Dr. Gemma VilahurCardiovascular Research Center
CSIC-ICCCInstituto de Investigaciones Biomédicas Sant Pau (IIB-Sant Pau)
Barcelona
How big is the problem?
WORLDWHO 201217,5 million people died from CVD (mainly ISCHEMIC HEART DISEASE)LEADING CAUSE OF DEATH GLOBALLY(31% total)
G. Vilahur , L. Badimon et al Eur Heart J 2014
Thrombotic
occlusion
THROMBUS
Atheroscerotic plaque
From the culprit coronary thrombotic occlusion to platelets
Thrombus formation I: Platelet aggregationL. Badimon, R. Storey, G. Vilahur. Thromb Haemost. 2011
Foam Cell
GPIIb/IIIavWF
GPIb
Vitronectin
αvβ3
Collagen
GPVI
GPIa/IIavWF
AATXA2
ADP
P2Y
TF
Fg
PSGL
Platelet
vWF
Thrombi
n
GPIIb/IIIa PAR
Fg
CollagenP-selectin
Collagen
Initial contact
Firm adhesion
Activation
AggregationADP
vWF
Platelet recruitment: G protein–mediated signaling pathways
• ADP• Thrombin• Thromboxane A2
•Common activation of downstream effectors that lead to GPIIb/IIIa activation
L. Badimon & G. Vilahur. J Int Med. 2015
ADP
PAR-1PAR-4
TP
GPIIb/IIIa
PLC
Fibrinogen binding
Platelet aggregation
PLCPLC hydrolyzes PIP2 to
yield IP3 and DAG
G-protein mediatedactivation of PLC
ADP
ADP
P2Y1
P2Y12
GPIIb/IIIaactivation
PIP2PLC
VASP-P
Adenylyl cyclase (AC) inhibition
α AMPc
PKA
AC
VASP
AMPc decrease
Reduction PKA activation
DAG
PKC
PKC activation
GPIIb/IIIaactivation
Ca2+
Ca2+
Ca2+
Ca2+
IP3
GPIIb/IIIaactivation
Calcium cytosolincrease
TXA
2
TXA2
PLA2AA
PGH2
PGG2
PLA2 activation
TXA2 generation
PLATETL
DEGRANULATION
XIa
IX
IXa
XII PC
XI
Intrinsic pathway
Va
PLATELET ACTIVATION
X
XaVII
Extrinsic pathway
FT
VIIa
Prothrombin
TISSUE FACTOR
Fibrinogen FIBRIN
PAR-1/PAR-4
L.Badimon & G.Vilahur. Eur Heart J Vasc Biol 2015
Thrombus formation II: Activation of the coagulation cascade
VIIIa
SECONDARY PREVENTIONPrevent atherothrombosis
PRIMARY PREVENTION
What increases the risk of CVD?
YOU CAN HELP IT!
YOU CANNOT HELP IT!
Unhealthy eating
RISK RISK
Healthy eating
>45 yr >55 yr
Seven countries study
Ancel Benjamin Keys 1958Healthy diet = heatlhy heart
A food pattern shared by countires that rim the Mediterranean Sea
(no official Mediterranean diet)
Spain
Italy
GreeceTurkey
North Africa
Israel
Lebanon
What is the Mediterranean diet ?
