Vitamin D, cardiovascular disease and mortality

A poor vitamin D status, i.e. low serum levels of 25-hydroxyvitamin D [25(OH)D], is common in the general population. This finding is of concern not only because of the classic vitamin D effects on musculoskeletal outcomes, but also because expression of the vitamin D receptor (VDR) and vitamin D metabolizing enzymes in the heart and blood vessels suggests a role of vitamin D in the cardiovascular system. VDR-knockout mice suffer from cardiovascular disease (CVD), and various experimental studies suggest cardiovascular protection by vitamin D, including antiatherosclerotic, anti-inflammatory and direct cardio-protective actions, beneficial effects on classic cardiovascular risk factors as well as suppression of parathyroid hormone (PTH) levels. In epidemiological studies, low levels of 25(OH)D are associated with increased risk of CVD and mortality. Data from randomized controlled trials (RCTs) are sparse and have partially, but not consistently, shown some beneficial effects of vitamin D supplementation on cardiovascular risk factors (e.g. arterial hypertension). We have insufficient data on vitamin D effects on cardiovascular events, but meta-analyses of RCTs indicate that vitamin D may modestly reduce all-cause mortality. Despite accumulating data suggesting that a sufficient vitamin D status may protect against CVD, we still must wait for results of large-scale RCTs before raising general recommendations for vitamin D in the prevention and treatment of CVD. In current clinical practice, the overall risks and costs of vitamin D supplementation should be weighed against the potential adverse consequences of untreated vitamin D deficiency.     

Vitamin D Deficiency and Its Implications on Cardiovascular Disease

Vitamin D is widely known for its important role in bone health. More recent evidence suggests that vitamin D may also play a protective role in many chronic conditions, including cancer, autoimmune, kidney, and cardiovascular diseases (CVDs). Observational studies have associated low vitamin D levels with CVD risk factors, including hypertension, hyperlipidemia, diabetes, and metabolic syndrome, as well as with cardiovascular events, including stroke, myocardial infarction, and congestive heart failure. Much less evidence is available from clinical trials of vitamin D supplementation. It remains to be determined whether the vitamin D–CVD relationship is causal, and what dosing of vitamin D supplementation would be adequate for prevention. Large-scale randomized controlled trials with adequate vitamin D dosing are needed before treatment strategies can be implemented. The purpose of this article is to review the scientific evidence linking vitamin D deficiency with CVD, including explanations of potential biologic mechanisms.     

Can vitamin D deficiency cause diabetes and cardiovascular diseases? Present evidence and future perspectives

Several studies have shown that vitamin D may play a role in many biochemical mechanisms in addition to bone and calcium metabolism. Recently, vitamin D has sparked widespread interest because of its involvement in the homeostasis of the cardiovascular system. Hypovitaminosis D has been associated with obesity, related to trapping in adipose tissue due to its lipophilic structure. In addition, vitamin D deficiency is associated with increased risk of cardiovascular disease (CVD) and this may be due to the relationship between low vitamin D levels and obesity, diabetes mellitus, dyslipidaemia, endothelial dysfunction and hypertension. However, although vitamin D has been identified as a potentially important marker of CVD, the mechanisms through which it might modulate cardiovascular risk are not fully understood. Given this background, in this work we summarise clinical retrospective and prospective observational studies linking vitamin D levels with cardio-metabolic risk factors and vascular outcome. Moreover, we review various randomised controlled trials (RCTs) investigating the effects of vitamin D supplementation on surrogate markers of cardiovascular risk. Considering the high prevalence of hypovitaminosis D among patients with high cardiovascular risk, vitamin D replacement therapy in this population may be warranted; however, further RCTs are urgently needed to establish when to begin vitamin D therapy, as well as to determine the dose and route and duration of administration.     

