Inflammation, Thrombosis,
Infection, and CARDIOVASCULAR
DISEASE
Nathan D Wong, PhD, FACC
Professor and Director
Heart Disease Prevention Program
University of California, Irvine
Beyond Cholesterol: Predicting
Cardiovascular Risk In the 21st Century
Cardiovascular Risk
Lipids
HTN
Diabetes
Behavioral
Hemostatic
Inflammatory Genetic
Thrombotic
Inflammation and Atherosclerosis
 Inflammation may determine plaque stability
- Unstable plaques have increased leukocytic infiltrates
- T cells, macrophages predominate rupture sites
- Cytokines and metalloproteinases influence both stability
and degradation of the fibrous cap
 Lipid lowering may reduce plaque inflammation
- Decreased macrophage number
- Decreased expression of collagenolytic enzymes (MMP-1)
- Increased interstitial collagen
- Decreased expression of E-selectin
- Reduced calcium deposition
Libby P. Circulation 1995;91:2844-2850.
Ross R. N Engl J Med 1999;340:115-126.
Is there clinical evidence that
inflammatory markers predict future
coronary events and provide additional
predictive information beyond traditional
risk factors?
Evaluating Novel Risk Factors for
CAD
 Consistency of
prospective data
 Strength of association
 Low variability
 High
reproducibility
 Independence of
association
 Biologic
 Improve predictive
value
 Low cost
 Standardized measure
plausibility
 Modifiable
Biomarkers for Venous and Arterial
Thrombosis
Parameter
Venous
Arterial
Fibrinogen
–
+++
Factor VII
–
+
vWF: ag
–
++
tPA: ag
–
+++
PAI-1: ag
–
++
Platelet function
–
++
Lp(a)
–
+
hs-CRP / SAA / IL-6 / TNF
–
+++
Biomarkers for Venous and Arterial
Thrombosis (cont’d)
Parameter
Venous
Arterial
+++
–
++
–
+
–
Factor VIII
++
–
Anti-thrombin III
++
–
Protein C
+
–
Protein S
+
–
Homocysteine
++
++
D-dimer
++
++
Factor V Leiden
Prothrombin mutation
Prothrombin
Thrombosis and Cardiovascular Risk
 Thrombus formation is a crucial factor in the
precipitation of unstable angina or myocardial
infarction, as well as occlusion during or following
angioplasty.
 Often preceded by platelet aggregation and
activation of the coagulation system.
 A thrombus may develop at sites of only mild to
moderate coronary stenosis. The majority of
coronary events occur where there is less than
70% stenosis.
 Occlusive coronary thrombosis plays a role in over
80% of myocardial infarctions and about 95% of
sudden death victims.
Fibrinogen and Atherosclerosis
 Promotes atherosclerosis
 Essential component of platelet aggregation
 Relates to fibrin deposited and the size of the
clot
 Increases plasma viscosity
 May also have a proinflammatory role
 Measurement of fibrinogen, incl. Test
variability, remains difficult.
 No known therapies to selectively lower
fibrinogen levels in order to test efficacy in
CHD risk reduction via clinical trials.
Fibrinogen and CHD Risk:
Epidemiologic Studies
 Recent meta-analysis of 18 studies involving 4018
CHD cases showed a relative risk of CHD of 1.8
(95% CI 1.6-2.0) comparing the highest vs lowest
tertile of fibrinogen levels (mean .35 vs. .25 g/dL)
 ARIC study in 14,477 adults aged 45-64 showed
relative risks of 1.8 in men and 1.5 in women,
attenuated to 1.5 and 1.2 after risk factor
adjustment.
 Scottish Heart Health Study of 5095 men and 4860
women showed fibrinogen to be an independent
risk factor for new events--RRs 2.2-3.4 for
coronary death and all-cause mortality.
Fibrinogen and CHD Risk Factors
 Fibrinogen levels increase with age and body
mass index, and higher cholesterol levels
 Smoking can reversibly elevated fibrinogen
levels, and cessation of smoking can lower
fibrinogen.
 Those who exercise, eat vegetarian diets, and
consume alcohol have lower levels. Exercise
may also lower fibrinogen and plasma
viscosity.
 Studies also show statin-fibrate combinations
(simvastatin-ciprofibrate) and estrogen
therapy to lower fibrinogen.
Other Thrombotic Factors and CHD
 Mixed reports of coagulation factor VIIc in
cardiovascular disease. PROCAM study
showed no association with CHD events, CHS
also showed no relation to subclinical CVD.
