In human plasma, platelet activating factor (PAF)-degrading acetylhydrolase (acetylhydrolase) is principally transported in association with LDLs and HDLs; this enzyme hydrolyzes PAF and short-chain forms of oxidized phosphatidylcholine, transforming them into lyso-PAF and lysophosphatidylcholine, respectively. We have examined the distribution, catalytic characteristics, and transfer of acetylhydrolase activity among plasma lipoprotein subspecies separated by isopycnic density gradient ultracentrifugation; the possibility that the plasma enzyme may be partially derived from adherent monocytes has also been evaluated. In normolipidemic subjects with Lp(a) levels < 0.1 mg/mL, acetylhydrolase was associated preferentially with small, dense LDL particles (LDL-5; d = 1.050 to 1.063 g/mL) and with the very-high-density lipoprotein-1 subfraction (VHDL-1; d = 1.156 to 1.179 g/mL), representing 23.9 +/- 1.7% and 20.6 +/- 3.2%, respectively, of total plasma activity. The apparent Km values for PAF of the enzyme associated with such lipoproteins were 89.7 +/- 23.4 and 34.8 +/- 4.5 mumol/L for LDL-5 and VHDL-1, respectively: indeed, the Km value for LDL-5 was some 10-fold higher than that of the light LDL-1, LDL-2, and LDL-3 subspecies, whereas the Km of VHDL-1 was some twofold greater than those of the HDL-2 and HDL-3 subspecies. Furthermore, when expressed on the basis of unit plasma volume, the Vmax of the acetylhydrolase associated with LDL-5 was some 150-fold greater than that in LDL-1 (d = 1.019 to 1.023 g/mL). No significant differences in the pH dependence of enzyme activity or in sensitivity to protease inactivation, sulfydryl reagents, the serine protease inhibitor Pefabloc, or the PAF antagonist CV 3988 could be detected between apo B-containing and apo A-I-containing lipoprotein particle subspecies. Incubation of LDL-1 (Km = 8.4 +/- 2.6 mumol/L) and LDL-2 (d = 1.023 to 1.029 g/mL; Km = 8.4 +/- 3.3 mumol/L) subspecies with LDL-5, in which acetylhydrolase had been inactivated by pretreatment with Pefabloc, demonstrated preferential transfer of acetylhydrolase to LDL-5. Acetylhydrolase transferred to LDL-5 from the light LDL subspecies exhibited a Km of 9.4 +/- 2.2 mumol/L, a value characteristic of the particle donors. Finally, acetylhydrolase (Km = 23.4 +/- 7.6 mumol/L) released by adherent human monocytes in culture was found to bind preferentially to small, dense LDL subspecies upon incubation of Pefabloc-inactivated plasma with monocyte supernatant.(ABSTRACT TRUNCATED AT 400 WORDS)

Background-Obesity-associated dyslipidemia in humans is associated with increased low-density lipoprotein (LDL) oxidation. Mice with combined leptin and LDL receptor deficiency are obese and show severe dyslipidemia and insulin resistance. We investigated the association between oxidation of apolipoprotein B-containing lipoproteins, high-density lipoprotein (HDL) antioxidant defense, and atherosclerosis in these mice. Methods and Results-LDL receptor knockout (LDLR Ϫ/Ϫ ), leptin-deficient (ob/ob), double-mutant (LDLR Ϫ/Ϫ ;ob/ob), and C57BL6 mice were fed standard chow. Double-mutant mice had higher levels of non-HDL (PϽ0.001) and HDL (PϽ0.01) cholesterol and of triglycerides (PϽ0.001). They also had higher oxidative stress, evidenced by higher titers of autoantibodies against malondialdehyde-modified LDL (PϽ0.001). C57BL6 and ob/ob mice had no detectable lesions. Lesions covered 20% of total area of the thoracic abdominal aorta in double-mutant mice compared with 3.5% in LDLR Ϫ/Ϫ mice (PϽ0.01). Higher macrophage homing and accumulation of oxidized apolipoprotein B-100 -containing lipoproteins were associated with larger plaque volumes in the aortic root of double-mutant mice (PϽ0.01). The activity of the HDL-associated antioxidant enzymes paraoxonase and lecithin:cholesterol acyltransferase (LCAT) (ANOVA; PϽ0.0001 for both) was lower in double-mutant mice. Adenovirus-mediated LCAT gene transfer in double-mutant mice increased plasma LCAT activity by 64% (PϽ0.01) and reduced the titer of autoantibodies by 40% (PϽ0.01) and plaque volume in the aortic root by 42% (PϽ0.05) at 6 weeks. Conclusions-Dyslipidemia and insulin resistance in obese LDL receptor-deficient mice are associated with increased oxidative stress and impaired HDL-associated antioxidant defense, evidenced by decreased paraoxonase and LCAT activity. Transient LCAT overexpression was associated with a reduction of oxidative stress and atherosclerosis.

