1. Introduction
Stress is highly prevalent in today’s society and has been closely associated with both psychological and physical health. Single episodes of stress have been shown to increase the risk of acute cardiovascular events [
1,
2,
3]. Furthermore, it is well established that stress perturbs cardiovascular activity [
4,
5], and such responses can contribute to stress-induced cardiovascular events. Indeed, laboratory-based stress tasks have been shown to evoke temporary myocardial ischaemia [
6], with peripheral vascular responses during mental stress associated with mental stress-induced ischaemia [
7].
Mental stress induces immediate increases in heart rate and blood pressure (BP) in healthy adults and this is accompanied by peripheral arterial vasodilation, as measured by forearm blood flow (FBF) [
8,
9,
10]. In addition to acute responses to stress, studies show that mental stress can lead to post-stress impairments in vascular function [
11]. Specifically, transient declines in endothelial function (measured by brachial flow-mediated dilatation; FMD) occur from 15 to 90 min after stress in healthy young adults [
12,
13,
14,
15,
16], in older adults, postmenopausal women with diabetes and depression, and populations with high cholesterol and metabolic syndrome [
17,
18,
19]. Importantly, reductions in FMD observed following mental stress (1–3% FMD) can be considered significant in the context of future cardiovascular risk, with 1% FMD reduction leading to a 13% increase in cardiovascular risk [
20]. Currently, the mechanisms underpinning stress-induced impairments in endothelial function have not been established but studies suggest that reductions in bioavailability of Nitric Oxide (NO), inflammatory markers, and cortisol, as well as increases in vasoconstrictors, such as endothelin-1 (ET-1), may play an important role [
21,
22,
23].
Despite the high prevalence of stress in today’s society, there is limited research investigating the efficacy of strategies to protect the vascular system from the deleterious impact of psychological stress. A 16-week intervention, combining a hypocaloric diet and exercise training in obese children, was shown to reduce BP and increase peripheral vasodilation (as measured by forearm vascular conductance, FVC) during mental stress [
24]. This suggests that lifestyle factors, such as diet and exercise, can protect the vasculature from acute mental stress.
Diets rich in fruits and vegetables have been shown to reduce the risk of cardiovascular disease (CVD), myocardial infarction, cardiovascular mortality, and all-cause mortality [
25,
26]. Flavonoids, a group of small molecules present in most fruits and vegetables, have gained increasing attention in the last 25 years, with epidemiological evidence suggesting significant associations between high intake of flavonoids and a lower CVD-related mortality and morbidity [
27,
28,
29,
30]. In particular, flavanols, a sub-group of flavonoids that can be found in cocoa, berries, grapes, apples, and tea, improve human endothelial function (brachial FMD) within 1–3 h of intake [
31,
32,
33,
34]. Acute effects of flavanols on endothelial function have been shown to translate into chronic benefits in FMD [
35,
36,
37,
38], as well as arteriolar and microvascular vasodilator capacity [
37] and BP [
36,
38,
39,
40,
41]. Circulating flavanol metabolites, particularly (−)-epicatechin-derived [
42,
43], are believed to drive the beneficial effects of cocoa flavanols on the endothelium by increasing NO bioavailability [
34,
44,
45,
46], and reducing ET-1 [
47]. Therefore, foods rich in flavanols have the potential to be effective as a dietary strategy to counteract the deleterious impact of mental stress on vascular function. In the current study, we used a randomised, placebo-controlled, double-blind, cross-over intervention design to assess whether the acute intake of cocoa flavanols can improve endothelial function (as measured by brachial FMD) following an episode of mental stress in healthy young adults. We further evaluated the effects of dietary flavanols on peripheral vasodilation (measured by FBF) and BP during mental stress, making this the first study to assess vascular function both during and following mental stress.
