Nieuwe CVD Risicofactoren: Wat kunnen we zien?

Transcription

1 Nieuwe CVD Risicofactoren: Wat kunnen we zien? Michel Langlois, MD, PhD Department of Clinical Chemistry AZ Sint-Jan Brugge & University Hospital Gent Belgium

2 Laboratory parameters for cardiovascular risk stratification Unalterable risk factors Genetic markers Lp(a) > 30 mg/dl Alterable risk factors Cholesterol > 190 mg/dl LDL > 115 mg/dl HDL < 40 mg/dl Triglycerides > 180 mg/dl Homocysteine> 15 µmol/l Alterable or unalterable CRP, IL-6 elevated

3 Prospective Study TC and CHD 4 16 CHD Risk Ratio CHD Mortality Rate Serum chol Serum chol. Framingham Study Pooling Project Israeli Prosp. Study MRFIT-Study n = males

4 Misleading cholesterol! Atherogenic & less atherogenic lipid particles Pre-analytical & biological variation

5 Not all LDL are equally atherogenic Highly atherogenic Small dense LDL (1.040 < d < g/ml): more susceptible to oxidation less readily cleared by LDL receptor OxLDL: direct uptake by monocyte / macrophage Lp (a) Less atherogenic Normal LDL ( < d < g/ml) «Lowering the concentration of normal LDL may have a limited effect on CHD risks»

6 Misleading cholesterol LDL cholesterol 135 mg/dl LDL cholesterol 135 mg/dl LDL cholesterol distribution LDL cholesterol distribution 55 mg/dl 60 mg/dl 25 mg/dl 30 mg/dl 40 mg/dl 65 mg/dl LDL I light LDL II intermediate LDL IIII dense LDL I light LDL II intermediate LDL IIII dense

7 PRE-ANALYTICAL FACTORS VARIATION IN PATIENT PREPARATION Fasting status Posture VARIATION DUE TO SAMPLE PROCESSING Venous occlusion Fingerprick vs. venous blood Serum vs. plasma Anticoagulant Storage temperature

8 BIOLOGICAL VARIATION IN SERUM LIPIDS AND LIPOPROTEINS PHYSIOLOGICAL VARIATION Temporal Seasonal Age Menstrual cycle Pregnancy BEHAVIOURAL VARIATION CLINICAL VARIATION

9 BIOLOGICAL VARIATION IN SERUM LIPIDS AND LIPOPROTEINS PHYSIOLOGICAL VARIATION BEHAVIOURAL VARIATION Diet Obesity Exercise Smoking Alcohol Coffee CLINICAL VARIATION

10 BIOLOGICAL VARIATION IN SERUM LIPIDS AND LIPOPROTEINS PHYSIOLOGICAL VARIATION BEHAVIOURAL VARIATION CLINICAL VARIATION Drugs Myocardial infarction Infection and inflammation Trauma and surgery Malignancy Secondary hyperlipidaemia

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12 Laboratory parameters for cardiovascular risk stratification Unalterable risk factors Alterable risk factors Genetic markers Lp(a) > 30 mg/dl Cholesterol LDL HDL Triglycerides Homocysteine Alterable or unalterable CRP, IL-6

13 Triglycerides & Metabolic Syndrome Synonyms Insulin resistance syndrome Syndrome X Dysmetabolic syndrome

14 Metabolic syndrome Central (abdominal) obesity, BMI > 25 Atherogenic dyslipidemia: high TG, sd-ldl, low HDL Hypertension Insulin resistance/glucose intolerance Hyperuricemia Prothrombotic state: fibrinogen, PAI-1 inhibitor Pro-inflammatory state: hs-crp, IL-6 Endothelial dysfunction: microalbuminuria, hyperhomocysteinemia

15 National Cholesterol Education Program (NCEP), NIH Adult Treatment Panel (ATP III) criteria Waist circumference >102 cm (men), >88 cm (women) HDL-C <40 mg/dl (men), <50 mg/dl (women) TG >150 mg/dl Blood pressure >130/85 mmhg Fasting glucose >110 mg/dl 3 features = Metabolic syndrome

16 Metabolic Syndrome (continued) Therapeutic Objectives (ATP III guidelines) To reduce underlying causes Overweight and obesity Physical inactivity To treat associated risk factors Hypertension Atherogenic dyslipidemia (lipid triad) Inflammation? Hyperhomocysteinemia?