Mediterranean
dietCheese and yogurt
Moderate in Dairy products
Beef or pork (small amounts)
Rabbit and chicken (lean meat)
more recommended
Poor in Meat
Nuts (i.e.,almonds and hazelnuts) Olive oil
Rich in ω-3/ω-6 fatty acids (PUFA)
Rich in ω -9 fatty acids (MUFA)
Walnuts
Vegetables
Fruit
Fatty fish
(anchovy, sardine, mackerel,
red tuna, etc)
Legumes
Fermented beverage
(moderate amounts)
Rich in polyphenols
Olive oil
Grains (especially wheat)
Rich in fiber
Diet characteristics of the Mediterranean dietL.Badimon & G.Vilahur. In press 2015
Secondary Prevention(AMI-patients)
Lyon Diet Heart StudyDe Lorgeril et al Lancet 1994
FREE CANOLA OIL BASED MARGARINE: high omega-3 PUFA
Mediterrenan dietFrench diet (control)
Primary Prevention(Asymptomatic high-risk patients)
PREDIMEDRamon Estruch et al New Eng J Med 2013
PR
IMA
RY
EN
D-P
OIN
T:
AM
I, s
tro
ke o
r d
eath
fro
m C
VD
(3
0%
↓)
PRIMARY PREVENTION: Beneficial effects of a healthy food
IMPROVED CARDIOVASCULAR HEALTHREDUCED CVD MORBI/MORTALITY
Improves vascular functionReduced inflammationEnhance NO availabilityImproved reactive hyperemea indices
Reduces blood pressure
Improved vascular functionReduces inflammationEnhanceNO availabilityReduces ROS
Reduces platelet reactivity
Improved platelet functionEnhance NO availability
Improves lipid profiles
Reduces LDL-cholesterolReduces TGReduces total Cholesterol
Reduces oxidative stress
Improves anti-oxidant capacityReduces LDL-oxidation
Reduces weight
Reduces ROSRuduce lipid profilesReduce blood pressureImproves exercise capacity
Adapted from R. Jay Widmer et al American Journal Med 2015
THROMBOTIC RISK
REDUCTION
G.Vilahur & L.Badimon Vasc Pharmacol 2014
Food stuff that known to reduce platelet reactivity
Energy 885 kcal
Carbohydrates 0 g
Protein 0 g
Fat
saturated 14 g
polyunsaturated 11 gMonounsaturated
(oleic acid) 73 g
Heatlhy properties of olive oil: MUFA and PHENOLYIC COMPOUNDS
Control cholesterolImproves blood pressureImproves lipid profileAnti-inflammatory effects
Badimon & Vilahur . Cardiovasc Therapeut 2010
Delgado Lista et al. Curr Pharnaceut Des 2011
Covas MI. Pharmacol Res 2007
Improves endothelial functionReduces LDL oxidationeImproves lipid profile
Lowers platelet reactivity
Polyphenols
A3A1
A3A1 RGD
EPINEPHRIN
vWF
Fibrinogen
PAR-1/PAR-4
P2Y12
GPIIb/IIIa
GPIIb/IIIa
TXA2
PLA2
AA
PGH2
PGG2
GPVI
GPIb-IX-V
GPIIb/IIIa
SEROTONIN
5-HT2
α cAMP
PKAVASP-P
VASP
β
ACCa2+
Ca2+
Ca2+
Ca2+
PLCβ
PIP2
IP3
DAG
PKC
PI3K
A3A1
A3A1 RGD
PLATELET
GRANULES
ADP
vWF
Fibrinogen
ADP
PLATETL
DEGRANULATION
PLCβ
Ca2+
Ca2+
Ca2+
VITRONECTIN
GPIa/IIa
ATP
COLLAGEN
MAPK
MAPK
TP
OLIVE OIL
GPIIb/IIIa
inhibition
activation
vWF
OLIVE OIL?
