Human Cytomegalovirus Infection and Cardiovascular Disease: Current Perspectives

Abstract

Infections with human cytomegalovirus (HCMV) are often asymptomatic in healthy adults but can be severe in people with a compromised immune system. While several studies have demonstrated associations between cardiovascular disease in older adults and HCMV seropositivity, the underlying mechanisms are unclear. We review evidence published within the last 5 years establishing how HCMV can contribute directly and indirectly to the development and progression of atherosclerotic plaques. We also discuss associations between HCMV infection and cardiovascular outcomes in populations with a high or very high burden of HCMV, including patients with renal or autoimmune disease, transplant recipients, and people living with HIV.

Introduction

Human cytomegalovirus (HCMV) is a beta-herpesvirus that commonly establishes asymptomatic latent infections with intermittent reactivations throughout life (Forte et al, 2020). HCMV can generate diverse clinical syndromes in those with a weakened immune system (e.g. transplant recipients, people living with HIV [PLWH], and neonates). Epidemiological studies have associated persistent HCMV infection with age-related diseases, such as cardiovascular disease (CVD) in individuals with no history of acute HCMV disease.

Atherosclerosis is a common cause of CVD and is characterized by the development of plaques in the arterial walls (Xu et al, 2019). Atherosclerosis begins with the activation of endothelial cells triggering the transmigration of inflammatory monocytes from the circulation into the intima, where they differentiate into macrophages and form foam cells after uptake of oxidized low-density lipoproteins (oxLDL) (reviewed in Lee et al, 2020). These steps are mediated primarily by intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) expressed on circulating leukocytes and on the vascular endothelium. Plaque formation narrows the arteries, leading to coronary artery disease (CAD) and stroke.

Atherosclerosis is now considered a chronic inflammatory disease and several pathogens, including HCMV, have been proposed as triggers (Li et al, 2020a). Smooth muscle cells (SMC), endothelial cells, and monocytes/macrophages support HCMV replication (Forte et al, 2020) and contribute to atherogenesis (Li et al, 2020a). Hence, we have proposed that CVD consequent to atherosclerosis should be defined as a clinical “footprint” of HCMV infection (Waters et al, 2018). This review of studies published since 2017 begins with direct and indirect mechanisms by which HCMV may promote atherosclerosis, followed by clinical studies that illuminate the underlying pathology.

HCMV Affects Natural Killer Cells, αβ T Cells, γδ T Cells

Natural killer (NK) cells can recognize and kill tumor and virus-infected cells without the need for prior antigen stimulation (reviewed in Barnes et al, 2020). NK cell activity is controlled by signals received via activating and inhibitory receptors interacting with major histocompatibility complex class I and class I-like ligands on target cells (Barnes et al, 2020).

NK cells expressing the activating receptor, NKG2C, killer immunoglobulin-like receptors or the inhibitory receptor, leukocyte immunoglobulin-like receptor 1 (LIR-1; or CD85j) circulate at higher frequencies in HCMV-seropositive individuals and individuals with discernible HCMV disease (Barnes et al, 2020). HCMV infections have also been associated with upregulation of the terminal differentiation marker CD57 and loss of FcɛRγ expression on NK cells. These long-lived, highly differentiated cells have been termed “adaptive NK cells” and possess an enhanced capacity for cytokine production and antibody-mediated cellular cytotoxicity (Barnes et al, 2020). The role of these cells in CVD is unclear, but a recent study of PLWH suggests they may be protective in relation to coronary atherosclerosis (Alsulami et al, 2022).

After HCMV infection, populations of HCMV-reactive αβ T cells expand and persist (reviewed in van den Berg et al, 2019), so up to 10% of all memory T cells in healthy and immunocompromised adults are specific for HCMV. These cells acquire a terminally differentiated phenotype characterized by the expression of CD57 and loss of the costimulatory receptor CD28 (CD28⁻), with an increased capacity to produce inflammatory cytokines (interferon [IFN]γ, tumor necrosis factor [TNF]α), and expression of cytotoxic molecules (perforin and granzyme B). Terminally differentiated memory T cells may contribute directly to CVD because they upregulate the expression of CX3CR1 (Chen et al, 2020), a chemokine receptor involved in the migration of activated macrophages and T cells within the atherosclerotic lesions (reviewed in Skoda et al, 2018).

γδ T cells comprise 1–5% of T cells in the peripheral blood (reviewed in Gaballa et al, 2021; Waters et al, 2018). Most γδ T cells (∼70%) express a T cell receptor (TCR) containing δ-chain variable region 2 (Vδ2) in association with γ-chain variable region 9 (Vγ9) and possess innate-like properties (Khairallah et al, 2017). HCMV infections in the months after renal transplantation have been associated with increased proportions of circulating γδ T cells lacking Vδ2 TCR (i.e., Vδ2⁻) (Kaminski et al, 2021).

Direct Effects of HCMV on Atherosclerosis

HCMV antigens have been identified in resected atherosclerotic plaques in several studies over the last 30 years (reviewed in Lee et al, 2020). More recently, peripheral arterial tissues from 11/15 patients with atherosclerosis undergoing vascular surgery expressed HCMV phosphoprotein 65 (pp65) (Wang et al, 2016). However, there was no histological evidence of active HCMV replication (HCMV DNA or immediate-early 1 [IE-1] protein) and no tissues from patients without atherosclerosis were included for comparison. Accordingly, immunohistochemical analysis identified HCMV antigens more frequently in carotid atherosclerotic plaques obtained from patients undergoing coronary artery bypass graft surgery than in ascending aorta specimens with no evidence of atherosclerosis (Cao et al, 2017).

In a more detailed study, HCMV DNA was identified by polymerase chain reaction in 82% of plaques, with positive correlations demonstrated between HCMV viral load and proportions of intra-plaque CD4⁺ and CD8⁺ T cells displaying a differentiated (CD45RA⁻CD197⁻CD27^+/−CD28⁺) phenotype (Nikitskaya et al, 2016). Single-cell sequencing showed that CD8⁺ T cells with TCR recognizing HCMV pp65 were abundant in vulnerable plaques and in many cases cross-reacted with proteins found on smooth muscle and endothelial cells (Chowdhury et al, 2022). However, the function of these cells in vivo is unclear. Schafer and Zernecke (2020) reviewed mechanisms by which regulatory CD8⁺ T cells with immunosuppressive properties may inhibit atherosclerosis while cytotoxic CD8⁺ T cells may promote the rupture of unstable plaques.

Effects of HCMV on endothelial cells

Endothelial cells in healthy blood vessels maintain vascular homeostasis and exert anticoagulant, antiplatelet, fibrinolytic, and anti-inflammatory properties. Direct effects of HCMV replication in these cells (Fig. 1) have been investigated in vitro. Infection with HCMV caused endothelial mesenchymal transition of human umbilical vein endothelial cells (HUVEC) in the presence of transforming growth factor-β and mediated by metalloproteinase-2 (Chen et al, 2019a). HCMV infection can promote the translation of mitochondrial calcium uniporter mRNA and protein expression to increase apoptosis of human aortic endothelial cells (Zhu et al, 2022). Furthermore, HCMV infection of HUVEC downregulated mRNA of regulator of G-protein signaling 5 via DNA hypermethylation, and so promoted proliferation of endothelial cells (Zhang et al, 2020c). Endothelial-mesenchymal transition (Souilhol et al, 2018) and endothelial cell apoptosis (Li et al, 2022) and proliferation (Theodorou and Boon, 2018) are processes that occur throughout the different stages of atherosclerotic plaque formation and progression.

FIG. 1.

Direct and indirect effects of HCMV on atherosclerosis. During HCMV infection, the virus can directly infect vascular endothelial and SMC. HCMV infection can increase local production of chemotactic factors and adhesion molecules on vascular endothelial cells that promote inflammatory cell recruitment in atherosclerosis. These proteins can also affect proliferation and apoptosis of vascular endothelial cells and SMC, enhance monocyte transendothelial migration, and promote foam cell formation. Together, these processes can influence atherosclerotic plaque development and progression. CRP, C-reactive protein; EMT, endothelial mesenchymal transition; HCLS1, hematopoietic lineage cell-specific protein 1; HCMV, human cytomegalovirus; HHEX, hematopoietically expressed homeobox; HMG-CoA, β-hydroxy β-methylglutaryl-CoA; ICAM, intercellular adhesion molecule; SMC, smooth muscle cell; sTNFRII, soluble tumor necrosis factor receptor 2; VCAM, vascular cell adhesion molecule; VSP, vasodilator-stimulated phosphoprotein.

HCMV infection of endothelial cells may affect other cells via cytokines. For example, infection of HUVEC can stimulate the production of interleukin (IL)-11, a member of the IL-6 family (Gustafsson et al, 2018), with anti-inflammatory and pro-inflammatory effects (Fig. 1) (reviewed in Nguyen et al, 2019). Serum levels of IL-11 were lower in patients with CAD than in healthy subjects (Liu et al, 2019), but high plasma levels of IL-11 were linked to an increased risk of cardiac events in patients with chronic heart failure (Ye et al, 2019). Further studies should address their burden of HCMV.

