**5. Association of Lp-PLA2 with cardiovascular disease risk**

Lp-PLA2 is an emerging inflammatory biomarker, characterized by high vascular specificity and low biovariability. High Lp-PLA2 levels are indicative of rupture-prone plaque [22]. Numerous epidemiological and clinical studies examined association between Lp-PLA2 concentration/enzyme activity and cardiovascular disease risk in apparently healthy individ‐ uals*,* in subjects with stable cardiovascular disease, acute coronary syndrome (ACS), heart failure (HF), stroke, transient ischemic attack (TIA) support, trying to answer in which patients would determination of Lp-PLA2 be the most valuable. There are novel data about the prognostic significance of Lp-PLA2 as a predictor of short-term or long-term outcome in patients with cardiovascular disease. To evaluate this, systematic literature review concerning the association of Lp-PLA2 with cardiovascular disease risk and prognostic implications was done. The studies on this issue were extracted from relevant electronic databases (Medline (http://www.ncbi.nlm.nih. gov/pubmed/), Embase (http://www. embase.com/), Google Scholar (http://scholar. google.com/), Yahoo (http:// www. yahoo. com/), Kobson (http:// www.kobson.nb.rs/), ClinicalTrails.gov.) and the obtained results were included in the text.

In 2008, the panel's recommendation [23] incorporated Lp-PLA2 testing as an adjunct to traditional risk factors in assessing future cardiovascular risks. It endorses Lp-PLA2 testing in moderate-risk persons determined as having simply two risk factors. An Lp-PLA2 >200 ng/mL warrants reclassification of the moderate-risk patient as high cardiovascular risk and should prompt reduction of the LDL-C target from 130 mg/dL to 100 mg/dL. The panel also recom‐ mends Lp-PLA2 testing for patients with coronary artery disease (CAD) or CAD risk equiva‐ lents (diabetes, ischemic stroke, etc.) be considered at very high risk when Lp-PLA2 is elevated, warranting reduction in the LDL-C target from 100 mg/dL to 70 mg/dL. Today, Lp-PLA2 measurement for cardiovascular disease risk stratification of patients is included in four international clinical guidelines: 2012 European Guidelines on CVD Prevention in Clinical Practice European Society of Cardiology – Lp-PLA2 may be measured as a part of a refined risk assessment in patients at high risk of a recurrent acute atherothrombotic event. Class IIb; 2010 ACCF/AHA Guideline for Assessment of Cardiovascular Risk in Asymptomatic Adults – Lp-PLA2 testing may be considered in intermediate-risk asymptomatic adults; 2011 AHA/ASA Guidelines for the Primary Prevention of Stroke Measurement of inflammatory markers such as high sensitive C-reactive protein (hs-CRP) or Lp-PLA2 in patients without CVD may be considered to identify patients who may be at increased risk of stroke; 2012 AACE Guidelines for Management of Dyslipidemia and Prevention of Atherosclerosis Test for LpPLA2, which in some studies has demonstrated more specificity than highly sensitive CRP, when it is necessary to further stratify a patient's CVD risk [40-43].

These finding has been observed in previous studies [31,32,33,34]. Lower Lp-PLA2 levels in women could be explained by estrogen-mediated down-regulation of Lp-PLA2 expression, due to lower concentrations of LDL cholesterol in women or estrogen-related decrease in platelet activating factor acetyl hydrolase activity [35,36]. Estrogen-replacement therapy can significantly reduce Lp-PLA2 activity in healthy postmenopausal women [37], while admin‐ istration of steroids with progesterone-like activity increases Lp-PLA2 activity [38]. Smoking

