Exercise and microvascular health in an ageing population : the EXAMIN AGE study

Background Cardiovascular (CV) disease remains a major health care burden worldwide. Exercise has a preventive effect on CV disease risk. Risk factors for CV disease include higher age, higher inactivity levels as well as reduced cardiorespiratory fitness. The retinal microcirculation is a valid vascular bed to detect vascular alterations in an early and subclinical stage. Alterations in retinal microvascular phenotype, defined as narrower central retinal arteriolar equivalents(CRAE), wider central retinal venular equivalents (CRVE) as well as reduced flicker light induced dilatation (FID), are associated with increased CV risk. Reactive oxygen species (ROS)production, modulated by DNA methylation of p66Shc, is a key driver for vascular alterations. To date no study exists that investigates the influence of physical activity (PA) or the effect of an exercise intervention on the ageing process of the retinal microcirculation in healthy individuals and patients with increased CV risk. Aims The aims of my PhD project were: 1) to investigate the association of long-term PA or inactivity on retinal microvascular phenotype in healthy older individuals, 2) to investigate the association of CV risk on retinal microvascular phenotype in long-term physical inactive older individuals and 3) to investigate the effects of twelve-weeks HIIT on retinal microvascular phenotype in older CV risk patients. Methods This PhD based on the “Exercise, Arterial Crosstalk-Modulation, and Inflammation in an Ageing Population” study (EXAMIN AGE). This study investigated the exercise effects in a systems physiology approach with a cross-sectional and an interventional study design. In the crosssectional approach 38 healthy active (HA), 36 healthy sedentary (HS) and 84 sedentary individuals at increased CV risk (SR) were included. SR were randomised into a twelve-week high-intensity interval training (HIIT) or a control condition with standard PA recommendations after the baseline assessment. The Retinal Vessel Analyser was used to measure the retinal microvascular phenotype. Enzyme-linked immunosorbent assay kits were used to analyse plasma 3-nitrotyrosine (3-NT) as a marker of oxidative stress. Gene expression of p66Shc and DNA methylation analysis were assessed in mononuclear cells by real-time quantitative polymerase chain reaction and Methylminer quantitative polymerase chain reaction to detect the epigenetic pathway of oxidative stress, one potential mechanism that affect retinal microvascular phenotype. Results Our results demonstrated wider CRAE and narrower CRVE in HA compared to HS resulting in a higher arteriolar-to-venular diameter ratio (AVR). By contrast, SR showed narrower CRAE and wider CRVE compared to HS resulting in a lower AVR compared to HS and HA. HS showed higher FID compared to SR and HA. FID in SR and HA did not significantly differ. A significant correlation between CRVE and maximal oxygen consumption (VO2peak) as well as between AVR and VO2peak were observed. In both sedentary groups, higher p66Shc expression and increased plasma levels of 3-NT were associated with hypomethylation of p66Shc promoter. HIIT reduced body mass index, fat mass, low-density lipoprotein and increased muscle mass and VO2peak. HIIT increased CRAE, decreased CRVE and increased arteriolar FID compared to the control group. A significant association between ΔCRAE and ΔVO2peak, ΔAVR and ΔVO2peak as well as between Δarteriolar FID and ΔVO2peak were observed. HIIT restored promoter methylation, blunting p66Shc expression and 3-NT levels. Conclusion Higher PA seems to be associated with favourable microvascular phenotype compared to sedentary individuals, with a further decline in sedentary individuals with increased CV risk. However, the use of FID seems to be limited in highly active individuals, eventually due to predilated arterioles. Therefore, our recommendation is to combine FID with analysis of retinal vessel diameters to differentiate functional non-responders from manifest microvascular endothelial dysfunction and thereby improve individual microvascular risk stratification. Exercise treatment has the potential to counteract microvascular dysfunction in older patients at increased CV risk. Exercise-induced reprogramming of DNA methylation on p66Shc gene promoter may represent a putative mechanistic link whereby exercise protects against age-related oxidative stress. Retinal vessel analysis seems to be a sensitive tool for detecting longterm PA as well as short-term exercise effects on retinal microvascular health in an ageing population

