Platelets: From Formation to Function

Platelets are small, anucleate cells that travel as resting discoid fragments in the circulation. Their average circulating life span is 8–9 days, and their formation is an elegant and finely orchestrated series of cellular processes known as megakaryocytopoiesis and thrombopoiesis. This involves the commitment of haematopoietic stem cells, proliferation, terminal differentiation of megakaryocytic progenitors and maturation of megakaryocytes to produce functional platelets. This complex process occurs in specialised endosteal and vascular niches in the bone marrow where megakaryocytes form proplatelet projections, releasing platelets into the circulation. Upon contact with an injured blood vessel, they prevent blood loss through processes of adhesion, activation and aggregation. Platelets play a central role in cardiovascular disease (CVD), both in the development of atherosclerosis and as the cellular mediator in the development of thrombosis. Platelets have diverse roles not limited to thrombosis/haemostasis, also being involved in many vascular inflammatory conditions. Depending on the physiological context, platelet functions may be protective or contribute to adverse thrombotic and inflammatory outcomes. In this chapter, we will discuss platelets in context of their formation and function. Because of their multifaceted role in maintaining physiological homeostasis, current and development of platelet function testing platforms will be discussed

Platelets: Functional Biomarkers of Epigenetic Drift

Cardiovascular disease (CVD) risk factors can be classed as modifiable or non-modifiable. Physical inactivity and obesity represent major behavioural risk factors for the initiation, development and progression of CVD. Platelet dysfunction is pivotal to the aetiology of CVD, a chronic vascular inflammatory condition, which is characterised by a lag time between onset and clinical manifestation. This indicates the role of epigenetic drift, defined by stochastic patterns of gene expression not dependent on dynamic changes in coding DNA. The epigenome, a collection of chemical marks on DNA and histones, is established during embryogenesis and modified by age and lifestyle. Biogenesis and effector function of non-coding RNA, such as microRNA, play a regulatory role in gene expression and thus the epigenetic mechanism. In this chapter, we will focus on the effect of the modifiable risk factors of physical activity/inactivity and overweight/obesity on platelet function, via epigenetic changes in both megakaryocytopoiesis and thrombopoiesis. We will also discuss the role of acute exercise on platelet function and the impact of cardiorespiratory fitness (CRF) on platelet responses to acute exercise. This chapter will highlight the potential role of platelets as circulating functional biomarkers of epigenetic drift to implement, optimise and monitor CVD preventive management strategies

The pathogenic c.1171A>G (p.Arg391Gly) and c.2359G>A (p.Val787Ile) ABCC6 variants display incomplete penetrance causing pseudoxanthoma elasticum in a subset of individuals

ABCC6 promotes ATP efflux from hepatocytes to bloodstream. ATP is metabolized to pyrophosphate, an inhibitor of ectopic calcification. Pathogenic variants of ABCC6 cause pseudoxanthoma elasticum, a highly variable recessive ectopic calcification disorder. Incomplete penetrance may initiate disease heterogeneity, hence symptoms may not, or differently manifest in carriers. Here, we investigated whether incomplete penetrance is a source of heterogeneity in pseudoxanthoma elasticum. By integrating clinical and genetic data of 589 patients, we created the largest European cohort. Based on allele frequency alterations, we identified two incomplete penetrant pathogenic variants, c.2359G>A (p.Val787Ile) and c.1171A>G (p.Arg391Gly), with 6.5% and 2% penetrance, respectively. However, when penetrant, the c.1171A>G (p.Arg391Gly) manifested a clinically unaltered severity. After applying in silico and in vitro characterization, we suggest that incomplete penetrant variants are only deleterious if a yet unknown interacting partner of ABCC6 is mutated simultaneously. The low penetrance of these variants should be contemplated in genetic counseling

Plasma Inorganic Pyrophosphate Deficiency Links Multiparity to Cardiovascular Disease Risk.