4. Discussion
The present study shows that cocoa flavanols prevented the decline in brachial FMD 30 min post-stress, and FMD remained significantly higher following high-flavanol cocoa compared to low-flavanol cocoa 90 min post-stress. Furthermore, high-flavanol cocoa increased FBF at rest and during stress in comparison to low-flavanol cocoa. Perceptions of the stress task and performance on the stress task were not significantly different between conditions, suggesting a consistent stress experience across interventions. Mental-stress induced changes in HR, HRV, PEP, and BP were not affected by the intake of flavanols. To our knowledge, this is the first study to show that plant-derived flavonoids are effective at counteracting mental-stress induced endothelial dysfunction and improving vascular function during mental stress.
Cocoa flavanols were shown to be effective at preventing the decline in brachial FMD experienced 30 min following mental stress. This is in line with previous work, showing benefits of flavanols on endothelial function 2 h post-intake [
31,
33,
34,
62,
63,
64]. Furthermore, the observed decline in FMD (approx. 1.4% FMD) is in agreement with the literature, reporting a 1–3% impairment in endothelial function post-stress in healthy adults [
12,
13,
14,
15,
16]. Similarly to the current work, one recent study investigated the efficacy of acute intake of Vitamin C (a strong antioxidant) in the context of mental stress, yet reported no effects on endothelial function [
21]. However, in contrast to the current findings, their stress task did not induce significant impairments in brachial FMD (at 30 and 90 min post-stress), making the interpretation of this data difficult. Therefore, to our knowledge, this is the first study to show that one dose of dietary flavanols can effectively eliminate post-stress impairments in endothelial function.
As expected, the physiological responses to stress were indicative of both sympathetic activation and parasympathetic withdrawal [
8,
9]. Whilst flavanols did not influence HR, HRV, PEP, and BP during stress, they led to increased peripheral vasodilation. This confirms previous research showing flavanols’ benefits in FBF at rest [
37], and adds to the literature by demonstrating that these benefits persist when the vasculature is challenged by stress. Despite a significantly higher vasodilatation during mental stress following high-flavanol intake, no attenuation in the rise of BP during stress was observed. This disconnection between FBF and BP responses during stress has been reported before. For example, inflammation-induced changes in FBF during acute mental stress were also not accompanied by modifications in BP [
9]. In addition, acute intake of cocoa flavanols does not typically lead to changes in BP in young healthy adults [
33,
65,
66,
67,
68], but only in older individuals [
37] or after chronic flavanol intake in overweight [
62], hypertensive [
36,
39,
69], and other at-risk populations [
54]. In line with our data, previous chronic interventions combining a chronic hypocaloric diet and exercise training (16 weeks) in obese children, have similarly been shown to be effective at increasing FBF and FVC at rest and during stress, but no changes in BP and other cardiac parameters were observed [
24]. Whilst flavanols did not impact BP in this study, the positive effect on vasodilation during stress-induced increases in BP, suggests a protective effect on the vasculature and perhaps a more efficient way of responding to stress. More importantly, flavanol-induced vasodilation during stress may have clinical relevance as associations between peripheral vasodilatory responses and myocardial ischaemia have been previously reported [
7].
In the current study, flavanol intake did not affect measures of sympathetic and parasympathetic activity during stress. While this is an area of limited research, previous human studies show that grape polyphenols supplementation do not induce changes in muscle sympathetic nervous activity, but attenuate increases in HR during mental stress in hypertensive adults [
70]. This may suggest that flavanols’ effects on HR may vary depending on the target population. On the other hand, in vivo rodent studies report that acute intake of pure flavanols ((−)-epicatechin, procyanidins) can increase plasma catecholamines, adrenaline and noradrenaline with varying efficacy [
71,
72], but to what extent such results translate to humans is currently unclear.
