by the Alzheimer\u27s Association and held in Washington, DC, in May 2019. Results: Our framework delineates specific imaging protocols and methods of analysis for imaging structural changes in retinal neuronal layers, with optional add‐on procedures of fundus autofluorescence to examine beta‐amyloid accumulation and optical coherence tomography angiography to examine AD‐related changes in the retinal vasculature. Discussion: This minimum data set represents a first step toward the standardization of retinal imaging data acquisition and analysis in cognitive aging and AD. A standardized approach is essential to move from discovery to validation, and to examine which retinal AD biomarkers may be more sensitive and specific for the different stages of the disease severity spectrum. This approach has worked for other biomarkers in the AD field, such as magnetic resonance imaging; amyloid positron emission tomography; and, more recently, blood proteomics. Potential context of use for retinal AD biomarkers is discussed

Age effects on retinal blood flow assessed using spectral-domain optical coherence tomography doppler

Purpose: : To determine the impact of healthy aging upon total retinal blood flow as derived by Spectral-Domain, Optical Coherence Tomography Doppler SD-OCT. Methods: : SD-OCT Doppler blood flow was non-invasively measured using the RTVue system (Optovue Inc., USA). One eye of each of 6 healthy young (mean age 25.7; SD 3.5 years) and 6 healthy elderly (mean age 63.5; SD 1.8 years) subjects was randomly selected for the study and dilated using Mydriacyl 1%. A double circular scanning pattern was employed. A minimum of six separate SD-OCT Doppler measurements (i.e. each separate measurement comprising a superior nasal pupil scan and an inferior nasal pupil scan) were acquired. Total retinal blood flow was calculated, using data from valid scans only, by summing flow from all detectable venules. Results: : 117 of 170 images (68.82%) were determined to be valid using the DOCTORC software (Centre for Ophthalmic Optics and Lasers, CA, USA). Mean total retinal blood flow for the young group was 44.96±15.59 µl/min, while for the elderly group it was 39.45±13.20 µl/min. Two-sided t-test showed no significant difference between both groups (p=0.52). Linear regression analysis showed no significant correlation between total retinal blood flow and age (r=-0.31, p=0.33). Conclusions: : Preliminary data acquired using SD-OCT Doppler shows no significant difference in retinal blood flow between a young and elderly group

Age effects on retinal blood flow assessed using spectral-domain optical coherence tomography doppler

Purpose: To determine the impact of healthy aging upon total retinal blood flow as derived by Spectral-Domain, Optical Coherence Tomography Doppler SD-OCT. Methods: SD-OCT Doppler blood flow was non-invasively measured using the RTVue system (Optovue Inc., USA). One eye of each of 6 healthy young (mean age 25.7; SD 3.5 years) and 6 healthy elderly (mean age 63.5; SD 1.8 years) subjects was randomly selected for the study and dilated using Mydriacyl 1%. A double circular scanning pattern was employed. A minimum of six separate SD-OCT Doppler measurements (i.e. each separate measurement comprising a superior nasal pupil scan and an inferior nasal pupil scan) were acquired. Total retinal blood flow was calculated, using data from valid scans only, by summing flow from all detectable venules. Results: 117 of 170 images (68.82%) were determined to be valid using the DOCTORC software (Centre for Ophthalmic Optics and Lasers, CA,USA). Mean total retinal blood flow for the young group was 44.96±15.59μl/min, while for the elderly group it was 39.45±13.20 μl/min. Two-sided t-test showed no significant difference between both groups (p=0.52). Linear regression analysis showed no significant correlation between total retinal blood flow and age (r=-0.31, p=0.33). Conclusions: Preliminary data acquired using SD-OCT Doppler shows no significant difference in retinal blood flow between a young and elderly group

Variability and repeatability of quantitative, SD-OCT doppler blood flow in young and elderly healthy subjects

Purpose: The purpose was to determine the within-session variability and between-session repeatability of SD-OCT Doppler measurement of retinal blood flow in young and elderly healthy subjects. Methods: SD-OCT Doppler blood flow was non-invasively measured using the RTVue system (Optovue Inc., USA). One eye of each of 6 healthy young (mean age 25.7; SD 3.5 years) and 6 healthy elderly (mean age 63.1; SD 1.8 years) subjects was randomly selected for the study and the pupil was dilated using Mydriacyl 1%. A double circular scanning pattern was employed. A minimum of six separate SD-OCT Doppler measurements (i.e. each separate measurement comprising an upper nasal pupil scan and a lower nasal pupil scan) were acquired at each session. Measurements were repeated on a second day. Retinal blood flow was calculated, using data from valid scans only, by summing flow from all detectable venules. The coefficient of variation and the coefficient of repeatability were calculated for each individual. Results: The individual COVs for retinal blood flow for young subjects ranged from 2 to 51.9% (median 28.8%) and for the elderly subjects ranged from 0.6 to 81.2% (median 9.9%). The group mean CORs for retinal blood flow for young subjects were 30.5 μl/min (median 29.5 μl/min, relative to a mean effect 39.8 μl/min) and for elderly subjects were 16.9 μl/min (median 7.2 μl/min, relative to a mean effect 34.7 μl/min). Conclusions: The Doppler SD-OCT in general gave consistent measurements of retinal blood flow in normal subjects, while the data was far more repeatable for the elderly group. The relatively small sample size needs to be considered when interpreting these results