 Endogenous tissue-type plasminogen activator
(tPA) shown in some studies to relate to
increased cardiovascular risk--Physician’s
Health Study showed RR for MI 2.8, stroke 3.5
in those in 5th vs. 1st quintile of tPA.
 Plasminogen activitor inhibitor type 1 (PAI-1)
shown associated with increased
cardiovascular risk, esp in diabetic patients.
Aspirin and Cardiovascular Risk:
Clinical Trial Evidence for Primary
Prevention
 US Physician’s Health Study- 22,071 male physicians 44% reduction in MI risk, 13% nonsignificant increase
in risk of stroke
 British Doctor’s Study of 5139 male physicians showed
nonsignificant 3% reduction in MI risk,13%
nonsignificant increase in stroke
 Hypertension Optimal Treatment (HOT) study among
18,790 pts w/htn showed 15% reduction in CVD events,
36% reduction in MI
 Women’s Health Study (n=39,876 women aged 45+)
randomized to 100 mg asprin/day vs placebo, 10 years
follow-up – results recently released and asprin
preventive only for stroke (17% reduction overall,
p=0.04; 24% ischemic stroke, p<.001); nonfatal MI
RR=1.02, CVD death 0.95, ns) (NEJM 2005; 352: 13668).
Aspirin and Cardiovascular Risk:
Clinical Trial Evidence for Secondary
Prevention
 Antiplatelet Trialists Collaboration of 54,000
patients with cardiovascular disease (10 trials
post-MI) showed 31% reduction in MI, 42%
reduction in stroke, 13% reduction in total
vascular mortality
 International Study of Infarct Survival of
17,187 pts w/evolving MI showed 49%
reduction in reinfarction, 26% reduction in
nonfatal stroke, and 23% reduction in total
vascular mortality
Antiplatelet Therapy: Targets
dipyridamole
clopidogrel bisulfate
ticlopidine hydrochloride
phosphodiesterase
ADP
ADP
Gp 2b/3a Inhibitors
Gp IIb/IIIa
(Fibrinogen
Receptor)
Activation
COX
Collagen
Thrombin
TXA2
TXA2
aspirin
ADP = adenosine diphosphate, TXA2 = thromboxane A2, COX = cyclooxygenase
Schafer AI. Am J Med 1996;101:199–209
Antiplatelet Therapy: Common Oral Agents
Trade Name
Class
Formulation
Maintenance Dose
Major Bleeding Risk
(%)
Acetylsalicylic acid
(ASA)
Clopidogrel bisulfate*
Ticlopidine
hydrochloride*
Aspirin
Plavix®
Ticlid®
Salicylate
Thienopyridine
Thienopyridine
Active Drug
Pro-Drug
Active Drug
75-325 mg daily
75 mg daily
250 mg twice daily
2.4-3.3%1
1.0-3.7% alone2,3
3.0-4.9% w/ ASA4
1.0% alone5
1.7-5.5% w/ ASA6,7
1Topol
*Clopidogrel is generally given preference over
2Diener
Ticlopidine because of a superior safety profile
EJ et al. Circulation. 2003;108:399-406
H-C et al. Lancet 2004;364;331-7
3Plavix® package insert. www.sanofi-synthelabo.us
4Peters RJ et al. Circulation 2003;108:1682-7
5Hass WK. NEJM 1989;321:501-7
6Urban P. Circulation. 1998;98:2126-32
7Ticlid® package insert. www.rocheusa.com
Aspirin: Mechanism of Action
Membrane Phospholipids
ARACHIDONIC ACID
COX-1
Aspirin
Prostaglandin H2
Thromboxane A2
 Platelet Aggregation
Vasoconstriction
Prostacyclin
 Platelet Aggregation
Vasodilitation
Aspirin Recommendations
I IIa IIb III
Primary Prevention
Aspirin (75-162 mg daily) for intermediate risk men
with a 10 year risk of CHD >10%.
I IIa IIb III
Aspirin (75-162 mg daily) for intermediate risk
women with a 10 year risk of CHD >10%.
Aspirin for low risk women with a 10 year risk of
I IIa IIb III CHD<10%.
Aspirin (75-325 mg daily) for those with known CHD.
I IIa IIb III
Secondary Prevention
Rader, NEJM 2000; 343: 1181.
P. Ridker
CRP vs hs-CRP
 CRP is an acute-phase protein produced by the liver
in response to cytokine production (IL-6, IL-1,
tumor necrosis factor) during tissue injury,
inflammation, or infection.
 Standard CRP tests determine levels which are
increased up to 1,000-fold in response to infection
or tissue destruction, but cannot adequately assess
the normal range
 High-sensitivity CRP (hs-CRP) assays (i.e. Dade
Behring) detect levels of CRP within the normal
range, levels proven to predict future cardiovascular
events.