Lp-PLA2 is an emerging inflammatory biomarker, characterized by high vascular specificity and low biovariability. High Lp-PLA2 levels are indicative of rupture-prone plaque [22]. Numerous epidemiological and clinical studies examined association between Lp-PLA2 concentration/enzyme activity and cardiovascular disease risk in apparently healthy individ‐ uals*,* in subjects with stable cardiovascular disease, acute coronary syndrome (ACS), heart failure (HF), stroke, transient ischemic attack (TIA) support, trying to answer in which patients would determination of Lp-PLA2 be the most valuable. There are novel data about the prognostic significance of Lp-PLA2 as a predictor of short-term or long-term outcome in patients with cardiovascular disease. To evaluate this, systematic literature review concerning the association of Lp-PLA2 with cardiovascular disease risk and prognostic implications was done. The studies on this issue were extracted from relevant electronic databases (Medline (http://www.ncbi.nlm.nih. gov/pubmed/), Embase (http://www. embase.com/), Google Scholar (http://scholar. google.com/), Yahoo (http:// www. yahoo. com/), Kobson (http:// www.kobson.nb.rs/), ClinicalTrails.gov.) and the obtained results were included in the text. In 2008, the panel's recommendation [23] incorporated Lp-PLA2 testing as an adjunct to traditional risk factors in assessing future cardiovascular risks. It endorses Lp-PLA2 testing in moderate-risk persons determined as having simply two risk factors. An Lp-PLA2 >200 ng/mL warrants reclassification of the moderate-risk patient as high cardiovascular risk and should prompt reduction of the LDL-C target from 130 mg/dL to 100 mg/dL. The panel also recom‐ mends Lp-PLA2 testing for patients with coronary artery disease (CAD) or CAD risk equiva‐ lents (diabetes, ischemic stroke, etc.) be considered at very high risk when Lp-PLA2 is elevated, warranting reduction in the LDL-C target from 100 mg/dL to 70 mg/dL. Today, Lp-PLA2 measurement for cardiovascular disease risk stratification of patients is included in four international clinical guidelines: 2012 European Guidelines on CVD Prevention in Clinical Practice European Society of Cardiology – Lp-PLA2 may be measured as a part of a refined risk assessment in patients at high risk of a recurrent acute atherothrombotic event. Class IIb; 2010 ACCF/AHA Guideline for Assessment of Cardiovascular Risk in Asymptomatic Adults – Lp-PLA2 testing may be considered in intermediate-risk asymptomatic adults; 2011 AHA/ASA Guidelines for the Primary Prevention of Stroke Measurement of inflammatory markers such as high sensitive C-reactive protein (hs-CRP) or Lp-PLA2 in patients without CVD may be considered to identify patients who may be at increased risk of stroke; 2012 AACE Guidelines for Management of Dyslipidemia and Prevention of Atherosclerosis Test for Lp-

may increase the carrier (LDL) and the substrate (oxidized LDL) for Lp-PLA2 [39].

**5. Association of Lp-PLA2 with cardiovascular disease risk**

116 Lipoproteins - From Bench to Bedside

Some studies suggested that Lp-PLA2 predicts risk complementary to hs-CRP, although when compared with hs-CRP, Lp-PLA2 in some studies is suggested to be a more promising marker of risk prediction. In multiple previous studies, correlation analysis revealed no association of Lp-PLA2 with CRP [28,31,44-47]. It can be expected because CRP is an acute-phase reactant and its elevation can be caused by a wide range of inflammatory conditions. CRP shows intraindividual variability of ∼40%. On the other hand, Lp-PLA2 is not affected by systemic inflammation; it is a specific marker of vascular inflammation. Also, it shows significantly lower biologic variability than CRP, and higher stability in states of myocardial ischemia [48].

Lp-PLA2 is suggested to be a more promising marker of risk prediction than CRP [49]. Winkler et al. [50] showed that increased Lp-PLA2 levels in moderate-risk patients with hs-CRP<3 mg/ L doubled the risk for cardiac death. The WOSCOPS study [44] found that Lp-PLA2 was significantly associated with cardiovascular risk, compared with hs-CRP. Stankovic et al.'s [32] results confirmed that Lp-PLA2 have better risk prediction than CRP.