HCMV infection can also reduce the expression of vasodilator-stimulated phosphoprotein in HUVEC, potentially impairing barrier function (Tian et al, 2018) and allowing monocyte transendothelial migration (Fig. 1). Infiltrated monocytes can differentiate into M1 or M2 macrophages, which can drive plaque progression and/or instability depending on the predominant subset. M2 macrophages display atheroprotective and anti-inflammatory effects, whereas M1 macrophages can promote atherosclerotic plaque enlargement and progression (Bartlett et al, 2019).

Effects of HCMV on vascular SMC

Vascular SMC produces extracellular matrix to maintain elasticity and stability of vessel walls and participate during all stages of plaque development (reviewed in Basatemur et al, 2019). During the early stages of atherogenesis, vascular SMC proliferates and migrates, thickening the intimal layer of the vessel walls. In the mature plaque, enhanced apoptosis and necrosis of vascular SMC can destabilize the plaque and increase the risk of rupture.

Infection of vascular SMC with HCMV in vitro increased expression of hematopoietically expressed homeobox (HHEX), a member of the HOX gene family promoting proliferation and reducing apoptosis (Fig. 1) (Li et al, 2016b). Expression of HHEX is high in atherosclerotic tissues, advanced plaques and oxLDL-treated SMC (Zhang et al, 2020a). HCMV infection of vascular SMC can also affect cellular lipid metabolism by increasing expression of β-hydroxy β-methylglutaryl-CoA (HMG-CoA) synthase and HMG-CoA reductase—increasing the cholesterol content of cultured cells (Fig. 1) (Li et al, 2016a).

Indirect Effects of HCMV on Atherosclerosis: Potential Mechanisms

The indirect effects of HCMV on atherosclerosis reflect the induction of inflammatory mediators and increased cell trafficking.

Micro RNAs encoded by HCMV and the host can influence atherogenesis

Micro RNAs (miRNA) are short non-coding RNA that affect gene silencing via mRNA degradation or post-translational repression. They are implicated in many pathologies including CVD (reviewed in Kalayinia et al, 2020). Infection of human HUVEC with HCMV increased expression of Homo sapiens- (has-) miR-138 and promoted endothelial cell motility (Fig. 1) (Zhang et al, 2017). The authors implicated reduced expression of histone deacetylases (notably SIRT1) and upregulation of the transcription factor, p-STAT3, in this change. This could promote atherosclerosis development and progression because SIRT1 can inhibit inflammation and oxidative stress (Wang and Chen, 2020), whereas STAT3 promotes macrophage polarization, pro-inflammatory cytokine production, and endothelial cell dysfunction (Chen et al, 2019b).

Potential roles of host-encoded miRNA have been studied with murine CMV (MCMV). Infection of C57BL/6J mice with MCMV increased blood pressure and arterial pressure, and reduced expression of Mus musculus- (mmu-) miR-1929-3p in the aorta (Zhou et al, 2021). The study overexpressed mmu-miR-1929-3p using an adeno-associated virus system, decreasing expression of endothelial injury factor (endothelin 1), endogenous nitric oxide synthase activity, and nitric oxide levels, thereby preventing MCMV-induced endothelial dysfunction (Zhou et al, 2021).

Overexpression of mmu-miR-1929-3p also reduced MCMV-induced expression of NLRP3 (NLR Family Pyrin Domain Containing 3) inflammasome and IL-1β in the aorta (Zhou et al, 2021). miRNA may also influence CVD via SMC function. Overexpression of mmu-miR-1929-3p in MCMV-infected mice suppressed MCMV-induced vascular remodeling as evidenced by reduced thickness of the aortic media, decreased collagen expression, and increased expression of α-smooth muscle actin (Zhou et al, 2021). These changes may affect the structural stability of atherosclerotic plaques (Harman and Jorgensen, 2019).

The HCMV genome also encodes several miRNAs that can regulate host and viral genes (reviewed in Zhang et al, 2020b). Among 2,763 participants of the Framingham Heart Study, the presence of hcmv-miR-US25-2-3p in plasma was associated with higher levels of proinflammatory cytokines, soluble TNF receptor II and IL-6, and lower levels of P-selectin, even though hcmv-miR-US25-2-3p was not associated with hypertension or coronary heart disease (CHD) (Koupenova et al, 2018).

Overexpression of hcmv-miR-UL112 in HUVEC can modulate genes encoding mediators of endothelial cell activation and damage, such as the mitogen-activated protein kinase and chemokine signaling pathways, cytokine-receptor interactions, and adhesion molecules (Shen et al, 2018). This may be important in vivo because we demonstrated that saliva from several renal transplant recipients (RTR) and healthy adults contained detectable hcmv-encoded miRNAs, most commonly miR-US5-2-3p (Waters et al, 2020). Furthermore, the presence of any hcmv-encoded miRNA (notably miR-US5-2-3p or miRUS25-2-3p) in saliva from RTR was associated with increased circulating T cell responses to IE-1, the first protein expressed during viral replication (Waters et al, 2020).

HCMV carries genes encoding chemokine receptors and IL-10

HCMV encodes US28, a G protein-coupled receptor with homology to endogenous human chemokine receptors. US28 expression is described in vascular SMC from renal allograft biopsies and is implicated in cell-to-cell spread of the virus (Fig. 1) (Lollinga et al, 2017). Chemokines that interact with US28 include CCL2, CCL3, CCL5, and CX3CL1. The expression of these chemokines is upregulated in cells of the arterial wall during atherogenesis. This can promote the recruitment of HCMV-infected cells to the endothelium facilitating viral dissemination and the movement of HCMV-infected SMC into atherosclerotic lesions (reviewed in Lee et al, 2017b).

Latent infection of monocytes with HCMV can increase the expression of HCLS1 (hematopoietic lineage cell-specific protein 1) in a US28-dependent manner (Aslam et al, 2019). Infected monocytes demonstrated increased adherence to and transmigration through the endothelial cell layers (Aslam et al, 2019), which contributed to the establishment of atherosclerotic plaques and disease progression (Jaworowski et al, 2019). Next-generation sequencing of US28 derived from clinical specimens revealed an association between the R267K variant and higher flow-mediated dilation (FMD), a measure of endothelial dysfunction (Waters et al, 2021). Furthermore, protein modeling indicated that R267K may influence the presentation of the chemokine receptor to ligands and therefore indirectly affect ligand binding (Waters et al, 2021). These studies support a role for US28 in cellular recruitment during atherosclerosis.

HCMV also carries a gene (UL111A) encoding a functional homolog of human IL-10 (cmvIL-10), which may help the virus to avoid immune clearance (reviewed in Patro, 2019). In the presence of cmvIL-10, primary monocytes displayed increased CXCR4 signaling and migration toward CXCL12 (Tu et al, 2018). This may drive atherosclerosis, as CXCL12 mRNA is highly expressed in carotid plaques, especially unstable plaques (Merckelbach et al, 2018). In an in vitro model of atherosclerosis, oxLDL induced the expression of CXCL12 in macrophages and promoted foam cell formation and proliferation of human aortic vascular SMC (Fig. 1) (Li et al, 2020d).

In the breast cancer cell line MDA-MB-231, cmvIL-10 can increase expression of matrix metalloproteinase-3 (MMP-3) and plasminogen activator inhibitor-1 (PAI-1) (Valle Oseguera and Spencer, 2017). MMP-3 and PAI-1 are both involved in atherogenesis and are highly expressed in several cells associated with the plaque (Olejarz et al, 2020; Sillen and Declerck, 2020). Levels of MMP-3 were found to be independent predictors of cardiovascular outcomes in patients with CAD (Guizani et al, 2019). Moreover, inhibition of PAI-1 prevented macrophage accumulation in atherosclerotic plaques from Ldlr ^−/− mice fed a “Western” diet (Khoukaz et al, 2020). Studies using endothelial cells, SMC, and macrophages are needed to define the role of cmvIL-10 in atherosclerotic plaques. In RTR, we linked carriage of the P122S variant of the UL111A gene with lower FMD, marking inferior vascular endothelial function (Waters et al, 2022).

HCMV induces the production of inflammatory mediators by men and mice

In a study of 694 participants (aged 18–85 years), HCMV seropositivity aligned with increased levels of the inflammatory marker C-reactive protein (CRP), but not with serum amyloid A, after adjusting for age, body mass index (BMI), smoking status, gender, and ethnicity (Styles et al, 2020). Furthermore, high HCMV antibody levels associated with increased levels of ICAM-1 and VCAM-1 (Styles et al, 2020). Levels of HCMV DNA in blood were higher in patients with acute coronary syndrome than in controls and correlated with plasma levels of CRP (Nikitskaya et al, 2016).

