12-hydroxy-5-8-10-14-eicosatetraenoic-acid and Arteriosclerosis

We previously showed that vascular smooth muscle cells and endothelial cells cultured under high glucose conditions produced more 12(S)-hydroxyeicosatetraenoic acid (12-HETE), the 12-lipoxygenase (12-LO) product of arachidonate metabolism, relative to normal glucose. Because the lipoxygenase (LO) pathway has been associated with oxidant stress and the pathogenesis of atherosclerosis, we now examined 12-LO activation in vivo in a pig model of diabetes-induced accelerated atherosclerosis which displays human characteristics.. The animal model was developed in pigs who were fed a normal or high fat diet and given streptozotocin injections to produce normolipemic-normoglycaemic (NLNG), normolipemic-hyperglycaemic (NLHG), hyperlipemic-normoglycaemic (HLNG) and hyperlipemic-hyperglycaemic (HLHG) pigs. Tissue samples were obtained from key arterial beds to examine 12-LO expression at 20 weeks after the pigs began their diet.. All HG pigs maintained threefold higher serum glucose concentrations. The HL groups developed atherosclerosis but diabetic HLHG pigs showed markedly accelerated atherosclerosis (twofold) relative to non-diabetic HLNG pigs. Immunostaining showed progressive increases in 12-LO in arteries in the order NLNG, NLHG, HLNG and HLHG. Leukocyte-type 12-LO protein (immuno-blotting) as well as mRNA expression (by competitive PCR) in abdominal and coronary arteries were significantly greater in HLHG pigs than in all the other three groups. Furthermore, increased oxidant stress was observed in monocytes from NLHG and HLNG pigs, and greatly potentiated in HLHG pigs.. These results are consistent with the hypothesis that 12-LO activation plays a key role in accelerated atherosclerosis due to diabetes and hyperlipemia.

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; Animals; Aorta, Abdominal; Arachidonate 12-Lipoxygenase; Arteriosclerosis; Blood Glucose; Cholesterol; Coronary Vessels; Diabetes Mellitus, Experimental; Diet, Atherogenic; Disease Progression; Hyperlipidemias; Male; Oxidative Stress; Polymerase Chain Reaction; Swine; Triglycerides

In this paper, three compounds were isolated and identified from the leaves of Salix matsudana. They are apigenin-7-0-beta-D-glucopyranside(I), luteolin-7-0-beta-D-glucopyranside(II), compound III. Compound I and II are isolated firstly from Salix spp., compound III is found firstly in the world. Furthermore, study on effect of arachidonic acid metabolisin in rat platelets by them with radio-chromatography found that they can significantly inhibit the production of 12-HETE(12-hydroxy-5,8, 10,14-eicosatetraenoic acid), which can induce allergy and arteriosclerosis. The production of apigenin-7-0-beta-D-glucopyranside being hydrolyzed was apigenin, it can inhibit TXB2(thromoxane B2) which can induce platelet aggregation.

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; Animals; Apigenin; Arachidonic Acid; Arteriosclerosis; Blood Platelets; Drugs, Chinese Herbal; Fibrinolytic Agents; Glucosides; Luteolin; Plant Leaves; Platelet Aggregation; Rats; Salix; Thromboxane B2

The development of atherosclerosis is accelerated in individuals with type 2 diabetes. Adhesion of monocytes to the vascular endothelium is a key initial step in atherogenesis. We have previously shown that monocyte adhesion to human aortic endothelial cells (HAECs) cultured long-term in high-glucose medium (25 mmol/L, 2 passages) is increased compared with cells grown in normal glucose (5 mmol/L). One potential mechanism for increased monocyte adhesion to HAECs under hyperglycemic conditions is via the 12-lipoxygenase (12-LO) pathway. In this study, we demonstrated in HAECs that the major LO metabolite of arachidonic acid was the 12-LO product, 12(S)-hydroxyeicosatetraenoic acid [12(S)-HETE], which was increased severalfold in HAECs cultured under high-glucose conditions. Furthermore, treatment of HAECs with 12(S)-HETE induced monocyte, but not neutrophil, adhesion an average of 3-fold (range of 1.5- to 5-fold) compared with untreated cells (75+/-5 versus 26+/-1 monocytes per field, respectively, P<0.001). Expression of the adhesion molecules vascular cell adhesion molecule-1, E-selectin, and intercellular adhesion molecule-1 was not significantly increased. However, both glucose and 12(S)-HETE induced a 60% increase in HAEC surface expression of connecting segment-1 (ie, CS-1) fibronectin, a ligand for very late-acting antigen-4 (VLA-4). The antibodies used to block monocyte integrin VLA-4 and leukocyte function-related antigen-1, a monocytic counterreceptor for intercellular adhesion molecule-1, inhibited the ability of both 12-LO products and high glucose to induce monocyte adhesion. These results definitively demonstrate for the first time in HAECs that the 12-LO pathway can induce monocyte-endothelial cell interaction and that the effects of glucose may be mediated, at least in part, through this pathway. Thus, these results suggest that the 12-LO pathway may play a role in the increased susceptibility of diabetics to atherosclerosis.