Serum and dietary vitamin D and cardiovascular disease risk in elderly men: a prospective cohort study

Background and aimRecent research suggests that low vitamin D may be associated with cardiovascular disease (CVD).Methods and resultsWe prospectively evaluated the association of dietary plus supplemental vitamin D intake and serum 25(OH) vitamin D with CVD incidence in the Osteoporotic Fractures in Men (MrOS) Study. Vitamin D intake was measured using a food frequency questionnaire and self-reported supplemental intake in 3094 men (mean age 76.4 years). From a subset of this population, we measured 25(OH) vitamin D in 813 men. Median 25(OH) vitamin D was 25.3 ng/mL. During a median follow-up of 4.4 years, there were 472 CVD cases, including 371 from coronary heart disease (CHD) and 101 from cerebrovascular attack (CVA). In the 25(OH) vitamin D sub-cohort, there were 140 cases of CVD including 115 from CHD and 25 from CVA. We used a Cox proportional hazards regression to calculate hazard ratios (HR) for CVD by vitamin D quartile. After adjusting for age, season, and standard CVD risk factors, the lowest quartile of 25(OH) vitamin D (HR, 1.18; 95% CI, 0.69–2.03) and vitamin D intake (HR, 0.76; 95% CI, 0.56–1.04) were not significantly associated with CVD incidence, compared to the highest vitamin D quartiles. When 25(OH) vitamin D was further analyzed by sufficiency (≥30 ng/mL), insufficiency (≥15–29.9 ng/mL), and deficiency (<15 ng/mL), vitamin D deficiency was not significantly associated with CVD incidence compared to sufficiency (HR 1.34; 95% CI 0.65–2.77).ConclusionVitamin D intake and serum 25(OH) vitamin D were not associated with CVD risk.     

Hypovitaminosis D and peripheral arterial disease: Emerging link beyond cardiovascular risk factors

Vitamin D has received increasing interest for its beneficial effect on health. Beyond its conventional role in bone metabolism, emerging evidence suggests a possible link between low vitamin D levels and cardiovascular disease (CVD), including peripheral arterial disease (PAD), and cardiovascular risk factors. Vitamin D interacts either directly with the vascular tree or indirectly through its association with cardiovascular risk factors, but the exact mechanism remains controversial. This review outlines the association between hypovitaminosis D and PAD. Both entities are quite prevalent in the general population and, therefore, their potential association might have important clinical implications. Whether vitamin D deficiency represents a novel risk factor for PAD/CVD, and whether vitamin D supplementation would reduce the burden of CVD still remains to be answered. Until then, vitamin D intake is not recommended for PAD/CVD prevention. Outdoor physical activity, coupled with adequate but safe sun exposure, is a healthy lifestyle practice suggested for the prevention of both PAD and hypovitaminosis D.     

Vitamin D and Cardiovascular Risk

Vitamin D deficiency is a common condition that has well-documented effects on musculoskeletal health. A growing body of literature has related vitamin D deficiency to other chronic disorders, including cardiovascular disease. Several plausible biological mechanisms have been postulated to explain this association, including the effect of poor vitamin D status on intermediate risk factors (eg, hypertension and diabetes), neurohormonal activation, inflammation, and cardiac remodeling. These mechanisms have been explored in experimental and animal studies, as well as several small interventional studies. The results of the controlled trials have not been conclusive to date. In this review, we summarize the existing studies investigating the effects of vitamin D on cardiovascular health, and propose that additional well-designed, prospective, randomized controlled trials are necessary to delineate the appropriate role of vitamin D supplementation in reducing the burden of cardiovascular disease.     

Association of vitamin D with cardiometabolic risk factors in rheumatoid arthritis

Objective

Individuals with rheumatoid arthritis (RA) are at a greater risk for cardiovascular disease (CVD). Vitamin D deficiency is a potential risk factor for CVD and metabolic syndrome. Since patients with RA have a high prevalence of vitamin D deficiency, we investigated the association of vitamin D levels with cardiometabolic risk factors in a cohort of RA patients with no prior history of CVD.

Methods

Serum 25‐hydroxyvitamin D (25[OH]D) levels were measured among RA patients enrolled in a cohort study of subclinical CVD. The cross‐sectional associations of 25(OH)D levels with traditional CVD risk factors such as insulin resistance (IR; estimated using the homeostatic model assessment [HOMA]), adipokines, markers of systemic inflammation, and endothelial activation were explored, adjusting for pertinent sociodemographic, lifestyle, and RA characteristics.

Results

Among 179 RA patients, 73 (41%) had a 25(OH)D level <30 ng/ml. Only 23 patients (13%) had a 25(OH)D level ≥45 ng/ml. After adjusting for demographics and body mass index (BMI), 25(OH)D remained significantly associated with high‐density lipoprotein (HDL) cholesterol and inversely associated with HOMA‐IR, fibrinogen, E‐selectin, and soluble intercellular adhesion molecule 1 (sICAM‐1). Significant associations with HDL cholesterol, E‐selectin, and sICAM‐1 were maintained after adjusting for the Disease Activity Score in 28 joints (DAS28) and autoantibody status. These associations were similar between the groups subdivided by sex, ethnicity, BMI, DAS28 level, and autoantibody status.