Retinal endothelial function, physical fitness and cardiovascular risk: a diagnostic challenge

Introduction: Dynamic retinal vessel analysis (DVA) is a new non-invasive method to quantify microvascular endothelial dysfunction by flicker light-induced dilatation (FID). FID has been shown to be impaired in type 2 diabetes as well as heart failure. The aim of the study was to analyze FID in healthy active versus healthy sedentary and cardiovascular (CV) risk patients in addition to corresponding static vessel diameters. Methods: Thirty-one healthy active (HA, mean age 60 +/- 8 years), 33 healthy sedentary individuals (HS, 59 +/- 7 years) and 76 sedentary patients with increased CV risk (SR, 58 +/- 6 years) were included in this cross-sectional study. Group differences in CV risk factors and cardiorespiratory fitness, maximal arteriolar (ADmax) and venular (VDmax) dilatation as well as the arteriolar (AFarea) and venular (VFarea) area under the flicker curve were analyzed. The central retinal arteriolar and venular diameters were used to calculate the arteriolar-to-venular diameter ratio (AVR). Results: HS [ADmax = 3.5 (2.1)%; AFarea = 48.2 (31.9)%*s] showed higher FID compared to SR [ADmax = 2.7 (1.8)%, p = 0.021; AFarea = 34.5 (26.5)%*s, p = 0.006] and HA [AFarea = 32.8 (23.1)%*s, p = 0.029]. HA and SR did not significantly differ. HA had a higher AVR (0.87 +/- 0.05) compared to HS (0.83 +/- 0.04, p < 0.001) with further deterioration in SR (0.79 +/- 0.05, p < 0.001). Interestingly, 28 participants had impaired FID but normal AVR and 43 participants had normal FID but impaired AVR. Discussion: FID can differentiate between sedentary low and high risk individuals. However, FID in healthy active persons (HA) seemed impaired with a concomitant higher AVR. We postulate that lower FID in HA may be explained by predilatated arterioles and a reduced dilatation reserve. We recommend combination of FID with analysis of retinal vessel diameters to differentiate functional non-responders from manifest microvascular endothelial dysfunction and, thereby, improve microvascular risk stratification in a personalized medicine approach. Clinical Trial Registration: ClinicalTrials.gov: NCT02796976 (https://clinicaltrials.gov/ct2/show /NCT02796976)

Exercise, Arterial Crosstalk-Modulation, and Inflammation in an Aging Population: The ExAMIN AGE Study

Background: Age is a key determinant for the development of cardiovascular disease and higher age coincides with an increased prevalence of obesity and physical inactivity. The study examines the influence of physical activity on aging processes of physiological systems focusing on the mechanisms of vascular aging. Methods/Design: The study consists of two parts. The cross-sectional approach aims at examining the association of physical fitness and cardiovascular risk with large and small artery function in healthy older active (HOA, n = 40) and sedentary (HOS, n = 40) persons as well as older sedentary individuals with increased cardiovascular risk (OSR, n = 80) aged 50–80 years. In the interventional approach, the OSR group is randomized into a 12-week walking-based high intensity interval training (HIIT) group or a control condition, aiming at examining the effects of HIIT on arterial function in diseased older adults. Active lifestyle is defined as &gt;9 metabolic equivalent of task (MET) per week and sedentary as ≤3 MET/week. Inclusion criteria for OSR are overweight or obesity (body mass index ≥30 kg/m2) plus at least one additional cardiovascular risk factor. The primary outcome is arterial stiffness as determined by aortic pulse wave velocity (PWV). The secondary outcomes are retinal arterial and venous diameters. Further cardiovascular assessments include peripheral PWV, central haemodynamics, retinal endothelial function, carotid intima media thickness, cardiac strain and diastolic function as well as autonomic function and inflammation. Physical fitness is measured by a treadmill-based spiroergometry to determine peak oxygen uptake. Discussion: The aim of the study is to demonstrate the importance of and need for specific physical activity programs for seniors to achieve healthier aging as a longterm goal. Vascular function defines disease- and age-related end organ damage and represents the potential to contain health at older age. This research will identify cardiovascular biomarkers that best resemble underlying cardiovascular risk in age and disease. The integrated approach will help define new recommendations for treatment guidance of exercise therapy in an aging population