Potential Mechanisms Linking CRP
to Atherothrombosis
 Confounding by cigarette
consumption
 Innocent bystander
- Acute phase response
 Cytokine surrogate
- IL-6, TNF-, IL-1
 Direct effects of CRP
- Innate immunity
- Complement activation
- CAM induction
 Prior infection
- Chlamydia, H pylori, CMV
 Marker for subclinical
atherosclerosis
- EBCT / IMT / ABI
 Marker for insulin
resistance/ obesity
 Marker for endothelial
dysfunction
 Marker for dysmetabolic
syndrome
 Marker for plaque
vulnerability
hs-CRP and Risk of Future MI in
Apparently Healthy Men
P Trend <0.001
P<0.001
3
Relative Risk of MI
P<0.001
2
P=0.03
1
0
1
2
3
Quartile of hs-CRP
Ridker et al, N Engl J Med. 1997;336:973–979.
4
hs-CRP and Risk of Future Stroke in
Apparently Healthy Men
P Trend <0.03
Relative Risk of
Ischemic Stroke
2
P=0.02
P<0.02
3
4
1
0
1
2
Quartile of hs-CRP
Ridker et al, N Engl J Med. 1997;336:973–979.
hs-CRP as a Risk Factor For Future CVD :
Primary Prevention Cohorts
Kuller MRFIT 1996
CHD Death
Ridker PHS 1997
MI
Ridker PHS 1997
Stroke
Tracy CHS/RHPP 1997
CHD
Ridker PHS 1998,2001
PAD
Ridker WHS 1998,2000,2002
CVD
Koenig MONICA 1999 CHD
Roivainen HELSINKI 2000
CHD
Mendall CAERPHILLY 2000
CHD
Danesh BRHS 2000
CHD
Gussekloo LEIDEN 2001
Fatal Stroke
Lowe SPEEDWELL 2001
CHD
Packard WOSCOPS 2001
CV Events*
Ridker AFCAPS 2001
CV Events*
Rost FHS 2001
Stroke
Pradhan WHI 2002
MI,CVD death
Albert PHS 2002
Sudden Death
Sakkinen HHS 2002
MI
0
Ridker PM. Circulation 2003;107:363-9
1.0
2.0
3.0
4.0
5.0
Relative Risk (upper vs lower quartile)
6.0
hs-CRP Adds to Predictive Value of TC:HDL
Ratio in Determining Risk of First MI
Relative Risk
5.0
4.0
3.0
2.0
1.0
0.0
High
Medium
High
Medium
Low
Total Cholesterol:HDL Ratio
Ridker et al, Circulation. 1998;97:2007–2011.
Low
Risk Factors for Future Cardiovascular
Events: WHS
Lipoprotein(a)
Homocysteine
IL-6
TC
LDLC
sICAM-1
SAA
Apo B
TC: HDLC
hs-CRP
hs-CRP + TC: HDLC
0
1.0
2.0
4.0
6.0
Relative Risk of Future Cardiovascular Events
Ridker et al, N Engl J Med. 2000;342:836-43
Is there clinical evidence that
inflammation can be modified by
preventive therapies?
Percent with CRP 0.22
mg/dL
Elevated CRP Levels in Obesity:
NHANES 1988-1994
25
20
15
10
5
0
Normal
Overweight
Visser M et al. JAMA 1999;282:2131-2135.
Obese
Effects of Weight Loss on CRP
Concentrations in Obese Healthy
Women
 83 women (mean BMI 33.8, range 28.2-43.8 kg/m2) placed
on very low fat, energy-restricted diet (6.0 MJ, 15% fat) for
12 weeks
 Baseline CRP positively associated with BMI (r=0.281,
p=0.01)
 CRP reduced by 26% (p<0.001)
 Average weight loss 7.9 kg, associated with change in CRP
 Change in CRP correlated with change in TC (r=0.240,
p=0.03) but not changes in LDL-C, HDL-C, or glucose
 At 12 weeks, CRP concentration highly correlated with TG
(r=0.287, p=0.009), but not with other lipids or glucose
Heilbronn LK et al. Arterioscler Thromb Vasc Biol
2001;21:968-970.
Effect of HRT on hs-CRP: the PEPI
Study
hs-CRP (mg/dL)
3.0
CEE + MPA cyclic
CEE + MPA
CEE
+ MP
continuous
CEE
2.0
Placebo
1.0
0
12
Months
36
Cushman M et al. Circulation 1999;100:717-722.