Although, the majority of published studies showed a significant relationship between Lp-PLA2 levels and cardiovascular events, there are several important differences across ethnic groups; for example, African-Americans and Caucasians with respect to Lp-PLA2. Activity of Lp-PLA2 was higher among African-Americans with CAD. The difference in Lp-PLA2 activity levels between CAD and non-CAD patients was higher among African-Americans. Also, the Lp-PLA2 index was independently associated with the extent of CAD among African-Americans [51-53].

From the WOSCOPS publication (West of Scotland Coronary Prevention Study, WOSCOPS) [44], which revealed a positive association between elevated circulating concentrations of Lp-PLA2 and the risk of coronary heart disease (CHD), the interest in Lp-PLA2 as a biomarker for cardiovascular disease rapidly increased. The vast body of evidence derived from prospective epidemiologic studies, two meta-analyses (79,036 participants in 32 prospective studies, and 52,995 subjects participated in 33 studies) and review revealed the positive association of elevated Lp-PLA2 with cardiovascular risk [24,54-56]. Furthermore, Lp-PLA2 has been confirmed to predict the presence of CAD, even among patients undergoing coronary angiography. Uniquely, Lp-PLA2 predicted the risk of CAD death, but not all cause death [57].

LpPLA2 has been considered as a prognostic marker in patients with CAD. Li et al. [58] identified prognostic value of Lp-PLA2 baseline measurement for major adverse cardiac event (MACE) (cardiovascular death, nonfatal myocardial infarction, and target vessel revasculari‐ zation) in ACS patients during follow-up for a median of 6 months. Mockel et al. [59] demon‐ strated that Lp-PLA2 levels in the first 7 hours after onset of symptoms may be an independent predictor of MACE within 42 days in ACS patients. PROVEIT-TIMI 22 study [25] has shown that Lp-PLA2 in patients randomized to atorvastatin or pravastatin measured 30 days after ACS are associated with an increased risk of cardiovascular events (death, myocardial infarction, unstable angina, revascularization, or stroke) over 24 months of follow-up. Gerber et al. MACE [48] found that Lp-PLA2 level measured early after myocardial infarction is strongly and independently associated with 1-year mortality. Stankovic et al. [32] demon‐ strated that the Lp-PLA2 may have short-term predictive value in pure STEMI patients treated by primary percutaneous coronary intervention (PCI). They concluded that pre-interventional plasma Lp-PLA2 level is an independent predictor of 30-day MACE in patients with first anterior STEMI treated by primary PCI, and suggested that Lp-PLA2 level could help in very early risk stratification of STEMI patients treated by PCI.

Previously published studies examined and suggested the association between Lp-PLA2 and heart failure (HF) incidence in a population-based cohort of healthy individuals, and in people older than 65 years [60]. Baseline Lp-PLA2 levels are associated with a high risk of developing heart failure in 3,991 adults older than 65 years, independent of coronary risk factors [61]. Lp-PLA2 activity is significantly associated with congestive heart failure in 5,531 persons older than 65 years [62]. Gerber et al. [63] evaluated the association of Lp-PLA2 with mortality in subjects with diagnosed HF. Lp-PLA2 was strongly and independently associated with mortality in patients under 80 years of age. Moldoveanu et al.'s study [64] in 208 patients with HF found significantly increased Lp-PLA2 activity in HF patients with preserved ejection fraction (EF) than in HF with reduced EF. The literature data about the association of Lp-PLA2 and heart failure on admission in patients with acute myocardial infarction are missing. Stankovic et al. [32] suggested that patients with the first anterior STEMI who had higher levels of Lp-PLA2 had a worse prognosis, but not with a greater probability of developing HF. In Raichlin et al.'s [65] study in heart transplant patients, Lp-PLA2 correlated with the progression of cardiac allograft vasculopathy and increased risk of cardiovascular events/death suggesting that it could be therapeutic target in heart transplant patients.