Epidemiological studies indicate that elevated alkaline phosphatase activity is associated with increased cardiovascular disease risk. Other epidemiological data demonstrate that mothers giving multiple childbirths (multipara) are also at increased risk of developing late-onset cardiovascular disease. We hypothesized that these two associations stem from a common cause, the insufficient plasma level of the ectopic mineralization inhibitor inorganic pyrophosphate, which is a substrate of alkaline phosphatase. As alkaline phosphatase activity is elevated in pregnancy, we hypothesized that pyrophosphate concentrations decrease gestationally, potentially leading to increased maternal vascular calcification and cardiovascular disease risk in multipara. We investigated plasma pyrophosphate kinetics pre- and postpartum in sheep and at term in humans and demonstrated its shortage in pregnancy, mirroring alkaline phosphatase activity. Next, we tested whether multiparity is associated with increased vascular calcification in pseudoxanthoma elasticum patients, characterized by low intrinsic plasma pyrophosphate levels. We demonstrated that these patients had increased vascular calcification when they give birth multiple times. We propose that transient shortages of pyrophosphate during repeated pregnancies might contribute to vascular calcification and multiparity-associated cardiovascular disease risk threatening hundreds of millions of healthy women worldwide. Future trials are needed to assess if gestational pyrophosphate supplementation might be a suitable prophylactic treatment to mitigate maternal cardiovascular disease risk in multiparous women

Contribution of Social Isolation, Restraint, and Hindlimb Unloading to Changes in Hemodynamic Parameters and Motion Activity in Rats

The most accepted animal model for simulation of the physiological and morphological consequences of microgravity on the cardiovascular system is one of head-down hindlimb unloading. Experimental conditions surrounding this model include not only head-down tilting of rats, but also social and restraint stresses that have their own influences on cardiovascular system function. Here, we studied levels of spontaneous locomotor activity, blood pressure, and heart rate during 14 days under the following experimental conditions: cage control, social isolation in standard rat housing, social isolation in special cages for hindlimb unloading, horizontal attachment (restraint), and head-down hindlimb unloading. General activity and hemodynamic parameters were continuously monitored in conscious rats by telemetry. Heart rate and blood pressure were both evaluated during treadmill running to reveal cardiovascular deconditioning development as a result of unloading. The main findings of our work are that: social isolation and restraint induced persistent physical inactivity, while unloading in rats resulted in initial inactivity followed by normalization and increased locomotion after one week. Moreover, 14 days of hindlimb unloading showed significant elevation of blood pressure and slight elevation of heart rate. Hemodynamic changes in isolated and restrained rats largely reproduced the trends observed during unloading. Finally, we detected no augmentation of tachycardia during moderate exercise in rats after 14 days of unloading. Thus, we concluded that both social isolation and restraint, as an integral part of the model conditions, contribute essentially to cardiovascular reactions during head-down hindlimb unloading, compared to the little changes in the hydrostatic gradient

MECANISMES DE LA DYSFONCTION VASCULAIRE INDUITE PAR DES MODELES DE MICROGRAVITE

Gravitational stimulation is an inherent part of terrestrial environment. Modified gravity changes functioning of numerous body systems, including cardiovascular system. The aim of this work was to study the mechanisms of vascular modifications induced by modeled microgravity. The human studies were performed using dry immersion. This model is considered as one of the most appropriate to reproduce the effects of prolonged microgravity and especially to mimic the lack of support. We found that dry immersion induces microcirculatory impairment with endothelial dysfunction (study of circulating microparticles) and impaired skin vasodilation (study by iontophoresis of vasoactive substances coupled with laser Doppler flowmetry). This vascular impairment is integrated to more general cardiovascular deconditioning syndrome. The global deconditioning was also considered in this work, including the rapid reset of water and sodium balance to a new steady state. During the recovery period, we observed a significant increase of plasmatic NT-proBNP which might reflect the degree of cardiovascular deconditioning. Our studies with animal models of horizontal and antiorthostatic hypokinesia suggest that increased endotoxin translocation and reduced energy metabolism in microgravity could contribute to endothelial impairment. Endothelial dysfunction may be regarded as one of the therapeutic targets for pathologies associated with physical inactivity and microgravity.La gravité est une stimulation permanente pour l'organisme sur terre. La microgravité induit des modifications de nombreuses fonctionnalités, y compris celles du système cardio-vasculaire. Le but de ce travail était d‟étudier les mécanismes des modifications vasculaires induites par les modèles de microgravité. Nous avons conduit nos études chez l‟homme à l‟aide de l‟immersion sèche qui est considérée comme un des modèles reproduisant au mieux la microgravité. L'immersion sèche est le seul modèle de microgravité prolongée à imiter l'absence d‟appui. Notre travail montre que l‟immersion sèche induit une atteinte au niveau microcirculatoire avec une dysfonction endothéliale (étude des microparticules circulantes) et une atteinte de la fonction vasodilatatrice cutanée (étude par iontophorèse couplée au laser Doppler). Cette atteinte vasculaire s'intègre dans un syndrome de déconditionnement cardiovasculaire plus global que nous avons également étudié. Il comprend un nouvel équilibre hydro-sodé qui survient très rapidement, dans les 24 premières heures. Pendant la période de récupération, nous avons observé une augmentation significative du NT-proBNP qui pourrait être un marqueur biologique du déconditionnement cardio-vasculaire. Nos études sur l‟animal avec les modèles d‟hypokinésie et d'inclinaison tête en bas discontinues chroniques suggèrent qu'une translocation augmentée d‟endotoxine et une réduction du métabolisme énergétique en microgravité pourraient participer aux atteintes endothéliales. La dysfonction endothéliale deviendrait ainsi une cible thérapeutique pour des atteintes liées à l‟inactivité physique et à la microgravité

