Tijs Louwies

Retinal Microvascular Responses to Short-Term Changes in Particulate Air Pollution in Healthy Adults

Background: Microcirculation plays an important role in the physiology of cardiovascular health. Air pollution is an independent risk factor for the development and progression of cardiovascular diseases, but the number of studies on the relation between air pollution and the microcirculation is limited. Objectives: We examined the relationship between short-term changes in air pollution and microvascular changes. Methods: We measured retinal microvasculature using fundus image analysis in a panel of 84 healthy adults (52% female), 22–63 years of age, during January–May 2012. Blood vessels were measured as central retinal arteriolar/venular equivalent (CRAE/CRVE), with a median of 2 measurements (range, 1–3). We used monitoring data on particulate air pollution (PM10) and black carbon (BC). Mixed-effect models were used to estimate associations between CRAE/CRVE and exposure to PM10 and BC using various exposure windows. Results: CRAE and CRVE were associated with PM10 and BC concentrations, averaged over the 24 hr before the retinal examinations. Each 10-µg/m3 increase in PM10 was associated with a 0.93-µm decrease (95% CI: –1.42, –0.45; p = 0.0003) in CRAE and a 0.86-µm decrease (95% CI: –1.42, –0.30; p = 0.004) in CRVE after adjusting for individual characteristics and time varying conditions such as ambient temperature. Each 1-µg/m3 increase in BC was associated with a 1.84-µm decrease (95% CI: –3.18, –0.51; p < 0.001) in CRAE. Conclusions: Our findings suggest that the retinal microvasculature responds to short-term changes in air pollution levels. These results support a mechanistic pathway through which air pollution can act as a trigger of cardiovascular events at least in part through effects on the microvasculature. Citation: Louwies T, Int Panis L, Kicinski M, De Boever P, Nawrot TS. 2013. Retinal microvascular responses to short-term changes in particulate air pollution in healthy adults. Environ Health Perspect 121:1011–1016; http://dx.doi.org/10.1289/ehp.1205721

Blood pressure changes in association with black carbon exposure in a panel of healthy adults are independent of retinal microcirculation

Exposure to ambient particulate matter and elevated blood pressure are risk factors for cardiovascular morbidity and mortality. Microvascular changes might be an important pathway in explaining the association between air pollution and blood pressure. The objective of the study was to evaluate the role of the retinal microcirculation in the association between black carbon (BC) exposure and blood pressure. We estimated subchronic BC exposure based on 1-week personal measurements (μ-Aethalometer, AethLabs) in 55 healthy nurses. Blood pressure and retinal microvasculature were measured on four different days (range: 2-4) during this week. Subchronic BC exposure averaged (± SD) 1334±631ng/m(3) and ranged from 338ng/m(3) to 3889ng/m(3). An increased exposure of 631ng/m(3) BC was associated with a 2.77mmHg (95% CI: 0.39 to 5.15, p=0.027) increase in systolic blood pressure, a 2.35mmHg (95% CI: 0.52 to 4.19, p=0.016) increase in diastolic blood pressure and with 5.65μm (95% CI: 1.33 to 9.96, p=0.014) increase in central retinal venular equivalent. Mediation analysis failed to reveal an effect of retinal microvasculature in the association between blood pressure and subchronic BC exposure. In conclusion, we found a positive association between blood pressure and subchronic black carbon exposure in healthy adults. This finding adds evidence to the association between black carbon exposure and cardiovascular health effects, with elevated blood pressure as a plausible intermediate effector. Our results suggest that the changes in a persons blood pressure as a result of subchronic black carbon exposure operate independently of the retinal microcirculation.

Short-term fluctuations in personal black carbon exposure are associated with rapid changes in carotid arterial stiffening