1999 Lippincott Williams & Wilkins.
Long-Term Effect of Statin Therapy on
hs-CRP: Placebo and Pravastatin Groups
Placebo
0.25
Median hs-CRP
Concentration
(mg/dL)
0.24
0.23
-21.6%
(P=0.004)
0.22
0.21
0.20
Pravastatin
0.19
0.18
Baseline
Ridker et al, Circulation. 1999;100:230-235.
5 Years
Effect of Statin Therapy on hs-CRP Levels
at 6 Weeks
hs-CRP (mg/L)
6
*p<0.025 vs. Baseline
5
*
4
*
*
3
2
1
0
Baseline
Prava
Simva
Atorva
(40 mg/d) (20 mg/d) (10 mg/d)
Jialal I et al. Circulation 2001;103:1933-1935.
2001 Lippincott Williams & Wilkins.
Change at 24 weeks, %
Effect of Bezafibrate with and without
Fluvastatin on Plasma Fibrinogen, PAI-1, and
CRP in Patients with CAD and Mixed
Hyperlipidemia
15
10
5
0
-5
-10
-15
-20
Fibrinogen
PAI-1
CRP
70 72 63
83 80 75
n: 81 80 74
*
*
Beza
400
mg/d
P<0.05 vs. baseline
*
Beza 400 mg/d
+ fluva 20 mg/d
Cortellaro M et al. Thromb Haemost 2000;83:549-553.
Beza 400 mg/d
+ fluva 40 mg/d
JUPITER
Randomized Trial of Rosuvastatin in the Primary
Prevention of Cardiovascular Events Among
Individuals with Low Levels of LDL-C
and Elevated Levels of hs-CRP
Rosuvastatin (N =7500)
No History of CAD
Men > 55, Women > 65
LDL-C <130 mg/dL
hs-CRP >2 mg/L
Screening Screening
Visit 1
Visit 2
hs-CRP
Lipids
hs-CRP
LFTs
HbA1C
4 week
Run-in
Randomization
Visit
Placebo (N =7500)
Safety
Visit
hs-CRP
LFTs
UA
Bi-Annual
Follow-Up Visits
MI
Stroke
Unstable
Angina
CVD Death
CABG/PTCA
End of Study
Visit
hs-CRP
LFTs
UA
Lipids
HbA1C
N Engl J Med. 2002;347:1157-1165
Event-Free Survival According to Baseline Quintiles of
C-Reactive Protein and LDL Cholesterol
Quintiles of LDL
0.99
0.99
1
1
3
2
0.98
0.98
2
3
4
0.97
0.97
4
5
0.96
5
0.96
CVD Event-Free Survival Probability
1.00
1.00
Quintiles of CRP
0
2
4
6
Years of Follow-Up
Ridker et al, N Engl J Med. 2002;347:1157-1165.
8
0
2
4
6
Years of Follow-Up
8
CV Event-Free Survival Using Combined
hs-CRP and LDL-C Measurements
1.00
Probability of Event-free Survival
Median LDL 124 mg/dl
Median CRP 1.5mg/l
Low CRP-low LDL
0.99
Low CRP-high LDL
0.98
High CRP-low LDL
0.97
0.96
High CRP-high LDL
0.00
0
2
4
6
Years of Follow-up
Ridker et al, N Engl J Med. 2002;347:1157-1165.
8
hs-CRP Adds Prognostic Information at all Levels of
LDL-C and at all Levels of the Framingham Risk Score
<1.0
1.0-3.0
>3.0
C-Reactive Protein (mg/L)
3
Multivariable relative risk
25
Relative risk
20
15
10
5
0
<1.0
0-1
2-4
5-9
10-20
Framingham estimate of 10-year risk (%)
Ridker et al, N Engl J Med. 2002;347:1557.
1.0-3.0
>3.0
C-Reactive Protein (mg/L)
2
1
0
<130
130-160
>160
LDL cholesterol (mg/dL)
What is the role of hs-CRP with
regard to diabetes and the
metabolic syndrome?
Circulation. 2003;107:391-397.