Rubio-Senent Eur J Nutr 2014
Mezzano et al Pathophysiol Thromb Haemost 2003
Turpeinen et al. Net Met Cardiovasc Dis 1999
COLLAGEN
OLIVE OIL
OLIVE OIL
OLIVE OIL
Differential proteomics: ADP-stimulated platelets
Alperujo extractNon-alperujo
Phenolyc compounds in olive oil protect against platelet activation
Alperujo extract
GPIIb
Rho GDP-dissociationinhibitor
Annexin A5
1. Regulation of platelet structure2. Anticoagulant effects3. GPIIb/IIIa
Roos et al Eur J Nutr 2011
- Hydroxytyrosol (HT)
- Dihydroxyphenylglycol
(DHPG)
Platelet aggregationPlatelet activation (AE: 40mg/mL)
Platelet inhibition
MUFAs reduce the activation of the coagulation cascadeL. Badimon & G.Vilahur. Eur Heart Journal ACC 2012 (minimally modified)
Francisco Perez-Jime nez et al Public Health Nutr 2006 MUFA vs SAFA
In a prospective study of 43,757 male health professionals followed for 6 years, a relatively small
increase in ALA intake (1% of total energy) was associated with a 59% decrease in the risk of acute MI
PUFA: Essential fatty acids
Alpha-linolenic acid (ALA)
Omega-3 Fatty Acids: ALA, EPA, DHA
Eicosapentaenoic acid (EPA)
Docosahexaenoic acid (DHA)
Must be obtained from the diet
Omega-6 Fatty Acids
Linoleic acid (LA)
Arachidonic acid (AA)
Weak anti-inflammatory & anti-thromboticmediators
PG3 LTB5 TXA3
Potent Pro-inflammatorypro-thromboticmediators
PG2 LTB4 TXA2
Elongase
6-desaturase
5-desaturase
COX
Omega-6
Linoleic acid
Gamma linoleic acid
Arachidonic acid
(AA)
Dihomo GLA
Omega-3
Alpha-linoleic acid
Stearic acid
Eicosatetraenoic acid
Eicosapentaenoic acid
(EPA)
Preferred
pathway
As the Omega-6/Omega-3 ratio decreases, so does the platelet aggregationFreese et al Thromb & Haemost 1994
The OMEGA-PCI (OMEGA-3 fatty acids after PCI to modify
responsiveness to dual antiplatelet therapy) study
Addition of omega-3 ethyl esters to aspirin and
clopidogrel therapy modifies antiplatelet
response in patients with stable CAD
undergoing PCI
63 patients- Stable CAD- Elective PCI- Aspirin and clopidogrel on board
Platelet function test
ASA+clopidogrel (600mg) +EPA+DHA (1gr)ASA+clopidogrel (600mg) +placebo
Platelet function test(12h)
ASA+clopidogrel +EPA+DHAASA+clopidogrel +placebo
Platelet function test3-5 days1month
Platelet function test3-5 days1month
G. Gajos, et al JACC 2010
1 month
Lycopene? Adenosine?
Tomatoes : red, green, sauce, boiled…are they all the same?
MaximalAggregationADP Collagen
MaximalAggregation TRAP6
MaximalAggregation AA
MaximalAggregation
E. Fuentes et al 2013
Green vs red tomatoes
Fresh vs pomace (heated)
Do tomatoes exert antiplatelet effects via lycopene-related effects?
Red-colored pulp more than 40 times higher lycopene content than that in the green pulp (lycopene is directly related to color)
MaximalAggregationADP Collagen
MaximalAggregation
1. Lycopene partially contributes to
inhibit platelet activation
2. Heat does not affect antiplatlet
function of tomatoes
GPIIb/IIIa
P2Y12
Platelets
A2A
Platelet inhibition
A1A2aA2bA3
Adenosine receptors
- Vasodilation- Cardioprotection
(post-conditioning)- Modulation of inflammation
ADENOSINE
Potential adenosine-related antiplatelet effects
1 kg of fresh tomato contains about 30-times more adenosine than lycopene.
Garlic and onions
Antiplatelet properties are lost with cooking (aliinase is deactivated by heat)
Bordia A, et al. Effect of garlic (Allium sativum) on blood lipids, blood sugar, fibrinogen and fibrinolytic activity in patients with
coronary artery disease.
Prostaglandins Leukot Essent Fatty Acids. 1998.
>>
Kiesewetter H, et al Effect of garlic on thrombocyte aggregation, microcirculation, and other risk factors.