17 Laboratory parameters for cardiovascular risk stratification Unalterable risk factors Genetic markers Lp(a) > 30 mg/dl Alterable risk factors Cholesterol LDL HDL Triglycerides Homocysteine Alterable or unalterable CRP, IL-6

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19 Hyperhomocysteinemia - Independent CVD risk factor - Atherogenic mechanism? Endothelial dysfunction, NO release Cytotoxic effects ROS production (oxidative stress) Potentiation of LDL oxidation Smooth muscle cell proliferation Impaired platelet function

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22 thcy Immunoassays 1. Enzymatic conversion to S-adenosyl-L-homocysteine (SAH) 2. Immunochemical quantification with anti-sah mab

23 thcy Methods Need for standardization (no certified reference material) Inaccuracy (bias) < 10% Imprecision < 5% Enzyme & Immunoassays 5-20% HPLC methods LC-MS/MS < 10% (plasma) 8-12% (dried blood spots)

24 Pre-analytical factors - Overnight fasting preferred. Large, protein-rich meal: 10-15% thcy by after 6-8 h. - Supine vs. sitting position: 10% thcy ( albumin). - EDTA plasma recommended. - Release of Hcy from RBC: immediate centrifugation (< 1h), or keep specimen cooled on ice until centrifugation (< 8h). - Hcy in plasma or serum is stable for 4d. at 21 C. - Storage at -20 C. Freeze-thaw cycles are tolerated.

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26 Determinants of thcy - Homocystinuria (CBS, MTHFR, ) - Age, male sex - Pregnancy ( ) - Renal failure, dialysis - Folate, Vit B12, Vit B6 (dose-response relationship). - Lifestyle factors: smoking, coffee, sedentary lifestyle - Drugs: oral contraceptives/hormonal replacement ( ), corticosteroids, cyclosporine, methotrexate, theophylline, anticonvulsants (carbamazepine, phenytoine)

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29 Hyperhomocysteinemia 1. Moderate: µmol/l Unhealthy lifestyle, poor diet, drugs, renal impairment, preanalytical error 2. Intermediate: µmol/l Folate/B12 deficiency, renal failure 3. Severe: >100 µmol/l Homocystinuria

30 AACC Expert Opinion for thcy - Main indications: homocystinuria, preclinical folate/b12 deficiency, CVD risk stratification. - Single thcy measurement is adequate. - Repeat thcy 3-5 years apart in the elderly (>75 y) (high prevalence of B12 deficiency). - thcy change >25-30% between sampling is significant - Methionine-loading test: not recommended in routine clinical setting

31 thcy for CVD risk assessment AACC Expert Opinion NOT recommended : - General population screening for CVD risk - Blind folate/b12 supplementation without thcy testing

32 thcy for CVD risk assessment Recommended: - Young CVD patients (<40 y), family history: to exclude homocystinuria - CVD patients & subjects at high CVD risk: thcy > 15 µmol/l = high risk group change lifestyle, treatment for causal factors (e.g. vitamin deficiency, renal failure) - ESRD, dialysis patients: thcy > 30 µmol/l = high CVD mortality

33 thcy for CVD risk assessment

34 Laboratory parameters for cardiovascular risk stratification Unalterable risk factors Genetic markers Lp(a) Alterable risk factors Cholesterol LDL HDL Triglycerides Homocysteine Alterable or unalterable CRP, IL-6

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39 IL-6 Factors influencing the measurement of cytokines

40 IL-6 Pre-analytical limitations Cytokine production continues after blood collection by activated immune cells (coagulation, syringe contact) Cytokines can be degraded in the collection tube Cytokines bind to cell receptors during storage Endotoxin-free heparin sampling Rapid centrifugation (within 2h). Until then, keep tube cooled. Analysis within 5 h. Storage at -80 C. Avoid freezing/thawing cycles.

