Toll-Like Receptor-4 Is Expressed by Macrophages in Murine and Human Lipid-Rich Atherosclerotic Plaques and Upregulated by Oxidized LDL

Abstract

Background-Inflammation is implicated in atherogenesis and plaque disruption. Toll-like receptor 2 (TLR-2) and TLR-4, a human homologue of drosophila Toll, play an important role in the innate and inflammatory signaling responses to microbial agents. To investigate a potential role of these receptors in atherosclerosis, we assessed the expression of TLR-2 and TLR-4 in murine and human atherosclerotic plaques. Methods and Results-Aortic root lesions of high-fat diet-fed apoE-deficient mice (nϭ5) and human coronary atherosclerotic plaques (nϭ9) obtained at autopsy were examined for TLR-4 and TLR-2 expression by immunohistochemistry. Aortic atherosclerotic lesions in all apoE-deficient mice expressed TLR-4, whereas aortic tissue obtained from control C57BL/6J mice showed no TLR-4 expression. All 5 lipid-rich human plaques expressed TRL-4, whereas the 4 fibrous plaques and 4 normal human arteries showed no or minimal expression. Serial sections and double immunostaining showed TLR-4 colocalizing with macrophages both in murine atherosclerotic lesions and at the shoulder region of human coronary artery plaques. In contrast to TLR-4, none of the plaques expressed TLR-2. Furthermore, basal TLR-4 mRNA expression by human monocyte-derived macrophages was upregulated by ox-LDL in vitro. A potential role for infection in the development of atherosclerosis has been considered for several decades, but interest in this topic has recently reemerged because of several recent observations. Accumulating evidence has implicated specific infectious agents, including Chlamydia pneumoniae, in the progression and/or destabilization of atherosclerosis. [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17] Recent studies suggest that chlamydia lipopolysaccharide (LPS) induces foam-cell formation, whereas its heat-shock protein (chlamydia HSP60) induces oxidative modification of LDL. 5,18 Chlamydia HSP60 has been implicated in the induction of deleterious immune responses in human chlamydial infection and has been found to colocalize with infiltrating macrophages in the atheroma lesions. 19 Collectively, these data support a potential role for C pneumoniae in the development and progression of atherosclerosis and suggest that this organism may indeed play an active role in atheroma development. Available data, however, also underscore the current lack of a complete understanding of the molecular mechanisms that link C pneumoniae infection to innate immunity and trigger the signals for enhanced inflammation and atherogenesis. Conclusions-OurLPS, a major component of the outer surface of Gramnegative bacteria, activates the proinflammatory transcription factor nuclear factor (NF)-B in endothelial cells and macrophages. 20,21 Recently, human Toll-like receptor-4 (TLR-4), a human homologue of drosophila Toll, has been identified as Currently, more than 10 human TLRs have been identified, and at least 10 human homologues of drosophila Toll have been sequenced. Whereas TLR-4 is used by enteric Gramnegative bacteria and LPS, TLR-2 is use...

Synergistic Proinflammatory Effects of the Antiviral Cytokine Interferon-α and Toll-Like Receptor 4 Ligands in the Atherosclerotic Plaque

“…IFN-␣-facilitated recognition of TLR4 ligands may affect inflammatory activation not only in response to lipopolysaccharide and other microbial molecules 33 but also to (modified) endogenous molecules including heat-shock proteins, modified lipids, fibronectin, biglycan, and hyaluronan oligosaccharides, all abundantly present in the atherosclerotic lesion microenvironment. 14,[17][18][19] Amplified effector cytokine production has severe consequences for atherosclerotic plaque stability. 34 TNF-␣ destabilizes plaque tissue integrity by promoting macrophageinduced vascular smooth muscle cell apoptosis.…”

“…8 Most likely, simultaneous triggering of different TLRs is caused by a combination of exogenous and endogenous stimuli available in the plaque. Viral infection inducing IFN-␣ production should be sufficient to intensify and sustain endogenous TLR4 triggering, transforming chronic lowgrade inflammation in the atherosclerotic lesion 14,19 into acute inflammatory activation with abundant effector molecule production, potentially leading to plaque rupture.…”

“…Circulating monocytes overexpress TLR4 during acute coronary syndrome, 12 and TLR4 is expressed in atherosclerotic plaques. 13,14 Apolipoprotein E knockout mice lacking TLR4 develop reduced atherosclerotic lesions, 15 and a TLR4 genetic variation is associated with diminished risk of atherosclerosis. 16 Apart from microbial molecules such as lipopolysaccharides, autoantigens found in the plaque environment are also candidates for TLR4-dependent activation of APCs.…”

“…16 Apart from microbial molecules such as lipopolysaccharides, autoantigens found in the plaque environment are also candidates for TLR4-dependent activation of APCs. 14,[17][18][19] When activated by TLR ligands, macrophages and mDCs release powerful proinflammatory cytokines, such as TNF-␣ and interleukin (IL)-12. TNF-␣ activates every cellular component of the atheroma.…”

The Role of Human Antigen R, an RNA-binding Protein, in Mediating the Stabilization of Toll-Like Receptor 4 mRNA Induced by Endotoxin

“…7 The functional expression of TLR4 and subsequent augmentation of intimal hyperplasia have recently been described. 8 Hypoxia diminishes TLR4 expression through reactive oxygen species (ROS) generated by mitochondria.…”