Int J Clin Pharmacol Ther Toxicol. 1991
Kiesewetter H, et al Effects of garlic coated tablets in peripheral arterial occlusive disease.
Clin Investig. 1993
Steiner M, Lin RS. Changes in platelet function and susceptibility of lipoproteins to oxidation associated with administration of aged
garlic extract.
J Cardiovasc Pharmacol. 1998
Barrie SA, et al. Effects of garlic oil on platelet aggregation, serum lipids and blood pressure in humans.
J Orthomolecular Med. 1987
Two cloves/day
Garlic and onions majorly interfere with AA metabolism
Sulfur compounds from gralic and onions
Reduces platelet
aggregation
induced by ALL
agonists
Decreases
functional
GPIIb/IIIa
receptors
Interferes with
membrane viscosity
(inner layer)
Rendu et al Biochem Pharmacol 1989
FRAMINGHAM
Heart study
Friedman, 1986
Honolulu Heart
Study
Blackwelder,1980
Caerphilly Scotish
Study
Renaud, 1993
The Copenhagen City
Heart Study
Tjionneland,1999
Thun et al N Eng J Med 1997
Rimm et al. Br Med J 1999
Hennekens et al, JAMA 1979
Fuchs et al N Eng J Med 1995
Camargo et al Arch Intern Med 1997
Stampfer et al N Eng J Med 1988
Yano et al N Eng J Med 1977
Cardiovascular effects of moderate intake of fermented beverages
Mortality reduction in around 33%
Coronary and cerebrovascular risk reduction in around 20-40%
U or J - SHAPE
Benefitial effects of wine intake
Moderate alcohol consumption:30g/day men: 2-3 beers, 2-3 glasses of wine20g/day women: 1-2 beers, 1-2 glass of wine
•↓ Coagulation(PAI) •↓ Fibrinogen, VWF•↑ Fibrinolysis (tPA)•↑ HDL•↓ LDL oxidation•↓ ICAM, VCAM, E-selectin•↓ PCR y IL-6•↑ VEGF y adiponectin•↓ Homocysteinemia↑ PKC-ε•↑ Paraoxanase-1•↑ Insulin sensitivity
Platelet
aggregation
NO
PGI2
TxA2
MCP-1 VEGF
TF
TF
MMPs
CRP
LDLox
LDLox
LDLox
HDLHDL
+
Monocyte
adhesion
LDL
Lipoprotein
oxidation
Wine
WineBadia,2004
++ Wine
Leikert,2002
Wine
Kondo,1994
Gaziano,1993
Guivernau,1989
Wine
LDLox
Beneficial effects of moderate intake of red wineModified B.C.Choi, G.Vilahur, JJ. Badimon. Current Molecular Medicine, 2006
Hyperlipemiccontrol
Hyperlipemic + HD wine(40g alcohol/día)
Hyperlipemic + LD wine(20g alcohol/día)
Platelets
Fibrin
Wine reduces thrombus formation and fibrin depositionL.Casani, G. Vilahur, L. Badimon et al Circulation 2004
GDP
GDPInactive Rho A
GEF
GTPActive Rho A
GTP
resveratrol
PLATELET AGGREGATION
eNOS
PLATELET AGGREGATION
[AMPc][GMPc]
PDEAMPGMP
Resveratrol-related antiplatelet mechanisms of actionL.Casani, G. Vilahur, L. Badimon et al Circulation 2004
Dark chocolate: the “sweetest” antiplatelet food
The fruit of the Theobroma cacao tree
Source Flavanol Content(mg/kg or mg/L)
Chocolate 460–610
Green tea 100–800
Red wine 80–300
Black tea 60–500
Corti R et al. Circulation 2009Catehins and Epicatechins
Dark chocolate: Polyphenol content 30% to 80% cocoaMilk chocolate: Polyphenol content 7% to 35% cocoaWhite chocolate: does not contain polyphenols.
