LUNet: deep learning for the segmentation of arterioles and venules in high resolution fundus images.

Objective.This study aims to automate the segmentation of retinal arterioles and venules (A/V) from digital fundus images (DFI), as changes in the spatial distribution of retinal microvasculature are indicative of cardiovascular diseases, positioning the eyes as windows to cardiovascular health.Approach.We utilized active learning to create a new DFI dataset with 240 crowd-sourced manual A/V segmentations performed by 15 medical students and reviewed by an ophthalmologist. We then developed LUNet, a novel deep learning architecture optimized for high-resolution A/V segmentation. The LUNet model features a double dilated convolutional block to widen the receptive field and reduce parameter count, alongside a high-resolution tail to refine segmentation details. A custom loss function was designed to prioritize the continuity of blood vessel segmentation.Main Results.LUNet significantly outperformed three benchmark A/V segmentation algorithms both on a local test set and on four external test sets that simulated variations in ethnicity, comorbidities and annotators.Significance.The release of the new datasets and the LUNet model (www.aimlab-technion.com/lirot-ai) provides a valuable resource for the advancement of retinal microvasculature analysis. The improvements in A/V segmentation accuracy highlight LUNet's potential as a robust tool for diagnosing and understanding cardiovascular diseases through retinal imaging.

PKSEA-Net: A prior knowledge supervised edge-aware multi-task network for retinal arteriolar morphometry.

Retinal fundus images serve as a non-invasive modality to obtain information pertaining to retinal vessels through fundus photography, thereby offering insights into cardiovascular and cerebrovascular diseases. Retinal arteriolar morphometry has emerged as the most convenient and fundamental clinical methodology in the realm of patient screening and diagnosis. Nevertheless, the analysis of retinal arterioles is challenging attributable to imaging noise, stochastic fuzzy characteristics, and blurred boundaries proximal to blood vessels. In response to these limitations, we introduce an innovative methodology, named PKSEA-Net, which aims to improve segmentation accuracy by enhancing the perception of edge information in retinal fundus images. PKSEA-Net employs the universal architecture PVT-v2 as the encoder, complemented by a novel decoder architecture consisting of an Edge-Aware Block (EAB) and a Pyramid Feature Fusion Module (PFFM). The EAB block incorporates prior knowledge for supervision and multi-query for multi-task learning, with supervision information derived from an enhanced Full Width at Half Maximum (FWHM) algorithm and gradient map. Moreover, PFFM efficiently integrates multi-scale features through a novel attention fusion method. Additionally, we have collected a Retinal Cross-Sectional Vessel (RCSV) dataset derived from approximately 200 patients in Quzhou People's Hospital to serve as the benchmark dataset. Comparative evaluations with several state-of-the-art (SOTA) networks confirm that PKSEA-Net achieves exceptional experimental performance, thereby establishing its status as a SOTA approach for precise boundary delineation and retinal vessel segmentation.

Network response of brain microvasculature to neuronal stimulation.

Neurovascular coupling (NVC), or the adjustment of blood flow in response to local increases in neuronal activity is a hallmark of healthy brain function, and the physiological foundation for functional magnetic resonance imaging (fMRI). However, it remains only partly understood due to the high complexity of the structure and function of the cerebrovascular network. Here we set out to understand NVC at the network level, i.e. map cerebrovascular network reactivity to activation of neighbouring neurons within a 500×500×500 µm3 cortical volume (∼30 high-resolution 3-nL fMRI voxels). Using 3D two-photon fluorescence microscopy data, we quantified blood volume and flow changes in the brain vessels in response to spatially targeted optogenetic activation of cortical pyramidal neurons. We registered the vessels in a series of image stacks acquired before and after stimulations and applied a deep learning pipeline to segment the microvascular network from each time frame acquired. We then performed image analysis to extract the microvascular graphs, and graph analysis to identify the branch order of each vessel in the network, enabling the stratification of vessels by their branch order, designating branches 1-3 as precapillary arterioles and branches 4+ as capillaries. Forty-five percent of all vessels showed significant calibre changes; with 85 % of responses being dilations. The largest absolute CBV change was in the capillaries; the smallest, in the venules. Capillary CBV change was also the largest fraction of the total CBV change, but normalized to the baseline volume, arterioles and precapillary arterioles showed the biggest relative CBV change. From linescans along arteriole-venule microvascular paths, we measured red blood cell velocities and hematocrit, allowing for estimation of pressure and local resistance along these paths. While diameter changes following neuronal activation gradually declined along the paths; the pressure drops from arterioles to venules increased despite decreasing resistance: blood flow thus increased more than local resistance decreases would predict. By leveraging functional volumetric imaging and high throughput deep learning-based analysis, our study revealed distinct hemodynamic responses across the vessel types comprising the microvascular network. Our findings underscore the need for large, dense sampling of brain vessels for characterization of neurovascular coupling at the network level in health and disease.

