Involvement of lipoxygenase in lysophosphatidic acid-stimulated hydrogen peroxide release in human HaCaT keratinocytes.
Although it is now recognized that low levels of reactive oxygen species (ROS) are required for the mitogenic response, mitogen-induced signalling pathways that regulate ROS generation in non-phagocytic cells remain largely uncharacterized. Using a real-time assay for measuring hydrogen peroxide (H(2)O(2)) formation, we analysed H(2)O(2) release in human HaCaT keratinocytes in response to lysophosphatidic acid (LPA), a mitogen for keratinocytes. LPA rapidly increased H(2)O(2) release in HaCaT cells. Unlike LPA-induced mitogen-activated protein (MAP) kinase activation, LPA-stimulated H(2)O(2) release was independent of the tyrosine kinase activity of the epidermal growth factor (EGF) receptor. Calcium chelators, phospholipase A(2) inhibitors, and lipoxygenase inhibitors effectively blocked LPA-stimulated H(2)O(2) release, whereas cyclooxygenase inhibitors were without effect. Addition of 5-lipoxygenase products 5-hydroperoxyeicosatetraenoic acid (5-HPETE) and leukotriene B(4), but not 5-hydroxyeicosatetraenoic acid (5-HETE) and leukotriene C(4), restored LPA-stimulated H(2)O(2) release in cells treated with the lipoxygenase inhibitors nordihydroguaiaretic acid and Zileuton. These results suggest that the lipoxygenase products 5-HPETE and leukotriene B(4) are required for LPA-stimulated H(2)O(2) release in HaCaT cells.
Sekharam
M
M
Molecular Oncology Program, H. Lee Moffitt Cancer Center and Research Institute, University of South Florida, Tampa, FL 33612, USA.
Cunnick
J M
JM
Wu
J
J
D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D004261:DNA Replication / Q000187:drug effects; D066246:ErbB Receptors / Q000378:metabolism; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism*; D015603:Keratinocytes / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D008084:Lipoxygenase / Q000378:metabolism*; D008246:Lysophospholipids / Q000494:pharmacology*
D008246:Lysophospholipids; D006861:Hydrogen Peroxide; D008084:Lipoxygenase; D066246:ErbB Receptors; D002118:Calcium