Cocoa and chocolate have higher levels of polyphenolsthan in many fruits:
20 times higher than in tomatoes2 times higher than in garlic3 times higher than in grapes
Dark chocolate: is it all about flavanols?Rull et l Vasc Pharmacol 2015
6 weeks intake 50g/day
Improves vasodilation
PDE inhibitor
AMPc
GMPc
PDE
AMPGMP
Platelet inhibition
Kamruzzaman SM et al Phytochem Res 2011
Kamruzzaman SM et al Vasc Phamacol 2013
Mechanisms involved in platelet inhibition
G.Vilahur. & L.Badimon Vasc Pharmacol 2014
The impact of foods and dietary supplements onnormal hemostasis and antithrombotic therapyshould be taken into consideration in patients
prescribed antithrombotic drugs
Many foods may exert beneficial effects in primaryprevention by modulating platelet
function…moreover…foods may offer new leads forthe development of antiplatelet agents..yet…
1.- En el proceso de adhesión y activación plaquetar:
1. La exposición de contenido aterogénico al torrente circulatorio es el único desencadenante de la
adhesión / activación plaquetar
2. El factor de Von Willebrand juega un papel clave en la adhesión plaquetar bajo cualquier condición de
flujo sanguíneo
3. El receptor plaquetar GPIIb/IIIa es el efector final en el que convergen todas las vías de señalización
plaquetar
4. El ADP liberado por las células blancas induce activación plaquetaria
2.- En cuanto a la influencia de la dieta y/o nutrientes sobre la función plaquetar:
1. Todos los alimentos poseen propiedades antiplaquetares
2. Las propiedades antiplaquetares de determinados alimentos (cacao, vino, aceite) viene determinada
exclusivamente por su contenido en antioxidantes
3. Los ácidos grasos monoinsaturados (MUFA) w-6 presentes en el pescado desplazan los ácidos grasos
poliinsaturados (PUFA) w-3 de la membrana de las plaquetas lo que deriva en última instancia en una
menor reactividad plaquetaria.
4. Sólo las formas crudas del ajo y la cebolla ejercen efectos antipalquetares.
Las preguntas……..
3.- En el contexto del vino como producto natural capaz de inhibir la activación plaquetar:
1. El contenido en alcohol modula de manera dosi-dependiente la función plaquetaria
2. El vino interfiere en múltiples vías de activación plaquetar
3. Su contenido en resveratrol (polifenol presente en la pulpa de la uva) es quien determina el grado de
inhibición plaquetar.
4. Sólo el vino blanco ejerce efectos antiplaquetares.
4.- Los siguientes componentes bioactivos ejercen efectos antiplaquetares:
1. Hidroxitirosol presente en el aceite de pescado
2. Los ácido eicosapentanoico y ácido docosahexaenoico presentes en el aceite de pescado
3. Compuestos ricos en sulfuro presentes en las setas
4. El licopeno presente en el chocolate negro
EPINEFRINA
FvW
Fibrinógeno
PAR-1/PAR-4
P2Y1
P2Y12
GPIIb/IIIa
GPIIb/IIIa
TP
ATP
TXA2
PLA2
AA
PGH2
PGG2
GPVI
GPIb-IX-V
SEROTONINA
5-HT2
P2X1
α cAMP
PKAVASP-P
VASP
β
ACCa2+
Ca2+
Ca2+
Ca2+
PLCβPIP2
IP3
DAG
PKC
PI3K
A3A1
A3A1 RGD
Gránulos
plaquetares
ADP
FvW
Fibrinógeno
FvW
ADP
Degranulación
plaquetar
PLCβ
Ca2+
Ca2+
Ca2+
COLLAGEN
VITRONECTIN
GPIa/IIa
ATP
COLLAGEN
MAPK (JNK,
p38MAPK)
MAPK (Erk)TOMATOES
TOMATOES
GPIIb/IIIaTOMATOES
inhibición
activación
MAPK
(Erk)
PLCγ