Prognostic significance of the wall to lumen ratio of retinal arterioles evaluated by adaptive optics.

OBJECTIVE: Microvascular structural alterations may be considered an important form of hypertension-mediated organ damage. An increased media-to-lumen ratio of subcutaneous small arteries evaluated with locally invasive techniques (micromyography) predicts the development of cardiovascular (CV) events. However, it is not known whether retinal arteriole structural alterations evaluated with a noninvasive approach (Adaptive Optics) may have a prognostic significance. DESIGN AND METHODS: Two-hundred and thirty-seven subjects (mean age 58.7 ± 16.1 years, age range 13-89 years; 116 males) were included in the study: 65 normotensive subjects (27.4 %) and 172 patients with essential hypertension or primary aldosteronism (72.6 %). All subjects underwent a non-invasive evaluation of retinal arteriolar wall-to-lumen ratio (WLR) by Adaptive Optics. Subjects were re-evaluated after an average follow-up time of 4.55 years in order to assess the occurrence of clinical events (non CV and/or CV death or events). RESULTS: Fifty-four events occurred in the study population:26 were cardio-cerebrovascular events (ischemic or hemorragic stroke, atrial fibrillation, heart failure, coronary artery disease, peripheral artery disease, cardiac valvular disease) while the remaining were deaths for any cause, or neoplastic diseases. Subjects with events were older and had a WLR of retinal arterioles significantly greater than those without events. The event-free survival was significantly worse in those with a baseline WLR above the median value of the population (0.28) according to Kaplan-Mayer survival curves and multivariate analysis (Cox's proportional hazard model). The evidence was confirmed after restricting the analysis to CV events. CONCLUSIONS: Structural alterations of retinal arterioles evaluated by Adaptive Optics may predict total and CV events.

Retinal Arteriolar Wall Remodeling in Diabetes Captured With AOSLO.

Purpose: Adaptive optics scanning laser ophthalmoscopy (AOSLO) enables the visualization and measurement of the retinal microvasculature structure in humans. We investigated the hypothesis that diabetes mellitus (DM) induces remodeling to the wall structure in small retinal arterioles. These alterations may allow better understanding of vascular remodeling in DM. Methods: We imaged retinal arterioles in one eye of 48 participants (26 with DM and 22 healthy controls) with an AOSLO. Structural metrics of 274 arteriole segments (203 with DM and 71 healthy controls) ≤ 50 µm in outer diameter (OD) were quantified and we compared differences in wall thickness (WT), wall-to-lumen ratio (WLR), inner diameter (ID), OD, and arteriolar index ratio (AIR) between controls and participants with DM. We also compared the individual AIR (iAIR) in groups of individuals. Results: The WLR, WT, and AIRs were significantly different in the arteriole segments of DM participants (P < 0.001). The iAIR was significantly deviated in the DM group (P < 0.001) and further division of the participants with DM into groups revealed that there was an effect of the presence of diabetic retinopathy (DR) on the iAIR (P < 0.001). Conclusions: DM induces remodeling of wall structure in small retinal arterioles and in groups of individuals. The use of AIR allows us to assess remodeling independently of vessel size in the retina and to compute an index for each individual subject. Translational Relevance: High-resolution retinal imaging allows noninvasive assessment of small retinal vessel remodeling in DM that can improve our understanding of DM and DR in living humans.