Cardiorespiratory fitness and development of childhood cardiovascular risk: The EXAMIN YOUTH follow-up study

Background: Obesity- and hypertension-related cardiovascular (CV) risk has been shown to originate in childhood. Higher body mass index (BMI) and blood pressure (BP) have been associated with increased large artery stiffness and a lower microvascular arteriolar-to-venular diameter ratio (AVR) in children. This study aimed to investigate the association of cardiorespiratory fitness (CRF) with development of BMI, BP and vascular health during childhood.Methods: In our prospective cohort study, 1,171 children aged 6–8 years were screened for CRF, BMI, BP, retinal vessel diameters and pulse wave velocity using standardized protocols. Endurance capacity was assessed by 20 m shuttle run test. After 4 years, all parameters were assessed in 664 children using the same protocols.Results: Children with a higher CRF at baseline developed a significantly lower BMI (β [95% CI] −0.09 [−0.11 to −0.06] kg/m2, p &lt; 0.001), a lower systolic BP (β [95% CI] −0.09 [−0.15 to −0.03] mmHg, p = 0.004) and a higher AVR (β [95% CI] 0.0004 [0.00004 to 0.0007] units, p = 0.027) after 4 years. The indirect association of CRF with development of retinal arteriolar diameters was mediated by changes in BMI.Conclusion: Our results identify CRF as a key modulator for the risk trajectories of BMI, BP and microvascular health in children. Obesity-related CV risk has been shown to track into adulthood, and achieving higher CRF levels in children may help counteract the development of CV risk and disease not only in pediatric populations, but may also help reduce the burden of CVD in adulthood.Registration:http://www.clinicaltrials.gov/ (NCT02853747)

Short-term high-intensity interval training improves micro- but not macrovascular function in hypertensive patients

Arterial hypertension is a global health burden that affects vascular structure and function. Assessment of endothelial function can improve cardiovascular (CV) risk stratification. Exercise treatment reduces over all CV risk and improves vascular health. However, it is still not clear which part of the vascular bed is most sensitive to exercise treatment in patients with CV risk. This study aimed to investigate the effects of an 8-week walking based and supervised high-intensity interval training (HIIT) on macro- and microvascular endothelial function as add-on therapy in patients with arterial hypertension.; Forty patients (mean age 58 ± 7 years) treated for arterial hypertension were randomized in the HIIT (3×/week) or control group (CG) receiving standard physical activity recommendations. Arteriolar (aFID) and venular (vFID) flicker light-induced dilatation for retinal microvascular and flow-mediated dilatation (FMD) for macrovascular endothelial function were assessed. In addition, standardized assessments of patients' characteristics were performed before and after 8 weeks.; Both groups reduced weight and body mass index but only the HIIT group reduced body fat, visceral fat, and increased peak oxygen uptake after 8 weeks. The control group reduced diastolic blood pressure. No blood pressure changes were found in the HIIT group. Arteriolar FID increased in the HIIT group independently of confounders (pre: 2.40 ± 0.98%, post: 3.19 ± 1.31%, p < 0.001) but not in the control group (pre: 3.06 ± 1.50%, post: 2.90 ± 1.46%, p = 0.280). No changes were found for FMD in either group.; Arteriolar FID was found to be a sensitive vascular biomarker to assess exercise-induced microvascular improvements even in a short time setting of an 8-week exercise therapy with HIIT. Short-term exercise training affects microvascular endothelial function but not large artery endothelial function. Thus, retinal aFID appears to be a sensitive biomarker to detect short-term exercise efficacy on a vascular level. Dynamic retinal vessel analysis as a diagnostic approach may prove to be an ideal candidate vascular biomarker to monitor treatment effects of exercise in patients with hypertension on top of standard clinical care and may support clinical decision-making in the future

Untargeted sequencing of circulating microRNAs in a healthy and diseased older population

