Depletion of phospholipid hydroperoxide glutathione peroxidase up-regulates arachidonate metabolism by 12S-lipoxygenase and cyclooxygenase 1 in human epidermoid carcinoma A431 cells.
Phospholipid hydroperoxide glutathione peroxidase (PHGPx), a selenium-dependent glutathione peroxidase, can interact with lipophilic substrates, including the phospholipid hydroperoxides, fatty-acid hydroperoxides, and cholesteryl ester hydroperoxides, and reduce them to hydroxide compounds. We studied the functional role of endogenous PHGPx in regulation of 12(S)-lipoxygenase and cyclooxygenase 1 activities in human epidermoid carcinoma A431 cells by using a cell system overexpressing anti-PHGPx mRNA. A retroviral expression vector designated as L1-3, wherein cDNA of PHGPx was reversely inserted into pFB-ERV in antisense orientation, was constructed. A number of stable transfectants of A431 cells with PHGPx depletion were generated from virions containing plasmid L1-3. In an intact cell assay system, the metabolism of arachidonic acid to prostaglandin E2 and 12(S)-hydroxyeicosatetraenoic acid was significantly enhanced in stable L1-3 transfectants compared with that in vector-control cells. Flow cytometric analysis revealed a significant elevated level of intracellular hydroperoxides in stable L1-3 transfectants. Treatment of stable L1-3 transfectants with 50 microM arsenite induced more significant formation of intracellular hydroperoxides than that of vector-control cells. Taken together, these results support the notion that the endogenous PHGPx plays a pivotal role in the regulation of 12(S)-lipoxygenase and cyclooxygenase 1 activities by reducing the level of intracellular lipid hydroperoxides in arachidonate metabolism in A431 cells.
Chen
Ching-Jiunn
CJ
Department of Pharmacology, College of Medicine, National Cheng Kung University, 138 Sheng-Li Road, Tainan 704, Taiwan.
Huang
Huei-Sheng
HS
Chang
Wen-Chang
WC
D001092:Arachidonate 12-Lipoxygenase / Q000378:metabolism*; D016718:Arachidonic Acid / Q000378:metabolism*; D002294:Carcinoma, Squamous Cell / Q000201:enzymology* / Q000378:metabolism; D051545:Cyclooxygenase 1; D005979:Glutathione Peroxidase / Q000235:genetics / Q000502:physiology*; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism*; D008054:Lipid Peroxides / Q000096:biosynthesis; D008565:Membrane Proteins; D008954:Models, Biological; D000080662:Phospholipid Hydroperoxide Glutathione Peroxidase; D011451:Prostaglandin-Endoperoxide Synthases / Q000378:metabolism*; D016372:RNA, Antisense / Q000235:genetics; D014162:Transfection; D014407:Tumor Cells, Cultured; D015854:Up-Regulation
D007527:Isoenzymes; D008054:Lipid Peroxides; D008565:Membrane Proteins; D016372:RNA, Antisense; D016718:Arachidonic Acid; D000080662:Phospholipid Hydroperoxide Glutathione Peroxidase; D005979:Glutathione Peroxidase; D001092:Arachidonate 12-Lipoxygenase; D051545:Cyclooxygenase 1; C496511:PTGS1 protein, human; D011451:Prostaglandin-Endoperoxide Synthases