Imaging the time course, morphology, neuronal tissue compression, and resolution of cerebral microhemorrhages in mice using intravital two-photon microscopy: insights into arteriolar, capillary, and venular origin.

Cerebral microhemorrhages (CMHs, microbleeds), a manifestation of age-related cerebral small vessel disease, contribute to the pathogenesis of cognitive decline and dementia in older adults. Histological studies have revealed that CMHs exhibit distinct morphologies, which may be attributed to differences in intravascular pressure and the size of the vessels of origin. Our study aimed to establish a direct relationship between the size/morphology of CMHs and the size/anatomy of the microvessel of origin. To achieve this goal, we adapted and optimized intravital two-photon microscopy-based imaging methods to monitor the development of CMHs in mice equipped with a chronic cranial window upon high-energy laser light-induced photodisruption of a targeted cortical arteriole, capillary, or venule. We assessed the time course of extravasation of fluorescently labeled blood and determined the morphology and size/volume of the induced CMHs. Our findings reveal striking similarities between the bleed morphologies observed in hypertension-induced CMHs in models of aging and those originating from different targeted vessels via multiphoton laser ablation. Arteriolar bleeds, which are larger (> 100 µm) and more widely dispersed, are distinguished from venular bleeds, which are smaller and exhibit a distinct diffuse morphology. Capillary bleeds are circular and smaller (< 10 µm) in size. Our study supports the concept that CMHs can occur at any location in the vascular tree, and that each type of vessel produces microbleeds with a distinct morphology. Development of CMHs resulted in immediate constriction of capillaries, likely due to pericyte activation and constriction of precapillary arterioles. Additionally, tissue displacement observed in association with arteriolar CMHs suggests that they can affect an area with a radius of ~ 50 µm to ~ 100 µm, creating an area at risk for ischemia. Longitudinal imaging of CMHs allowed us to visualize reactive astrocytosis and bleed resolution during a 30-day period. Our study provides new insights into the development and morphology of CMHs, highlighting the potential clinical implications of differentiating between the types of vessels involved in the pathogenesis of CMHs. This information may help in the development of targeted interventions aimed at reducing the risk of cerebral small vessel disease-related cognitive decline and dementia in older adults.

In vivo detection of penetrating arteriole alterations in cerebral white matter in patients with diabetes with 7 T MRI.

Cerebral small vessel disease (SVD) is responsible for primary intracerebral hemorrhages, lacunar infarcts and white matter hyperintensity in T2 weighted images. While the brain lesions attributed to small vessel disease can be characterized by conventional MRI, it remains challenging to noninvasively measure the early pathological changes of the small underlying vessels. We evaluated the feasibility of detecting alterations in white matter penetrating arterioles (PA) in patients with diabetes with ultra-high field 7 T MRI. 19 participants with diabetes mellitus (DM) and 19 age- and sex-matched healthy controls were scanned with whole brain T2 and susceptibility weighted MRI and a single slice phase contrast MRI 15 mm above the corpus callosum. The PC-MRI scans were repeated three times. PA masks were manually drawn on the first images after anonymization or automatically segmented on all three images. For each PA, lumen diameter, flow velocity and volume flow rate were derived by model-based analyses of complex difference images. Quasi-Poisson regression was performed for PA count using disease condition, age, and sex as independent variables. Linear mixed effect model analyses were performed for the other measurements using disease condition and age as fixed effect and participant pair specific disease condition as random effect. No severe radiological features of SVD were observed in T2 and susceptibility weighted images in any of the participants except for white matter hyperintensities with Fazekas score of 1 or 2 in 68% and 26% of patients and controls, respectively. The minimum diameter of visible PA was 78 µm and the majority had diameters <250 µm. Among the manually segmented PA with tilt angle less than 30o from the slice normal direction, flow velocities were lower in the DM group (1.9 ± 0.6 vs. 2.2 ± 0.6; p = 0.022), while no significant difference was observed in count, diameter, or volume flow rate. Similar results were observed in the automatically segmented PA. We also observed significantly increased diameter or decreased velocity with age in some of the scans. This study suggests that early PA alterations that are discriminative of disease state and age might be detectable in human cerebral white matter with 7 T MRI in vivo.

