Mechanisms for tissue specific expression and cell growth of 12/15-lipoxygenase

脂氧合酶(LOX)将分子氧结合到不饱和脂肪酸的特定碳原子中。根据花生四烯酸的氧化部位不同，哺乳动物组织中存在5-、8-、12-和15-LOX。根据它们的cDNA一级结构和酶学性质，它们主要分为5-LOX和12/15-LOX亚家族。5-LOX催化白三烯的第一步生成，白三烯在即刻过敏和变态反应中具有很强的生物活性。12/15-LOX有多种亚型：白细胞、血小板和表皮。尽管12/15-LOX已被证明在动脉粥样硬化和神经传递等多个系统中发挥作用，但其病理生理功能仍然是研究和讨论的主题。有大量间接证据表明LOX在低密度脂蛋白(LDL)的氧化修饰中发挥作用。本研究的目的是研究Intra…的作用。更多的细胞12-LOX在低密度脂蛋白的氧化修饰中导致泡沫细胞的形成。我们在巨噬细胞样细胞系J774A.1中过表达了12-脂氧合酶，该细胞系最初没有显示出可检测到的酶活性。当表达12-脂氧合酶的细胞与低密度脂蛋白在37℃的改良Eagle培养液中孵育12h时，硫代巴比妥酸反应物测定的低密度脂蛋白氧化作用明显增强。在碱解前后用高效液相色谱检测修饰低密度脂蛋白中的氧化产物。大多数含氧衍生物为酯化产物，主要产物为13S-羟基十八碳-9Z，11E-二烯酸。这些结果清楚地表明，低密度脂蛋白中的酯化脂肪酸是由J774A.1细胞中表达的12-脂氧合酶氧化的。此外，由细胞内12-脂氧合酶产生的氧化低密度脂蛋白被清道夫受体识别，通过巨噬细胞降解试验进行评估。较少

Lipoxygenase (LOX) incorporates a molecular oxygen into a specific carbon atom of unsaturated fatty acids. There are 5-, 8-, 12- and 15-LOXs in mammalian tissues according to the oxygenation site of arachidonic acid. Based upon the primary structures deduced from their cDNAs and enzymological properties, they are principally classified into 5-LOX and 12/15-LOX subfamilies. The 5-LOX catalyzes the first step in the generation of leukotrienes which have potent biological activities in the immediate hypersensitivity and allergy. There are number of isoforms of 12/15-LOXs : leukocyte, platelet and epidermis. Although the 12/15-LOXs have been shown to play roles in several systems such as atherosclerosis and neurotransmission, their pathophysiological functions are still subjects of investigation and discussion.There are a body of circumstantial evidences for a role of LOX in oxidative modification of low density lipoprotein (LDL). The aim of this study was to investigate the role of intrac … More ellular 12-LOX in the oxidative modification of LDL that leads to foam cell formation. We overexpressed 12-lipoxygenases in a macrophage-like cell line J774A.1 which originally did not show the detectable enzyme activity. When the 12-lipoxygenase-expressing cells were incubated with LDL in Dulbecco's modified Eagle medium at 37 ℃ for 12 h, LDL oxidation as determined by thiobarbituric acid reactive substance was markedly increased over the mock-transfected cells. Oxygenated products in the modified LDL were examined by high performance liquid chromatography before and after alkaline hydrolysis. Most of the oxygenated derivatives were of an esterified form, and the major product was identified to be 13S-hydroxyoctadeca-9Z, 11E-dienoic acid. These results clearly demonstrated that esterified fatty acids in LDL were oxygenated by the 12-lipoxygenases expressed in the J774A.1 cells. Furthermore, the oxidized LDL generated by intracellular 12-lipoxygenases was recognized by a scavenger receptor as assessed by macrophage degradation assay. Less