Iversen L, Ziboh V A, Shimizu T, Ohishi N, Rådmark O, Wetterholm A, Kragballe K
Department of Dermatology, Marselisborg Hospital, University of Aarhus, Denmark.
J Dermatol Sci. 1994 Jun;7(3):191-201. doi: 10.1016/0923-1811(94)90095-7.
The purpose of this study was to determine whether normal human epidermis could produce leukotriene B4 (LTB4) from leukotriene A4 (LTA4) ex vivo, and to localize this LTA4-hydrolase activity. Epidermis obtained by suction blister technique incubated with human polymorphonuclear cells, resulted in a 54% increase in LTB4 formation when compared to polymorphonuclear cells incubated alone. Furthermore, human epidermis transformed exogenous LTA4 into LTB4, and this reaction obeyed Michaelis-Menten kinetics with an apparent Km of 6 microM. Subcellular fractionation of homogenized epidermis localized the LTA4-hydrolase activity mainly in the 105,000 x g supernatant fraction (cytoplasmic fraction). This activity was inhibited by two inhibitors of LTA4-hydrolase (bestatin and captopril). Western blot analysis of the 105,000 x g fraction of homogenized epidermis and cultured keratinocytes supported the presence of a LTA4-hydrolase. Thus, normal human epidermis possesses LTA4-hydrolase activity which can transform exogenous LTA4 and polymorphonuclear cell-derived LTA4 into LTB4. The identification of LTA4-hydrolase in the cytoplasmic fraction of human epidermis indicates that epidermal cells may play a more active role in the enzymatic process leading to formation of the proinflammatory compound LTB4 than previously expected.
本研究的目的是确定正常人表皮能否在体外将白三烯A4（LTA4）转化为白三烯B4（LTB4），并对这种LTA4水解酶活性进行定位。通过水疱抽吸技术获取的表皮与人类多形核细胞共同孵育，与单独孵育的多形核细胞相比，LTB4生成增加了54%。此外，人表皮可将外源性LTA4转化为LTB4，该反应符合米氏动力学，表观Km为6微摩尔。对匀浆后的表皮进行亚细胞分级分离，结果显示LTA4水解酶活性主要位于105,000×g上清液组分（细胞质组分）中。该活性受到两种LTA4水解酶抑制剂（贝司他汀和卡托普利）的抑制。对匀浆后的表皮和培养的角质形成细胞的105,000×g组分进行蛋白质免疫印迹分析，证实存在LTA4水解酶。因此，正常人表皮具有LTA4水解酶活性，可将外源性LTA4和多形核细胞衍生的LTA4转化为LTB4。在人表皮细胞质组分中鉴定出LTA4水解酶，表明表皮细胞在导致促炎化合物LTB4形成的酶促过程中可能发挥比先前预期更积极的作用。