Mecanismes de la dysfonction vasculaire induite par des modèles de microgravité

La gravité est une stimulation permanente pour l'organisme sur terre. La microgravité induit des modifications de nombreuses fonctionnalités, y compris celles du système cardio-vasculaire. Le but de ce travail était d étudier les mécanismes des modifications vasculaires induites par les modèles de microgravité. Nous avons conduit nos études chez l homme à l aide de l immersion sèche qui est considérée comme un des modèles reproduisant au mieux la microgravité. L'immersion sèche est le seul modèle de microgravité prolongée à imiter l'absence d appui. Notre travail montre que l immersion sèche induit une atteinte au niveau microcirculatoire avec une dysfonction endothéliale (étude des microparticules circulantes) et une atteinte de la fonction vasodilatatrice cutanée (étude par iontophorèse couplée au laser Doppler). Cette atteinte vasculaire s'intègre dans un syndrome de déconditionnement cardiovasculaire plus global que nous avons également étudié. Il comprend un nouvel équilibre hydro-sodé qui survient très rapidement, dans les 24 premières heures. Pendant la période de récupération, nous avons observé une augmentation significative du NT-proBNP qui pourrait être un marqueur biologique du déconditionnement cardio-vasculaire. Nos études sur l animal avec les modèles d hypokinésie et d'inclinaison tête en bas discontinues chroniques suggèrent qu'une translocation augmentée d endotoxine et une réduction du métabolisme énergétique en microgravité pourraient participer aux atteintes endothéliales. La dysfonction endothéliale deviendrait ainsi une cible thérapeutique pour des atteintes liées à l inactivité physique et à la microgravité.Gravitational stimulation is an inherent part of terrestrial environment. Modified gravity changes functioning of numerous body systems, including cardiovascular system. The aim of this work was to study the mechanisms of vascular modifications induced by modeled microgravity. The human studies were performed using dry immersion. This model is considered as one of the most appropriate to reproduce the effects of prolonged microgravity and especially to mimic the lack of support. We found that dry immersion induces microcirculatory impairment with endothelial dysfunction (study of circulating microparticles) and impaired skin vasodilation (study by iontophoresis of vasoactive substances coupled with laser Doppler flowmetry). This vascular impairment is integrated to more general cardiovascular deconditioning syndrome. The global deconditioning was also considered in this work, including the rapid reset of water and sodium balance to a new steady state. During the recovery period, we observed a significant increase of plasmatic NT-proBNP which might reflect the degree of cardiovascular deconditioning. Our studies with animal models of horizontal and antiorthostatic hypokinesia suggest that increased endotoxin translocation and reduced energy metabolism in microgravity could contribute to endothelial impairment. Endothelial dysfunction may be regarded as one of the therapeutic targets for pathologies associated with physical inactivity and microgravity.ANGERS-BU Médecine-Pharmacie (490072105) / SudocSudocFranceF

Comparing physical activity prescription with verbal advice for general practice patients with cardiovascular risk factors: results from the PEPPER randomised controlled trial