The mechanisms by which flavanols affect the vasculature are not yet well established, but there is evidence from human [
34,
47,
73] and cellular models [
44,
45,
46,
74,
75] to suggest that the flavanol (−)-epicatechin and its metabolites can increase bioavailability of NO by enhancing eNOS activation through calcium-mediated activation of signalling pathways, such as PI3K, Akt and PKA. As such, it is likely that similar mechanisms underlie the beneficial effects of flavanols on vasodilation and endothelial function during and following stress. Indeed, there is strong evidence to suggest that NO contributes to the rise in FBF during mental stress [
8], as pharmacologically inhibiting NO synthase (with NG-monomethyl-L-arginine and atropine) during stress attenuates increases in blood flow [
76].
Human studies also show that flavanols can down-regulate bioavailability of the vasoconstrictor ET-1 [
47,
77], and this may also be relevant in the context of vascular responses to stress. There is in vitro evidence from in vitro studies that cortisol and corticotropin-releasing hormone (CRH)-induced increases in ET-1, known to reduce NO bioavailability, may contribute to the impairment in FMD following mental stress [
11]. Furthermore, increases in cortisol have been associated with post-stress impairments in FMD [
21] and, similarly, inflammatory markers remain elevated following mental stress [
78]. Conversely, previous evidence suggests that cocoa flavanols can attenuate IL-6 production, as well as reduce reactive oxygen species (ROS), promoting resilience to social stress in mouse models [
79]. Whilst we did not assess circulating levels of inflammatory and vascular biomarkers in the present study, we hypothesise that flavanols may reduce the stress-induced inflammatory and/or cortisol response, resulting in a reduction in ET-1 and increase in NO bioavailability during stress. The molecular mechanisms underpinning the effects of flavanols during stress should be addressed in future studies.
Interestingly, brachial systolic BP, but not diastolic, remained significantly elevated at 30 and 90 min following stress, while flavanols had no effect. Previous studies have reported elevated systolic and diastolic BP up to 20 min following mental stress [
12,
13,
80], with SBP returning to baseline at a slower rate [
80]. We have also observed a significant increase in resting arterial diameter at 30 and 90 min post-stress, suggesting that arteries are more dilated following stress, so increases in BP are not driven by vasoconstriction. To the best of our knowledge, this is the first time that SBP is shown to remain elevated for prolonged periods of time after stress has ceased, and this might have further implications for the vascular system. Whilst this may not be problematic in a healthy population, for patients with hypertension or at risk of CVD, prolonged stress-induced increases in SBP are of great significance for vascular health.
Limitations
One of the main limitations of the present study is the exclusion of females, which makes the applicability of the present results limited to healthy men. Future studies should focus on the impact of flavanols on stress-responses in women, particularly given the evidence suggesting that there are gender differences in vascular responses to stress [
81]. Secondly, previous evidence has shown that polyphenol microbial derived metabolites can be detected in the blood for up to 48 h [
48], whilst in the present study we only restricted polyphenol intake for 24 h prior to the study visits. More recent evidence also shows a large reduction in urinary polyphenol metabolites from 24 to 48 h [
42], indicating that most metabolites are excreted within the first 24 h. Whilst previous studies suggest that flavanols can modulate NO, ET-1 and IL-6, in the present study we have not assessed these biomarkers and cannot conclude that flavanol-induced vascular responses to mental stress are specifically linked to these mechanisms. A more in-depth investigation of mechanisms of action underlying these responses should be the focus of future work, particularly in regard to modulation of cortisol levels post-stress. Although the efficacy of flavanol intake in improving vascular responses to stress demonstrated in the present study is of value, extending this work to populations at higher risk of cardiovascular diseases (e.g., hypertensives) or mental stress (e.g., care workers) should be addressed in the future. For example, flavanols might be more effective at modulating blood pressure responses during mental stress in these populations.
Finally, we have a lower number of volunteers (n = 10/13) with a complete set of cardiovascular data, due to a malfunction in the Ambulatory Monitoring System. Despite a lower n, we still detected significant changes in these variables during stress (effect sizes of time effect = 0.598–0.751). In contrast, the effect sizes for the intervention and intervention × time interaction effects (effect sizes = 0.0001–0.107) are considerably lower. It is therefore unlikely that the non-significant results between flavanol interventions are due to low sample size.