title: Combined incubation of colon carcinoma cells with phorbol ester and mitochondrial uncoupling agents results in synergic elevated reactive oxygen species levels and increased γ-glutamyltransferase expression.
abstract: The NADPH oxidase (NOX) is a significant determinant for the expression and activity of γ-glutamyltransferase (GGT), which is frequently upregulated after increased levels of reactive oxygen species (ROS) and oxidative stress. Earlier studies on human colon carcinoma HT-29 cells have shown that treatment with phorbol 12-myristate 13-acetate (PMA) activates NOX thus increasing the intracellular level of ROS and upregulating GGT. Another important source of cellular ROS is the mitochondria, and treatment with the mitochondria uncoupler carbonylcyanide-4-(trifluoromethoxy)-phenylhydrazone (FCCP) results in increased ROS levels. The present study shows that when HT-29 cells were simultaneously treated with both agents, a significant and synergic increase in intracellular ROS was detected. NOX activity contributed at least 50 % of this increase as inhibiting NOX activity with apocynin or downregulating the NOX activity using siRNA against p22 phox reduced the synergic ROS production. The combined FCCP and PMA treatment also provoked highly increased GGT mRNA levels after 24 h whereas only minor and delayed increases in GGT protein and enzyme activity levels were detected. The results strongly indicate that ROS production by both mitochondria and NOX is involved in the regulation of GGT expression in colon carcinoma cells.
authors.affiliation: Tumor Biology Research Group, Department of Medical Biology, Faculty of Health Sciences, University of Tromsø, 9037, Tromsö, Norway.
authors.forename: Seila
authors.initials: S
authors.lastname: Pandur
authors.forename: Chandra
authors.initials: C
authors.lastname: Ravuri
authors.forename: Ugo
authors.initials: U
authors.lastname: Moens
authors.forename: Nils-Erik
authors.initials: NE
authors.lastname: Huseby
mesh_terms: D000098:Acetophenones / Q000494:pharmacology; D001578:Benzopyrans / Q000494:pharmacology; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone / Q000494:pharmacology; D045744:Cell Line, Tumor; D003110:Colonic Neoplasms / Q000378:metabolism*; D004338:Drug Combinations; D004357:Drug Synergism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D019073:HT29 Cells; D006801:Humans; D008928:Mitochondria / Q000378:metabolism; D019255:NADPH Oxidases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D034622:RNA Interference; D012333:RNA, Messenger / Q000096:biosynthesis; D034741:RNA, Small Interfering; D017382:Reactive Oxygen Species / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014475:Uncoupling Agents / Q000494:pharmacology*; D005723:gamma-Glutamyltransferase / Q000096:biosynthesis* / Q000235:genetics
chemical_list: D000098:Acetophenones; D001578:Benzopyrans; D004338:Drug Combinations; D004791:Enzyme Inhibitors; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D017382:Reactive Oxygen Species; D014475:Uncoupling Agents; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; C056165:acetovanillone; C085746:rottlerin; D019255:NADPH Oxidases; C086986:CYBA protein, human; D005723:gamma-Glutamyltransferase; D013755:Tetradecanoylphorbol Acetate