Gene transfer of PAF-AH inhibited injury-induced neointima formation and spontaneous atherosclerosis in apolipoprotein E-deficient mice. Our data indicate that PAF-AH, by reducing oxidized lipoprotein accumulation, is a potent protective enzyme against atherosclerosis.

Background-Weight loss in obese insulin-resistant but not in insulin-sensitive persons reduces coronary heart disease risk. To what extent changes in gene expression are related to atherosclerosis and cardiovascular function is unknown. Methods and Results-We studied the effect of diet restriction-induced weight loss on gene expression in the adipose tissue, the heart, and the aortic arch and on atherosclerosis and cardiovascular function in mice with combined leptin and LDL-receptor deficiency. Obesity, hypertriglyceridemia, and insulin resistance are associated with hypertension, impaired left ventricular function, and accelerated atherosclerosis in those mice. Compared with lean mice, peroxisome proliferator-activated receptors (PPAR)-␣ and PPAR-␥ expression was downregulated in obese double-knockout mice. Diet restriction caused a 45% weight loss, an upregulation of PPAR-␣ and PPAR-␥, and a change in the expression of genes regulating glucose transport and insulin sensitivity, lipid metabolism, oxidative stress, and inflammation, most of which are under the transcriptional control of these PPARs. Changes in gene expression were associated with increased insulin sensitivity, decreased hypertriglyceridemia, reduced mean 24-hour blood pressure and heart rate, restored circadian variations of blood pressure and heart rate, increased ejection fraction, and reduced atherosclerosis. PPAR-␣ and PPAR-␥ expression was inversely related to plaque volume and to oxidized LDL content in the plaques. Conclusions-Induction of PPAR-␣ and PPAR-␥ in adipose tissue, heart, and aortic arch is a key mechanism for reducing atherosclerosis and improving cardiovascular function resulting from weight loss. Improved lipid metabolism and insulin signaling is associated with decreased tissue deposition of oxidized LDL that increases cardiovascular risk in persons with the metabolic syndrome. Key Words: atherosclerosis Ⅲ circadian rhythm Ⅲ genes Ⅲ lipoproteins Ⅲ obesity I nsulin resistance is now receiving increasing attention not only as a precursor to type 2 diabetes but also as a predictor of increased risk of cardiovascular disease. 1 Fat distributed in the abdominal region is a risk factor for type 2 diabetes and cardiovascular disease and is associated closely with insulin resistance. 2 Weight loss in insulin-resistant but not in insulinsensitive obese persons reduces their risk of coronary heart disease (CHD). 3 It is not known, however, to what extent changes in the intra-abdominal adipose gene expression profile are important for the reduction of the risk. 4 Several adipokines, and more specifically peroxisome proliferator-activated receptors (PPARs), regulate a number of the processes that contribute to the development of atherosclerosis, including dyslipidemia, arterial hypertension, endothelial dysfunction, insulin resistance, and vascular remodeling. Adipokines are preferentially expressed in intraabdominal adipose tissue, and the secretion of proinflammatory adipokines is elevated with increasing adiposity. Approaches to re...

Immunohistochemical analyses of human atherosclerotic plaques revealed the expression of TIMP-1 in some but not all macrophage-rich and IL-8-rich areas. Therefore, IL-8 may play a potential atherogenic role by inhibiting local TIMP-1 expression, thereby leading to an imbalance between MMPs and TIMPs at focal sites in the atherosclerotic plaque.

