Endogenous Bioactive Peptides as Potential Biomarkers for Atherosclerotic Coronary Heart Disease

Cardiovascular disease is the leading cause of death worldwide, with high medical costs and rates of disability. It is therefore important to evaluate the use of cardiovascular biomarkers in the early diagnosis of coronary artery disease (CAD). We have screened a variety of recently identified bioactive peptides candidates in anticipation that they would allow detection of atherosclerotic CAD. Especially, we have focused on novel anti-atherogenic peptides as indicators and negative risk factors for CAD. In vitro, in vivo and clinical studies indicated that human adiponectin, heregulin-β1, glucagon-like peptide-1 (GLP-1), and salusin-α, peptides of 244, 71, 30, and 28 amino acids, respectively, attenuate the development and progression of atherosclerotic lesions by suppressing macrophage foam cell formation via down-regulation of acyl-coenzyme A: cholesterol acyltransferase-1. Circulating levels of these peptides in the blood are significantly decreased in patients with CAD compared to patients without CAD. Receiver operating characteristic analyses showed that salusin-α is a more useful biomarker, with better sensitivity and specificity, compared with the others for detecting CAD. Therefore, salusin-α, heregulin-β1, adiponectin, and/or GLP-1, alone or in various combinations, may be useful as biomarkers for atherosclerotic CAD.

This review focuses on the protective roles of adiponectin, heregulin-β 1 , GLP-1, and salusin-α in atherosclerotic cardiovascular diseases and their emerging roles for biomarkers and therapeutic targets for CAD.

Presence in Coronary Artery Atherosclerosis and Circulating Blood
Immunohistochemical analyses of human coronary arteries from patients with CAD using anti-heregulin-β 1 or anti-salusin-α antibodies show faint staining in advanced coronary atherosclerotic lesions, suggesting decreased expression at their protein levels [16,39]. The expression of adiponectin at mRNA levels is also significantly lower in epicardial adipose tissues in CAD [46]. These findings strongly suggest that a decline in anti-atherogenic peptides may be associated with the progression of atherosclerotic lesions in human coronary arteries.
Circulating markers are more convenient for diagnosis of CAD. As specific antibodies against these peptides have been developed, their concentrations in blood samples could be quantified using radioimmunoassay and enzyme-linked immunosorbent assay (ELISA). Serum levels of total and high-molecular weight adiponectin and plasma heregulin-β 1 levels were measured by ELISA, and their accuracy and precision were comparable among several studies [16,47−53]. The accuracy and precision of serum salusin-α and plasma GLP-1 levels measured by radioimmunoassay and ELISA were identical among each several studies [39,54−59].
To assess essential levels of peptide hormones, the factors that influence peptide production must be taken into consideration. In general, these include food intake, smoking, gender, and the presence of diabetes, hypertension or obesity. Serum salusin-α and plasma heregulin-β 1 levels have been demonstrated to be unaffected by a number of physiological stimuli [8,9]. Since adiponectin is known to show sexual dimorphism with higher levels in women than men, von Eynatten et al. [48] have studied serum adiponectin levels in the limited male subjects. Because GLP-1 is temporary increased after food intake, plasma GLP-1 levels have been measured in the fasting state and/or after 75-g oral glucose tolerance test [60−62]. Similar to adiponectin measurements, we determined serum salusin-α and plasma heregulin-β 1 levels in the fasting state [16,39].

Biomarkers for CAD
Matsubara et al. [60] reported that fasting plasma GLP-1 levels are significantly lower in CAD patients than in non-CAD patients  [60]. However, Nathanson et al. [62] reported that impaired GLP-1 secretion after oral glucose load does not predict CAD in the presence of diabetes. El-Menyar et al. [63] reported that serum levels of high-molecular weight adiponectin are significantly decreased in CAD patients compared with those in non-CAD patients (1.9 ± 0.2 versus 3.1 ± 0.3 μg/mL, P = 0.003). Serum levels of high-molecular weight adiponectin were shown to be inversely correlated with angiographic severity of coronary artery lesions in patients with CAD [48]. Circulating levels of heregulin-β or salusin-α are also significantly decreased ( Figure 1) and inversely correlated with angiographic severity of coronary artery lesions in patients with CAD [16,39,53].
Receiver operating characteristic (ROC) curves were plotted and the area under the curve (AUC) was analyzed to compare the predictive power of high-molecular weight adiponectin, heregulin-β 1 , and salusin-α. The optimal cut-off values of these peptides for detecting CAD were set at the point showing a higher true-positive rate (sensitivity) with a low false-positive rate (1-specifity) on the respective ROC curve. von Eynatten et al. [48] reported that the AUC value for high-molecular weight adiponectin was 0.673. The ROC curve was slightly improved by using the high-molecular weight adiponectin/total adiponectin ratio, and its AUC value became 0.718. On the basis of our previous studies [16,39], we also performed ROC analyses and calculated the AUC values in the present time.
The AUC values of heregulin-β 1 and salusin-α are 0.706 and 0.916, respectively (Figure 2). In addition, the cut-off levels were 2.4 ng/mL for heregulin-β 1 with sensitivity and specificity of 76.6% and 58.1%, respectively, and that for salusin-α was 8.5 pM with sensitivity and specificity of 81.5% and 92.7%, respectively. In comparisons among these peptides, the AUC values for high-molecular weight adiponectin and the high-molecular weight adiponectin/total adiponectin ratio were similar to that of heregulin-β 1 but inferior to that of salusin-α Therefore, serum salusin-α level has higher diagnostic value in detecting CAD compared with the other three peptides.   [52,66]. Thus, further studies are required to determine their utility as biomarkers in predicting atherosclerotic CAD in the presence or absence of severe heart failure. Recently, several studies have shown that administration of heregulin-β 1 or GLP-1 improves cardiac dysfunction in patients with heart failure [67−71]. Intracoronary administration of adiponectin led to a reduction in myocardial infarct size and improvement of left ventricular function after ischemia/reperfusion injury in pigs [72]. However, effects of adiponectin and salusin-α on cardiac function have not yet been reported in humans. These findings provide insights into the potential use of heregulin-β 1 or GLP-1 as an extended therapeutic window for combating refractory heart failure.

Conclusions
Adiponectin, heregulin-β 1 , GLP-1, and salusin-α could contribute to the early diagnosis and therapeutic efficacy of atherosclerosis. Decreased levels of adiponectin, heregulin-β 1 , GLP-1 and salusin-α in circulating blood and/or cardiovascular tissues are closely linked with human atherosclerosis. Thus, adiponectin, heregulin-β 1 , GLP-1, and/or salusin-α, alone or in various combinations are candidate biomarkers for predicting CAD, which may be useful for the earlier detection of atherosclerotic cardiovascular diseases.