BACKGROUND Vascular changes may underpin the association between airborne black carbon (BC) and cardiovascular events. Accurate assessment of personal exposure is a major challenge in epidemiological research. BC concentrations are strongly related to time-activity patterns, which is particularly relevant when investigating short-term effects. We investigated associations between arterial stiffness and personal short-term BC exposure. METHODS This panel study included 54 healthy adults (92% women, mean age 40.7years). BC exposure was monitored individually with a micro-aethalometer during one workweek. Functional and structural properties of the carotid artery were examined ultrasonographically on two separate days. The effect of different short-term personal BC exposure windows (1, 2, 4, 6, 8, 24 and 48h before the ultrasound examination) on carotid artery stiffness was estimated using mixed models while adjusting for other known correlates of arterial stiffness. RESULTS Median personal BC exposures within the same day ranged from 599.8 to 728.9ng/m(3) and were associated with carotid arterial stiffness measures. Youngs elastic modulus and pulse wave velocity, both measures of stiffness, were positively associated with BC exposure, while the distensibility and compliance coefficient, measures of elasticity, were negatively associated with BC exposure. The strongest associations were observed with BC exposure 8h before the clinical examination. For each 100ng/m(3) increase in exposure within this time window, Youngs elastic modulus increased by 2.38% (95% CI: 0.81 to 3.97; P=0.0033), while the distensibility coefficient decreased by 2.27% (95% CI: -3.62 to -0.92; P=0.0008). CONCLUSIONS Short-term elevations in personal BC exposure, even within hours, are associated with increased arterial stiffness. This response may reflect a pathway by which air pollution triggers cardiovascular events.

miRNA expression profiles and retinal blood vessel calibers are associated with short-term particulate matter air pollution exposure.

INTRODUCTION Air pollution, a risk factor for cardiovascular diseases, can exert its effects through the microcirculation. Retinal blood vessel width is considered a marker for microvascular health and is associated with short-term PM10 exposure. microRNAs are key regulators of complex biological processes in cardiovascular health and disease and miRNA expression can be affected by air pollution exposure. Studies investigating the effect of ambient air pollution exposure on miRNA expression in combination with an assessment of the microvasculature do not exist. METHODS 50 healthy adults (50% women, 23-58 years old) were examined once a month from December 2014 until April 2015 in Flanders (Belgium). Fundus photos and venous blood samples were collected during the study visits. PM10 data were obtained from a nearby monitoring station. Image analysis was used to calculate the width of retinal blood vessels, represented as the Central Retinal Arteriolar/Venular Equivalent (CRAE/CRVE). Total miRNA was isolated from blood and the expression of miR-21, -146a and, -222 were measured using quantitative real-time PCR. Mixed models were used for statistical analysis. RESULTS Each short-term increase of 10µg/m(3) PM10 during the 24h preceding the study visit was associated with a 0.58µm decrease (95% CI: -1.16, -0.0005; p=0.056) in CRAE, a 0.99µm increase (95% CI: 0.18, 1.80; p=0.021) in CRVE, a 6.6% decrease (95% CI: -11.07, -2.17; p=0.0038) in miR-21 expression and a 6.7% decrease (95% CI: -10.70, -2.75; p=0.0012) in miR-222 expression. Each 10% increase in miR-21 was associated with a 0.14µm increase (95% CI: 0.0060, 0.24; p=0.046) in CRAE whereas a similar increase in miR-222 expression was associated with a 0.28µm decrease (95% CI: -0.50, -0.062; p=0.016) in CRVE. These associations were also found in exposure windows ranging from 2h to 1 week. Finally, we observed that the association between PM10 exposure and CRAE was mediated by miRNA-21 expression. CONCLUSION PM10 exposure was associated with retinal arteriolar narrowing and venular widening. PM10 exposure affected miRNAs that are involved in inflammatory and oxidative stress pathways. We suggest that miRNA changes may be relevant to explain the association between PM10 and retinal vessel calibers.

Fundus Photography as a Convenient Tool to Study Microvascular Responses to Cardiovascular Disease Risk Factors in Epidemiological Studies

The microcirculation consists of blood vessels with diameters less than 150 µm. It makes up a large part of the circulatory system and plays an important role in maintaining cardiovascular health. The retina is a tissue that lines the interior of the eye and it is the only tissue that allows for a non-invasive analysis of the microvasculature. Nowadays, high-quality fundus images can be acquired using digital cameras. Retinal images can be collected in 5 min or less, even without dilatation of the pupils. This unobtrusive and fast procedure for visualizing the microcirculation is attractive to apply in epidemiological studies and to monitor cardiovascular health from early age up to old age. Systemic diseases that affect the circulation can result in progressive morphological changes in the retinal vasculature. For example, changes in the vessel calibers of retinal arteries and veins have been associated with hypertension, atherosclerosis, and increased risk of stroke and myocardial infarction. The vessel widths are derived using image analysis software and the width of the six largest arteries and veins are summarized in the Central Retinal Arteriolar Equivalent (CRAE) and the Central Retinal Venular Equivalent (CRVE). The latter features have been shown useful to study the impact of modifiable lifestyle and environmental cardiovascular disease risk factors. The procedures to acquire fundus images and the analysis steps to obtain CRAE and CRVE are described. Coefficients of variation of repeated measures of CRAE and CRVE are less than 2% and within-rater reliability is very high. Using a panel study, the rapid response of the retinal vessel calibers to short-term changes in particulate air pollution, a known risk factor for cardiovascular mortality and morbidity, is reported. In conclusion, retinal imaging is proposed as a convenient and instrumental tool for epidemiological studies to study microvascular responses to cardiovascular disease risk factors.