Normative data of retinal arteriolar and venular calibre measurements determined using confocal scanning laser ophthalmoscopy system - Importance and implications for study of cardiometabolic disorders.

Purpose: To determine and validate retinal vascular caliber measurements by using the confocal scanning laser ophthalmoscopy system. Retinal vasculature changes are often regarded as clinical markers for systemic disease. Methods: It was a prospective observational study conducted on 600 eyes of 300 normal subjects with no systemic or ocular illness from January 1, 2016 to June 30, 2017 in a tertiary referral eye center. Non-mydriatic infrared reflectance, blue reflectance, and blue peak blue autofluorescence fundus imaging were done on the confocal scanning laser ophthalmoscopy system. The dimensions of the retinal vessels were measured using inbuilt calipers at 1800 µm from the center of the optic disc. Internal and external dimensions were measured. Observer variation and its comparison using Image J software were assessed. Results: The median age was 29 years (18-50 years). Mean internal and external diameters for arterioles were 85.1 ± 12.4 µm and 105.0 ± 12.0 µm, and for venules were 133.8 ± 16.6 µm and 145.4 ± 16.1 µm, respectively. The mean internal and external wall thicknesses were 19.7 ± 8.0 µm and 11.0 ± 5.6 µm, and wall thickness-to-lumen ratios were 0.3 ± 0.1 and 0.1 ± 0.1, respectively. Arteriolar-to-venular ratio for lumen and vessel was 0.66 ± 0.1 and 0.74 ± 0.1, respectively. There was no statistically significant difference between age groups. Both inter- and intra-observer reproducibility was >95%. The Bland-Altman plot showed that the difference between measurements using both confocal scanning laser ophthalmoscopy and Image J software lies within the limits of agreement approximately 95% of the time. Conclusion: This is the first effort to develop a normative database by using a simple non-invasive confocal scanning laser ophthalmoscopy system with high observer reproducibility.

Tissue sodium content correlates with hypertrophic vascular remodeling in type 2 diabetes.

BACKGROUND: Prospective studies describe a linkage between increased sodium intake and higher incidence of cardiovascular organ damage and end points. We analyzed whether tissue sodium content in the skin and muscles correlate with vascular hypertrophic remodeling, a risk factor for cardiovascular disease. METHODS: In patients with type 2 diabetes we assessed tissue sodium content and vascular structural parameters of the retinal arterioles. The structural parameters of retinal arterioles assessed by Scanning Laser Doppler Flowmetry were vessel (VD) and lumen diameter (LD), wall thickness (WT), wall-to-lumen ratio (WLR) and wall cross sectional area (WCSA). Tissue sodium content was measured with a 3.0 T clinical 23Sodium-Magnetic Resonance Imaging (23Na-MRI) system. RESULTS: In patients with type 2 diabetes (N = 52) we observed a significant correlation between muscle sodium content and VD (p = 0.005), WT (p = 0.003), WCSA (p = 0.002) and WLR (p = 0.013). With respect to skin sodium content a significant correlation has been found with VD (p = 0.042), WT (p = 0.023) and WCSA (p = 0.019). Further analysis demonstrated that tissue sodium content of skin and muscle is a significant determinant of hypertrophic vascular remodeling independent of age, gender, diuretic use and 24-hour ambulatory BP. CONCLUSION: With the 23Na-MRI technology we could demonstrate that high tissue sodium content is independently linked to hypertrophic vascular remodeling in type 2 diabetes. TRIAL REGISTRATION: Trial registration number: NCT02383238 Date of registration: March 9, 2015.