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; Antibodies, Blocking; Antigens, CD; Aorta; Arteriosclerosis; CD18 Antigens; Cell Adhesion; Cells, Cultured; E-Selectin; Endothelium, Vascular; Glucose; Humans; Hydroxyeicosatetraenoic Acids; Hyperglycemia; Integrin alpha4; Intercellular Adhesion Molecule-1; Lipoxygenase; Monocytes; Neutrophils; Vascular Cell Adhesion Molecule-1

In summary, we suggest that hyperglycemia causes upregulation of 12-lipoxygenase activity. The increased production of 12-LO products, 12(S) and 15(S)-HETE, activates monocyte integrins which result in enhanced adhesion of monocytes to endothelium. The binding of monocytes to endothelium is a key early event in development of atherosclerosis. Upregulation of this process by vascular cells exposed to chronic elevations in glucose may be one explanation for the accelerated atherosclerosis observed in patients with Type 2 diabetes.

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; Arachidonate 12-Lipoxygenase; Arteriosclerosis; Cell Adhesion; Cell Communication; Cells, Cultured; Diabetic Angiopathies; Endothelium, Vascular; Glucose; Humans; Hydroxyeicosatetraenoic Acids; Hyperglycemia; Lipoxygenase Inhibitors; Monocytes; Signal Transduction

Cytokines such as tumor necrosis factor (TNF), interleukin-1 (IL-1), and gamma-interferon (IF) are produced by activated hematopoietic cells. They possess antiviral activity and have other biological activities such as induction of cell proliferation and hemorrhagic necrosis of tumors. Since herpes simplex virus (HSV) infection of human vascular cells is known to produce a biochemical and cytopathological effect virtually indistinguishable from atherosclerosis, we hypothesized that these cytokines many prevent cholesteryl ester (CE) accumulation in arterial smooth muscle cells (SMC) that is seen with herpesvirus infection. We now report that TNF and IL-1 but not gamma-IF prevent CE accumulation in HSV-infected arterial SMC by induction of cyclic AMP-dependent CE hydrolysis. This effect is mediated through the arachidonate 12-lipoxygenase pathway via 12-HETE since pretreatment of cells with several lipoxygenase inhibitors abolishes the antiviral effect and 12-HETE is the major (greater than 99%) lipoxygenase metabolite produced by these cells. This conclusion is further based on our observations that TNF and IL-1 enhance 12-HETE production in SMC and that 12-HETE significantly increases both intracellular cyclic AMP and lysosomal CE hydrolysis. Moreover, dibutyryl cyclic AMP restored a normal phenotype in these virally infected cells. Collectively, these findings identify for the first time a biochemical mechanism involved in the reduction of lipid accumulation in virally infected arterial SMC by these potent cytokines.

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; Animals; Arachidonate 12-Lipoxygenase; Arachidonate Lipoxygenases; Arteriosclerosis; Biological Factors; Cells, Cultured; Cholesterol; Cytokines; Cytopathogenic Effect, Viral; Drug Interactions; Herpes Simplex; Hydroxyeicosatetraenoic Acids; Interferon-gamma; Interleukin-1; Muscle, Smooth, Vascular; Tumor Necrosis Factor-alpha

The formation of the major metabolic products of endogenous arachidonic acid (AA) via cyclooxygenase and lipoxygenase pathways in platelets from normal and type IIA hypercholesterolemic subjects was evaluated. 12-Hydroxyeicosatetraenoic acid (12-HETE) and thromboxane B2(TXB2) were determined by selected ion monitoring (SIM) after extraction and purification of collagen stimulated platelet-rich plasma (PRP). The levels of both arachidonic acid metabolites in the non-stimulated PRP of control and type IIA subjects were below the detection limit of the method, rising significantly after collagen stimulation. Both 12-HETE and TXB2 levels in collagen-stimulated PRP samples from the patients were significantly higher than levels in controls (P less than 0.001). In view of the key role of 12-HETE in mediating smooth muscle cell migration and proliferation and in stimulating macrophage activity, these data may provide information for the understanding of the elevated incidence of thrombosis and atheromatous lesion in patients with type IIA hypercholesterolemia.