GLI2 promoter hypermethylation in saliva of children with a respiratory allergy

BackgroundThe prevalence of respiratory allergy in children is increasing. Epigenetic DNA methylation changes are plausible underlying molecular mechanisms.ResultsSaliva samples collected in substudies of two longitudinal birth cohorts in Belgium (FLEHS1 & FLEHS2) were used to discover and confirm DNA methylation signatures that can differentiate individuals with respiratory allergy from healthy subjects. Genome-wide analysis with Illumina Methylation 450K BeadChips revealed 23 differentially methylated gene regions (DMRs) in saliva from 11y old allergic children (N=26) vs. controls (N=20) in FLEHS1. A subset of 7 DMRs was selected for confirmation by iPLEX MassArray analysis. First, iPLEX analysis was performed in the same 46 FLEHS1 samples for analytical confirmation of the findings obtained during the discovery phase. iPLEX results correlated significantly with the 450K array data (P <0.0001) and confirmed 4 out of the 7 DMRs. Aiming for additional biological confirmation, the 7 DMRs were analyzed using iPLEX in a substudy of an independent birth cohort (FLEHS2; N=19 cases vs. 20 controls, aged 5 years). One DMR in the GLI2 promoter region showed a consistent statistically significant hypermethylation in individuals with respiratory allergy across the two birth cohorts and technologies. In addition to its involvement in TGF-β signaling and T-helper differentiation, GLI2 has a regulating role in lung development.ConclusionGLI2 is considered an interesting candidate DNA methylation marker for respiratory allergy.

Recent versus chronic fine particulate air pollution exposure as determinant of the retinal microvasculature in school children

Background: Microvascular changes may represent an underlying mechanism through which exposure to fine particulate matter with a diameter ≤ 2.5 &mgr;m (PM2.5) contributes to age‐related disease development. We investigated the effect of recent and chronic exposure to PM2.5 on the microcirculation, exemplified by retinal vessel diameters, using repeated measurements in 8‐ to 12‐year‐old children. Methods: 221 children (49.1% girls; mean age 9.9 years) were examined repeatedly (25 one, 124 two, and 72 three times) adding up to 489 retinal vessel examinations. Same‐day exposure to PM2.5 was measured at school. In addition, recent (same and previous day) and chronic (yearly mean) exposure was modelled at the childs residence using a high‐resolution interpolation model. Residential proximity to major roads was also assessed. Changes in retinal vessel diameters associated with recent and chronic exposures were estimated using mixed models, while adjusting for other known covariates such as sex, age, BMI, blood pressure and birth weight. Results: Each 10 &mgr;g/m³ increment in same‐day exposure to PM2.5 measured at school was associated with 0.35 &mgr;m (95% CI: 0.09–0.61 &mgr;m) narrower retinal arterioles and 0.35 &mgr;m (−0.03 to 0.73 &mgr;m) wider venules. Children living 100 m closer to a major road had 0.30 &mgr;m (0.05–0.54 &mgr;m) narrower arterioles. Conclusions: Blood vessel diameters of the retinal microcirculation of healthy school‐aged children respond to same‐day PM2.5 exposure. Furthermore, children living closer to major roads had smaller arteriolar diameters. Our results suggest that the microcirculation, with retinal microvasculature as a proxy in this study, is a pathophysiological target for air pollution in children. HighlightsChildrens retinal blood vessel diameters respond to recent PM2.5 exposure.Repeated design with multiple retinal microvascular examinations in each child.Integrative exposure assessment at school and at home.Microvascular changes may be an early phenotypic marker for disease development.