We performed untargeted profiling of circulating microRNAs (miRNAs) in a well characterized cohort of older adults to verify associations of health and disease-related biomarkers with systemic miRNA expression. Differential expression analysis revealed 30 miRNAs that significantly differed between healthy active, healthy sedentary and sedentary cardiovascular risk patients. Increased expression of miRNAs miR-193b-5p, miR-122-5p, miR-885-3p, miR-193a-5p, miR-34a-5p, miR-505-3p, miR-194-5p, miR-27b-3p, miR-885-5p, miR-23b-5b, miR-365a-3p, miR-365b-3p, miR-22-5p was associated with a higher metabolic risk profile, unfavourable macro- and microvascular health, lower physical activity (PA) as well as cardiorespiratory fitness (CRF) levels. Increased expression of miR-342-3p, miR-1-3p, miR-92b-5p, miR-454-3p, miR-190a-5p and miR-375-3p was associated with a lower metabolic risk profile, favourable macro- and microvascular health as well as higher PA and CRF. Of note, the first two principal components explained as much as 20% and 11% of the data variance. miRNAs and their potential target genes appear to mediate disease- and health-related physiological and pathophysiological adaptations that need to be validated and supported by further downstream analysis in future studies.Clinical Trial Registration: ClinicalTrials.gov: NCT02796976 ( https://clinicaltrials.gov/ct2/show/NCT02796976 )

How Ceramides Orchestrate Cardiometabolic Health-An Ode to Physically Active Living

Cardiometabolic diseases (CMD) represent a growing socioeconomic burden and concern for healthcare systems worldwide. Improving patients' metabolic phenotyping in clinical practice will enable clinicians to better tailor prevention and treatment strategy to individual needs. Recently, elevated levels of specific lipid species, known as ceramides, were shown to predict cardiometabolic outcomes beyond traditional biomarkers such as cholesterol. Preliminary data showed that physical activity, a potent, low-cost, and patient-empowering means to reduce CMD-related burden, influences ceramide levels. While a single bout of physical exercise increases circulating and muscular ceramide levels, regular exercise reduces ceramide content. Additionally, several ceramide species have been reported to be negatively associated with cardiorespiratory fitness, which is a potent health marker reflecting training level. Thus, regular exercise could optimize cardiometabolic health, partly by reversing altered ceramide profiles. This short review provides an overview of ceramide metabolism and its role in cardiometabolic health and diseases, before presenting the effects of exercise on ceramides in humans

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

Investigating the circulating sphingolipidome response to a single high-intensity interval training session within healthy females and males in their twenties (SphingoHIIT): Protocol for a randomised controlled trial [version 2; peer review: 2 approved]

Introduction: Growing scientific evidence indicates that sphingolipids predict cardiometabolic risk, independently of and beyond traditional biomarkers such as low-density lipoprotein cholesterol. To date, it remains largely unknown if and how exercise, a simple, low-cost, and patient-empowering modality to optimise cardiometabolic health, influences sphingolipid levels. The SphingoHIIT study aims to assess the response of circulating sphingolipid species to a single session of high-intensity interval training (HIIT). Methods: This single-centre randomised controlled trial (RCT) will last 11 days per participant and aim to include 32 young and healthy individuals aged 20-29 (50% females). Participants will be randomly allocated to the HIIT (n= 16) or control groups (physical rest, n= 16). Participants will self-sample fasted dried blood spots for three consecutive days before the intervention (HIIT versus rest) to determine baseline sphingolipid levels. Dried blood spots will also be collected at five time points (2, 15, 30, 60min, and 24h) following the intervention (HIIT versus rest). To minimise the dietary influence, participants will receive a standardised diet for four days, starting 24 hours before the first dried blood sampling. For females, interventions will be timed to fall within the early follicular phase to minimise the menstrual cycle's influence on sphingolipid levels. Finally, physical activity will be monitored for the whole study duration using a wrist accelerometer. Ethics and dissemination: The Ethics Committee of Northwest and Central Switzerland approved this protocol (ID 2022–00513). Findings will be disseminated in scientific journals and meetings. Trial Registration The trial was registered on www.clinicaltrials.gov (NCT05390866, https://clinicaltrials.gov/ct2/show/NCT05390866) on May 25, 2022