MCMV infection accelerated the progression of atherosclerosis in ApoE^−/− mice on high-fat diet in parallel with increased levels of intracellular reactive oxygen species, TNFα, IL-6, VCAM-1, and ICAM-1, and components of the HMGB1-TLRS-NFκB signaling pathways (Lv et al, 2020). The natural dietary compound curcumin inhibited MCMV replication in vivo and in vitro and reduced HCMV replication in HUVEC (Lv et al, 2020).

Associations Between HCMV and Clinical Outcomes in Different Patient Populations

Most early studies stratified individuals as HCMV seropositive or seronegative, but this does not address differences in the burden of HCMV or the presence of active viral replication. The burden of HCMV in individuals who are not acutely immunodeficient can be ascertained by monitoring levels of HCMV IgG antibody or HCMV DNA in biological specimens (Waters et al, 2018). The presence of HCMV DNA in plasma, blood leukocytes, saliva, or urine indicates current viral replication and may be short-lived. In contrast, HCMV-reactive antibodies and T cells plausibly reflect the cumulative viral burden and may rise in response to waves of viral replication (van den Berg et al, 2018). This can provide insight into immune system dysfunction and its clinical implications but may not provide meaningful quantitation of the viral burden if other factors are impacting upon immune responsiveness.

Recent studies linking HCMV and CVD in different populations are discussed below and summarized in Table 1. Factors that must be considered in the evaluation of these data include the age of the subjects, and whether the assays detect active replication or persistent (potentially quiescent) infections.

Table 1.

Studies Linking Human Cytomegalovirus Infection with Cardiovascular Disease in Different Populations

Study groups	Findings	Ref.
General population
Patients with atherosclerosis (n = 3,328), controls (n = 2,090)	• HCMV infection (HCMV IgG and IgM, HCMV DNA, and HCMV pp65) associated with ↑ risk (OR 2.02–8.92) of atherosclerosis	Jia et al (2017)
Participants (n = 34,564), CVD patients (n = 4,789)	• HCMV seropositivity associated with 22% ↑ risk of future CVD events	Wang et al (2017)
Participants (n = 268)	• HCMV seropositivity and ↑ HCMV antibody levels associated with prevalence of CVD	Samson et al (2020)
Participants (n = 10122)	• HCMV seropositivity or high HCMV IgG quartiles not associated with ↑ risk of cardiovascular death	Chen et al (2020)
Participants (n = 806)	• HCMV seropositivity not associated with ↑ risk of cardiovascular death	Nenna et al (2020)
Patients with STEMI (n = 33), no CVD (n = 33)	• ↑ Frequency of detection and plasma HCMV DNA load in patients with STEMI (vs. no CVD) • Detection of HCMV DNA was an independent predictor for vascular endothelial dysfunction	Lebedeva et al (2020)
Patients undergoing carotid endarterectomy (n = 36)	• ↑ Frequency of HCMV DNA detection in patients with carotid artery stenosis versus no stenosis	Beyaz et al (2019)
Participants (n = 6,101)	• HCMV seropositivity was associated with stroke in women but not in men	Zhen et al (2022)
CHD (n = 192), controls (n = 79)	• ↑ Frequency of HCMV IgM and IgG seropositivity in CHD patients versus controls	Li et al (2020c)
Patients with atherosclerosis (n = 42), healthy subjects (n = 30)	• ↑ Frequency of HCMV seropositivity (not HCMV DNA) in patients with carotid atherosclerosis versus healthy subjects • HCMV seropositivity associated with unstable plaques	Li et al (2020b)
Hypertensive patients (n = 207)	• Proportions of HCMV pp65-specific IFNγ-producing CD8⁺ T cells and CD57⁺ and CD28⁻ CD8⁺ T cells positively associated with PWV	Youn et al (2018)
Healthy subjects, hypertension, CAD, diabetes (n = 415)	• HCMV pp65-specific CD8⁺ T cell responses but not HCMV IgG antibody levels associated with PWV	Yu et al (2017)
Patients with renal disease
Patients with ESRD (n = 408), healthy subjects (n = 57)	• ↑ Levels of HCMV IgG antibody levels compared to healthy subjects • ↑ HCMV IgG antibody levels associated with prevalent CAD	Yang et al (2018)
Patients with ESRD (n = 60), Healthy subjects (n = 30)	• HCMV seropositivity and frequency of CD28⁻CD4⁺ T cells were independent predictors of cIMT	Okba et al (2019)
Patients with non-dialysis chronic kidney disease (n = 764)	• HCMV seropositivity associated with ↑ systolic blood pressure, prevalent CVD, ischemic heart disease, and cerebrovascular disease in univariate analyses • HCMV seropositivity associated with ↑ prevalence of CVD in multivariate analysis	Karangizi et al (2020)
RTR
RTR (n = 82), healthy subjects (n = 81)	• HCMV IgG antibody levels associated with FMD but not cIMT in RTR	Lee et al (2017a)
RTR (n = 82)	• Presence of HCMV DNA in saliva or plasma associated with ↑ plasma ICAM-1 and VCAM-1	Waters et al (2019)
RTR (n = 45), healthy subjects (n = 58)	• Detectable HCMV DNA in saliva (but not sIFNAR2, sTNFR1, sCD14, CRP, P-selectin, ICAM-1, VCAM-1) predicted impaired FMD in RTR (and not healthy subjects)	Affandi et al (2020)
RTR (n = 27), healthy subjects (n = 28)	• Proportions of LIR-1⁺ and/or FcRγ ⁻ NK cells correlated inversely with FMD	Lee et al (2019b)
RTR (n = 82), healthy subjects (n = 81)	• Genetic variants of LILRB1, NKG2C and HLA-G were associated with FMD and cIMT	Waters et al (2017)
Patients with autoimmune diseases
ANCA-associated vasculitis (n = 40), healthy subjects (n = 38)	• HCMV infection and CD4⁺CD28⁻ T cells did not correlate with cIMT or PWV	Slot et al (2017)
ANCA-associated vasculitis (n = 53), healthy subjects (n = 30)	• Frequency of CD28⁻ CD4⁺ T cells (predominantly HCMV-specific, Th1 phenotype, CX3CR1⁺) associated with arterial stiffness	Chanouzas et al (2018)
New-onset rheumatoid arthritis (n = 79), healthy subjects (n = 44)	• HCMV-seropositive patients experienced a rapid ↑ in cIMT and ↑ frequencies of CD4⁺ and CD8⁺ CD28⁻ (vs. HCMV seronegative patients)	Wahlin et al (2021)
PLWH
PLWH (n = 105), healthy subjects (n = 105)	• ↑ HCMV IgG antibody levels associated with ↑ risk of coronary artery calcium score and ↑ cIMT in PLWH but not in healthy donors	Knudsen et al (2019)
PLWH (n = 94)	• Cardiac microvascular dysfunction associated with ↑ HCMV IgG in women with HIV	Knudsen et al (2018)
PLWH (n = 79)	• HCMV IE-1 antibody correlated with the right retinal artery calibers (3 months on ART)	Edwar et al (2019)
PLWH (n = 67)	• HCMV IE-1 antibody levels positively correlated with right cIMT (12 months on ART)	Karim et al (2017)
PLWH (n = 82)	• HCMV IgG antibody levels predicted ↑ left and right cIMT (5 years on ART)	Wulandari et al (2020)
PLWH (n = 60), healthy subjects (n = 31)	• HCMV-specific CD8⁺ T cells (but not CD4⁺ T cells or HCMV IgG) associated with pulse pressure	Ballegaard et al (2020)
PLWH (n = 70)	• ↑ Proportions of CX3CR1⁺GPR56⁺CD57⁺CD4⁺ T cells in individuals with carotid plaque (15% of CX3CR1⁺GPR56⁺CD57⁺ CD4⁺ T cells were HCMV-specific)	Wanjalla et al (2021)

ANCA, anti-neutrophil cytoplasmic autoantibodies; ART, antiretroviral therapy; CAD, coronary artery disease; CHD, coronary heart disease; cIMT, carotid intima-media thickness; CRP, C-reactive protein; CVD, cardiovascular disease; ESRD, end-stage renal disease; FMD, flow-mediated dilation; HCMV, human cytomegalovirus; ICAM-1, intercellular adhesion molecule-1; IE-1, immediate-early 1; IFN, interferon; LIR-1, leukocyte immunoglobulin-like receptor 1; NK, natural killer; OR, odds ratio; PLWH, people living with HIV; pp65, phosphoprotein 65; PWV, pulse wave velocity; RTR, renal transplant recipients; sIFNAR2, soluble interferon alpha receptor 2; STEMI, ST-elevation myocardial infarction; VCAM-1, vascular cell adhesion molecule-1.