Retinal diffusion restrictions in acute branch retinal arteriolar occlusion.

This study sought to investigate the occurrence of retinal diffusion restrictions (RDR) in branch retinal arteriolar occlusion (BRAO) using standard brain diffusion-weighted imaging (DWI). Two radiologists assessed DWI MRI scans of BRAO patients for RDR in a retrospective cohort study. Inter- and intrarater reliability were calculated using Kappa statistics. Detection rates of RDR were compared among MRI scans with varying field strength, sequence type and onset-to-DWI time intervals. 85 BRAO patients (63.1 ± 16.5 years) and 89 DWI scans were evaluated. Overall sensitivity of RDR in BRAO was 46.1% with visually correlating low ADC signal in 56.1% of cases. Localization of RDR matched distribution of fundoscopic retinal edema in 85% of patients. Inter- and intra-rater agreement for RDR in BRAO was κinter = 0.64 (95% CI 0.48-0.80) and κintra = 0.87 (95% CI 0.76-0.96), respectively. RDR detection rate tended to be higher for 3T, when compared to 1.5T MRI scans (53.7% vs. 34.3%%; p = 0.07). RDR were identified within 24 h up to 2 weeks after onset of visual impairment. RDR in BRAO can be observed by means of standard stroke DWI in a substantial proportion of cases, although sensitivity and interrater reliability were lower than previously reported for complete central retinal artery occlusion.

Normative data and standard operating procedures for static and dynamic retinal vessel analysis as biomarker for cardiovascular risk.

Retinal vessel phenotype is predictive for cardiovascular outcome. This cross-sectional population-based study aimed to quantify normative data and standard operating procedures for static and dynamic retinal vessel analysis. We analysed central retinal arteriolar (CRAE) and venular (CRVE) diameter equivalents, as well as retinal endothelial function, measured by flicker light-induced maximal arteriolar (aFID) and venular (vFID) dilatation. Measurements were performed in 277 healthy individuals aged 20 to 82 years of the COmPLETE study. The mean range from the youngest compared to the oldest decade was 196 ± 13 to 166 ± 17 µm for CRAE, 220 ± 15 to 199 ± 16 µm for CRVE, 3.74 ± 2.17 to 3.79 ± 2.43% for aFID and 4.64 ± 1.85 to 3.86 ± 1.56% for vFID. Lower CRAE [estimate (95% CI): - 0.52 (- 0.61 to - 0.43)], CRVE [- 0.33 (- 0.43 to - 0.24)] and vFID [- 0.01 (- 0.26 to - 0.00)], but not aFID, were significantly associated with older age. Interestingly, higher blood pressure was associated with narrower CRAE [- 0.82 (- 1.00 to - 0.63)] but higher aFID [0.05 (0.03 to 0.07)]. Likewise, narrower CRAE were associated with a higher predicted aFID [- 0.02 (- 0.37 to - 0.01)]. We recommend use of defined standardized operating procedures and cardiovascular risk stratification based on normative data to allow for clinical implementation of retinal vessel analysis in a personalized medicine approach.

Baseline extended zone retinal vascular calibres associate with sensory nerve abnormalities in adolescents with type 1 diabetes: A prospective longitudinal study.

