Identification and subcellular localization of leukotriene A 4-hydrolase activity in human epidermis

Abstract

The purpose of this study was to determine whether normal human epidermis could produce leukotriene B 4 (LTB 4) from leukotriene A 4 (LTA 4) ex vivo; and to localize this LTA 4-hydrolase activity. Epidermis obtained by suction blister technique incubated with human polymorphonuclear cells, resulted in a 54% increase in LTB 4 formation when compared to polymorphonuclear cells incubated alone. Furthermore, human epidermis transformed exogenous LTA 4 into LTB 4, and this reaction obeyed Michaelis-Menten kinetics with an apparent K m of 6 μM. Subcellular fractionation of homogenized epidermis localized the LTA 4-hydrolase activity mainly in the 105 000 × g supernatant fraction (cytoplasmic fraction). This activity was inhibited by two inhibitors of LTA 4-hydrolase (bestatin and captopril). Western blot analysis of the 105 000 × g fraction of homogenized epidermis and cultured keratinocytes supported the presence of a LTA 4-hydrolase. Thus, normal human epidermis possesses LTA 4-hydrolase activity which can transform exogenous LTA 4 and polymorphonuclear cell-derived LTA 4 into LTB 4. The identification of LTA 4-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 LTB 4 than previously expected.