Glasgow W C, Hill E M, McGown S R, Tomer K B, Eling T E
Laboratory of Molecular Biophysics, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina 27709, USA.
Mol Pharmacol. 1996 Jun;49(6):1042-8.
Metabolism of arachidonic and linoleic acid can be regulated by polypeptide growth factors in a variety of cell types. In Syrian hamster embryo (SHE) fibroblasts, epidermal growth factor (EGF) stimulates the conversion of exogenous linoleic acid to 13(S)-hydroxyoctadecadienoic acid (HODE). Inhibition of 13-HODE biosynthesis blocks the EGF-mitogenic response in SHE cells, and 13-HODE and its hydroperoxy precursor are potent and highly specific enhancers of EGF-dependent DNA synthesis. We demonstrated that EGF stimulates a biphasic production and release of endogenous 13-HODE. Through development of a stable isotope-dilution GC/MS assay for 13-HODE, we observed 13-HODE production as early as 5 min after EGF stimulation, and this initial phase peaked at 1 hr. A second rise in 13-HODE formation was seen at 2-4 hr, and this phase plateaued at 4-6 hr at a level of 30-40 ng/10(6) cells. EGF stimulation of 13-HODE biosynthesis is not mediated by transcriptional or translational regulation of the inducible form of prostaglandin H synthase. Based on enzyme inhibitor studies and structural characterization of products, the linoleate metabolite is apparently formed by an n-6 lipoxygenase that remains to be characterized. EGF stimulation of 13-HODE formation is linked with activation of the EGF receptor tyrosine kinase. Inhibition of EGF receptor tyrosine kinase activity with methyl-2,5-dihydroxycinnamate blocked EGF-dependent linoleic acid metabolism and EGF-regulated DNA synthesis. Potentiation of the EGF receptor tyrosine phosphorylation cascade through treatment of SHE cells with the tyrosine phosphatase inhibitor vanadate resulted in a 3-fold increase in EGF-stimulated 13-HODE production and a corresponding enhancement of the EGF mitogenic response. The coupling of EGF-regulated linoleic acid metabolism with the EGF receptor tyrosine kinase activity suggests the importance of specific linoleate compounds in mediating mitogenic signal transduction.
花生四烯酸和亚油酸的代谢可在多种细胞类型中受多肽生长因子调控。在叙利亚仓鼠胚胎（SHE）成纤维细胞中，表皮生长因子（EGF）刺激外源性亚油酸转化为13(S)-羟基十八碳二烯酸（HODE）。抑制13-HODE生物合成可阻断SHE细胞中的EGF促有丝分裂反应，且13-HODE及其氢过氧化物前体是EGF依赖性DNA合成的强效且高度特异性增强剂。我们证明EGF刺激内源性13-HODE的双相产生和释放。通过开发一种用于13-HODE的稳定同位素稀释气相色谱/质谱分析法，我们观察到在EGF刺激后5分钟最早出现13-HODE产生，且这个初始阶段在1小时达到峰值。在2 - 4小时观察到13-HODE形成的第二次升高，且这个阶段在4 - 6小时达到平稳，水平为30 - 40 ng/10(6)个细胞。EGF对13-HODE生物合成的刺激不是由前列腺素H合酶诱导形式的转录或翻译调控介导的。基于酶抑制剂研究和产物的结构表征，亚油酸代谢产物显然是由一种有待鉴定的n-6脂氧合酶形成的。EGF对13-HODE形成的刺激与EGF受体酪氨酸激酶的激活相关。用甲基-2,5-二羟基肉桂酸抑制EGF受体酪氨酸激酶活性可阻断EGF依赖性亚油酸代谢和EGF调节的DNA合成。通过用酪氨酸磷酸酶抑制剂钒酸盐处理SHE细胞来增强EGF受体酪氨酸磷酸化级联反应，导致EGF刺激的13-HODE产生增加3倍以及相应增强EGF促有丝分裂反应。EGF调节的亚油酸代谢与EGF受体酪氨酸激酶活性的偶联表明特定亚油酸化合物在介导有丝分裂信号转导中的重要性。
