Dark Chocolate and Platelet Function in Humans

Cardiovascular disease (CVD) is a primary cause of premature deaths worldwide, with incidence rates in the United Kingdom, particularly in Scotland, being amongst the highest worldwide. Thus identification of dietary components that most effectively prevent CVD is potentially of wide public health benefit.

Consumption of diets rich in plant-based products protects against the development of CVD. Such effects have been ascribed in part to polyphenols, which are non-nutritive but, potentially bioactive secondary metabolites ubiquitous found in fruits, vegetables, herbs, spices, teas and wines. The beneficial effects of polyphenols on CVD is believed to be mediated, at least in part, though improving platelet function. At least 10 human intervention studies found a consistent and robust beneficial effect of cocoa products on platelet function, but unfortunately all of these studies used only one or two methods to assess platelet function, therefore only getting limited insights into the complex physiological behavior of platelets. In addition, none of these studies assessed potential mechanisms by which flavan-3-ols may inhibit platelet function. Schramm et al. have shown that consumption of chocolate rich in flavan-3-ols and their oligomers (procyanidins) lead to increased production of prostacyclin, a strong platelet inhibitor. This finding has also been observed when aortic endothelial cells are treated with procyanidins in vitro. Thus the stimulation of prostacyclin production in endothelial cells may reflect one pathway by which flavan-3-ols indirectly inhibit platelet activation. Many other potential mechanisms are discussed in the literature but so far the evidence for such mechanisms is limited or non-existing.

In this study we assess effects of consumption of chocolate enriched in flavan-3-ols on platelet function by measuring not only platelet aggregation, but also in vitro coagulation and platelet activation in healthy humans. In addition, we examine the effects of consumption of flavan-3-ols on the regulation of the platelet proteome to elucidate pathways by which these bioactive cocoa compounds affect platelet function.

HYPOTHESIS

Acute consumption of a moderate amount of dark chocolate enriched in flavan-3-ols results in decreased platelet activation and aggregation by decreasing the levels of thromboxane A2 produced by endothelial cells.

OBJECTIVES

The main objective of the proposed study is to determine whether consumption of 60 g dark chocolate enriched in flavan-3-ols results in decreased platelet activation and aggregation by decreasing levels of thromboxane A2, as well as assessing what other mechanisms could be involved.

The specific objectives of the proposed study are to determine:

1. whether acute intake of 60 g dark chocolate enriched in flavan-3-ols, as compared with standard dark chocolate low in flavan-3-ols and white chocolate containing no flavan-3-ols, affects platelet aggregation, thromboxane A2 formation upon aggregation, in vitro bleeding time, P-selectin expression, and activation of the fibrinogen receptor;

2. whether and how acute intake of 60 g dark chocolate enriched in flavan-3-ols, as compared with standard dark chocolate and white chocolate, affects the platelet proteome, and thereby potential new biomarkers of platelet function, as well as protein levels of anti-oxidant enzymes;

3. identities and concentrations of flavan-3-ols and their metabolites in plasma and/ or urine 2 and 6 h after acute intake of 60 g dark chocolate enriched in flavan-3-ols, as compared with standard dark chocolate and white chocolate.