Plasma hs-CRP Levels According to
Severity of the Metabolic Syndrome
C-reactive protein (mg/L)
8
6
4
2
0
0
1
2
3
4
5
Number of Components of the Metabolic Syndrome
Ridker et al, Circulation 2003;107:391-7
Event Free Survival According to hs-CRP Levels: Analysis
Limited to Participants with Metabolic Syndrome at Baseline
1.00
CVD Event-Free
Survival Probability
0.99
0.98
CRP <1 mg/L
0.97
CRP 1-3 mg/L
0.96
CRP >3 mg/L
0.95
0
2
4
6
Years of Follow-Up
Ridker et al, Circulation 2003;107:391-7
8
AHA / CDC Scientific Statement
Markers of Inflammation and Cardiovascular Disease:
Applications to Clinical and Public Health Practice
Circulation January 28, 2003
“Measurement of hs-CRP is an independent marker of risk
and may be used at the discretion of the physician as part
of global coronary risk assessment in adults without known
cardiovascular disease. Weight of evidence favors use
particularly among those judged at intermediate risk by
global risk assessment”.
Clinical Application of hs-CRP for
Cardiovascular Risk Prediction
1 mg/L
Low
Risk
3 mg/L
Moderate
Risk
Ridker PM. Circulation 2003;107:363-9
10 mg/L
High
Risk
>100 mg/L
Acute Phase Response
Ignore Value, Repeat Test in 3 weeks
Inflammatory and Infections Agents in CHD
 Belgian epidemiologic study included 446 of
16307 male workers aged 35-39 who had
evidence of CHD vs. 892 controls.
 CRP, but none of the infectious agents (H.
pylori, C. pneumoniae, CMV, and EBV) were
associated with CHD, even after adjustment
for other risk factors.
De Backer et al. Atherosclerosis 2002; 160: 457-63.
Infection and CHD - is there a
connection?
 Local or systemic infections resulting from
gram negative bacteria such as Chlamydia
pneumoniae and Helicobacter pylori, including
cytomegalovirus (CMV) have been implicated
in atheroscelosis
 While several case control studies have shown
increased titers of C.pneumoniae and H. Pylori
in those with vs. without CHD, convincing
evidence from prospective studies is lacking.
Prospective Studies of CHD and
Infectious Pathogens
 Physician’s Health Study (nested case-control)
shows RR 1.1 (0.8-1.5) for C. Pneumoniae,
0.94 (0.7-1.2) for cytomegalovirus, and 0.72
(0.6-0.9) for Herpes simplex virus.
 H. pylori also shows mixed results. Whincup
showed a nonsignificant 1.3 OR when adjusted
for other risk factors, the large ARIC study
showed no relation, and the Caerphilly
Prospective study showed RR=1.05 in 1796
men followed 14 years.
Other Studies of Infectious Agents
 In South Asian persons with CHD vs. controls, C. pneumoniae
specific IgG antibody was seropositive in similar proportions; risk
factors appeared to mediate any relations (Mendis et al. Int J
Cardiol 2001; 79: 191-6).
 Cross-sectional survey of 704 individuals of C. pneumoniae and
CMV with risk factors did nto show significant associations
(Danesh et al., J Cardiovasc Risk 1999; 6: 387-90).
 Meta-analysis of 24 articles involving H. pylori infection and CHD
showed a pooled odds ratio of 1.55 (95% CI: 1.38-1.74)
(p<0.001), suggested a weak relation, but high hetrogeneity
between studies precludes clear demonostration (Pellicano et al.,
Eur J Epidemiol 1999; 15: 611-9).
 ARIC Study failed to show clear relation between IgG antibodies
for C. pneumoniae and incident CHD occurring over average 3.3
years. (Nieto et al. Am J Epidemiol 1999; 150: 149-56).
Clinical Trial Evidence for Antibiotic
Treatment and Prevention of CVD
 ACADEMIC Study of 302 patients with CHD seropositive to
C. Pneumoniae randomized to azithromycin 500 mg/wk or
placebo for 3 months showed no significant treatment
difference (HR=0.89, p=0.74) for recurrent events
(Muhlestein et al., Circulation 2000; 102: 1755-60).
 AZACS Multicenter study of 1439 pts with unstable angina
randomized to 250 mg azithromycin/day for up to 6
months showed no significant benefit for death, recurrent
MI, or recurrent ischemia (Cercek et al., Lancet 2003;
361: 809-13).
 WIZARD trial of 7,747 pts post-MI randomized to 12 week
of therapy with azithromycin or placebo showed no
significant reduction in reinfarction, revascularization,
hospitalization for angina, or death (O’Connor et al., JAMA
2003; 290: 1459-66).
Infectious Agents and the Future
 Individuals with greater infectious burdens
may be at greater risk, because they are older,
have poorer health habits, less access to care.
 Observed associations often may be due to
selection biases or confounding from age and
other factors
 Prospective clinical trials under way examining
role of certain antibiotics such as azithromycin
on reduction of recurrent events in CHD
patients.
 Until these data are available, no role for
measurement or treatment of infectious
burden.