OBJECTIVE: The relationship between retinal vascular calibres (RVCs) and diabetic neuropathy is unclear. We investigated associations between RVCs and sensory nerve abnormality in adolescents with type 1 diabetes. RESEARCH DESIGN AND METHODS: In a prospective longitudinal study of 889 adolescents with type 1 diabetes with baseline mean (±SD) age 14.1 ± 1.5 years and HbA1c IFCC 69.4 ± 14.1 mmol/mol (8.6 ± 1.3%), RVCs were assessed from baseline retinal photographs: 'central zone' calibres, summarized as central retinal arteriolar (CRAE) and venular equivalents (CRVE) and 'extended zone' calibres: mean width of arterioles (MWa) and venules (MWv). Sensory nerve abnormality was defined as at least one abnormal sensory quantitative testing from two thermal and two vibration threshold tests measured at foot every 1-2 years. Associations between baseline RVC and sensory nerve function were examined using generalized estimating equations and cumulative risk by Cox regression analyses. RESULTS: During a median study follow-up of 6.2 [IQR 3.7-10.4] years, sensory nerve abnormality was found in 27% of adolescents. Narrower extended zone calibre quartiles but not CRAE or CRVE quartiles were independently associated with sensory nerve abnormality: MWa (Q1 vs. Q2-4: OR 1.35 (95% CI 1.02, 1.61) and MWv (Q1 vs. Q2-4: 1.31 (1.03, 1.7)), after adjusting for HbA1c , duration and blood pressure. Similarly, in Cox regression, the narrowest quartiles were associated with sensory nerve abnormality: MWa hazard ratio (HR) 1.5 (1.3, 1.8) and MWv 1.6 (1.4, 1.9). CONCLUSIONS: Narrower extended zone retinal calibres were associated with sensory nerve abnormality in adolescents with type 1 diabetes and may present useful biomarkers to understand the pathophysiology of neuropathy.

Imaging the construction of capillary networks in the neonatal mouse brain.

Capillary networks are essential for distribution of blood flow through the brain, and numerous other homeostatic functions, including neurovascular signal conduction and blood-brain barrier integrity. Accordingly, the impairment of capillary architecture and function lies at the root of many brain diseases. Visualizing how brain capillary networks develop in vivo can reveal innate programs for cerebrovascular growth and repair. Here, we use longitudinal two-photon imaging through noninvasive thinned skull windows to study a burst of angiogenic activity during cerebrovascular development in mouse neonates. We find that angiogenesis leading to the formation of capillary networks originated exclusively from cortical ascending venules. Two angiogenic sprouting activities were observed: 1) early, long-range sprouts that directly connected venules to upstream arteriolar input, establishing the backbone of the capillary bed, and 2) short-range sprouts that contributed to expansion of anastomotic connectivity within the capillary bed. All nascent sprouts were prefabricated with an intact endothelial lumen and pericyte coverage, ensuring their immediate perfusion and stability upon connection to their target vessels. The bulk of this capillary expansion spanned only 2 to 3 d and contributed to an increase of blood flow during a critical period in cortical development.

Retinal Arteriolar Occlusions and Exudative Retinal Detachments in Malignant Hypertension: More Than Meets the Eye.

BACKGROUND: Malignant hypertension is macrovascular and microvascular endothelial injury responsible for multiple organ damage. Considering the anatomical and functional homologies between the posterior pole of the eye and the kidney, ophthalmological explorations may inform clinicians on the mechanisms underpinning concurrent kidney injury in this condition. More specifically, we investigated whether the wall-to-lumen ratio (WLR) of retinal arterioles measured by adaptive optics ophthalmoscopy could be correlated to WLR of kidney arterioles as determined by pathology. We sought to estimate the incidence of retinal arteriole occlusion a supposedly uncommon complication of malignant hypertension. METHODS: All patients hospitalized in our renal Intensive Care Unit for malignant hypertension between 2016 and 2019 were referred to ophthalmological examinations. RESULTS: Twenty-seven patients were included. Median retinal WLR was 0.39 [0.31-0.47] and was correlated with initial systolic (r = 0.56, P = 0.003) and mean blood pressure (r = 0.46, P = 0.02) upon admission. The retinal WLR was not correlated to renal pathological findings, as assessed by juxtaglomerular WLR (r = 0.38, P = 0.2), ratio of glomerulosclerosis (r = -0.39, P = 0.2), or tubulointerstitial fibrosis (r = -0.45, P = 0.08). Retinal WLR was not associated with neurological or cardiovascular end-organ damage. Branch retinal artery occlusion was detected in 18.5% of patients and exudative retinal detachment (ERD) in 29.6% of patients, without any significant correlation with canonical signs of retinal hypertension including optic disc swelling. CONCLUSIONS: In the setting of malignant hypertension, we failed to demonstrate a significant relationship between WLR and other meaningful end-organ injuries. However, branch retinal artery occlusion and ERD may have been hitherto underestimated.