Links between HCMV and CVD in the general population

A meta-analysis of 30 studies (3,328 patients with atherosclerosis and 2,090 controls) demonstrated an association between HCMV infection (defined as the detection of HCMV antibody or DNA) with increased risk (odds ratio 2.02–8.92) of atherosclerosis (Jia et al, 2017). Furthermore, this association was greater among Asian populations. This may reflect the high HCMV seroprevalence and antibody levels observed in Asia (Zuhair et al, 2019). Another meta-analysis of 10 community-based prospective studies (34,564 participants and 4,789 patients with CVD [ischemic heart disease, stroke, cardiovascular death]) linked HCMV seropositivity with a 22% increase in the relative risk of future CVD events (Wang et al, 2017). Subgroup analyses revealed a 16% increase in the relative risk of stroke or ischemic heart disease and a 30% higher relative risk of cardiovascular mortality in HCMV-seropositive individuals.

A longitudinal study linked elevated levels of HCMV antibody with prevalence of CVD, but not with frailty, in 268 individuals (average age of 43 years) followed for 27 years (Samson et al, 2020). Another recent study of five longitudinal cohorts of Caucasian community-dwelling older adults (aged 59–93 years, followed for 2.8–11.4 years) initially linked HCMV seropositivity or high HCMV IgG quartiles with cardiovascular mortality. The association was lost after adjustment for confounders (e.g., age, sex, BMI, smoking status, comorbidities, medications, education and plasma CRP) (Chen et al, 2021)—perhaps because comorbidities and inflammation can reflect HCMV status (Waters et al, 2018). Since HCMV antibody levels rise with age but eventually plateau, the consequence of adjustments for age must depend on the age range of participants.

Among 6,101 participants (20–49 years old), HCMV seropositivity was associated with stroke in women but not in men after adjustment for age, BMI, and other markers of cardiovascular risk (Zhen et al, 2022). HCMV IgM and IgG seropositivity were more common in patients with CHD than healthy controls. The seropositivity rate was highest in patients with acute myocardial infarction, followed by individuals with angina pectoris and latent CHD (Li et al, 2020c). Hypertension, diabetes, family history, and HCMV IgM and IgM seropositivity were independent predictors of CHD (Li et al, 2020c). Li et al (2020b) reported a higher frequency of HCMV seropositivity but not HCMV DNA load in patients with carotid atherosclerosis (aged 64.9 ± 12.2 years) compared to healthy controls. Furthermore, the authors reported higher levels of MMP-9, TNFα, and lectin-like oxidized low-density lipoprotein receptor-1, and the presence of unstable carotid plaques in HCMV-seropositive patients (Li et al, 2020b).

Active HCMV replication appears to mark all stages of CVD. The frequency of detection and plasma HCMV DNA load were higher in patients with ST-elevation myocardial infarction (STEMI) compared to individuals without CVD (Lebedeva et al, 2020). In multiple regression models, the presence of HCMV DNA (but not age, sex, hypertension, or elevated CRP level) was an independent predictor for the early development of vascular endothelial dysfunction determined using FMD of the brachial artery (Lebedeva et al, 2020). In 36 patients who underwent carotid endarterectomy, HCMV DNA detection in plaques was more frequent in patients with bilateral carotid artery stenosis (Beyaz et al, 2019).

Among 207 hypertensive patients (aged 63 ± 8 years), proportions of HCMV pp65-specific IFNγ-producing CD8⁺ T cells and CD57⁺ and CD28⁻ CD8⁺ T cells were positively associated with pulse wave velocity (PWV) (Youn et al, 2018), a non-invasive measure of arterial stiffness. Yu et al (2017) also linked HCMV pp65-specific CD8⁺ T cell responses with PWV in 415 Koreans (aged 20–82 years) after adjustment for traditional cardiovascular risk factors but found no association between HCMV IgG antibody levels and PWV. A study of 27 healthy young participants and 215 healthy older individuals found HCMV infection to be the major risk factor for accumulation of senescent CD28⁻CD4⁺ T cells, whereas aging contributed little (Pera et al, 2018).

Links between HCMV and CVD in patients with renal disease

CVD remains the major cause of morbidity and mortality in patients with end-stage renal disease. Traditional CVD risk factors including hypertension, dyslipidemia, and diabetes only explain a small proportion of the high burden of CVD in this population, so other factors may play a role (Jankowski et al, 2021). Persistent low-grade inflammation may link HCMV reactivation and CVD. In 408 patients with end-stage renal disease, elevated HCMV IgG antibody levels correlated with higher prevalence of chronic heart failure and myocardial infarction, and with prevalent CAD after adjusting for age, gender, and other cardiovascular risk factors (Yang et al, 2018). As expected, levels of HCMV IgG antibody also correlated with proportions of terminally differentiated CD8⁺ and CD4⁺ T cells (Yang et al, 2018). In a similar study, HCMV seropositivity, CD28^-CD4⁺ T cells, serum cholesterol, and triglycerides were independent predictors of carotid intima-media thickness (cIMT), an early indicator of atherosclerosis (Okba et al, 2019).

HCMV seropositivity was associated with higher systolic blood pressure, prevalent CVD, ischemic heart disease, and cerebrovascular disease in patients with non-dialysis chronic kidney disease (Karangizi et al, 2020). These findings are broadly similar to the associations seen in RTR discussed below.

Links between HCMV and CVD in transplant recipients

HCMV infection has been implicated in increased atherosclerotic events observed in RTR (Rodriguez-Goncer et al, 2020). In our cohort of 82 RTR studied >2 years post-transplant, HCMV IgG antibody levels were independently associated with FMD but not cIMT (Lee et al, 2019a). The presence of HCMV DNA in saliva or plasma from RTR was associated with increased plasma levels of ICAM-1 and VCAM-1, biomarkers implicated in atherogenesis (Waters et al, 2020). Furthermore, detectable HCMV DNA in saliva (and not soluble interferon alpha receptor 2 [sIFNAR2], sTNFR1, sCD14, CRP, P-selectin, ICAM-1, VCAM-1) predicted impaired FMD measured 3 years later (Affandi et al, 2020).

As described earlier, HCMV infection alters the phenotype of NK cells, with characteristic loss of FcRγ expression and expression of the inhibitory receptor LIR-1 (Barnes et al, 2020). Of the seven RTR with detectable HCMV DNA in plasma, we observed the highest frequency of NK cells expressing the activating receptor (NKG2C) and LIR-1, and lacking FcRγ in an individual with the highest burden of HCMV (Makwana et al, 2019). Accordingly, proportions of LIR-1⁺ and/or FcRγ ⁻ NK cells induced by HCMV correlated inversely with FMD (Lee et al, 2019b). This in itself does not imply causation—the NK cell profiles may simply mark a high burden of HCMV. However, in the same cohorts of RTR and healthy adults, polymorphisms in genes encoding LIR-1, NKG2C, and HLA-G were associated with FMD and cIMT (Waters et al, 2017). This suggests a direct role for the NK cell profiles.

HCMV-seropositive RTR with stable graft function retained higher frequencies of circulating Vδ2⁻ γδ T cells than HCMV-seropositive healthy controls or HCMV-seronegative individuals (Lee et al, 2017a). Frequencies of circulating Vδ2⁻ but not Vδ2⁺ γδ T cells were also lower in the seropositive RTR with carotid plaques than those without (Lee et al, 2017a). This may reflect selective recruitment of these cells to the plaques, so a direct role for γδ T cell subsets in atherosclerotic plaques warrants investigation.

Cardiac allograft vasculopathy is an accelerated form of CAD and is one of the leading causes of death after heart transplantation (Habibi et al, 2020). Cardiac allograft vasculopathy and atherosclerosis share several characteristics including endothelial dysfunction and inflammation. In addition to immunologic factors (e.g., HLA mismatch), non-immunologic factors such as hyperlipidemia and HCMV infection may drive cardiac allograft vasculopathy. While HCMV seropositivity was associated with an increased (23–41%) risk of all-cause mortality 5 years after heart transplantation when compared with HCMV seronegative recipients, no association was evident with allograft vasculopathy in a U.S. study following 21,878 patients (Suarez-Pierre et al, 2020). The authors also found that the use of ganciclovir alone was also associated with increased risk of mortality (Suarez-Pierre et al, 2020). Interestingly, a murine aortic allograft model showed that deletion of the viral G protein-coupled receptor M33, sharing biological functions with US28, associated with reduced SMC proliferation, luminal occlusion, and expression of ICAM-1 and VCAM-1 (Fritz et al, 2021).

Links between HCMV and CVD in patients with autoimmune diseases

HCMV may also promote CVD in patients with autoimmune diseases. Several studies have addressed the role of CD28⁻ T cell populations. HCMV infection was associated with increased frequencies of CD28⁻CD4⁺ T cells in patients with anti-neutrophil cytoplasmic autoantibody (ANCA)-associated vasculitis and percentages were higher compared to HCMV-seropositive healthy subjects. However, neither CD28⁻CD4⁺ T cells nor HCMV infection correlated with cIMT or PWV (Slot et al, 2017). In contrast, another study of HCMV-seropositive patients with ANCA-associated vasculitis reported an association between percentage of CD28^-CD4⁺ T cells with PWV that was independent of age, proteinuria, peripheral arterial blood pressure, and plasma TNFα levels. Furthermore, the CD28⁻CD4⁺ T cells were predominantly HCMV-specific, possessed a Th1 phenotype, and expressed CX3CR1 and cytotoxic molecules (Chanouzas et al, 2018).