41 BIOASSAYS IL-6 dependent cell lines < 1 pg/ml Low specificity CV ~ % 1-4 d. Biologically active IL-6 IMMUNOASSAYS Automated 1-10 pg/ml High specificity CV ~ 5-10% Short assay time Antigenic presence Cross-reactivity: precursors & degradation products Interference: soluble receptors & protein binding (α2-macroglobulin, IgM)

42 From CRP to hs-crp : Latex-Enhanced Immuno-Turbidity Test + Ab bound to latex Ag from the sample Measurement of turbidity or light scattering

43 Latex-enhanced CRP Ultra Sensitive nephelometry / turbidimetry

44 Within-run CVs for different CRP assays

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46 The quintile concept Risk 5th quintile: th quintile: rd quintile: nd quintile: st quinntile: < hs-crp concentration [mg/l]

47 MONICA (Monitoring Trends and Determinants in Cardiovascular Disease) Augsburg Cohort Study, 1984 to 1992

48 hs-crp for predicting cardiovascular risk 5 Myocardial infarction 4 Ischemic stroke Relative risk 3 2 Venous thrombosis 1,7 1,7 2,6 2,9 1,9 1,9 1 1,0 1,0 1,0 1,1 1,2 1, Quartile of CRP

49 Aspirin decreases coronary risk 5 Relative risk for myocardial infarction ,16 Placebo Aspirin 1 2,07 2,59 1,37 1,39 4,16 1, Quartile of CRP Baseline CRP can predict success of aspirin therapy.

50 hs-crp in patients with unstable angina Cumulative Survival (no death, no admission for MI or unstable angina) hs-crp > 3 mg/l hs-crp < 3 mg/l Months

51 hs-crp and lipid profile for predicting cardiovascular risk 95% confidence interval Lp(a) Homocysteine Total Cholesterol (TC) LDL Chol Apo B TC / HDL Chol hs-crp hs-crp & TC/HDL Chol Relative risk for future cardiovascular events (highest quartile / lowest quartile) Ridker et al.: N Engl J Med 2000; 342:

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53 hs-crp and lipid profile for predicting cardiovascular risk C Relative Risk Rifai N, Ridker PM: Clin Chem 2001; 47: 28-30

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55 New AHA (American Heart Association) & Centers for Disease Control (CDC) Recommendations, 2003 High-risk patients would be missed by just measuring cholesterol levels Major risk factor scoring underestimates the risk of the metabolic syndrome

56 AHA/CDC Recommendations, 2003 hs-crp recommended in 1. moderate risk patients based on multiple risk factor scoring 2. Metabolic syndrome 3. Prognosis of events (death, AMI, restenosis) in patients with stable & acute coronary syndromes (secondary prevention)

57 AHA/CDC Recommendations, 2003 hs-crp NOT recommended in - high CVD risk patients based on multiple risk factor scoring - Management of acute coronary syndromes - Monitoring effects of treatment - General population screening Other inflammatory markers & cytokines should not be measured in addition to hs-crp

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59 hs-crp for predicting cardiovascular risk Don t measure hs-crp after recent infection, inflammation or trauma (CRP is often > 10 mg/l). In this case wait 2 weeks. If, unexplained, hs-crp >10 mg/l persists after repeated testing, examine patient for sources of infection and inflammation The predictive value is greatly improved if 2 measurements are taken 2 weeks apart

60 hs-crp for predicting cardiovascular risk Serum or heparin plasma; no EDTA (-16%). Fasting before sampling is recommended (turbidimetric & nephelometric assays) Take lower value (AACC) or average (AHA/CDC) of the 2 values for risk assessment Categorize patients in relative risk tertiles: < 1 mg/dl : LOW 1-3 mg/dl: average > 3 mg/dl: HIGH (2-fold higher risk than low-risk tertile)

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62 ACC (American College of Cardiology) & AHA Scientific Statement There is no evidence that lowering thcy will necessarily lower CVD risk There is no evidence that lowering CRP (aspirin?) will necessarily lower CVD risk hs-crp is no target of therapeutic intervention only to motivate patients to improve lifestyle or to comply with (drug) therapies