Decreased Naive and Increased Memory CD4⁺ T Cells Are Associated with Subclinical Atherosclerosis: The Multi-Ethnic Study of Atherosclerosis

<div><p>Background</p><p>Adaptive immunity has been implicated in atherosclerosis in animal models and small clinical studies. Whether chronic immune activation is associated with atherosclerosis in otherwise healthy individuals remains underexplored. We hypothesized that activation of adaptive immune responses, as reflected by higher proportions of circulating CD4⁺ memory cells and lower proportions of naive cells, would be associated with subclinical atherosclerosis.</p><p>Methods and Findings</p><p>We examined cross-sectional relationships of circulating CD4⁺ naive and memory T cells with biomarkers of inflammation, serologies, and subclinical atherosclerosis in 912 participants of the Multi-Ethnic Study of Atherosclerosis (MESA). Circulating CD4⁺ naive cells were higher in women than men and decreased with age (all p-values <0.0001). European-Americans had higher levels of naive cells and lower levels of memory cells compared with African-Americans and Hispanic-Americans (all p-values ≤0.0005). Lower naive/higher memory cells were associated with interleukin-6 levels. In multivariate models, cytomegalovirus (CMV) and <i>H. Pylori</i> titers were strongly associated with higher memory and lower naive cells (all p-values <0.05). Higher memory cells were associated with coronary artery calcification (CAC) level in the overall population [β-Coefficient (95% confidence interval (CI))  = 0.20 (0.03, 0.37)]. Memory and naive (inversely) cells were associated with common carotid artery intimal media thickness (CC IMT) in European-Americans [memory: β =  0.02 (0.006, 0.04); naive: β = −0.02 (−0.004, −0.03)].</p><p>Conclusions</p><p>These results demonstrate that the degree of chronic adaptive immune activation is associated with both CAC and CC IMT in otherwise healthy individuals, consistent with the known role of CD4⁺ T cells, and with innate immunity (inflammation), in atherosclerosis. These data are also consistent with the hypothesis that immunosenescence accelerates chronic diseases by putting a greater burden on the innate immune system, and suggest the importance of prospective studies and research into strategies to modulate adaptive immune activation in chronic disease states such as atherosclerosis.</p></div

Regression models for CD4⁺ naive and memory cell subpopulations.

<p>Backward elimination regression was used to develop multivariate models for CD4⁺ T cell subpopulations. Independent variables were divided by their standard deviations (shown in parentheses). Age, gender, race/ethnicity, seasonality, BMI, IL-6, and CMV and <i>H. pylori</i> titers were included as the candidate starting variables. Only significant variables (p<0.05) were retained in the final models. BMI: Body mass index; CMV: Cytomegalovirus; IL-6: Interleukin-6; ns: non-significant.</p

Baseline characteristics of participants with intervening pneumonia in-between CAC measurements and matched controls without the infection.

<p>Baseline characteristics of participants with intervening pneumonia in-between CAC measurements and matched controls without the infection.</p

Trajectories of change (panel A) and absolute magnitude of change (panel B) of CAC scores in participants with and without intervening hospitalization with pneumonia stratified by their baseline risk of arteriosclerotic cardiovascular disease (ASCVD).

<p>Trajectories of change (panel A) and absolute magnitude of change (panel B) of CAC scores in participants with and without intervening hospitalization with pneumonia stratified by their baseline risk of arteriosclerotic cardiovascular disease (ASCVD).</p

Selection of pneumonia cases and controls.

<p>Selection of pneumonia cases preceded the selection of controls. MESA denotes the Multi-Ethnic Study of Atherosclerosis. CAC denoted coronary artery calcium. CRP denotes serum C-reactive protein. ASCVD denotes atherosclerotic cardiovascular disease. ^<b>1</b> Matching was by age (±2.5 years), sex, CRP values (±0.5 mg/dL), and 10-year risk of ASCVD events (±2.5% as per the ACC/AHA ASCVD risk-equation). When there were multiple potential matches, we followed a greedy algorithm to select the closest match.</p

Characteristics of the MESA sample population being studied.

<p>Data are from MESA exam 4 (2005–2007) unless otherwise noted. *: Data are from MESA baseline (exam 1; 2000–2002). **: Includes all participants with incident cardiovascular events (myocardial infarction, resuscitated cardiac arrest, definite or probable angina, and stroke) from baseline through the start of exam 4. AU: Agatston units; BMI: Body mass index; CAC: Coronary artery calcification; CMV: Cytomegalovirus; CRP: C-reactive protein; CVD: Cardiovascular disease; HSV: Herpes simplex virus; IL-6: Interleukin-6; IMT: Intimal media thickness.</p

Final regression model for coronary artery calcification.

<p>Backward elimination regression was used to develop multivariate models for coronary artery calcification (CAC) level. CAC was analyzed using the ln-agatston score in individuals with a score >0. Independent variables were divided by their standard deviations (shown in parentheses). The candidate starting variables were: age, gender, race/ethnicity, IL-6, BMI, systolic BP, use of BP lowering medication, smoking status, total-cholesterol, HDL-cholesterol, use of lipid lowering medication, type 2 diabetes status, CMV and <i>H. pylori</i> titers, and CD4⁺ memory cell proportions or, in separate analyses, CD4⁺ naive cell proportions. Only significant variables (p<0.05) were retained in the final model to obtain the model's R². ns: non-significant.</p

Typical flow cytometric data for lymphocyte, CD4⁺ memory, and CD4⁺ naive cell populations.

<p>At least 30,000 lymphocytes were evaluated and were gated based on their forward (FCS; X-axis) and side (SCS; Y-axis) scatter (Panels A&B). Memory and naive cell subpopulations were gated by positive surface staining for CD4 (Y-axis, panels C–F); memory cells were gated by positive surface staining for CD45RO (X-axis, panels C&D); naive cells were gated by positive surface staining for CD45RA (X-axis, panels E&F). Typical data from respective isotype controls (Panels A, C, and E) and fluorescently labeled samples (Panels B, D, and F) are shown.</p

Study sample.

<p>MESA = Multi- Ethnic Study of Atherosclerosis; MRI = magnetic resonance imaging; RV = right ventricle.</p

Multivariable nonparametric smoothed relationship between pericardial fat volume in cm3 and right ventricular (RV) parameters with adjustment for age, gender, race, height, weight, study site, education level, exercise habits, and smoking status including pack years smoked (black line).

<p>Gray lines represent 95% Confidence intervals. The relationship between pericardial fat and (A) RV mass, (B) RV end diastolic volume (EDV), (C) RV end systolic volume, (D) RV stroke volume (SV), (E) ejection fraction (EF).</p