Analysis of the three dimensional structure of the kidney glomerulus capillary network.

The capillary network of the kidney glomerulus filters small molecules from the blood. The glomerular 3D structure should help to understand its function, but it is poorly characterized. We therefore devised a new approach in which an automated tape collecting microtome (ATUM) was used to collect 0.5 µm thick serial sections from fixed mouse kidneys. The sections were imaged by scanning electron microscopy at ~ 50 nm/pixel resolution. With this approach, 12 glomeruli were reconstructed at an x-y-z resolution ~ 10 × higher than that of paraffin sections. We found a previously undescribed no-cross zone between afferent and efferent branches on the vascular pole side; connections here would allow blood to exit without being adequately filtered. The capillary diameters throughout the glomerulus appeared to correspond with the amount of blood flow within them. The shortest path (minimum number of branches to travel from afferent to efferent arterioles) is relatively independent of glomerular size and is present primarily on the vascular pole size. This suggests that new branches and longer paths form on the urinary pole side. Network analysis indicates that the glomerular network does not form by repetitive longitudinal splitting of capillaries. Thus the 3D structure of the glomerular capillary network provides useful information with which to understand glomerular function. Other tissue structures in the body may benefit from this new three dimensional approach.

Mutant G2019S-LRRK2 Induces Abnormalities in Arteriolar Cerebral Blood Volume in Mouse Brains: An MRI Study.

BACKGROUND: Parkinson's disease (PD) is the second most common neurodegenerative disease and the most common movement disorder characterized by motor impairments resulting from midbrain dopamine neuron loss. Abnormalities in small pial arteries and arterioles, which are the primary pathways of local delivery of nutrients and oxygen in brain tissue, have been reported in many neurodegenerative diseases including PD. Mutations in LRRK2 cause genetic PD and contribute to sporadic PD. The most common PD-linked mutation LRRK2 G2019S contributes 20-47% of genetic forms of PD in Caucasian populations. The human LRRK2 G2019S transgenic mouse model displays PD-like movement impairment and was used to identify novel LRRK2 inhibitors, which provides a useful model for studying microvascular abnormalities in PD. OBJECTIVES: To investigate abnormalities in arteriolar cerebral blood volume (CBVa) in various brain regions using the inflow-based vascular-space occupancy (iVASO) MRI technique in LRRK2 mouse models of PD. METHODS: Anatomical and iVASO MRI scans were performed in 5 female and 7 male nontransgenic (nTg), 3 female and 4 male wild-type (WT) LRRK2, and 5 female and 7 male G2019S-LRRK2 mice of 9 months of age. CBVa was calculated and compared in the substantia nigra (SN), olfactory cortex, and prefrontal cortex. RESULTS: Compared to nTg mice, G2019S-LRRK2 mice showed decreased CBVa in the SN, but increased CBVa in the olfactory and prefrontal cortex in both male and female groups, whereas WT-LRRK2 mice showed no change in CBVa in the SN (male and female), the olfactory (female), and prefrontal (female) cortex, but a slight increase in CBVa in the olfactory and prefrontal cortex in the male group only. CONCLUSIONS: Alterations in the blood volume of small arteries and arterioles (CBVa) were detected in the G2019S-LRRK2 mouse model of PD. The opposite changes in CBVa in the SN and the cortex indicate that PD pathology may have differential effects in different brain regions. Our results suggest the potential value of CBVa as a marker for clinical PD studies.

Dynamic Measurement and Imaging of Capillaries, Arterioles, and Pericytes in Mouse Heart.

