Characterization of an 8-lipoxygenase activity induced by the phorbol ester tumor promoter 12-O-tetradecanoylphorbol-13-acetate in mouse skin in vivo

An enzymatic activity has been found in cytosolic preparations from mouse epidermis which catalyzes the formation of 8-hydroperoxyeicosatetraenoic acid/8-hydroxyeicosatetraenoic acid (8-HPETE/8-HETE) from arachidonate. In contrast to 12-lipoxygenase this enzyme activity was not detectable in normal (untreated) mouse skin but only after in vivo treatment with the phorbol ester tumor promoter TPA (12-O-tetradecanoylphorbol-13-acetate). The induction showed a maximum at 24 h after TPA treatment strictly depended on the age of the mice and the TPA dose and was prevented by cycloheximide. The primary product formed from arachidonic acid was 8-HPETE, and the enzyme seems not to possess a significant peroxidase activity. This result as well as studies with specific inhibitors and its cytosolic localization indicates this enzyme to be a member of the lipoxygenase family. Most of the 8-lipoxygenase activity is located in cells of the suprabasal compartment of the epidermis. In spite of being a cytosolic enzyme 8-lipoxygenase appeared to be lipophilic to some extent and was activated by lecithin. The enzyme did not require calcium ions or ATP and showed a pH optimum at 7.5-8.0. 8-HPETE/8-HETE levels in mouse epidermis in vivo were determined by gas chromatography-mass spectrometry and found to be strongly increased after phorbol ester treatment, in agreement with the induction of 8-lipoxygenase observed.