HCMV-seropositive patients with new-onset rheumatoid arthritis displayed a more rapid increase in cIMT after 1.5 years and higher frequencies of CD28⁻CD4⁺ and CD8⁺ T cells compared to HCMV-seronegative patients (Wahlin et al, 2021). Furthermore, the authors linked increased percentages of CD28⁻ T cells with cIMT after 11 years, after adjusting for systolic blood pressure (Wahlin et al, 2021). These studies support a link between HCMV and CVD in autoimmune diseases.

Links between HCMV and CVD in PLWH

Almost all PLWH are HCMV-seropositive (Freeman et al, 2016; Hoehl et al, 2020). HCMV retinitis was an AIDS-defining illness, but is now rare as patients begin antiretroviral therapy (ART) before their CD4⁺ T cell counts decline markedly (Ude et al, 2022). There is abundant evidence that PLWH have elevated rates of CVD (Shah et al, 2018). While it is plausible that HCMV may contribute to this increase, it is difficult to prove without matched cohorts of HCMV-seronegative PLWH.

In our JakCCANDO cohort, all PLWH began ART with very high burdens of HCMV evidenced by extremely high levels of HCMV-reactive antibody and the detection of HCMV DNA by a simple quantitative polymerase chain reaction in 52% of patients (Ariyanto et al, 2022). As the antibody levels were similar in PLWH with and without detectable HCMV DNA, we conclude that this grouping divides patients with a moderately high burden of HCMV from those with an extremely high burden. In this respect, the cohort is distinct from transplant recipients and PLWH from better resourced settings. A further complication is the observed rise in antibody levels on ART (Ariyanto et al, 2018), which we ascribe to immune recovery rather than an increase in the viral burden.

In this cohort, levels of HCMV IE-1 antibody correlated with the right retinal artery calibers, a non-invasive measure of microvasculopathy (Edwar et al, 2019). We also demonstrated a direct relationship between HCMV IE-1 antibody levels and right cIMT, whereas HCMV antibody levels inversely correlated with internal diameter of the right artery during 12 months of ART (Karim et al, 2017). HCMV IgG antibody levels and periodontal disease predicted increased left and right cIMT after 5 years on ART (Wulandari et al, 2020).

In settings that are less “resource constrained” (e.g., Europe and North America), high HCMV IgG antibody levels have been associated with increased risk of coronary artery calcium score and higher cIMT in PLWH but not in healthy donors (Knudsen et al, 2019). Positron emission tomography linked high HCMV IgG antibody levels with cardiac microvascular dysfunction in women living with HIV (Knudsen et al, 2018).

Among PLWH with no detectable HIV RNA on ART, frequencies of HCMV-specific cytokine-producing CD8⁺ T cells were associated with pulse pressure (difference between systolic and diastolic blood pressure) after adjustment for age, smoking, and LDL-cholesterol (Ballegaard et al, 2020). HIV-related factors, IL-6, and CD8⁺ T cell senescence did not mediate the associations, and no associations were found between HCMV-specific CD4⁺ T cell responses and blood pressure (Ballegaard et al, 2020). The authors speculate that the association with HCMV-specific CD8⁺ T cells may be causative.

Another study linked HCMV-specific IFNγ responses with frequencies of CD4⁺ T cells expressing CX3CR1 (Garg et al, 2019). CD4⁺ T cells from PLWH had reduced CD28 expression and high expression of CD2, CD57, and CX3CR1 (Chen et al, 2020). The authors reported higher expression of the activation marker CD69 on CD4⁺ T cells in atherosclerotic plaques from HIV-uninfected individuals and expression of CX3CL1 and LFA-3 on the vascular endothelium (Chen et al, 2020). The authors postulated that plaque formation may reflect recruitment of CX3CR1⁺CD57⁺CD28⁻ CD4⁺ T cells to CX3CL1-expressing endothelial cells via cytokine and release of lytic granules following CD2/LFA-3 interactions.

A study of 70 aviraemic PLWH with no known CVD demonstrated increased proportions of circulating CX3CR1⁺GPR56⁺ CD57⁺CD4⁺ T cells in individuals with carotid plaque. CX3CR1⁺CD4⁺ T cells were also demonstrated in coronary plaques from PLWH (Wanjalla et al, 2021). A role for HCMV in their accumulation is supported by the detection of CX3CR1⁺GPR56⁺CD57⁺CD4⁺ T cells in the aorta of HIV-uninfected individuals since ∼15% of the population recognized a single HCMV gB epitope (Wanjalla et al, 2021).

HCMV and cardiovascular outcomes in patients with COVID-19

Emerging data have associated infection with SARS CoV-2 (COVID-19) with poor CVD outcomes seen in some patients (reviewed in Liu and Zhang, 2020; Soumya et al, 2021). Increased systemic inflammation (cytokine storms) leading to endothelial dysfunction and direct infection of cardiovascular tissues have been proposed as plausible mechanisms. (Liu and Zhang, 2020; Soumya et al, 2021). The reactivation of HCMV has been reported in patients with COVID-19 infection (Le Balc'h et al, 2020), and HCMV seropositivity is a risk factor for severe COVID-19 disease and subsequent hospitalization (Alanio et al, 2022; Weber et al, 2022). The increasing reports of patients with long COVID-19 who still have persistent symptoms several months after infection (Brodin et al, 2022) highlight the need to understand the role of HCMV in this setting.

Concluding Remarks

Evidence from different populations has confirmed HCMV seropositivity or high antibody levels as significant risk factors for poorer vascular health and increased CVD events. HCMV contributes to CVD through several mechanisms (Fig. 1). HCMV infection can increase local production of chemotactic factors and adhesion molecules on vascular endothelial cells to promote inflammatory cell recruitment in atherosclerotic plaques. Furthermore, direct infection can affect proliferation and apoptosis of vascular endothelial cells and SMC, enhance monocyte transendothelial migration, and promote foam cell formation. HCMV-encoded miRNAs and proteins such as US28 and cmvIL-10 can alter vascular endothelial and SMC functions to promote plaque rupture, and the inflammatory responses induced by HCMV infection can destabilize plaques and cause acute coronary syndromes. Further studies are warranted to determine how HCMV contribute directly and indirectly to the pathogenesis of CVD. This will allow the identification of patients who will benefit from therapy to reduce their burden of HCMV.

Footnotes

Authors' Contributions

Conceptualization; writing—original draft, S.L. Review and editing, P.P., J.A., S.W. All authors have read and agreed to the published version of the article.

Author Disclosure Statement

No competing financial interests exist.

Funding Information

No funding was received for this article.

References

Affandi

, Lee

, Chih

, et al. Cytomegalovirus burden improves a predictive model identifying measures of vascular risk in renal transplant recipients and healthy adults. J Med Virol, 2020. [Epub ahead of print]; doi: 10.1002/jmv.25697

Alanio

, Verma

, Mathew

, et al. Cytomegalovirus latent infection is associated with an increased risk of COVID-19-related hospitalization. J Infect Dis, 2022.226(3):463–473; doi: 10.1093/infdis/jiac020

Alsulami

, Sadouni

, Tremblay-Sher

, et al. High frequencies of adaptive NK cells are associated with absence of coronary plaque in cytomegalovirus infected people living with HIV. Medicine (Baltimore), 2022; 101(38):e30794; doi: 10.1097/MD.0000000000030794

Ariyanto

, Estiasari

, Karim

, et al. Which NK cell populations mark the high burden of CMV present in all HIV patients beginning ART in Indonesia?. AIDS Res Ther, 2022; 19(1):16; doi: 10.1186/s12981-022-00439-2

Ariyanto

, Estiasari

, Waters

, et al. Active and persistent cytomegalovirus infections affect T cells in young adult HIV patients commencing antiretroviral therapy. Viral Immunol, 2018; 31(6):472–479; doi: 10.1089/vim.2018.0014

Aslam

, Williamson

, Romashova

, et al. Human cytomegalovirus upregulates expression of HCLS1 resulting in increased cell motility and transendothelial migration during latency. iScience, 2019; 20:60–72; doi: 10.1016/j.isci.2019.09.016

Ballegaard

, Pedersen

, Braendstrup

, et al. Cytomegalovirus-specific CD8+ T-cell responses are associated with arterial blood pressure in people living with HIV. PLoS One, 2020; 15(1):e0226182; doi: 10.1371/journal.pone.0226182

Barnes

, Schilizzi

, Audsley

, et al. Deciphering the immunological phenomenon of adaptive natural killer (NK) cells and cytomegalovirus (CMV). Int J Mol Sci, 2020; 21(22):8864; doi: 10.3390/ijms21228864

Bartlett

, Ludewick

, Misra

, et al. Macrophages and T cells in atherosclerosis: A translational perspective. Am J Physiol Heart Circ Physiol, 2019; 317(2):H375–H386; doi: 10.1152/ajpheart.00206.2019

10.

