Cardiovascular disease is the leading cause of morbidity and mortality in the developed world. Lipids have traditionally and solidly been linked to atherogenesis, but only lately has the role of inflammation in atheroma formation been given the attention it deserves.1–2 Clinical studies have provided the first evidence implicating inflammation in atherosclerosis. The experimental studies that followed have established the inflammatory nature of arterial lesions. Much research has been carried out on the diagnostic and clinical value of specific serum or tissue inflammatory biomarkers. Finally, the potential for medical preventative or therapeutic intervention in the inflammatory component of arterial disease is being fervently investigated.
Clinical Studies
All of the clinical manifestations of atherosclerosis have traditionally been associated with inflammation of the plaque. Acute coronary syndromes are usually the result of plaque rupture. Activated macrophages, T cells and mast cells at sites of rupture produce numerous inflammatory molecules and proteolytic enzymes that can destabilise lesions. They lyse cap collagen and halt collagen synthesis, inhibiting the formation of stable fibrous caps, and initiate thrombus formation. Unstable angina (UA) is associated with systemic inflammation and with expansion of interferon-(gamma)-producing T lymphocytes. UA is associated with the emergence of monoclonal T-cell populations. The unstable – but not the stable – plaque is invaded by clonally expanded T cells, suggesting a direct involvement of these lymphocytes in plaque disruption.3 Chronic low-grade inflammation documented by high levels of inflammation-sensitive plasma proteins is associated with the fatality of future coronary events. Men who have been exposed to a low-grade inflammation many years earlier have a higher proportion of coronary heart disease deaths and less nonfatal myocardial infarction.4
Carotid atherosclerosis and peripheral arterial disease have also been associated with inflammation and Chlamydia pneumoniae (Cp) infection. It seems that hypertensive, hypercholesterolaemic men are more likely to have Cp-infected carotid plaques and that these plaques have higher tumour necrosis factor-α TNF-α concentrations. Transient ischaemic attacks and strokes were strongly correlated with both Cp infection and TNF-α concentration of the atheroma.5 In our study of claudicants, we found that claudication was more severe in patients with elevated C-reactive protein (CRP) serum levels and titres of immunoglobin A (IgA) antibodies against Cp.6 CRP and anti-Cp IgA antibodies were inversely associated with the ankle-brachial index and the initial and absolute claudication distances.
Experimental Studies
Most evidence in favour of a significant role for inflammation in atherosclerosis is the product of studies in animal models using apolipoprotein E (apoE-/-) or low-density lipoprotein (LDL) receptor (LDL-R-/-)-deficient mice, which characteristically develop accelerated atherosclerosis with extensive lipid deposition. These atheromas have a high content of T cells. Whether plaque T-cell infiltration is a result or an integral part of atherosclerosis has been investigated by crossbreeding transgenic mice. Severe combined immunodeficiency mice lack T and B cells. Crossbreeding of these mice with apoE-/- mice led to a significant reduction of fatty streak lesions within the aorta. Reconstitution of the T-cell population by transfer of CD4+ T cells increased atherosclerotic lesion development by 164%.7 Proinflammatory cytokines also play a major role in plaque inflammation. Interferon-ψ (IFN-ψ) is mainly secreted by the T cells. Systemic application of IFN-ψ to apoE-/- mice has been shown to further accelerate atherosclerosis.8 Several interleukins (IL-12, IL-18) likewise enhance atherosclerotic lesions in apoE-/- mice. Another cytokine, TNF-α, is upregulated in atheromatic plaques. Blocking of TNF-α activity, or disruption of its gene expression, has led to diminished atherosclerosis development in apoE-/- mice.9 All of this evidence clearly shows that T cells and cytokines have a role to play in lesion progression.
Macrophages, like T cells, enter the vessel wall during atherogenesis, with the valuable aid of adhesion molecules and chemokines expressed on endothelial cells. Gene therapy directed against monocyte chemoattractant protein-1 (MCP-1 ) or lack of the corresponding receptor have minimised the progression of atherosclerosis in apoE-/- mice. When activated by T cells, many genes are induced in macrophages, leading to the production of tissue factor, cytokines and matrix metalloproteinases (MMPs). All of these molecules are associated with lesion progression and plaque instability. Crossbreeding op/op mice (which lack functional macrophages) with apoE-/- mice has elucidated the key role of macrophages in atherosclerosis. ApoE-/- mice with defective macrophages exhibited significantly reduced atherosclerotic plaques.10
Inflammatory Biomarkers
Research over the last 20 years has identified several inflammatory mediators that are involved in the atherosclerotic process. Many of these molecules can be measured systemically by sensitive assays, and their diagnostic and prognostic significance is an object of intensive research. The elevated serum levels of a few of them have been shown to be associated with future cardiovascular (CV) events.