Coronary arterial tone along with the opening or closing of the capillaries largely determine the blood flow to cardiomyocytes at constant perfusion pressure. However, it is difficult to monitor the dynamic changes of the coronary arterioles and the capillaries in the whole heart, primarily due to its motion and non-stop beating. Here we describe a method that enables monitoring of arterial perfusion rate, pressure and the diameter changes of the arterioles and capillaries in mouse right ventricular papillary muscles. The mouse septal artery is cannulated and perfused at a constant flow or pressure with the other dynamically measured. After perfusion with a fluorescently labeled lectin (e.g., Alexa Fluor-488 or -633 labeled Wheat-Germ Agglutinin, WGA), the arterioles and capillaries (and other vessels) in right ventricle papillary muscle and septum could be readily imaged. The vessel-diameter changes could then be measured in the presence or absence of heart contractions. When genetically encoded fluorescent proteins were expressed, specific features could be monitored. For examples, pericytes were visualized in mouse hearts that expressed NG2-DsRed. This method has provided a useful platform to study the physiological functions of capillary pericytes in heart. It is also suitable for studying the effect of reagents on the blood flow in heart by measuring the vascular/capillary diameter and the arterial luminal pressure simultaneously. This preparation, combined with a state-of-the-art optic imaging system, allows one to study the blood flow and its control at cellular and molecular level in the heart under near-physiological conditions.

Automatic analysis of normative retinal oximetry images.

Retinal oximetry is an important screening tool for early detection of retinal pathologies due to changes in the vasculature and also serves as a useful indicator of human-body-wide vascular abnormalities. We present an automatic technique for the measurement of oxygen saturation in retinal arterioles and venules using dual-wavelength retinal oximetry images. The technique is based on segmenting an optic-disc-centered ring-shaped region of interest and subsequent analysis of the oxygen saturation levels. We show that the two dominant peaks in the histogram of the oxygen saturation levels correspond to arteriolar and venular oxygen saturations from which the arterio-venous saturation difference (AVSD) can be calculated. For evaluation, we use a normative database of Asian Indian eyes containing 44 dual-wavelength retinal oximetry images. Validations against expert manual annotations of arterioles and venules show that the proposed technique results in an average arteriolar oxygen saturation (SatO2) of 87.48%, venular SatO2 of 57.41%, and AVSD of 30.07% in comparison with the expert ground-truth average arteriolar SatO2 of 89.41%, venular SatO2 of 56.32%, and AVSD of 33.09%, respectively. The results exhibit high consistency across the dataset indicating that the automated technique is an accurate alternative to the manual procedure.

Exploring Associations Between Cardiac Structure and Retinal Vascular Geometry.

Background Retinal arteriolar narrowing and venular widening has been widely suggested to be associated with subclinical changes in cardiac structure. The novel retinal vascular geometric indices might reflect more comprehensive information on microvasculature other than vascular caliber alone. However, the association between suboptimal retinal vascular geometry and cardiac structural alteration has not been studied. Methods and Results We recruited 50 participants without cardiovascular disease from the Cardiac Aging Study conducted between 2014 and 2016. We performed transthoracic echocardiography imaging to measure cardiac structure indices such as left ventricular internal diameter end diastole index, left ventricular internal diameter end systole index, left ventricular mass index, and left atrial volume index, and retinal imaging to measure retinal vascular geometric indices including branching angle, curvature tortuosity, and fractal dimension. We applied multiple linear regressions to examine associations between indices of cardiac structure and retinal vascular geometry, adjusting for age, sex, body mass index, mean blood pressure, and comorbidity. The average age of all participants was 62.54 years old and slightly more than half were male (27; 54%). Each unit increase in a set of cardiac structure indices was associated with larger retinal arteriolar branching angle (ß and 95% CI: for left ventricular internal diameter end systole index, 26.93°; 6.00-47.86; for left ventricular internal diameter end diastole index, 17.86°; 1.61-34.11; for left ventricular mass index, 0.39°; 0.10-0.67; for left atrial volume index, 0.91°; 0.24-1.58). Conclusions Adverse retinal arteriolar geometric morphology mirrored suboptimal cardiac structural alteration.