Conclusion

These data suggest that vitamin D deficiency is common in RA and may be independently associated with several cardiometabolic intermediates in this population.     

Circulating 25-hydroxy-vitamin D and the risk of cardiovascular diseases. Systematic review and meta-analysis of prospective cohort studies

AimsCirculating vitamin D is linked with the risk of cardiovascular disease (CVD). A meta-analysis has yet to explicitly explore correlation between vitamin D and the risk of CVD incidence and recurrent CVD. This meta-analysis examines the association between 25-hydroxy-vitamin D (25(OH)D) and the risk of CVD incidence (fatal, non-fatal, fatal and non-fatal combined events) and the risk of recurrent CVD (fatal, recurrent, and fatal and recurrent combined events). PROSPERO registration-CRD42021251483.Data synthesisA total of 79 studies (46 713 CVD cases in 1 397 831 participants) were included in the meta-analysis, of which 61 studies examined the risk of CVD incidence events, and 18 studies examined risk of recurrent CVD events. The risk of CVD incidence events (RR = 1.34, 95% CI: 1.26–1.43, p < 0.001) and recurrent CVD events (RR = 1.86, 95% CI: 1.46–2.36, p < 0.001) was higher in the lowest than the highest category of circulating 25(OH)D. Dose–response analysis reported a linear association for every 10 ng/ml increment of 25(OH)D and non-fatal CVD incidence events (RR = 0.94; 95% CI = 0.89–0.98, p = 0.005), lower fatal recurrent CVD events (RR = 0.45; 95% CI = 0.32–0.62, p < 0.001) and lower combined recurrent CVD events (RR = 0.80; 95% CI = 0.65–0.97, p = 0.023). A non-linear association was observed between higher 25(OH)D and lower fatal CVD incidence events (P-nonlinear<0.001), lower combined CVD incidence events (P-nonlinear = 0.001), and lower non-fatal recurrent CVD events (P-nonlinear = 0.044).ConclusionsThe lowest category of circulating 25(OH)D was associated with a higher risk of CVD incidence events and recurrent CVD events.     

Vitamin D and Cardiovascular Disease: Is There Evidence to Support the Bandwagon?

In the last 3?years, more evidence accumulated that vitamin D (vitD)deficiency associates with cardiovascular disease (CVD) and risk factors. The association with higher cardiovascular (CV) mortality was stronger than with nonfatal CVD events. A higher incidence of type 2 diabetes was also shown. Many factors related to lifestyle (physical activity in particular) influence both vitD levels and CVD, and may contribute to explain these observational data. Whether the association between vitD and CVD is causal can only be established through randomized controlled trials (RCTs), and to date the results of the randomized trials, which were not designed for investigating CV outcomes, do not support the association data. Answers on the effects of vitD supplementation on primary and secondary prevention of CV may be found in the specifically designed ongoing RCTs. In the mean time, low vitamin D levels should be regarded as a marker of unhealthy lifestyle, requiring a more aggressive attempt at modifying individual lifestyle.     

Future directions in vitamin D and cardiovascular research

AimsEvidence is accumulating that vitamin D status may influence the risk of cardiovascular disease (CVD). Final confirmation for a causal relationship between vitamin D and CVD is however still lacking. The present viewpoint article outlines several future research directions to close this gap.Data synthesisFuture directions include the need of performing large randomised controlled supplementation trials with vitamin D in specific risk groups. In addition, large register sets of data on vitamin D supplementation can be used, provided that adequate statistical methods such as propensity score modelled analysis are applied. To better understand vitamin D-mediated effects on CVD risk, the routine measurement of circulating levels of the hormonal vitamin D form, 1,25-dihydroxyvitamin D, is also necessary, in addition to the determination of its precursor 25-hydroxyvitamin D. Further, genetic association studies may help in clarifying the contribution of vitamin D to the development of CVD. Finally, the interrelationship of vitamin D with physical activity should be considered when studying CVD risk.ConclusionsOverall, it can be expected that the next 10–15 years will provide an increased clarity concerning the role of vitamin D in CVD.     

