(B) Representative Western blot analysis for Lp-PLA2 and the average quantification obtained by densitometric analysis of all the samples (20 subjects per group, randomly selected). in smokers (P<0.001), positively correlated with lysoPC (r=0.55, P<0.001). Then ininvitrostudy we demonstrated that both oxLDL (at concentrations similar to those found in smokers serum) and oxidized phospholipids contained in oxLDL, were able to up-regulate mRNA Lp-PLA2 in PBMC. This effect was likely due, at least in part, to the enrichment in oxidized phospholipids found in PBMC after exposure to oxLDL. Our results also showed that in human aortic smooth muscle cells lysoPC, at concentrations similar to those found in smokers, increased the expression of biglycan and versican, two main proteoglycans. Rabbit polyclonal to AMPK gamma1 == Conclusions == In smokers a further effect of raised oxidative stress is the up-regulation of Lp-PLA2 expression in PBMC with subsequent increase of plasma Lp-PLA2 and lysoPC. Moreover the correlation between lysoPC and CIMT together with the finding that lysoPC up-regulates proteoglycan synthesis suggests that lysoPC may be a link between smoking and intimal thickening. == Introduction == Cigarette smoking is a major risk factor for atherosclerotic vascular BTB06584 disease and it is an important factor contributing to premature vascular aging [1-3]. Emerging evidence demonstrates that oxidative stress and activation of inflammatory pathways contribute to many of the pathophysiologic manifestations of cigarette smoking [2-6]. Free radicals from cigarette smoking also cause peroxidation of the polyunsaturated fatty acids of cell membranes that amplify oxidative stress during smoking [5,6]. Lipoprotein-associated phospholipase A2 (Lp-PLA2) is an emerging BTB06584 and independent risk factor for coronary and carotid artery disease [7], that in animal studies has been shown to be involved in the causal pathway of the intimal oxidative-inflammatory cascade typical of atherogenesis [8]. In particular, a positive association between Lp-PLA2 activity and circulating oxidized low-density lipoprotein (oxLDL) has been shown in a hypercholesterolemic swine model of atherosclerosis [9]. Moreover, it has been recently reported that oxLDL can up-regulate the expression of Lp-PLA2 in THP1 monocyte-like cells and in human monocyte-derived macrophages [10]. Lp-PLA2 that is secreted predominantly by inflammatory cells, circulates in plasma mainly bound to LDL [8]. The oxidation of LDL within the arterial wall, a key factor in atherogenesis, yields oxidized phospholipids that are the substrate for Lp-PLA2 reaction, producing lysophosphatidylcholine (lysoPC) and oxidized free fatty acids both of which exhibit several pro-atherogenic effects [8,11].In vitroobservations have demonstrated that lysoPC is a major determinant of the pro-atherogenic properties of oxLDL and activates the redox-sensitive inflammatory pathways by increasing the recruitment of leukocytes and by inducing endothelial dysfunction and apoptosis of endothelial and vascular smooth muscle cells (SMCs) [8,11]. Ultrasound-measured carotid intima-media thickness (CIMT) represents a measure of early alterations of the arterial walls, that has been widely used as a markers of subclinical atherosclerosis and a predictor of future cardiovascular disease events [10,12,13]. In this context, however, the accuracy of CIMT as a marker of atherosclerosis is mired by the fact that main predictors of medial hypertrophy or intimal thickening are age and hypertension [14]. Moreover, in clinical cardiology, smokers often present (at a younger age) myocardial infarction without extensive visible coronary atherosclerosis [2,15]. Although epidemiological studies have consistently shown that cigarette smoking is associated with higher CIMT [16,17], the BTB06584 mechanisms are likely multifaceted and not yet fully understood. It has been previously suggested that cigarette smoking has a specific fibrogenic effect which causes intimal thickening [3,4]. Secretory vascular SMCs, the major cellular component of the neointima, secrete predominantly the small chondroitin sulfate/dermatan sulfate proteoglycans biglycan and decorin, as well as versican and the heparan sulfate proteoglycans perlecan [18,19]. The retention of LDL via their interaction with biglycan and versican at sites of intimal thickening is believed to be a key step in early atherogenesis [20-22]. Interestingly, oxLDL was demonstrated to up-regulate biglycan synthesis in vascular SMCs, resulting in enhanced lipoprotein-proteoglycans binding [23]. Moreover, BTB06584 among the several bioactive lipids contained in oxLDL, only lysoPC was shown to increase biglycan synthesis [24], through reactive oxygen species formation [25]. Then, sincein vitrostudies have suggested that oxLDL increase Lp-PLA2 expression and that lysoPC may have a role in intimal expansion [24], this study aimed to assess whether cigarette smoking-induced oxidative stress affects Lp-PLA2 expression in peripheral blood mononuclear cells (PBMC) and hence Lp-PLA2 and lysoPC plasma concentrations, as well as the relationship between lysoPC and.
(B) Representative Western blot analysis for Lp-PLA2 and the average quantification obtained by densitometric analysis of all the samples (20 subjects per group, randomly selected)