Abstract Background Regular physical activity improves health and quality of life for people with cardiovascular risk factors. However, few studies have demonstrated the applicability of strategies in health care to promote physical activity. Objective To evaluate if a written physical activity prescription combined with pedometer increases physical activity over one year compared with verbal advice in patients with cardiovascular disease risk in primary care. Methods The randomised-controlled, interventional, 12-month PEPPER study recruited patients aged 35 to 74 years, having quarterly followed-ups for hypertension, dyslipidaemia, or diabetes, and judged insufficiently active. Seventeen practices randomised patients into either the experimental group, who received a written, personalised prescription for daily step numbers, pedometer and logbook, or control group, who received verbal advice to do at least 15 min of rapid walking or equivalent daily. The primary outcome was the change in total weekly energy expenditure measured using an accelerometer at 3 months. The secondary outcomes were changes in step count, physical activity levels, quality of life, perceived obstacles to physical activity, and biomedical indicators at 3 and 12 months. Results One hundred and twenty-one participants were randomised. Although, weekly energy expenditure did not differ between the prescription and verbal instruction group, the estimated time spent doing moderate-intensity activity was significantly higher in the prescription group than the verbal group by an average of four minutes/week (p = 0.018)(95% CI [0.7 – 7.4]) reaching 48 min after 12 months (95% CI: 8 – 89). Similarly, this was associated with a clinically, higher average step number of 5256 steps/week increase over a year (95% CI: 660 – 9852). Among the most sedentary subgroup, walking less than 5000 steps/day at baseline, an 8868 steps/week (95% CI [2988 – 14700]) increase was observed in the prescription group. Conclusion Prescribing physical activity did not significantly modify total weekly energy expenditure, but slightly increased moderate-intensity activity duration and step counts, particularly among the most sedentary participants. Prescribing personalised physical activity goals encourages sedentary patients to engage in physical activity. Trial registration The PEPPER trial is registered in the US National Institutes of Health Clinical Trials Registry under number NCT02317003 (15/12/2014)

Early arterial calcification does not correlate with bone loss in pseudoxanthoma elasticum

International audienceThis article appeared in a journal published by Elsevier. The attached copy is furnished to the author for internal non-commercial research and education use, including for instruction at the author's institution and sharing with colleagues. Other uses, including reproduction and distribution, or selling or licensing copies, or posting to personal, institutional or third party websites are prohibited. In most cases authors are permitted to post their version of the article (e.g. in Word or Tex form) to their personal website or institutional repository. Authors requiring further information regarding Elsevier's archiving and manuscript policies are encouraged to visit: Background and aims: Pseudoxanthoma elasticum (PXE; OMIM 264800, prevalence 1/25,000 to 1/50,000) is an autosomal recessive multisystem disease due to deficiency in ABCC6, an ATP-binding cassette, sub-family C transporter. The PXE phenotype is mainly characterized by progressive ectopic calcification of connective tissues (namely skin, retinal Bruch's membrane and peripheral arteries) but the impact of PXE on bone structure is currently unknown. The present study sought to investigate bone mineralization and its potential link with vascular calcification in a large cohort of PXE patients with inherited mutations of the ABCC6 gene. Methods and results: 96 patients (61 women) matching the PXE criteria participated in this study. Their clinical history and status and bone biological markers were collected. Bone mineral density (BMD) was measured by dual energy X-ray absorptiometry and expressed as T-and Z-scores. Osteoporotic fractures were identified by X-ray, and coronary (CAC) and lower limb arterial calcification (LLAC) scores were determined by CT scan. Results: 44% of the women were menopausal. Osteopenia was disclosed in 46% (17 women) while 23% (9 women) exhibited osteoporosis, 3 with severe osteoporosis. Fractures of an osteoporotic nature were authenticated in 3 patients (1 woman). Markers of bone remodelling processes (CTX, BSAP and osteocalcin) were within the normal range for our laboratory standards. Severe vitamin D deficiency (b 25 nmol/L) was found in 15%, while 51% exhibited no vitamin D deficiency (vitamin D ≥ 50 nmol/L). LLAC and CAC scores were significantly higher in the patients with a low T-and/or Z-score, although this difference disappeared in multivariate analysis with age as a confounding factor. There was no significant difference in LLAC and CAC between PXE patients with and without osteoporotic fractures. There was no statistically significant association between BMD, LLAC and CAC and any of the bone remodelling factors. Conclusions: This is the first report on the bone mineralization process in PXE patients. Our data shows that PXE patients are not markedly prone to exaggerated bone demineralization and fracture risk, and prevalence of oste-oporosis remains within the normal range for the general population. Furthermore, the relationships between LLAC, but not CAC, and BMD with age are similar to those observed in the general population. Therefore, despite its pivotal role in ectopic calcification, ABCC6 deficiency does not interfere with the bone-vascular axis. The lack of PXE-related disturbances between BMD and arterial calcification also supports vitamin D supplementation in PXE patients with vitamin D deficiency. ClinicalTrials.gov Identifier: NCT01446393