Vitamin D and acute myocardial infarction

Vitamin D deficiency is a prevalent condition,cutting across all ethnicities and among all age groups,and occurring in about 30%-50% of the population. Besides vitamin D established role in calcium homeostasis,its deficiency is emerging as a new risk factor for coronary artery disease. Notably,clinical investigations have suggested that there is an association between hypovitaminosis D and acute myocardial infarction(AMI). Not only has it been linked to incident AMI,but also to increased morbidity and mortality in this clinical setting. Moreover,vitamin D deficiency seems to predispose to recurrent adverse cardiovascular events,as it is associated with post-infarction complications and cardiac remodeling in patients with AMI. Several mechanisms underlying the association between vitamin D and AMI risk can be involved. Despite these observational and mechanistic data,interventional trials with supplementation of vitamin D are controversial. In this review,we will discuss the evidence on the association between vitamin D deficiency and AMI,in terms of prevalence and prognostic impact,and the possible mechanisms mediating it. Further research in this direction is warranted and it is likely to open up new avenues for reducing the risk of AMI.     

Vitamin D deficiency in the aetiology of obesity‐related insulin resistance

The obese insulin‐resistant state is often associated with low circulating concentration of vitamin D 25‐hydroxyvitamin D₃ [25(OH)D₃]. Fat sequestration of vitamin D in the expanded obese adipose tissue mass has been pointed out as a plausible explanation for this circulating vitamin D deficiency. However, the putative mechanisms behind this hypovitaminosis D remain to be elucidated. The presence of vitamin D receptor and vitamin D–metabolizing enzymes in insulin‐sensitive organs suggests that vitamin D may be involved in glucose and lipid metabolism and may be related to insulin sensitivity. Indeed, mainly in vitro studies support a role of vitamin D in regulating glucose and lipid metabolism in several insulin‐sensitive tissues including adipose tissue, skeletal muscle, liver, as well as pancreatic insulin secretion. A potential role of vitamin D in gut barrier function and metabolism has also been suggested. This review summarizes recent knowledge on vitamin D deficiency in the aetiology of obesity‐related insulin resistance and discusses potential underlying mechanisms. Finally, the role of vitamin D supplementation on insulin sensitivity and glycaemic control is discussed.     

Renal osteodystrophy in children: A systemic disease associated with cardiovascular manifestations

The increasing prevalence of chronic kidney disease (CKD) in the United States demands a closer evaluation of and attention to associated morbidities, and, particularly, the rising mortality related to cardiovascular disease in all age groups. Patients with CKD demonstrate an increased risk of coronary artery disease due to calcium deposition and subsequent arterial stiffening, in addition to left ventricular dysfunction with associated heart failure and arrhythmias. While clearly impacted by the traditional risk factors for development of cardiovascular disease (CVD), patients with CKD are also affected by non-traditional risk factors, including calcium overloading related to aggressive management of secondary hyperparathyroidism.Recent data have shown that a substantial number of patients with CKD are deficient in vitamin D on a nutritional basis, in addition to the known decrease in the kidney-produced active metabolite during progressive CKD. Historically, vitamin D has been described as an endocrine hormone that regulates blood calcium and parathyroid hormone levels. It has become increasingly clear, through the recognition of a vitamin D receptor in most tissues, that vitamin D possesses functions well beyond calcium homeostasis, such that a deficiency may contribute to the development of CVD. In this brief review, the role of vitamin D activation through its vitamin D receptor will serve as an introduction to the magnitude of the nutritional deficits in children, adults, and those with CKD. As therapeutic entities in the management of renal osteodystrophy, vitamin D analogues play an important role in cardiovascular health that continues to evolve. Preliminary studies indicate that vitamin D therapy for control of secondary hyperparathyroidism may confer cardioprotection and reduce mortality. Attention to care of osteodystrophy in CKD must take into account heart health as well.     

