Cytomegalovirus infection is associated with an increase in aortic stiffness in older men which may be mediated in part by CD4 memory T-cells

Abstract

Human Cytomegalovirus (CMV) infection is associated with atherosclerosis, higher cardiovascular disease (CVD) risk, and an increase in memory T-cells (T_mem). T-cells have also been implicated in CVD, independently of CMV infection. To better understand the CMV-associated CVD risk, we examined the association between CMV (IgG) serostatus and central aortic (carotid-to-femoral) pulse wave velocity (cfPWV), an early, independent predictor of CVD. We also investigated if such an association might be reflected by the distribution of T_mem and/or other T-cell subsets. Methods: Healthy older volunteers (60-93 years) underwent routine clinical and laboratory evaluation, including assessment of cfPWV in eligible participants. Flow-cytometry was used to assess proportions of memory T-cells, CD28^null T-cells, and CMV-specific T-cells. The following associations were examined; CMV serostatus/cfPWV, CMV serostatus/proportion of T_mem, proportion of T_mem/cfPWV, CD28^null T-cells/cfPWV, and CMV-specific T-cells/cfPWV. Linear regression models were used to adjust for age, sex, socioeconomic status, smoking, waist-to-hip ratio, cholesterol, and blood pressure as required. Results: Statistically significant positive associations were found (P-values for the fully adjusted models are given); CMV serostatus/cfPWV in men (P ≤ 0.01) but not in women, CMV serostatus/proportions of CD4 T_mem in men (P ≤ 0.05) but not in women; proportions of CD4 T_mem/cfPWV among CMV seropositive (CMV+) people (P ≤ 0.05) but not CMV seronegative (CMV-) people. Conclusion: CMV infection increases the CVD risk of older men by increasing cfPWV. This may be mediated in part by increased proportions of CD4 T_mem, higher numbers of which are found in CMV+ older people and more so among men than women. Given the high prevalence of CMV worldwide, our findings point to a significant global health issue. Novel strategies to mitigate the increased CVD risk associated with CMV may be required.

Keywords: Memory T-cells; cardiovascular risk; central aortic stiffness; human Cytomegalovirus (CMV); pulse wave velocity.

Figure 1

Differential distribution of canonical memory T-cell subsets in different sub-groups defined by CMV serostatus and sex. A. Box plots show the distribution of CD4 T_NA, T_CM, T_EM, and T_EMRA subsets in CMV- and CMV+ individuals among the entire cohort (top), men (middle) and women (bottom). B. Box plots show the distribution of the corresponding CD8 T cell subsets in each group. Box plots show median and interquartile range, outlier limits (whiskers, LQ-1.5*IQR, UQ+1.5*IQR), outliers (o) and extreme values (_*). Significant differences are indicated by P-values.

Figure 2

The percentage of antigen-experienced CD4 T_mem cells is higher in CMV+ than CMV- individuals among men but not women. A. Box plots show the proportions of CD4 and CD8 T_mem in the entire cohort (top), men (middle), and women (bottom). B. Box plots show the proportions of CD4 and CD8 T_mem in the entire cohort (top), CMV- individuals (middle), and CMV+ individuals (bottom). Box plots show median and interquartile range, outlier limits (whiskers, LQ-1.5*IQR, UQ+1.5*IQR), outliers (o) and extreme values (_*). Significant differences are indicated by P-values.

Figure 3

The percentage of antigen-experienced CD4 T_mem-cells correlates positively with cfPWV in CMV+ but correlates negatively in CMV- people. Scatterplots show the relationships between the percentage of antigen-experienced CD4 T_mem-cells (A), or CD8 T-cells (B) and cfPWV. The entire cohort is shown on the left, CMV+ individuals in the middle, and CMV- individuals on the right. cfPWV was Log10-transformed to improve visualization. Regression lines denote the results of regression on T_mem. The Pearson correlation coefficient (R) is shown with each regression line.

Figure 4

The proportions of CD4 and CD8 CD28^null T-cells show a moderate correlation with T_mem among CMV+ people but do not correlate with cfPWV. CD28^null T-cells were measured by flow-cytometry (gated as CD28^null/CD27- T-cells). A. Correlations between CD28^null CD4 (top) and CD8 (bottom) T-cells and T_mem are shown separately for CMV+ and CMV- people. B. Correlations between CD28^null CD4 (top) and CD8 (bottom) T-cells and cfPWV. Regression lines denote the results of regression on T_mem or CD28^null T-cells. The Pearson correlation coefficient (R) is shown with each regression line.

Figure 5

The frequencies of CMV-specific CD4 and CD8 T-cells do not correlate with cfPWV. Responses were measured by intracellular cytokine staining following stimulation of fresh PBMC with 16 CMV protein-spanning peptide pools arranged in 14 stimulation pools. For each protein the percentage of cells expressing each activation marker (IL-2, TNF, or IFN-γ) or at least one of them was determined. The summated responses across all 14 CMV-peptide stimulation pools are plotted against cfPWV. Response size was determined based on the presence of at least one activation marker (A) or (the usually dominant) IFN-γ only (B).