Separate and Combined Effects of Hypoxia and Horizontal Bed Rest on Retinal Blood Vessel Diameters

Purpose To assess the separate and combined effects of exposure to prolonged and sustained recumbency (bed rest) and hypoxia on retinal microcirculation. Methods Eleven healthy male subjects (mean ± SD age = 27 ± 6 years; body mass index [BMI] = 23.7 ± 3.0 kg m-2) participated in a repeated-measures crossover design study comprising three 21-day interventions: normoxic bed rest (NBR; partial pressure of inspired O2, PiO2 = 133.1 ± 0.3 mm Hg); hypoxic ambulation (HAMB; PiO2 = 90.0 ± 0.4 mm Hg), and hypoxic bed rest (HBR; PiO2 = 90.0 ± 0.4 mm Hg). Central retinal arteriolar (CRAE) and venular (CRVE) equivalents were measured at baseline and at regular intervals during each 21-day intervention. Results Normoxic bed rest caused a progressive reduction in CRAE, with the change in CRAE relative to baseline being highest on day 15 (ΔCRAE = -7.5 μm; 95% confidence interval [CI]: -10.8 to -4.2; P < 0.0001). Hypoxic ambulation resulted in a persistent 21-day increase in CRAE, reaching a maximum on day 4 (ΔCRAE = 9.4 μm; 95% CI: 6.0-12.7; P < 0.0001). During HBR, the increase in CRAE was highest on day 3 (ΔCRAE = 4.5 μm; 95% CI: 1.2-7.8; P = 0.007), but CRAE returned to baseline levels thereafter. Central retinal venular equivalent decreased during NBR and increased during HAMB and HBR. The reduction in CRVE during NBR was highest on day 1 (ΔCRVE = -7.9 μm; 95 CI: -13.3 to -2.5), and the maximum ΔCRVE during HAMB (24.6 μm; 95% CI: 18.9-30.3) and HBR (15.2 μm; 95% CI: 9.8-20.5) was observed on days 10 and 3, respectively. Conclusions The diameters of retinal blood vessels exhibited a dynamic response to hypoxia and bed rest, such that retinal vasodilation was smaller during combined bed rest and hypoxia than during hypoxic exposure.

Microvascular responses in association with recent and chronic exposure to particulate air pollution in school children

Methods 225 children (49% girls; mean age 9.9 years) were recruited at two primary schools in Belgium. Participating children were examined three times at school over the course of the school year, during which the fundus of both eyes was photographed. The caliber of the retinal blood vessels was summarized as the central retinal arteriolar/venular equivalent (CRAE/CRVE). Recent exposure to PM2.5 was measured at the school prior to the examination. Residential proximity to major roads was used as a proxy for chronic exposure. The effect of recent and chronic exposure to PM on retinal vessel caliber was estimated using mixed models, while adjusting for gender, age, BMI, blood pressure, heart rate, birth weight, time and day of examination, mothers education and passive smoking.

Changes in miRNA expression and retinal blood vessels are associated with short-term air pollution exposure.

Air pollution, a cardiovascular risk factor, might exert its effects through the microcirculation. Fundus photography allows study of the retinal vasculature in vivo. Short-term PM10 exposure is associated with changes in retinal blood vessels, but the underlying mechanism remains unknown. Expression of MIR21, MIR222 and MIR146A, gene regulators involved in oxidative stress and inflammatory processes, can be changed by air pollution and might be a pathway explaining the association between PM10 and microvascular changes. 50 healthy adults (50% women, 50% men, 23-58 years old) were sampled once a month from December 2014 until April 2015. At each study visit fundus photos and venous blood samples were collected. PM10 data were obtained from a nearby monitoring station. Image analysis was used to calculate the width of retinal blood vessels, represented as the Central Retinal Arteriolar/Venular Equivalent (CRAE/CRVE). miRNA was isolated from blood and expression was measured using qRT-PCR. Mixed models were used for statistical analysis. Short-term changes in PM10 exposure were associated with changes in CRAE, CRVE and miRNA-expression. Each 10 μg/m3 increase in PM10 during the previous 24 hours was associated with a 0.58 μm decrease (95% CI: -1.16, -0.0005; p=0.056) in CRAE, a 0.99 μm increase (95% CI: 0.18, 1.80; p=0.021) in CRVE, a 6.6% decrease (95% CI: -11.07, -2.17; p=0.0038) in miR-21 expression and a 6.7% decrease (95% CI: -10.70, -2.75; p=0.0012) in miR-222 expression. miRNA expression was associated with CRAE and CRVE. Each 10% increase in miR-21 expression and miR-222 was associated with respectively a 0.14 μm increase (95% CI: 0.0060, 0.24; p=0.046) in CRAE and a 0.28 μm decrease (95% CI: -0.50, -0.062; p=0.016) in CRVE. PM10 exposure affects miRNAs involved in inflammation and oxidative stress. These changes may be an underlying mechanism for the association between PM10 exposure and retinal arteriolar narrowing and venular widening.