Toll-Like Receptor-2 Mediates Diet and/or Pathogen Associated Atherosclerosis: Proteomic Findings

BACKGROUND. Accumulating evidence implicates a fundamental link between the immune system and atherosclerosis. Toll-like receptors are principal sensors of the innate immune system. Here we report an assessment of the role of the TLR2 pathway in atherosclerosis associated with a high-fat diet and/or bacteria in ApoE+/- mice. METHODS AND RESULTS. To explore the role of TLR2 in inflammation- and infection-associated atherosclerosis, 10 week-old ApoE+/--TLR2+/+, ApoE+/--TLR2+/- and ApoE+/--TLR2-/- mice were fed either a high fat diet or a regular chow diet. All mice were inoculated intravenously, once per week for 24 consecutive weeks, with 50 μl live Porphyromonas gingivalis (P.g) (107 CFU) or vehicle (normal saline). Animals were euthanized 24 weeks after the first inoculation. ApoE+/--TLR2+/+ mice showed a significant increase in atheromatous lesions in proximal aorta and aortic tree compared to ApoE+/--TLR2+/- and ApoE+/--TLR2-/- mice for all diet conditions. They also displayed profound changes in plaque composition, as evidenced by increased macrophage infiltration and apoptosis, increased lipid content, and decreased smooth muscle cell mass, all reflecting an unstable plaque phenotype. SAA levels from ApoE+/--TLR2+/+ mice were significantly higher than from ApoE+/--TLR2+/- and ApoE+/--TLR2-/- mice. Serum cytokine analysis revealed increased levels of pro-inflammatory cytokines in ApoE+/--TLR2+/+ mice compared to ApoE+/--TLR2+/- and TLR2-/- mice, irrespective of diet or bacterial challenge. ApoE+/--TLR2+/+ mice injected weekly for 24 weeks with FSL-1 (a TLR2 agonist) also demonstrated significant increases in atherosclerotic lesions, SAA and serum cytokine levels compared to ApoE+/--TLR2-/- mice under same treatment condition. Finally, mass-spectrometry (MALDI-TOF-MS) of aortic samples analyzed by 2-dimentional gel electrophoresis differential display, identified 6 proteins upregulated greater than 2-fold in ApoE+/--TLR2+/+ mice fed the high fat diet and inoculated with P.g compared to any other group. CONCLUSION. Genetic deficiency of TLR2 reduces diet- and/or pathogen-associated atherosclerosis in ApoE+/- mice, along with differences in plaque composition suggesting greater structural stability while TLR-2 ligand-specific activation triggers atherosclerosis. The present data offers new insights into the pathophysiological pathways involved in atherosclerosis and paves the way for new pharmacological interventions aimed at reducing atherosclerosis.National Heart, Lung, and Blood Institute (R01 HL076801

Vascular Remodeling in Health and Disease

The term vascular remodeling is commonly used to define the structural changes in blood vessel geometry that occur in response to long-term physiologic alterations in blood flow or in response to vessel wall injury brought about by trauma or underlying cardiovascular diseases.1, 2, 3, 4 The process of remodeling, which begins as an adaptive response to long-term hemodynamic alterations such as elevated shear stress or increased intravascular pressure, may eventually become maladaptive, leading to impaired vascular function. The vascular endothelium, owing to its location lining the lumen of blood vessels, plays a pivotal role in regulation of all aspects of vascular function and homeostasis.5 Thus, not surprisingly, endothelial dysfunction has been recognized as the harbinger of all major cardiovascular diseases such as hypertension, atherosclerosis, and diabetes.6, 7, 8 The endothelium elaborates a variety of substances that influence vascular tone and protect the vessel wall against inflammatory cell adhesion, thrombus formation, and vascular cell proliferation.8, 9, 10 Among the primary biologic mediators emanating from the endothelium is nitric oxide (NO) and the arachidonic acid metabolite prostacyclin [prostaglandin I2 (PGI2)], which exert powerful vasodilatory, antiadhesive, and antiproliferative effects in the vessel wall

Pathways from rural schools : does school VET make a difference?

