title: Photodynamic therapy mediates the oxygen-independent activation of hypoxia-inducible factor 1alpha.
abstract: Photodynamic therapy (PDT) induces the expression of the hypoxia-inducible factor 1alpha (HIF-1alpha) subunit of the HIF-1 transcription factor and its target genes in vitro and in vivo. PDT also induces the expression of the enzyme cyclooxygenase-2 and its metabolite, prostaglandin E2 (PGE2). PGE2 and hypoxia act independently and synergistically to increase HIF-1alpha accumulation and nuclear translocation. To examine the expression of HIF-1 target genes in response to PDT-mediated oxidative stress and PGE2 under normoxic conditions, we established EMT6 cells transfected with a plasmid consisting of a hypoxia response element promoter and a downstream gene encoding for green fluorescent protein (GFP). To examine the temporal kinetics of HIF-1alpha nuclear translocation in response to PDT, we transfected a second line of EMT6 cells with a GFP-tagged HIF-1alpha fusion vector. Cell monolayers were incubated with 1 microg mL(-1) Photofrin for 24 h and irradiated with fluences of 1, 2.5, and 5 J cm(-2). Direct measurement of oxygen concentration during irradiation confirmed that cells remained well oxygenated. Cells were also exposed to 1 and 10 micromol/L PGE2 for 3 h. In normoxic conditions, Photofrin, PDT, and PGE2 treatment activated HIF-1alpha and induced its nuclear translocation. Maximal Photofrin-PDT-mediated HIF-1alpha activation was intermediate in magnitude between that induced by PGE2 and that by the hypoxia mimic cobalt chloride. This work establishes that PDT induces significant activation of the HIF-1alpha pathway in the absence of hypoxia and supports the interpretation that the induction of HIF-1 target genes by PDT may be mediated, at least in part, by the prostaglandin pathway.
authors.affiliation: Department of Imaging Sciences, University of Rochester, 601 Elmwood Avenue, Box 648, Rochester, NY 14642, USA.
authors.forename: Soumya
authors.initials: S
authors.lastname: Mitra
authors.forename: Scott E
authors.initials: SE
authors.lastname: Cassar
authors.forename: David J
authors.initials: DJ
authors.lastname: Niles
authors.forename: John A
authors.initials: JA
authors.lastname: Puskas
authors.forename: John G
authors.initials: JG
authors.lastname: Frelinger
authors.forename: Thomas H
authors.initials: TH
authors.lastname: Foster
mesh_terms: D000818:Animals; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D017323:Dihematoporphyrin Ether / Q000494:pharmacology; D015232:Dinoprostone / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D008325:Mammary Neoplasms, Experimental / Q000188:drug therapy / Q000235:genetics / Q000378:metabolism; D051379:Mice; D010100:Oxygen / Q000378:metabolism; D010778:Photochemotherapy / Q000379:methods*; D014162:Transfection
chemical_list: C497443:Hif1a protein, mouse; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; C485192:green fluorescent protein, Aequorea victoria; D049452:Green Fluorescent Proteins; D017323:Dihematoporphyrin Ether; D015232:Dinoprostone; D010100:Oxygen