title: Dark chocolate acutely improves walking autonomy in patients with peripheral artery disease.
abstract: BACKGROUND
  NOX-2, the catalytic subunit of NADPH oxidase, has a key role in the formation of reactive oxidant species and is implicated in impairing flow-mediated dilation (FMD). Dark chocolate exerts artery dilatation via down-regulating NOX2-mediated oxidative stress. The aim of this study was to investigate whether dark chocolate improves walking autonomy in peripheral artery disease (PAD) patients via an oxidative stress-mediated mechanism.
  
  
  RESULTS
  FMD, serum levels of isoprostanes, nitrite/nitrate (NOx) and sNOX2-dp, a marker of blood NOX2 activity, maximal walking distance (MWD) and maximal walking time (MWT) were studied in 20 PAD patients (14 males and 6 females, mean age: 69±9 years) randomly allocated to 40 g of dark chocolate (>85% cocoa) or 40 g of milk chocolate (≤35% cocoa) in a single blind, cross-over design. The above variables were assessed at baseline and 2 hours after chocolate ingestion. Dark chocolate intake significantly increased MWD (+11%; P<0.001), MWT (+15%; P<0.001), serum NOx (+57%; P<0.001) and decreased serum isoprostanes (-23%; P=0.01) and sNOX2-dp (-37%; P<0.001); no changes of the above variables were observed after milk chocolate intake. Serum epicatechin and its methylated metabolite significantly increased only after dark chocolate ingestion. Multiple linear regression analysis showed that Δ of MWD was independently associated with Δ of MWT (P<0.001) and Δ of NOx (P=0.018). In vitro study demonstrated that HUVEC incubated with a mixture of polyphenols significantly increased nitric oxide (P<0.001) and decreased E-selectin (P<0.001) and VCAM1 (P<0.001).
  
  
  CONCLUSIONS
  In PAD patients dark but not milk chocolate acutely improves walking autonomy with a mechanism possibly related to an oxidative stress-mediated mechanism involving NOX2 regulation.
  
  
  BACKGROUND
  http://www.clinicaltrials.gov. Unique identifier: NCT01947712.
authors.affiliation: Department of Internal Medicine and Medical Specialties, Sapienza University of Rome, Rome, Italy (L.L., L.P., E.C., P.P., M.B., C.N., S.B., R.C., F.V.).
authors.forename: Lorenzo
authors.initials: L
authors.lastname: Loffredo
authors.affiliation: Department of Internal Medicine and Medical Specialties, Sapienza University of Rome, Rome, Italy (L.L., L.P., E.C., P.P., M.B., C.N., S.B., R.C., F.V.).
authors.forename: Ludovica
authors.initials: L
authors.lastname: Perri
authors.affiliation: Department of Internal Medicine and Medical Specialties, Sapienza University of Rome, Rome, Italy (L.L., L.P., E.C., P.P., M.B., C.N., S.B., R.C., F.V.).
authors.forename: Elisa
authors.initials: E
authors.lastname: Catasca
authors.affiliation: Department of Internal Medicine and Medical Specialties, Sapienza University of Rome, Rome, Italy (L.L., L.P., E.C., P.P., M.B., C.N., S.B., R.C., F.V.).
authors.forename: Pasquale
authors.initials: P
authors.lastname: Pignatelli
authors.affiliation: Department of Internal Medicine and Medical Specialties, Sapienza University of Rome, Rome, Italy (L.L., L.P., E.C., P.P., M.B., C.N., S.B., R.C., F.V.).
authors.forename: Monica
authors.initials: M
authors.lastname: Brancorsini
authors.affiliation: Department of Internal Medicine and Medical Specialties, Sapienza University of Rome, Rome, Italy (L.L., L.P., E.C., P.P., M.B., C.N., S.B., R.C., F.V.).
authors.forename: Cristina
authors.initials: C
authors.lastname: Nocella
authors.affiliation: Department of Medical-Surgical Sciences and Biotechnologies, Sapienza University of Rome, Rome, Italy (E.D.F., G.F.).
authors.forename: Elena
authors.initials: E
authors.lastname: De Falco
authors.affiliation: Department of Internal Medicine and Medical Specialties, Sapienza University of Rome, Rome, Italy (L.L., L.P., E.C., P.P., M.B., C.N., S.B., R.C., F.V.).
authors.forename: Simona
authors.initials: S
authors.lastname: Bartimoccia
authors.affiliation: Department of Medical-Surgical Sciences and Biotechnologies, Sapienza University of Rome, Rome, Italy (E.D.F., G.F.) Department of AngioCardioNeurology, IRCCS NeuroMed, Pozzilli, Italy (G.F.).
authors.forename: Giacomo
authors.initials: G
authors.lastname: Frati
authors.affiliation: Department of Internal Medicine and Medical Specialties, Sapienza University of Rome, Rome, Italy (L.L., L.P., E.C., P.P., M.B., C.N., S.B., R.C., F.V.).
authors.forename: Roberto
authors.initials: R
authors.lastname: Carnevale
authors.affiliation: Department of Internal Medicine and Medical Specialties, Sapienza University of Rome, Rome, Italy (L.L., L.P., E.C., P.P., M.B., C.N., S.B., R.C., F.V.).
authors.forename: Francesco
authors.initials: F
authors.lastname: Violi
mesh_terms: D000368:Aged; D000975:Antioxidants; D002099:Cacao; D002182:Candy; D002392:Catechin / Q000097:blood; D018592:Cross-Over Studies; D005080:Exercise Test; D017079:Exercise Tolerance; D005260:Female; D006801:Humans; D028421:Isoprostanes / Q000097:blood; D008297:Male; D008562:Membrane Glycoproteins / Q000097:blood; D008875:Middle Aged; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases / Q000097:blood; D009566:Nitrates / Q000097:blood; D009573:Nitrites / Q000097:blood; D018384:Oxidative Stress; D058729:Peripheral Arterial Disease / Q000097:blood / Q000178:diet therapy* / Q000503:physiopathology; D016037:Single-Blind Method; D014664:Vasodilation; D016138:Walking
chemical_list: D000975:Antioxidants; D028421:Isoprostanes; D008562:Membrane Glycoproteins; D009566:Nitrates; D009573:Nitrites; D002392:Catechin; C056558:CYBB protein, human; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases
keywords: antioxidant; atherosclerosis; chocolate; oxidant stress; peripheral vascular disease