Heart doi:10.1136/heartjnl-2011-300304
  • Original article

NOX2-mediated artery dysfunction in smokers: acute effect of dark chocolate

  1. Francesco Violi
  1. I Clinica Medica, Sapienza University, Rome, Italy
  1. Correspondence to Professor Francesco Violi, I Clinica Medica, Viale del Policlinico 155, Roma 00161, Italy; francesco.violi{at}
  1. Contributors Conception and design: FV, LL. Analysis and interpretation of data: FV, LL, RC, PP. Flow-mediated dilation analysis: LL. Patient enrolment: LP, EC, TA, RC, FA. Laboratory analysis: RC, CN, CP. Drafting the article: FV, LL.

  • Accepted 30 June 2011
  • Published Online First 31 July 2011


Background Cocoa seems to exert artery dilatation via oxidative stress inhibition but the mechanism is still unclear.

Objectives To investigate whether in smokers, dark chocolate elicits artery dilatation via down-regulation of NOX2, the catalytic core of NADPH oxidase.

Methods Flow-mediated dilatation (FMD), oxidative stress (as assessed by urinary isoprostanes excretion), nitric oxide generation (as assessed by serum levels of nitrite/nitrate (NOx)), NOX2 activity (as assessed by blood levels of soluble NOX2 derived peptide (sNOX2-dp)) and serum epicatechin were studied in 20 smokers and 20 healthy subjects (HS) in a crossover, single-blind study. Patients were randomly allocated to 40 g dark chocolate (>85% cocoa) or 40 g of milk chocolate (≤35% cocoa). FMD, urinary isoprostanes, NOx and sNOX2-dp were assessed at baseline and 2 h after chocolate ingestion.

Results Smokers had lower FMD and NOx and higher sNOX2-dp compared to HS. After dark chocolate intake, urinary isoprostanes and sNOX2-dp significantly decreased and FMD and NOx significantly increased in smokers but not in HS. No changes of the above variables were observed after milk chocolate intake. Multiple linear regression analysis showed that in smokers the only independent predictive variable associated with a change in FMD was a change in sNOX2-dp. Serum epicatechin increased in either group only after dark chocolate intake, reaching values higher than 0.1 μM. Platelets from smokers (n=5), but not from HS (n=5), showed lower p47phox translocation to platelet membrane and higher NOx when incubated with 0.1–10 μM epicatechin.

Conclusion Results suggest that in smokers, cocoa enhances artery dilatation by lowering of NOX2 activation.


  • Funding This study was supported by a grant from the University of Rome “La Sapienza” (Ateneo Federato 2009) (funds to LL and FV).

  • Competing interests None.

  • Patient consent Obtained.

  • Ethics approval This study was conducted with the approval of the Sapienza University of Rome Ethics Committee.

  • Provenance and Peer review Not commissioned; externally peer reviewed.

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