CRP is the most important serum marker of inflammation because of its analytical stability, reproducibility of results and commercial availability of high-sensitivity assays. Results from more than 25 prospective studies have been reported and the vast majority of them clearly demonstrated a significant and independent association between high serum values of CRP and future CV events. A meta-analysis summarising the results of 14 prospective studies with a total of 2,557 cases and a mean follow-up period of eight years has found that people with a CRP value in the top tertile of the distribution had a relative risk of coronary heart disease of 1.9 compared with those whose CRP concentration was in the bottom tertile.11 The results of the Reykjavik study, a large prospective cohort, were not so strong, but still confirmed the predictive power of CRP (odds ratio (OR), 1.45).12 A subsequent meta-analysis of 22 population-based studies, including over 7,000 patients with incident coronary events, had similar results.12 The Cardiovascular Health Study found that elevated CRP levels improved the predictive power of the Framingham risk score.13
IL-6 is the principal procoagulant cytokine, but its most important function is the amplification of the inflammatory cascade. In a murine model of atherosclerosis, injection of excessive amounts of recombinant IL-6 resulted in enhanced fatty lesion development.14 Several prospective studies have consistently shown that IL-6 is a potent predictor of future CV end-points for the general population.15 IL-18 is also implicated in atherogenesis. TNF-α, lipoprotein-associated phospholipase A2, oxidised LDL, MCP-1 and many other inflammatory mediators are being investigated for their role in the atherosclerotic process.
Therapeutic Implications
There are examples of unexpected anti-inflammatory effects of existing therapies. Inhibition of the renin-angiotensin system is one of the main points of antihypertensive action. Inhibitors of angiotensin-converting enzyme (ACE) have been found to exert beneficial effects on plaque progression.16 One identified mechanism was the reduction in MCP-1 expression and concomitant macrophage plaque infiltration. Reduced MCP-1 levels have also been found in patients with MI treated with ACE inhibitors. Irbesartan, an inhibitor of the angiotensin II pathway, was used to treat patients with symptomatic carotid artery disease for four months before endarterectomy.17 Plaque analysis revealed decreased T-cell and macrophage inflammation and inhibited MMP activity in comparison with controls.
Statins are the cornerstone of treatment for cardiovascular disease. Their action is mainly due to serum cholesterol reduction. Statins can also prevent or even reverse ongoing inflammation and tissue damage. They have been found to inhibit the inflammatory activity of macrophages.18 In heritable hyperlipidaemic rabbits, cerivastatin decreased tissue factor (TF) and MMP expression in atheroma-associated macrophages.19 In the Atorvastatin and Thrombogenicity of the Carotid Atherosclerotic Plaque (ATROCAP) study, intermittent atorvastatin treatment between staged bilateral carotid endarterectomy reduced TF plaque activity when comparing specimens of the first and the second operation.20 In a retrospective study, it was shown that patients on statin treatment operated for symptomatic carotid stenoses and had a reduced in-hospital mortality and ischaemic stroke rate compared with patients who were not on statins.21 Carotid endarterectomy specimens from patients treated with statins had less macrophage and T-cell inflammation, reduced MMP immunoreactivity, increased expression of MMP tissue inhibitor (TIMP) and a higher collagen content.22
Apart from the existing therapies, new anti-inflammatory drugs are under investigation for use in patients with cardiovascular disease. Inhibitors of cyclo-oxygenase (COX), suppression of cytokine secretion or action, MMP inhibition and cannabinoid derivatives may all contribute to anti-atherosclerotic therapy in the future. COX-2 receptor is induced at sites of inflammation and is expressed in human atherosclerotic lesions.23 Aspirin, the oldest and most widely used COX inhibitor, has distinct anti-inflammatory actions that are independent and go beyond its antithrombotic effect.24 Unfortunately, the use of newer and COX-2-specific inhibitors are associated with increased cardiovascular risk.25 Suppressors of cytokine-signalling proteins could block cytokine-induced chronic inflammation of the vessel wall.26 MMPs contribute to atherosclerosis and destabilisation of plaques.27 Pre-treatment of patients scheduled for carotid endarterectomy with doxycycline, a nonselective MMP inhibitor, reduced MMP-1, but not MMP-2 and MMP-9, expression in carotid specimens.28 Finally, it was recently shown that the main cannabinoid receptor (CB2) is expressed in atherosclerotic plaques of humans and mice and that oral administration of low doses of tetrahydrocannabinol in mice inhibited atherosclerosis progression.29
Conclusions
New knowledge about inflammation in atherosclerosis has broadened our insight into its pathogenesis, provided us with new opportunities for diagnosis and cardiovascular risk prediction and may lead in the future to new alternative treatments for the most lethal disease of our times.