Background Higher Lp-PLA2 activity is associated with increased risk of coronary heart disease (CHD), making Lp-PLA2 a potential therapeutic target. PLA2G7 variants associated with Lp-PLA2 activity could evaluate whether this relationship is causal. Methods and Results A meta-analysis including a total of 12 studies (5 prospective, 4 case-control, 1 case-only and 2 cross-sectional, n=26,118) was undertaken to examine the association of: (i) LpPLA2 activity vs. cardiovascular biomarkers and risk factors and CHD events (two prospective studies; n=4884); ii) PLA2G7 SNPs and Lp-PLA2 activity (3 prospective, 2 case-control, 2 cross-sectional studies; up to n=6094); and iii) PLA2G7 SNPs and angiographic coronary artery disease (2 case-control, 1 case-only study; n=4971 cases) and CHD events (5 prospective, 2 case-control studies; n=5523). Lp-PLA2 activity correlated with several CHD risk markers. Hazard ratio for CHD events top vs. bottom quartile of Lp-PLA2 activity was 1.61 (95%CI: 1.31, 1.99) and 1.17 (95%CI: 0.91, 1.51) after adjustment for baseline traits. Of seven SNPs, rs1051931 (A379V) showed the strongest association with Lp-PLA2 activity, VV subjects having 7.2% higher activity than AAs. Genotype was not associated with risk markers, angiographic coronary disease (OR 1.03 (95%CI 0.80, 1.32), or CHD events (OR 0.98 (95%CI 0.82, 1.17). Conclusions Unlike Lp-PLA2 activity, PLA2G7 variants associated with modest effects on Lp-PLA2 activity were not associated with cardiovascular risk markers, coronary atheroma or CHD. Larger association studies, identification of SNPs with larger effects, or randomised trials of specific Lp-PLA2 inhibitors are needed to confirm/refute a contributory role for Lp-PLA2 in CHD.

Macrophage infiltration into the subendothelial space at lesion prone sites is the primary event in atherogenesis. Inhibition of macrophage homing might therefore prevent atherosclerosis. Since HDL levels are inversely correlated with cardiovascular risk, their effect on macrophage homing was assessed in apoE-deficient (apoE-/-) mice. Overexpression of human apolipoprotein AI in apoE-/- mice increased HDL levels 3-fold and reduced macrophage accumulation in an established assay of leukocyte homing to aortic root endothelium 3.2-fold (P<0.005). This was due to reduced in vivo betaVLDL oxidation, reduced betaVLDL triggered endothelial cytosolic Ca2+ signaling through PAF-like bioactivity, lower ICAM-1 and VCAM-1 expression, and diminished ex vivo leukocyte adhesion. Adenoviral gene transfer of human PAF-acetylhydrolase (PAF-AH) in apoE-/- mice increased PAF-AH activity 1.5-fold (P<0.001), reduced betaVLDL-induced ex vivo macrophage adhesion 3.5-fold (P<0.01), and reduced in vivo macrophage homing 2.6-fold (P<0.02). These inhibitory effects were observed in the absence of increased HDL cholesterol levels. In conclusion, HDL reduces macrophage homing to endothelium by reducing oxidative stress via its associated PAF-AH activity. This protective mechanism is independent of the function of HDL as cholesterol acceptor. Modulation of lipoprotein oxidation by PAF-AH may prevent leukocyte recruitment to the vessel wall, a key feature in atherogenesis.

Oxidation of low density lipoproteins is an initial step of atherogenesis that generates pro-inflammatory phospholipids, including platelet-activating factor (PAF) and its analogs. PAF is degraded by PAF-acetylhydrolase (PAF-AH), a circulating enzyme having both pro- and anti-inflammatory activities. PAF-AH activity has been postulated to be a risk factor for coronary artery disease (CAD); however, whether PAF-AH has a causal role or is simply a marker of risk is unclear. The aim of this study was to relate the variability of the genes encoding PAF-AH (PLA2G7) and the PAF-receptor (PTAFR) to the risk of CAD and its complications. All polymorphisms located in putatively functional regions were investigated in a prospective cohort of CAD patients (n = 1314) and a group of healthy controls (n = 485). The whole gene variability was investigated in relation to case-control status, prospective cardiovascular outcome and plasma PAF-AH levels by means of haplotype analyses. All analyses indicated an effect of the PLA2G7/A379V polymorphism independent of the other polymorphisms. The V379 allele was less frequent in CAD patients than in controls and was associated with a lower risk of future cardiovascular events, suggesting that this allele might be protective against the development of CAD. The V379 allele was also associated with a weak increase of plasma PAF-AH activity that was unlikely to explain the protective effect of the allele on risk. A more likely interpretation is that the A379V polymorphism might modify the enzyme function towards a more anti-atherogenic form. Polymorphisms of the PTAFR gene were not related to any phenotype.