Molecular biological studies on cyclooxygenases and lipoxygenases dynamics

本项目是对环氧合酶和脂氧合酶的分子生物学研究。环氧合酶是合成具有生物活性的花生四烯酸代谢产物的主要途径。(1)环氧合酶：小鼠成骨细胞系MC3T3-E1产生前列腺素E2，其环氧合酶被多种激素、生长因子和前列腺素诱导。进一步研究表明，两种环氧合酶同工酶参与了前列腺素E_2的产生。2脂氧合酶：12-脂氧合酶在底物特异性和免疫原性上存在差异。克隆了它们的cDNA和基因组DNA，并对它们的结构进行了分析。这种白细胞型酶不仅存在于白细胞中，也存在于猪脑垂体、牛气管、犬脑和大鼠松果体中。过去只在血小板中发现的血小板型12-脂氧合酶，最近在人类表皮细胞中被发现。建立了聚合酶链式反应扩增12-脂氧合酶基因的方法，发现3例骨髓增生性疾病患者的血小板中12-脂氧合酶表达水平较低，酶活性和蛋白含量降低。

This project is a molecular biological approach to cyclooxygenases and lipoxygenases which are responsible for the syntheses of bioactive arachidonate metabolites.(1) Cyclooxygenases : A murine osteoblastic cell line MC3T3-E1 produces prostaglandin E2, and its cyclooxygenase enzyme was shown to be induced by various hormones, growth factors and prostaglandins. Further studies demonstrated the involvement of two cyclooxygenase isozymes in the prostaglandin E2 production. Cyclooxygenase-1 was induced gradually only by 2-3 fold, whereas cyclooxygenase-2 appeared rapidly within 1-2 h and increased by 15-60 fold.(2) Lipoxygenases : The leukocyte-type and platelet-type of 12-lipoxygenase were distinguishied in terms of substrate specificity and immunogenicity. Their cDNAs and genomic DNAs were cloned, and their structures were elucidated. The leukocyte-type enzyme was found not only in leukocytes but also in porcine pituitary, bovine trachea, canine brain and rat pineal gland. The platelet-type 12-lipoxygenase, which had been found only in platelets, was recently found in human epidermal cells. A method for PCR-amplification of 12-lipoxygenase mRNA was established, and a low level of 12-lipoxygenase together with a decreased enzyme activity and protein was demonstrated in the platelets of three patients with myeloproliferative diseases.