This paper reports results from an NREC funded study of the medium term outcomes for youth and their communities of VET programs delivered by rural schools. The key findings relate to the outcomes of school VET programs in terms of education and training, employment, and community outcomes such as rural youth retention, and to the features that contribute to successful outcomes. Rural school VET students are less likely to continue with post school education and training in general, but more likely to go onto further vocational education and training than school non-VET students. The study reports strong links between industry area of school VET course, and further education and training, or employment, in the same industry area. It also indicates that school VET students are more likely to live in a rural area at some time during their working life. The features of rural school VET programs that influence outcomes include the purpose of the school VET program and work placements. Student motivation and gender also influence outcomes for individual students. Rural school VET courses represent pathways to related education and training for students who intend to live in a rural area during their working life, and for those who do not.<br /

Human Leukemia Inhibitory Factor Inhibits Development of Experimental Atherosclerosis

The effect of human leukemia inhibitory factor (hLIF) on the development of atherosclerosis was investigated in an experimental animal model. Two conditions were examined: one in which lesions could arise because of the influence of both 'injury' (cuffed vessel) and diet and one in which only the effect of diet could be significant in other areas of the vasculature (aorta). At time zero, the right carotid artery of rabbits (n = 32) was ensheathed in a soft Silastic cuff, and an osmotic minipump (2-mL capacity; 2.5 μL/h: 28 days) containing either hLIF or saline was inserted into the peritoneal cavity. Rabbits were divided into four groups (n=8): group 1 received normal diet/saline; group 2, normal diet/LIF (30 μg · kg · d); group 3, 1% cholesterol diet/saline; and group 4, 1% cholesterol diet/LIF (30 μg · kg · d). After 28 days, the cholesterol diet (group 3) resulted in a sixfold increase in plasma cholesterol level compared with group 1 rabbits on a normal diet (3.80 ± 0.50 versus 0.55 ± 0.01 mmol/L). This was significantly lower (P = .01) with hLIF treatment in group 4 rabbits (2.80 ± 0.44 mmol/L). Group 2 rabbits had higher aortic tissue cholesterol levels (1.40 ± 0.35 mg/g) compared with group 1 rabbits on a normal diet (0.10 ± 0.06 mg/g) (P = .01), whereas hLIF treatment decreased tissue cholesterol levels by 60% in group 4 rabbits (0.60 ± 0.05 mg/g) (P = .01). Group 3 rabbits developed lipid-filled lesions covering 63.25 ± 17.66% of the thoracic aorta surface, whereas lesions were significantly reduced (9.88 ± 8.79%) (P = .01) with LIF treatment (group 4). No thoracic aorta lesions were present in groups 1 and 2. Cuff placement around the right carotid artery in groups 1 and 3 resulted in neointimal formation (as percent total wall cross-sectional area) of 15.06 ± 8.83% and 19.30 ± 5.36%, respectively. In LIF-treated animals (groups 2 and 4), neointima composed only 2.12 ± 5.45% and 2.09 ± 4.32%, respectively, which represented a significant reduction (P = .01). Immunocytochemical analysis revealed no subendothelial macrophages in rabbits fed a normolipidic diet with or without hLIF treatment, but 153.60 ± 15.60 and 165.30 ± 22.40 (P = .677) macrophages per ring segment were found in animals fed a cholesterol diet in the presence and absence of hLIF, respectively. Together these results show that hLIF lowers plasma cholesterol and inhibits the development of aortic fatty streaks in cholesterol-fed rabbits. hLIF also inhibits the development of cuff-induced carotid neointimal thickening in rabbits fed either a normal or cholesterol diet but does not affect monocyte invasion of the vessel wall

Divergent effects of matrix metalloproteinases 3, 7, 9, and 12 on atherosclerotic plaque stability in mouse brachiocephalic arteries

Matrix metalloproteinases (MMPs) are thought to be involved in the growth, destabilization, and eventual rupture of atherosclerotic lesions. Using the mouse brachiocephalic artery model of plaque instability, we compared apolipoprotein E (apoE)/MMP-3, apoE/MMP-7, apoE/MMP-9, and apoE/MMP-12 double knockouts with their age-, strain-, and sex-matched apoE single knockout controls. Brachiocephalic artery plaques were significantly larger in apoE/MMP-3 and apoE/MMP-9 double knockouts than in controls. The number of buried fibrous layers was also significantly higher in the double knockouts, and both knockouts exhibited cellular compositional changes indicative of an unstable plaque phenotype. Conversely, lesion size and buried fibrous layers were reduced in apoE/MMP-12 double knockouts compared with controls, and double knockouts had increased smooth muscle cell and reduced macrophage content in the plaque, indicative of a stable plaque phenotype. ApoE/MMP-7 double knockout plaques contained significantly more smooth muscle cells than controls, but neither lesion size nor features of stability were altered in these animals. Hence, MMP-3 and MMP-9 appear normally to play protective roles, limiting plaque growth and promoting a stable plaque phenotype. MMP-12 supports lesion expansion and destabilization. MMP-7 has no effect on plaque growth or stability, although it is associated with reduced smooth muscle cell content in plaques. These data demonstrate that MMPs are directly involved in atherosclerotic plaque destabilization and clearly show that members of the MMP family have widely differing effects on atherogenesis