Cytomegalovirus-Productive Infection Is Associated With Acute Coronary Syndrome

Abstract

Background: Although an association between human herpesvirus (HHV) infection and atherosclerosis has been suggested, the data supporting such an association are controversial and, in most cases, are based on serological evidence or on the presence of cell-associated HHV DNA, which do not report about actual viral replication. We quantified the DNA of all 8 types of HHVs in plasma, in which their presence is evidence of viral replication.

Methods and results: Using quantitative real-time polymerase chain reaction, we evaluated the presence of HHV DNA in blood samples obtained at the time of hospitalization from 71 patients with acute coronary syndrome, 26 patients with stable coronary artery disease, and 53 healthy volunteers and in atherosclerotic plaques of 22 patients with peripheral artery disease who underwent endarterectomy. HHV-5 (cytomegalovirus [CMV]) was the only HHV with a level that was higher in acute coronary syndrome patients than in the control group and that correlated with the level of high-sensitivity C-reactive protein. The numbers of effector memory T cells positively correlated with the numbers of CMV genome copies in carotid arteries plaques, whereas the numbers of central memory T cells negatively correlated with CMV copy numbers.

Conclusions: Of all HHV levels, only CMV was higher in patients with stable coronary artery disease and acute coronary syndrome than in the healthy group, and its load correlated with the level of high-sensitivity C-reactive protein. The level of CMV in atherosclerotic plaques correlated with the state of immunoactivation of lymphocytes in plaques, suggesting that the reactivation of CMV may contribute to the immune activation associated with the progression of atherosclerosis.

Keywords: acute coronary syndrome; atherosclerosis; human herpesvirus; immune system; myocardial infarction; polymerase chain reaction; virus.

Figure 1

A, Standard's amplification for HHV‐1, HHV‐3, HHV‐5, HHV‐7, and ERV‐3 multiplex combination. B, Standard's curve for the first multiplex combination. FAM, HEX, Cy5, Quasar 705, and Cal Red 610 are fluorescent dyes.

Figure 2

A, Standard's amplification for HHV‐2, HHV‐4, HHV‐6, HHV‐8, and ERV‐3 multiplex combination. B, Standard's curve for the second multiplex combination. FAM, HEX, Cy5, Quasar 705, and Cal Red 610 are fluorescent dyes.

Figure 3

A flowchart of the experiment for HHV detection in plasma of healthy volunteers, in patients with CAD, and in atherosclerotic plaques from carotid arteries of patients undergoing CEA. Analysis of HHV DNA correlation with T‐cell subsets in atherosclerotic plaques. ACS indicates acute coronary syndrome; CAD, coronary artery disease; CEA, carotid endarterectomy; HHV, human herpesvirus; PCR, polymerase chain reaction; SCAD, stable coronary artery disease.

Figure 4

Frequencies (as percentages) of HHVs detected in blood plasma samples in groups of healthy volunteers (green), SCAD patients (blue), and ACS patients (red). Positivity was defined as HHV DNA at amounts >100 copies per microliter of plasma. *Differences are statistically significant at P<0.05. ACS indicates acute coronary syndrome; HHV, human herpesvirus; SCAD, stable coronary artery disease.

Figure 5

Receiver operating characteristic (ROC) curve for the cytomegalovirus (CMV) DNA load in patients with acute coronary syndrome compared with patients with stable coronary artery disease and healthy controls. The area under the ROC curve was 0.662 (95% CI 0.56–0.77, P=0.004). With the threshold of 142 copies of CMV DNA per microliter of plasma, specificity was 64.9% and sensitivity was 64.6%.

Figure 6

Receiver operating characteristic (ROC) curve for the cytomegalovirus (CMV) DNA load in all patients with coronary artery disease compared with healthy controls. The area under the ROC curve was 0.646 (95% CI 0.53–0.76, P=0.013). With the threshold of 114 copies of CMV DNA per microliter of plasma, specificity was 64.1% and sensitivity was 65.2%.

Figure 7

Odds ratios for binomial coronary artery disease risk factors (with age ≥55 years for men and ≥65 years for women), high‐sensitivity CRP level (≥2 mg/L), and CMV presence in the multiple logistic regression model for ACS patients and healthy volunteers. *Differences are statistically significant at P<0.05. ACS indicates acute coronary syndrome; CMV, cytomegalovirus; CRP, C‐reactive protein; OR, odds ratio.

Figure 8

Frequencies (as percentages) of HHVs detected in carotid artery plaque samples. Positivity was defined as HHV DNA at amounts >100 copies per 10⁶ cells. HHV indicates human herpesvirus.

Figure 9

Correlation coefficient with 95% CI according to Spearman rank test between CMV DNA loads and fractions of intermediately differentiated CD4⁺ Tem cells in atherosclerotic plaques. Correlation is statistically significant at P<0.05. CMV indicates cytomegalovirus; ID, intermediately differentiated; Tem, effector memory T cells.

Figure 10

Correlation coefficient with 95% CI according to Spearman rank test between CMV DNA loads and fractions of intermediately differentiated CD8⁺ Tem cells in atherosclerotic plaques. Correlation is statistically significant at P<0.05. CMV indicates cytomegalovirus; ID, intermediately differentiated; Tem, effector memory T cells.

Figure 11

Correlation coefficient with 95% CI according to Spearman rank test between CMV DNA load and fraction of CD4⁺ Tcm cells in atherosclerotic plaques. Correlation is statistically significant at P<0.05. CMV indicates cytomegalovirus; Tcm, central memory T cells.

Figure 12

Correlation coefficient with 95% CI according to Spearman rank test between CMV DNA load and fraction of CD8⁺ Tcm cells in atherosclerotic plaques. Correlation is statistically significant at P<0.05. CMV indicates cytomegalovirus; Tcm, central memory T cells.