Basatemur

, Jorgensen

, Clarke

MCH

, et al. Vascular smooth muscle cells in atherosclerosis. Nat Rev Cardiol, 2019; 16(12):727–744; doi: 10.1038/s41569-019-0227-9

11.

Beyaz

, Ugurlucan

, Oztas

, et al. Evaluation of the relationship between plaque formation leading to symptomatic carotid artery stenosis and cytomegalovirus by investigating the virus DNA. Arch Med Sci Atheroscler Dis, 2019; 4:e19–e24; doi: 10.5114/amsad.2019.83304

12.

Brodin

, Casari

, Townsend

, et al. Studying severe long COVID to understand post-infectious disorders beyond COVID-19. Nat Med, 2022; 28(5):879–882.

13.

Cao

, Mao

, Dong

, et al. Detection of specific Chlamydia pneumoniae and cytomegalovirus antigens in human carotid atherosclerotic plaque in a Chinese population. Oncotarget, 2017; 8(33):55435–55442; doi: 10.18632/oncotarget.19314

14.

Chanouzas

, Sagmeister

, Dyall

, Sharp

, Powley

, Johal

, et al. The host cellular immune response to cytomegalovirus targets the endothelium and is associated with increased arterial stiffness in ANCA-associated vasculitis. Arthritis Res Ther, 2018; 20(1):194; doi: 10.1186/s13075-018-1695-8

15.

Chen

, Morris

, Panigrahi

, et al. Cytomegalovirus coinfection is associated with increased vascular-homing CD57(+) CD4 T cells in HIV infection. J Immunol, 2020; 204(10):2722–2733; doi: 10.4049/jimmunol.1900734

16.

Chen

, Yang

, Wang

, et al. Human cytomegalovirus promotes the activation of TGF-beta1 in human umbilical vein endothelial cells by MMP-2 after endothelial mesenchymal transition. Adv Clin Exp Med, 2019a;28(11):1441–1450; doi: 10.17219/acem/109199

17.

Chen

, Lv

, Yang

, et al. Targeted inhibition of STAT3 as a potential treatment strategy for atherosclerosis. Theranostics, 2019b;9(22):6424–6442; doi: 10.7150/thno.35528

18.

Chen

, Pawelec

, Trompet

, et al. Associations of cytomegalovirus infection with all-cause and cardiovascular mortality in multiple observational cohort studies of older adults. J Infect Dis, 2021; 223(2):238–246; doi: 10.1093/infdis/jiaa480

19.

Chowdhury

, D'Addabbo

, Huang

, et al. Human coronary plaque T cells are clonal and cross-react to virus and self. Circ Res, 2022; 130(10):1510–1530; doi: 10.1161/CIRCRESAHA.121.320090

20.

Edwar

, Karim

, Wijaya

, et al. Factors affecting the health of retinal vessels in human immunodeficiency virus patients beginning anti-retroviral therapy. AIDS Res Hum Retroviruses, 2019; 35(6):529–535; doi: 10.1089/AID.2018.0251

21.

Forte

, Zhang

, Thorp

, et al. Cytomegalovirus latency and reactivation: An intricate interplay with the host immune response. Front Cell Infect Microbiol, 2020; 10:130; doi: 10.3389/fcimb.2020.00130

22.

Freeman

, Lederman

, Gianella

. Partners in crime: The role of CMV in immune dysregulation and clinical outcome during HIV infection. Curr HIV/AIDS Rep, 2016; 13(1):10–19; doi: 10.1007/s11904-016-0297-9

23.

Fritz

, Stamminger

, Ramsperger-Gleixner

, et al. Cytomegalovirus chemokine receptor M33 knockout reduces chronic allograft rejection in a murine aortic transplant model. Transpl Immunol, 2021; 64:101359; doi: 10.1016/j.trim.2020.101359

24.

Gaballa

, Alagrafi

, Uhlin

, et al. Revisiting the role of gammadelta T cells in anti-CMV immune response after transplantation. Viruses, 2021; 13(6):1031; doi: 10.3390/v13061031

25.

Garg

, Gianella

, Nakazawa

, et al. Association of cytomegalovirus DNA and immunologic markers of cardiovascular disease. Open Forum Infect Dis, 2019; 6(5):ofz113; doi: 10.1093/ofid/ofz113

26.

Guizani

, Zidi

, Zayani

, et al. Matrix metalloproteinase-3 predicts clinical cardiovascular outcomes in patients with coronary artery disease: A 5years cohort study. Mol Biol Rep, 2019; 46(5):4699–4707; doi: 10.1007/s11033-019-04914-4

27.

Gustafsson

KLR

, Renne

, Soderberg-Naucler

, et al. Human cytomegalovirus replication induces endothelial cell interleukin-11. Cytokine, 2018; 111:563–566; doi: 10.1016/j.cyto.2018.05.018

28.

Habibi

, Ghaffarpasand

, Shojaei

, et al. Prognostic value of biomarkers in cardiac allograft vasculopathy following heart transplantation: A literature review. Cardiology, 2020; 145(11):693–702; doi: 10.1159/000509630

29.

Harman

, Jorgensen

. The role of smooth muscle cells in plaque stability: Therapeutic targeting potential. Br J Pharmacol, 2019; 176(19):3741–3753; doi: 10.1111/bph.14779

30.

Hoehl

, Berger

, Ciesek

, Rabenau

. Thirty years of CMV seroprevalence—A longitudinal analysis in a German university hospital. Eur J Clin Microbiol Infect Dis, 2020; 39(6):1095–1102; doi: 10.1007/s10096-020-03814-x

31.

Jankowski

, Floege

, Fliser

, et al. Cardiovascular disease in chronic kidney disease: Pathophysiological insights and therapeutic options. Circulation, 2021; 143(11):1157–1172; doi: 10.1161/circulationaha.120.050686

32.

Jaworowski

, Hearps

, Angelovich

, et al. How monocytes contribute to increased risk of atherosclerosis in virologically-suppressed HIV-positive individuals receiving combination antiretroviral therapy. Front Immunol, 2019; 10:1378; doi: 10.3389/fimmu.2019.01378

33.

Jia

, Liu

, Han

, et al. Cytomegalovirus infection and atherosclerosis risk: A meta-analysis. J Med Virol, 2017; 89(12):2196–2206; doi: 10.1002/jmv.24858

34.

Kalayinia

, Arjmand

, Maleki

, et al. MicroRNAs: Roles in cardiovascular development and disease. Cardiovasc Pathol, 2020; 50:107296; doi: 10.1016/j.carpath.2020.107296

35.

Kaminski

, Menard

, El Hayani

, et al. Characterization of a unique gammadelta T-cell subset as a specific marker of cytomegalovirus infection severity. J Infect Dis, 2021; 223(4):655–666; doi: 10.1093/infdis/jiaa400

36.

Karangizi

AHK

, Chanouzas

, Fenton

, et al. Cytomegalovirus seropositivity is independently associated with cardiovascular disease in non-dialysis dependent chronic kidney disease. QJM, 2020; 113(4):253–257; doi: 10.1093/qjmed/hcz258

37.

Karim

, Wijaya

, Rahmaniyah

, et al. Factors affecting affect cardiovascular health in Indonesian HIV patients beginning ART. AIDS Res Ther, 2017; 14(1):52; doi: 10.1186/s12981-017-0180-9

38.

Khairallah

, Dechanet-Merville

, Capone

. gammadelta T cell-mediated immunity to cytomegalovirus infection. Front Immunol, 2017; 8:105; doi: 10.3389/fimmu.2017.00105

39.

Khoukaz

, Ji

, Braet

, et al. Drug targeting of plasminogen activator inhibitor-1 inhibits metabolic dysfunction and atherosclerosis in a murine model of metabolic syndrome. Arterioscler Thromb Vasc Biol, 2020; 40(6):1479–1490; doi: 10.1161/ATVBAHA.119.313775

40.

Knudsen

, Kristoffersen

, Panum

, et al. Coronary artery calcium and intima-media thickness are associated with level of cytomegalovirus immunoglobulin G in HIV-infected patients. HIV Med, 2019; 20(1):60–62; doi: 10.1111/hiv.12672

41.

Knudsen

, Thorsteinsson

, Christensen

, et al. Cardiac microvascular dysfunction in women living with HIV is associated with cytomegalovirus immunoglobulin G. Open Forum Infect Dis, 2018; 5(9):ofy205; doi: 10.1093/ofid/ofy205

42.

Koupenova

, Mick

, Corkrey

, et al. Micro RNAs from DNA viruses are found widely in plasma in a large observational human population. Sci Rep, 2018; 8(1):6397; doi: 10.1038/s41598-018-24765-6

43.

Le Balc'h

, Pinceaux

, Pronier

, et al. Herpes simplex virus and cytomegalovirus reactivations among severe COVID-19 patients. Crit Care, 2020; 24(1):530; doi: 10.1186/s13054-020-03252-3

44.