The effect of physical activity on dose-relationship between serum 25-hydroxyvitamin D and cardiovascular health events in older adults

Background and aimsReverse J- or U-shaped associations between serum 25-hydroxyvitamin D (25[OH]D) concentrations and cardiovascular outcomes have been reported, which need clarifications in older adults. Physical activity, correlating with both serum 25[OH]D concentration and cardiovascular health, may have an effect on the dose-relationships.Methods and ResultsAt baseline, 2790 participants aged 65 years and over, free of vitamin D supplementation use, had assays for serum 25[OH]D concentrations and health related characteristics and measurements, were followed up for cardiovascular events and death by up to 7 and 15 years, respectively. The dose–response associations of serum 25[OH]D concentrations with cardiovascular events and mortality risk were examined using Cox regression models.After adjusting for physical activity and other covariates, serum 25[OH]D concentration was non-linearly associated with cardiovascular mortality risk (U-shaped, P = 0.009). According to the Institute of Medicine categories, the HR(95% CI) of cardiovascular mortality risk separately in deficient (<25 nmol/L), inadequate (25 to < 50 nmol/L) and potentially harmful (≥125 nmol/L) level was 1.67 (0.23, 12.01), 1.66 (1.25, 2.20) and 2.21 (0.30, 16.37), respectively. The risk of 25[OH]D inadequacy for cardiovascular mortality was significantly attenuated by increased physical activity, especially leisure activity (P for trend = 0.008 and 0.021, respectively). No significant finding was observed for incident cardiovascular events.ConclusionBoth lower and higher serum 25[OH]D concentrations were associated with risk of cardiovascular mortality in Chinese community-dwelling older adults. Physical activity may attenuate the cardiovascular mortality risk of vitamin D inadequacy, but its role in individuals with higher 25[OH]D concentrations remains unclear.     

Vitamin D deficiency plays an important role in cardiac disease and affects patient outcome: Still a myth or a fact that needs exploration?

There is increasing evidence that a low vitamin D status may be an important and hitherto neglected factor of cardiovascular disease. This review is an overview of the current body of literature, and presents evidence of the mechanisms through which vitamin D deficiency affects the cardiovascular system in general and the heart in particular. Available data indicate that the majority of congestive heart failure patients have 25-hydroxyvitamin D deficiency. Furthermore, the low serum 25-hydroxyvitamin D level has a higher impact on hypertension, coronary artery disease an on the occurrence of relevant cardiac events. A serum 25-hydroxyvitamin D level below 75 nmol/l (30 ng/l) is generally regarded as vitamin D insufficiency in both adults and children, while a level below 50 nmol/l (20 ng/l) is considered deficiency. Levels below 50 nmol/l (20 ng/l) are linked independently to cardiovascular morbidity and mortality.     

25‐hydroxyvitamin D and cardiovascular risk factors in women with systemic lupus erythematosus

Objective

Low serum levels of 25‐hydroxyvitamin D (25[OH]D; vitamin D) are associated with a higher frequency of cardiovascular disease and risk factors in the general population. Vitamin D deficiency has also been noted in systemic lupus erythematosus (SLE). The objective of this study was to evaluate the associations of serum 25(OH)D levels with cardiovascular risk factors in women with SLE.

Methods

Data collected in 181 women with SLE included demographics, SLE activity and damage assessments, cardiovascular risk factors, medications, and laboratory assessments of inflammatory markers and 25(OH)D levels. Multiple linear and logistic regressions were used to estimate the association of 25(OH)D levels with cardiovascular risk factors.

Results

The mean age and disease duration were 43.2 and 11.9 years, respectively. The mean 25(OH)D level was 27.1 ng/ml and 62.2% had 25(OH)D levels <30 ng/ml. In unadjusted analyses, lower 25(OH)D levels were significantly associated with higher diastolic blood pressure, low‐density lipoprotein cholesterol, lipoprotein(a), body mass index (BMI), and fibrinogen levels, as well as self‐reported hypertension and diabetes mellitus. Lower 25(OH)D levels were also significantly associated with higher SLE disease activity and damage scores. After adjustment for age, seasonal variation, and race/ethnicity, lower 25(OH)D levels were also significantly related to higher fasting serum glucose. With further adjustment for BMI, associations between 25(OH)D and cardiovascular risk factors were no longer significant.

Conclusion

This study demonstrates that vitamin D levels are low in women with SLE and significant associations exist with selected cardiovascular risk factors, although most of these associations can be explained by BMI.     