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; Adult; Arachidonic Acid; Arachidonic Acids; Arteriosclerosis; Blood Platelets; Female; Humans; Hydroxyeicosatetraenoic Acids; Hyperlipoproteinemia Type II; Male; Middle Aged; Platelet Aggregation; Thromboxane B2

We have studied generation of eicosanoids in blood vessels and platelets of rabbits in relation to platelet aggregation and plasma fibrinolytic activity at early stages of experimental atherosclerosis. A level of lipid peroxides in plasma augmented progressively from the 2-nd to 12-th week of atherogenic diet. In parallel, platelets generated more TXA2 and 12-HETE while in blood vessels a gradual reduction of prostacyclin formation was observed. Reduction in generation of PGI2 was accompanied by steady formation of PGE2. In the second week of the diet a slight decrease appeared of aggregatory response while later aggregability of platelets was increased. At first, plasma fibrinolytic activity was decreased, but it significantly augmented after 12 weeks of the diet. The above data indicate that changes of arachidonic acid metabolism in platelets and blood vessels appear at a very early stage of atherosclerosis and they influence platelet function and plasma fibrinolytic activity.

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; Animals; Aorta; Arteriosclerosis; Blood Platelets; Blood Vessels; Diet, Atherogenic; Dinoprostone; Eicosanoic Acids; Epoprostenol; Fibrinolysis; Hydroxyeicosatetraenoic Acids; Lipid Peroxides; Mesenteric Arteries; Platelet Aggregation; Prostaglandins E; Rabbits; Thromboxane A2

The metabolism of arachidonic acid by cholesterol-enriched resident mouse peritoneal macrophages was investigated. The amounts of monohydroxyeicosatetraenoic acid (mono-HETE) produced by the cholesterol-rich macrophages were 2.5-fold greater when compared to control macrophages. The major lipoxygenase product, identified by high-performance liquid chromatography in both macrophages was 12-HETE. Since macrophages are important participants in the formation of atheromatous lesions, the increased metabolism of arachidonic acid to HETE products by cholesterol-rich macrophages could contribute to the initiation and progression of the atherosclerotic process.

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; Animals; Arachidonic Acids; Arteriosclerosis; Cells, Cultured; Cholesterol; Chromatography, Thin Layer; Cyclooxygenase Inhibitors; Hydroxyeicosatetraenoic Acids; Ibuprofen; Indomethacin; Lipoxygenase; Macrophages; Male; Mice

Several eicosanoids were tested for ability to inhibit proliferation of cells in culture. In rabbit aortic smooth muscle cells and mouse B16BL6 melanoma cells, order of potency was: 12-HETE greater than PGJ2 greater than PGA1 greater than or equal to PGE1 greater than PGE2 greater than or equal to PGD2 greater than or equal to PGA2. PGB1 was active in smooth muscle cells (greater than PGD2) but not in B16 cells. 5-HETE and Leukotriene B4 were weakly active in smooth muscle cells, and PGB2, PGF2 alpha and TXB2 were inactive in both cells types. In Swiss albino mouse 3T3 fibroblasts, PGJ2 and PGE1 showed much lower relative potency than in the other two cell lines, although the profile was otherwise similar. These findings may be relevant to the anti-atherosclerotic (and perhaps anti-tumor activity) of some eicosanoids.

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; Animals; Arteriosclerosis; Cell Division; Cells, Cultured; Fibroblasts; Hydroxyeicosatetraenoic Acids; Melanoma; Mice; Muscle, Smooth, Vascular; Prostaglandins; Rabbits; Thromboxanes

We investigated the effects of mono-hydroxyeicosatetraenoic acids (HETEs) and N-formyl-methionyl-leucyl-phenylalanine (F-Met-Leu-Phe) on rat aortic smooth muscle cell migration in modified Boyden chambers. 12-HETE showed the most potent stimulatory effect on smooth muscle cell migration among the mono-HETEs tested. The optimal concentrations for cell migration were 3 X 10(-15) and 3 X 10(-13) g/ml for 12-HETE and 10(-8) g/ml for 15-HETE, 5-HETE and F-Met-Leu-Phe were inactive with these cells. As 12-HETE is biosynthesized from arachidonic acid by the 12-lipoxygenase pathway in platelets and macrophages, and 15-HETE by the 15-lipoxygenase pathway in granulocytes, the present results indicate an important role for such cells in the early phase of atherosclerosis.

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; Animals; Aorta; Arachidonate Lipoxygenases; Arachidonic Acids; Arteriosclerosis; Cell Movement; Granulocytes; Hydroxyeicosatetraenoic Acids; Lipoxygenase; Muscle, Smooth, Vascular; N-Formylmethionine; N-Formylmethionine Leucyl-Phenylalanine; Oligopeptides; Rats