Decreased inducibility of TNF expression in lipid-loaded macrophages

BACKGROUND: Inflammation and immune responses are considered to be very important in the pathogenesis of atherosclerosis. Lipid accumulation in macrophages of the arterial intima is a characteristic feature of atherosclerosis which can influence the inflammatory potential of macrophages. We studied the effects of lipid loading on the regulation of TNF expression in human monocyte-derived macrophages. RESULTS: In macrophages incubated with acetylated low density lipoprotein (ac-LDL) for 2 days, mRNA expression of TNF in cells stimulated with TNF decreased by 75%. In cell cultures stimulated over night with IL-1β, lipid loading decreased secretion of TNF into culture medium by 48%. These results suggest that lipid accumulation in macrophages makes them less responsive to inflammatory stimuli. Decreased basal activity and inducibility of transcription factor AP-1 was observed in lipid-loaded cells, suggesting a mechanism for the suppression of cytokine expression. NF-κB binding activity and inducibility were only marginally affected by ac-LDL. LDL and ac-LDL did not activate PPARγ. In contrast, oxidized LDL stimulated AP-1 and PPARγ but inhibited NF-κB, indicating that the effects of lipid loading with ac-LDL were not due to oxidation of lipids. CONCLUSIONS: Accumulation of lipid, mainly cholesterol, results in down-regulation of TNF expression in macrophages. Since monocytes are known to be activated by cell adhesion, these results suggest that foam cells in atherosclerotic plaques may contribute less potently to an inflammatory reaction than newly arrived monocytes/macrophages

Activation of calpain-1 in human carotid artery atherosclerotic lesions

<p>Abstract</p> <p>Background</p> <p>In a previous study, we observed that oxidized low-density lipoprotein-induced death of endothelial cells was calpain-1-dependent. The purpose of the present paper was to study the possible activation of calpain in human carotid plaques, and to compare calpain activity in the plaques from symptomatic patients with those obtained from patients without symptoms.</p> <p>Methods</p> <p>Human atherosclerotic carotid plaques (n = 29, 12 associated with symptoms) were removed by endarterectomy. Calpain activity and apoptosis were detected by performing immunohistochemical analysis and TUNEL assay on human carotid plaque sections. An antibody specific for calpain-proteolyzed α-fodrin was used on western blots.</p> <p>Results</p> <p>We found that calpain was activated in all the plaques and calpain activity colocalized with apoptotic cell death. Our observation of autoproteolytic cleavage of the 80 kDa subunit of calpain-1 provided further evidence for enzyme activity in the plaque samples. When calpain activity was quantified, we found that plaques from symptomatic patients displayed significantly lower calpain activity compared with asymptomatic plaques.</p> <p>Conclusion</p> <p>These novel results suggest that calpain-1 is commonly active in carotid artery atherosclerotic plaques, and that calpain activity is colocalized with cell death and inversely associated with symptoms.</p

Inflammatory effects of very low-density lipoprotein and fatty acids.

High plasma triacylglycerol (triglyceride, TG) levels is a risk factor for atherosclerosis. Very large lipoproteins, such as chylomicrons, alone are not considered atherogenic, but TG-rich remnant lipoproteins can penetrate into the vascular wall. Importantly, accumulating evidence suggests that all TG-rich lipoproteins stimulate cytokine expression in circulating monocytes. Very low-density lipoprotein (VLDL) stimulates monocyte adhesion to endothelial cells and expression of inflammatory genes in macrophages. Furthermore, fatty acids released from large lipoproteins can stimulate both vascular cells and circulating monocytes. It is likely that fatty acids released from TG-rich lipoproteins contribute to atherogenesis, but the role of fatty acids in ischemic heart disease is not as direct as that of cholesterol. Fatty acids influence plasma lipoprotein levels and either stimulate or suppress numerous cellular functions relevant to atherogenesis. While certain n-3 fatty acids are good for health, most other medium- to long-chain fatty acids appear to promote inflammation in cell culture studies and need to be studied further. Nevertheless, the existing evidence supports the general conclusion that TG-rich lipoproteins and fatty acids greatly accelerate the progression of atherosclerosis. This may be because of their inflammatory effects

Very low density lipoprotein potentiates tumor necrosis factor-α expression in macrophages

High levels of the triacylglycerol-rich lipoproteins, very low density lipoprotein (VLDL) and intermediate density lipoprotein (IDL) have been identified as independent risk factors for coronary heart disease, and inflammation is thought to contribute to atherosclerosis and its complications. To understand how dyslipidemia promotes inflammation, we have characterised the effects of VLDL treatment on production of tumor necrosis factor-α (TNF) by human monocyte-derived macrophages. VLDL strongly potentiated lipopolysaccharide (LPS)-induced expression of TNF mRNA and secretion of TNF protein. VLDL activated mitogen-activated protein kinase-ERK kinase 1/2 (MEK1/2), and potentiated LPS-induced MEK1/2 activation. The MEK1/2 inhibitor U0126 strongly diminished TNF expression, indicating that MEK1/2 plays a central role in the regulation of TNF expression. VLDL did not activate transcription factors NF-κB and PPAR-γ, but it activated AP-1 at least as potently as LPS, and potentiated LPS-induced activation of AP-1. The inhibitor U0126 completely prevented this potentiation. Inhibition of AP-1 by decoy oligonucleotides abolished potentiation of TNF secretion by VLDL. In conclusion, VLDL treatment potentiates TNF expression in macrophages by activation of MEK1/2 and AP-1. These findings suggest that triacylglycerol-rich lipoproteins are involved in inflammatory processes associated with atherosclerosis

Identification of immune responses against aldehyde-modified peptide sequences in apolipoprotein B associated with cardiovascular disease

Objective- Atherosclerosis is associated with an immune response against oxidized LDL, which modulates the progression of the disease process. Methods and Results- Using a library of polypeptides covering the complete sequence of apoB-100, the only major protein of LDL, we have identified over 100 different human antibodies reacting against aldehyde-modified apoB-100 sequences. IgM antibody titer levels decreased with age and were associated with the intima-media thickness of the carotid artery in subjects younger than 60 years. There were also inverse associations between IgM levels and oxidized LDL in plasma. In prospective clinical studies, antibody levels against several aldehyde-modified apoB-100 sites were associated with cardiovascular disease in this age group. Whether this immune response is adaptive (protective) or maladaptive (causal) in atherosclerosis requires further investigation. Conclusions- We have characterized a large number of epitopes within the apoB-100 component of oxidized LDL that provoke an immune response in humans. These findings will make it possible to study the role of immune responses against specific sites in oxidized LDL and to determine whether these immune responses influence the risk for future cardiac events