The effect of vitamin D supplementation on cardiovascular risk in patients with prediabetes: A secondary analysis of the D2d study

AimsLow blood 25(OH)D level is associated with increased cardiovascular disease (CVD) risk. Additionally, individuals with prediabetes are at higher risk for CVD than individuals with normoglycemia. We investigated the effects of vitamin D supplementation on CVD outcomes in the vitamin D and type 2 diabetes (D2d) study, a large trial among adults with prediabetes.Methods2423 participants were randomized to 4000 IU/day of vitamin D₃ or placebo and followed for median 3.0 years for new-onset diabetes. In pre-specified secondary analyses, we examined the effect of vitamin D supplementation on composite Major Adverse Cardiovascular Events (MACE); expanded MACE (MACE + revascularization); atherosclerotic CVD (ASCVD) risk score; and individual CVD risk factors (blood pressure, lipids, high-sensitivity C-reactive protein). Cox models compared hazard ratios (HR) between the two groups on MACE and expanded MACE.ResultsMean age was 60 years, 45 % were women, 13 % had history of CVD. Twenty-one participants assigned to vitamin D and 12 participants assigned to placebo met the MACE outcome (HR 1.81, 95%CI 0.89 to 3.69). There were 27 expanded MACE outcomes in each group (HR 1.02, 95%CI, 0.59 to 1.76). There were no significant differences between vitamin D and placebo in individual CVD risk factors, but change in ASCVD risk score favored the vitamin D group (?0.45 %, 95%CI -0.75 to ?0.15).ConclusionsIn people with prediabetes not selected for vitamin D insufficiency and with intermediate CVD risk, vitamin D supplementation did not decrease MACE but had a small favorable effect on ASCVD risk score.Trial registration: D2d ClinicalTrials.gov number, NCT01942694, prospectively registered September 16, 2013.     

Vitamin D: epidemiology of cardiovascular risks and events

Vitamin D may influence blood pressure through the renin-angiotensin system, parathyroid hormone levels, myocardial function, inflammation, and vascular calcification. In the past several years, a number of high-quality prospective studies have examined 25(OH)vitamin D (25(OH)D) levels in relation to risk of cardiovascular disease (CVD). Studies consistently show that levels of 25(OH)D below 20-25 ng/mL are associated with an increased risk of CVD incidence or mortality. Risk appears especially elevated at 25(OH)D levels below 10 or 15 ng/mL. It is unclear if levels higher that 25 ng/mL provide further benefits for CVD disease. Currently, results from randomized clinical trials are sparse and do not allow a definitive conclusion. Given other potential benefits of vitamin D, and low potential for toxicity, deficient levels below 25-30 ng/mL should be avoided and treated when identified. Further observational and randomized clinical trial data are important to better characterize the optimal range for 25(OH)D.     

Effect of one-anastomosis gastric bypass on cardiovascular risk factors in patients with vitamin D deficiency and morbid obesity: A secondary analysis

Background and aimsBariatric patients often suffer from vitamin D (VD) deficiency, and both, morbid obesity and VD deficiency, are related to an adverse effect on cardiovascular disease (CVD) risk. Therefore, we assessed the change of known CVD risk factors and its associations during the first 12 months following one-anastomosis gastric bypass (OAGB).Methods and resultsIn this secondary analysis, CVD risk factors, medical history and anthropometric data were assessed in fifty VD deficient (25-hydroxy-vitamin D (25(OH)D) <75 nmol/l) patients, recruited for a randomized controlled trial of VD supplementation. Based on previous results regarding bone-mass loss and the association between VD and CVD risk, the study population was divided into patients with 25(OH)D ≥50 nmol/l (adequate VD group; AVD) and into those <50 nmol/l (inadequate VD group; IVD) at 6 and 12 months (T6/12) postoperatively. In the whole cohort, substantial remission rates for hypertension (38%), diabetes (30%), and dyslipidaemia (41%) and a significant reduction in CVD risk factors were observed at T12. Changes of insulin resistance markers were associated with changes of total body fat mass (TBF%), 25(OH)D, and ferritin. Moreover, significant differences in insulin resistance markers between AVD and IVD became evident at T12.ConclusionThese findings show that OAGB leads to a significant reduction in CVD risk factors and amelioration of insulin resistance markers, which might be connected to reduced TBF%, change in 25(OH)D and ferritin levels, as an indicator for subclinical inflammation, and an adequate VD status.Registered at clinicaltrials.gov(Identifier: NCT02092376) and EudraCT (Identifier: 2013-003546-16).