Setty B N, Stuart M J, Walenga R W
Biochim Biophys Acta. 1985 Mar 6;833(3):484-94. doi: 10.1016/0005-2760(85)90106-7.
Human umbilical arteries convert arachidonic acid into three hydroxy-eicosatetraenoic acids as well as 6-ketoprostaglandin F1 alpha, prostaglandins E2, F2 alpha and D2 and thromboxane B2. Two of these hydroxy derivatives of arachidonic acid were purified by reverse-phase HPLC and identified by GC-MS as 11-hydroxyeicosatetraenoic acid (11-HETE) and 15-hydroxyeicosatetraenoic acid (15-HETE) while a third, presumed dihydroxy derivative has not yet been identified. Both the cyclooxygenase and HETE synthesizing activities were found to be localized mainly in the microsomal fraction (100 000 X g pellet) (51 and 61% of total, respectively), and approx. 25% of both activities was found in the 10 000 X g pellet. The formation of these HETEs was inhibited by the cyclooxygenase inhibitors indomethacin and aspirin but not by the lipoxygenase inhibitor nordihydroguaiaretic acid. Production of immunoreactive 15-HETE as well as 6-ketoprostaglandin F1 alpha were also decreased significantly when arterial segments were incubated in the presence of either indomethacin or aspirin. Indomethacin inhibited the formation of both prostanoids and HETEs by microsomes in a concentration-dependent and time-dependent manner. The ID50 values for indomethacin against HETE synthesizing activity and against cyclooxygenase were 4.5 and 3.8 microM, respectively. The inactivation constants were found to be 0.09 and 0.08 min-1 for HETE synthesizing activity and cyclooxygenase, respectively. These two microsomal activities were solubilized in parallel with Tween-20. Incubation with three distinct monoclonal antibodies against different epitopes on cyclooxygenase precipitated both cyclooxygenase and HETE synthesizing activity. Each of these activities was recovered in the immune pellets. These studies demonstrate that in human umbilical arteries 11-HETE, 15-HETE and a presumed di-HETE are the products of cyclooxygenase.
人脐动脉可将花生四烯酸转化为三种羟基二十碳四烯酸以及6-酮前列环素F1α、前列腺素E2、F2α和D2以及血栓素B2。其中两种花生四烯酸的羟基衍生物通过反相高效液相色谱法纯化，并通过气相色谱-质谱法鉴定为11-羟基二十碳四烯酸（11-HETE）和15-羟基二十碳四烯酸（15-HETE），而第三种推测的二羟基衍生物尚未鉴定出来。发现环氧化酶和HETE合成活性主要定位于微粒体部分（100000×g沉淀）（分别占总量的51%和61%），并且在10000×g沉淀中发现两种活性的约25%。这些HETEs的形成受到环氧化酶抑制剂吲哚美辛和阿司匹林的抑制，但不受脂氧合酶抑制剂去甲二氢愈创木酸的抑制。当动脉段在吲哚美辛或阿司匹林存在下孵育时，免疫反应性15-HETE以及6-酮前列环素F1α的产生也显著降低。吲哚美辛以浓度依赖性和时间依赖性方式抑制微粒体中前列腺素和HETEs的形成。吲哚美辛对HETE合成活性和环氧化酶的ID50值分别为4.5和3.8 microM。发现HETE合成活性和环氧化酶的失活常数分别为0.09和0.08 min-1。这两种微粒体活性与吐温-20同时溶解。用三种针对环氧化酶上不同表位的不同单克隆抗体孵育沉淀了环氧化酶和HETE合成活性。每种活性都在免疫沉淀中回收。这些研究表明，在人脐动脉中，11-HETE、15-HETE和一种推测的二-HETE是环氧化酶的产物。