In 2005, the US Food and Drug Administration (FDA) approved Lp-PLA2 blood test for assessing patients at risk for ischemic stroke. The Rotterdam Study was the first populationbased study that determined the impact of elevated Lp-PLA2 on stroke. It identified that Lp-PLA2 activity was an independent predictor of ischemic stroke in the middle-aged healthy men and women population [31]**.** In the ARIC study, healthy middle-aged adults with increased levels of both Lp-PLA2 and hs-CRP had an 11-fold higher incidence of stroke than individuals with low Lp-PLA2 and hs-CRP levels [45]. This association between Lp-PLA2 mass/activity and first ischemic stroke was confirmed in the Malmo Diet and Cancer Study [66], Bruneck Study [67], and Cardiovascular Health Study [28]. The Tsekepis et al. study [68] showed that Lp-PLA2 correlated with the intima-media thickness in patients with beta-thalassemia, suggesting that Lp-PLA2 may be implicated in premature carotid atherosclerosis.

Although some reports inconclusively found this positive association of Lp-PLA2 and firstever and recurrent stroke, little is known about its influence on stroke outcome. Elkind et al. [69] measured Lp-PLA2 mass and activity in relation to outcome in first ischemic stroke patients, determined as recurrent stroke, recurrence of vascular events, and mortality. Lp-PLA2 was a good predictor of recurrent stroke risk. Delgado et al. [70] investigated the temporal profile of Lp-PLA2 mass and activity within the first 24 hours after stroke and found significant changes in Lp-PLA2 concentrations early after stroke onset. Patients with higher Lp-PLA2 mass were more likely to be resistant to intravenous t-PA administration with very low early recanalization rates.

Lp-PLA2, an inflammatory biomarker, has been described as able to predict risk of first-ever or recurrent stroke and myocardial infarction [71]. Moreover, Lp-PLA2 may also have a role in the pathophysiology of cerebrovascular disease, particularly in strokes of atherosclerotic etiology, since its expression is enhanced in atherosclerotic carotid lesions together with markers of oxidative damage, inflammation, and instability [72].

Sarlon-Bartoli et al. [73] reported that Lp-PLA2 mass is increased in patients with high-grade carotid stenosis and unstable plaque and suggest that Lp-PLA2 could be an important bio‐ marker for classifying carotid plaque as vulnerable and predict neurological risk of a carotid stenosis in asymptomatic subjects. Although these findings have the potential to improve cerebrovascular disease stratification, correlation with ultrasonic or MRI markers of plaque instability or the presence of infarction on brain imaging must be performed [74].

Shoamanesh et al. [75] investigated the association between circulating biomarkers of inflam‐ mation including Lp-PLA2 and MRI markers of cerebral small vessel disease in 1,763 strokefree Framingham offspring. They observed higher levels of lipoprotein-associated phospholipase A2 mass in patients with greater white matter hyperintensity volumes and silent cerebral infarcts. These results could improve stroke risk prognosis.

The accumulating results of numerous studies demonstrate a significant positive association between Lp-PLA2 levels and incident cardiovascular disease and heart failure. A dissociation could be noted between mass and activity in terms of risk prediction. Several of the epidemi‐ ology studies have measured only Lp-PLA2 mass (26,30,33,44,47,57,63,65,69), whereas others measured only enzyme activity (31,50,62,64,67) and a few have measured both in the same study population (25,27,28, 76). Recently published meta-analysis assessed the 32 prospective studies of Lp-PLA2 and cardiovascular outcomes. For each standard deviation Lp-PLA2 increase, the relative risks for the primary endpoint of coronary heart disease were 1.10 (1.05– 1.16) and 1.11 (1.07–1.16) for Lp-PLA2 activity and mass, respectively. The relative adjusted risks for ischemic stroke were 1.08 (0.97–1.20) and 1.14 (1.02–1.27); vascular mortality 1.16 (1.09–1.24) and 1.13 (1.05–1.22); and nonvascular mortality 1.10 (1.04–1.17); and 1.10 (1.03–1.18) for Lp-PLA2 activity and mass, respectively. The final decision whether to measure the Lp-PLA2 mass or its enzyme activity may help transition of this biomarker from research to routine clinical practice.