Lebedeva

, Maryukhnich

, Grivel

, et al. Productive cytomegalovirus infection is associated with impaired endothelial function in ST-elevation myocardial infarction. Am J Med, 2020; 133(1):133–142; doi: 10.1016/j.amjmed.2019.06.021

45.

Lee

, Affandi

, Irish

, et al. Cytomegalovirus infection alters phenotypes of different gammadelta T-cell subsets in renal transplant recipients with long-term stable graft function. J Med Virol, 2017a;89(8):1442–1452; doi: 10.1002/jmv.24784

46.

Lee

, Bartlett

, Dwivedi

. Adaptive immune responses in human atherosclerosis. Int J Mol Sci, 2020; 21(23):9322; doi: 10.3390/ijms21239322

47.

Lee

, Brook

, Affandi

, et al. A high burden of cytomegalovirus marks poor vascular health in transplant recipients more clearly than in the general population. Clin Transl Immunology, 2019a;8(2):e1043; doi: 10.1002/cti2.1043

48.

Lee

, Chung

, Lee

. US28, a virally-encoded GPCR as an antiviral target for human cytomegalovirus infection. Biomol Ther (Seoul), 2017b;25(1):69–79.

49.

Lee

, Doualeh

, Affandi

, et al. Functional and clinical consequences of changes to natural killer cell phenotypes driven by chronic cytomegalovirus infections. J Med Virol, 2019b;91(6):1120–1127; doi: 10.1002/jmv.25401

50.

, Xia

, Hu

. Infection and atherosclerosis: TLR-dependent pathways. Cell Mol Life Sci, 2020a;77(14):2751–2769; doi: 10.1007/s00018-020-03453-7

51.

, Li

, Yang

, et al. Human cytomegalovirus infection is correlated with atherosclerotic plaque vulnerability in carotid artery. J Gene Med, 2020b:e3236; doi: 10.1002/jgm.3236

52.

, Wang

, Zhang

, et al. Onset of coronary heart disease is associated with HCMV infection and increased CD14 (+)CD16 (+) monocytes in a population of Weifang, China. Biomed Environ Sci, 2020c;33(8):573–582; doi: 10.3967/bes2020.076

53.

, Du

, Rong

, et al. Foam cells promote atherosclerosis progression by releasing CXCL12. Biosci Rep, 2020d;40(1):BSR20193267; doi: 10.1042/BSR20193267

54.

, Li

, Dai

, et al. Lipid metabolism in vascular smooth muscle cells influenced by HCMV infection. Cell Physiol Biochem, 2016a;39(5):1804–1812; doi: 10.1159/000447880

55.

, Liu

, Kang

, et al. HHEX: A crosstalker between HCMV infection and proliferation of VSMCs. Front Cell Infect Microbiol, 2016b;6:169; doi: 10.3389/fcimb.2016.00169

56.

, Wang

, Fang

, et al. Programmed cell death in atherosclerosis and vascular calcification. Cell Death Dis, 2022; 13(5):467; doi: 10.1038/s41419-022-04923-5

57.

Liu

, Zhang

. Vigilance on new-onset atherosclerosis following SARS-CoV-2 infection. Front Med (Lausanne), 2020; 7:629413; doi: 10.3389/fmed.2020.629413

58.

Liu

, Ji

, Yao

, et al. Serum Metrnl is associated with the presence and severity of coronary artery disease. J Cell Mol Med, 2019; 23(1):271–280; doi: 10.1111/jcmm.13915

59.

Lollinga

, de Wit

, Rahbar

, et al. Human cytomegalovirus-encoded receptor US28 is expressed in renal allografts and facilitates viral spreading in vitro. Transplantation, 2017; 101(3):531–540; doi: 10.1097/TP.0000000000001289

60.

, Jia

, Wan

, et al. Curcumin inhibits the formation of atherosclerosis in ApoE(-/-) mice by suppressing cytomegalovirus activity in endothelial cells. Life Sci, 2020; 257:117658; doi: 10.1016/j.lfs.2020.117658

61.

Makwana

, Waters

, Irish

, et al. Deciphering effects of uncontrolled cytomegalovirus replication on immune responses in cytomegalovirus DNA-positive renal transplant recipients. Viral Immunol, 2019; 32(8):355–360; doi: 10.1089/vim.2019.0014

62.

Merckelbach

, van der Vorst

EPC

, Kallmayer

, et al. Expression and cellular localization of CXCR4 and CXCL12 in human carotid atherosclerotic plaques. Thromb Haemost, 2018; 118(1):195–206; doi: 10.1160/TH17-04-0271

63.

Nenna

, Zhai

, Packard

, et al. High cytomegalovirus serology and subsequent COPD-related mortality: A longitudinal study. ERJ Open Res, 2020; 6(2):0062-2020; doi: 10.1183/23120541.00062-2020

64.

Nguyen

, Abdirahman

, Putoczki

. Emerging roles for Interleukin-11 in disease. Growth Factors, 2019; 37(1–2):1–11; doi: 10.1080/08977194.2019.1620227

65.

Nikitskaya

, Lebedeva

, Ivanova

, et al. Cytomegalovirus-productive infection is associated with acute coronary syndrome. J Am Heart Assoc, 2016; 5(8):e003759; doi: 10.1161/JAHA.116.003759

66.

Okba

, Abd El Raouf Raafat

, Nazmy Farres

, et al. Expanded peripheral CD4(+)CD28(null) T cells and its association with atherosclerotic changes in patients with end stage renal disease on hemodialysis. Hum Immunol, 2019; 80(9):748–754; doi: 10.1016/j.humimm.2019.03.008

67.

Olejarz

, Lacheta

, Kubiak-Tomaszewska

. Matrix metalloproteinases as biomarkers of atherosclerotic plaque instability. Int J Mol Sci, 2020; 21(11):3946; doi: 10.3390/ijms21113946

68.

Patro

ARK

. Subversion of immune response by human cytomegalovirus. Front Immunol, 2019; 10:1155; doi: 10.3389/fimmu.2019.01155

69.

Pera

, Caserta

, Albanese

, et al. CD28(null) pro-atherogenic CD4 T-cells explain the link between CMV infection and an increased risk of cardiovascular death. Theranostics, 2018; 8(16):4509–4519; doi: 10.7150/thno.27428

70.

Rodriguez-Goncer

, Fernandez-Ruiz

, Aguado

. A critical review of the relationship between post-transplant atherosclerotic events and cytomegalovirus exposure in kidney transplant recipients. Expert Rev Anti Infect Ther, 2020; 18(2):113–125; doi: 10.1080/14787210.2020.1707079

71.

Samson

, van den Berg

, Engelfriet

, et al. Limited effect of duration of CMV infection on adaptive immunity and frailty: Insights from a 27-year-long longitudinal study. Clin Transl Immunology, 2020; 9(10):e1193; doi: 10.1002/cti2.1193

72.

Schafer

, Zernecke

. CD8(+) T cells in atherosclerosis. Cells, 2020; 10(1):37; doi: 10.3390/cells10010037

73.

Shah

ASV

, Stelzle

, Lee

, et al. Global burden of atherosclerotic cardiovascular disease in people living with HIV: Systematic review and meta-analysis. Circulation, 2018; 138(11):1100–1112; doi: 10.1161/circulationaha.117.033369

74.

Shen

, Xu

, Chen

, et al. Human cytomegalovirus-encoded miR-UL112 contributes to HCMV-mediated vascular diseases by inducing vascular endothelial cell dysfunction. Virus Genes, 2018; 54(2):172–181; doi: 10.1007/s11262-018-1532-9

75.

Sillen

, Declerck

. Targeting PAI-1 in cardiovascular disease: Structural insights into PAI-1 functionality and inhibition. Front Cardiovasc Med, 2020; 7:622473; doi: 10.3389/fcvm.2020.622473

76.

Skoda

, Stangret

, Szukiewicz

. Fractalkine and placental growth factor: A duet of inflammation and angiogenesis in cardiovascular disorders. Cytokine Growth Factor Rev, 2018; 39:116–123; doi: 10.1016/j.cytogfr.2017.12.001

77.

Slot

, Kroon

, Damoiseaux

, et al. CD4(+)CD28(null) T cells are related to previous cytomegalovirus infection but not to accelerated atherosclerosis in ANCA-associated vasculitis. Rheumatol Int, 2017; 37(5):791–798; doi: 10.1007/s00296-016-3643-8

78.

Souilhol

, Harmsen

, Evans

, et al. Endothelial-mesenchymal transition in atherosclerosis. Cardiovasc Res, 2018; 114(4):565–577; doi: 10.1093/cvr/cvx253

79.

Soumya

, Unni

, Raghu

. Impact of COVID-19 on the cardiovascular system: A review of available reports. Cardiovasc Drugs Ther, 2021; 35(3):411–425; doi: 10.1007/s10557-020-07073-y

80.

Styles

, Converse

, Griffin

, et al. Human cytomegalovirus infections are associated with elevated biomarkers of vascular injury. Front Cell Infect Microbiol, 2020; 10:334; doi: 10.3389/fcimb.2020.00334

81.

Suarez-Pierre

, Giuliano

, Lui

, et al. Impact of cytomegalovirus serologic status on heart transplantation. J Card Surg, 2020; 35(7):1431–1438; doi: 10.1111/jocs.14588

82.

Theodorou

, Boon

. Endothelial cell metabolism in atherosclerosis. Front Cell Dev Biol, 2018; 6:82; doi: 10.3389/fcell.2018.00082

83.

Tian

, He

, Zhang

, et al. Role of vasodilator-stimulated phosphoprotein in human cytomegalovirus-induced hyperpermeability of human endothelial cells. Exp Ther Med, 2018; 16(2):1295–1303; doi: 10.3892/etm.2018.6332

84.

, Arnolds

, O'Connor

, et al. Human cytomegalovirus UL111A and US27 gene products enhance the CXCL12/CXCR4 signaling axis via distinct mechanisms. J Virol, 2018; 92(5):e01981-17; doi: 10.1128/JVI.01981-17

85.

Ude

, Yeh

, Shantha

. Cytomegalovirus retinitis in the highly active anti-retroviral therapy era. Ann Eye Sci, 2022; 7:5; doi: 10.21037/aes-21-18

86.

Valle Oseguera

, Spencer

. Human cytomegalovirus interleukin-10 enhances matrigel invasion of MDA-MB-231 breast cancer cells. Cancer Cell Int, 2017; 17:24; doi: 10.1186/s12935-017-0399-5

87.

van den Berg

SPH

, Pardieck

, Lanfermeijer

, et al. The hallmarks of CMV-specific CD8 T-cell differentiation. Med Microbiol Immunol, 2019; 208(3–4):365–373; doi: 10.1007/s00430-019-00608-7

88.

van den Berg

SPH

, Wong

, Hendriks

, et al. Negative effect of age, but not of latent cytomegalovirus infection on the antibody response to a novel influenza vaccine strain in healthy adults. Front Immunol, 2018; 9:82; doi: 10.3389/fimmu.2018.00082

89.

Wahlin

, Fasth

AER

, Karp

, et al. Atherosclerosis in rheumatoid arthritis: Associations between anti-cytomegalovirus IgG antibodies, CD4+CD28null T-cells, CD8+CD28null T-cells and intima-media thickness. Clin Exp Rheumatol, 2021; 39(3):578–586; doi: 10.55563/clinexprheumatol/gs3o43

90.

Wang

, Chen

. Histone deacetylase SIRT1, smooth muscle cell function, and vascular diseases. Front Pharmacol, 2020; 11:537519; doi: 10.3389/fphar.2020.537519

91.

Wang

, Peng

, Bai

, et al. Cytomegalovirus infection and relative risk of cardiovascular disease (ischemic heart disease, stroke, and cardiovascular death): A meta-analysis of prospective studies up to 2016. J Am Heart Assoc, 2017; 6(7):e005025; doi: 10.1161/JAHA.116.005025

92.

Wang

, Cai

, Zhang

, et al. Positive expression of human cytomegalovirus phosphoprotein 65 in atherosclerosis. Biomed Res Int, 2016; 2016:4067685; doi: 10.1155/2016/4067685

93.

Wanjalla

, Mashayekhi

, Bailin

, et al. Anticytomegalovirus CD4(+) T cells are associated with subclinical atherosclerosis in persons with HIV. Arterioscler Thromb Vasc Biol, 2021; 41(4):1459–1473; doi: 10.1161/ATVBAHA.120.315786

94.

Waters

, Agostino

, Lee

, et al. Sequencing directly from clinical specimens reveals genetic variations in HCMV-encoded chemokine receptor US28 that may influence antibody levels and interactions with human chemokines. Microbiol Spectr, 2021; 9(2):e0002021; doi: 10.1128/Spectrum.00020-21

95.

Waters

, Brook

, Lee

, et al. HIV patients, healthy aging and transplant recipients can reveal the hidden footprints of CMV. Clin Immunol, 2018; 187:107–112; doi: 10.1016/j.clim.2017.11.001

96.

Waters

, Lee

, Affandi

, et al. The effect of genetic variants affecting NK cell function on cardiovascular health and the burden of CMV. Hum Immunol, 2017; 78(11–12):747–751; doi: 10.1016/j.humimm.2017.10.003

97.

Waters

, Lee

, Ariyanto

, et al. Sequencing of the viral UL111a gene directly from clinical specimens reveals variants of HCMV-encoded IL-10 that are associated with altered immune responses to HCMV. Int J Mol Sci, 2022; 23(9):4644; doi: 10.3390/ijms23094644

98.

Waters

, Lee

, Lloyd

, et al. The detection of CMV in saliva can mark a systemic infection with CMV in renal transplant recipients. Int J Mol Sci, 2019; 20(20):5230; doi: 10.3390/ijms20205230

99.

Waters

, Lee

, Munyard

, et al. Human cytomegalovirus-encoded microRNAs Can be found in saliva samples from renal transplant recipients. Noncoding RNA, 2020; 6(4):50; doi: 10.3390/ncrna6040050

100.

Weber

, Kehl

, Erber

, et al. CMV seropositivity is a potential novel risk factor for severe COVID-19 in non-geriatric patients. PLoS One, 2022; 17(5):e0268530; doi: 10.1371/journal.pone.0268530

101.

Wulandari

EAT

, Wijaya

, Karim

, et al. Periodontitis and cytomegalovirus associate with atherosclerosis among HIV patients after 5years on ART. J Acquir Immune Defic Syndr, 2020; 85(2):195–200; doi: 10.1097/QAI.0000000000002417

102.

, Jiang

, Chen

, et al. Vascular macrophages in atherosclerosis. J Immunol Res, 2019; 2019:4354786; doi: 10.1155/2019/4354786

103.

Yang

, Shu

, Chen

, et al. Anti-cytomegalovirus IgG antibody titer is positively associated with advanced T cell differentiation and coronary artery disease in end-stage renal disease. Immun Ageing, 2018; 15:15; doi: 10.1186/s12979-018-0120-0

104.

, Wang

, Ye

, et al. Increased interleukin-11 levels are correlated with cardiac events in patients with chronic heart failure. Mediators Inflamm, 2019; 2019:1575410; doi: 10.1155/2019/1575410

105.

Youn

, Kim

, Jung

, et al. Analysis of cytomegalovirus-specific T-cell responses in patients with hypertension: Comparison of assay methods and antigens. Clin Hypertens, 2018; 24:5; doi: 10.1186/s40885-018-0090-8

106.

, Youn

, Kim

, et al. Arterial stiffness is associated with cytomegalovirus-specific senescent CD8(+) T cells. J Am Heart Assoc, 2017; 6(9):e006535; doi: 10.1161/JAHA.117.006535

107.

Zhang

, Qin

, Zhou

, et al. Analysis of genes and underlying mechanisms involved in foam cells formation and atherosclerosis development. PeerJ, 2020a;8:e10336; doi: 10.7717/peerj.10336

108.

Zhang

, Yu

, Liu

. MicroRNAs expressed by human cytomegalovirus. Virol J, 2020b;17(1):34; doi: 10.1186/s12985-020-1296-4

109.

Zhang

, Liu

, Wang

, et al. miR-138 promotes migration and tube formation of human cytomegalovirus-infected endothelial cells through the SIRT1/p-STAT3 pathway. Arch Virol, 2017; 162(9):2695–2704; doi: 10.1007/s00705-017-3423-0

110.

Zhang

, Tang

, Xi

, et al. Human cytomegalovirus promoting endothelial cell proliferation by targeting regulator of G-protein signaling 5 hypermethylation and downregulation. Sci Rep, 2020c;10(1):2252; doi: 10.1038/s41598-020-58680-6

111.

Zhen

, Zeng

, Zheng

, et al. Human cytomegalovirus infection is associated with stroke in women: The US National Health and Nutrition Examination Survey 1999–2004. Postgrad Med J, 2022; 98(1157):172–176; doi: 10.1136/postgradmedj-2020-139201

112.

Zhou

, Xi

, Shi

, et al. MicroRNA19293p participates in murine cytomegalovirusinduced hypertensive vascular remodeling through Ednra/NLRP3 inflammasome activation. Int J Mol Med, 2021; 47(2):719–731; doi: 10.3892/ijmm.2020.4829

113.

Zhu

, Zhang

, Wang

. Vitamin D3 suppresses human cytomegalovirus-induced vascular endothelial apoptosis via rectification of paradoxical m6a modification of mitochondrial calcium Uniporter mRNA, which is regulated by METTL3 and YTHDF3. Front Microbiol, 2022; 13:861734; doi: 10.3389/fmicb.2022.861734

114.

Zuhair

, Smit

GSA

, Wallis

, et al. Estimation of the worldwide seroprevalence of cytomegalovirus: A systematic review and meta-analysis. Rev Med Virol, 2019; 29(3):e2034; doi: 10.1002/rmv.2034