title: DHA-induced stress response in human colon cancer cells - Focus on oxidative stress and autophagy.
abstract: Polyunsaturated fatty acids (PUFAs) are important constituents of the diet and health benefits of omega-3/n-3 PUFAs, especially eicosapentaenoic acid (EPA, 20:5 n-3) and docosahexaenoic acid (DHA, 22:6 n-3) have been well documented in relation to several diseases. Increasing evidence suggests that n-3 PUFAs may have anticancer activity and improve the effect of conventional cancer therapy. The mechanisms behind these effects are still unclear and need to be elucidated. We have examined the DHA-induced stress response in two human colon cancer cell lines, SW620 and Caco-2. SW620 cells are growth-inhibited at early time points by DHA, while the growth of Caco-2 cells almost remains unaffected by the same treatment. Gene expression analysis of SW620 cells treated with DHA revealed changes at early time points; transcripts involved in oxidative stress and autophagy were among the first to be differentially expressed. We find that oxidative stress is induced in both cell lines, although at different time points and to different extent. DHA induced nuclear translocation of the oxidative stress sensor NFE2L2 in both cell lines, indicating an induction of an anti-oxidative response. However, vitamin E did not counteract ROS-production or the translocation of NFE2L2 to the nucleus. Neither vitamin E nor the antioxidants butylated hydoxyanisole (BHA) and butylated hydoxytoluene (BHT) did affect the growth inhibition in SW620 cells after DHA-treatment. Also, siRNA-mediated down-regulation of NFE2L2 did not sensitize SW620 and Caco-2 cells to DHA. These results indicate that oxidative stress response is not the cause of DHA-induced cytotoxicity in SW620 cells. Using biochemical and imaging based functional assays, we found a low basal level of autophagy and no increase in autophagic flux after adding DHA to the SW620 cells. However, Caco-2 cells displayed a higher level of autophagy, both in the absence and presence of DHA. Inhibition of autophagy by siRNA mediated knock down of ATG5 and ATG7 sensitized both SW620 and Caco-2 cells to DHA. Stimulation of autophagy by rapamycin in SW620 and Caco-2 cells resulted in decreased DHA-sensitivity and inhibition of autophagy in Caco-2 cells by chloroquine resulted in increased DHA-sensitivity. These results suggest that autophagy is important for the DHA sensitivity of colon cancer cells and imply possible therapeutic effects of this fatty acid against cancer cells with low autophagy.
authors.affiliation: Department of Laboratory Medicine, Children's and Women's Health, Faculty of Medicine, Norwegian University of Science and Technology, NTNU, 7006 Trondheim, Norway; Department of Technology, Erling Skjalgssons gt. 1, University College of Sør-Trøndelag, 7006 Trondheim, Norway; Centre of Molecular Inflammation Research, Department of Cancer Research and Molecular Medicine, Norwegian University of Science and Technology, 7491 Trondheim, Norway.
authors.forename: Kristine
authors.initials: K
authors.lastname: Pettersen
authors.affiliation: Department of Laboratory Medicine, Children's and Women's Health, Faculty of Medicine, Norwegian University of Science and Technology, NTNU, 7006 Trondheim, Norway.
authors.forename: Vivi Talstad
authors.initials: VT
authors.lastname: Monsen
authors.affiliation: Department of Laboratory Medicine, Children's and Women's Health, Faculty of Medicine, Norwegian University of Science and Technology, NTNU, 7006 Trondheim, Norway.
authors.forename: Caroline Hild
authors.initials: CH
authors.lastname: Hakvåg Pettersen
authors.affiliation: Department of Laboratory Medicine, Children's and Women's Health, Faculty of Medicine, Norwegian University of Science and Technology, NTNU, 7006 Trondheim, Norway; Central Norway Regional Health Authority, 7055 Stjørdal, Norway.
authors.forename: Hilde Bremseth
authors.initials: HB
authors.lastname: Overland
authors.affiliation: Department of Laboratory Medicine, Children's and Women's Health, Faculty of Medicine, Norwegian University of Science and Technology, NTNU, 7006 Trondheim, Norway.
authors.forename: Grete
authors.initials: G
authors.lastname: Pettersen
authors.affiliation: Department of Laboratory Medicine, Children's and Women's Health, Faculty of Medicine, Norwegian University of Science and Technology, NTNU, 7006 Trondheim, Norway.
authors.forename: Helle
authors.initials: H
authors.lastname: Samdal
authors.affiliation: Department of Laboratory Medicine, Children's and Women's Health, Faculty of Medicine, Norwegian University of Science and Technology, NTNU, 7006 Trondheim, Norway.
authors.forename: Almaz Nigatu
authors.initials: AN
authors.lastname: Tesfahun
authors.affiliation: Department of Laboratory Medicine, Children's and Women's Health, Faculty of Medicine, Norwegian University of Science and Technology, NTNU, 7006 Trondheim, Norway.
authors.forename: Anne Gøril
authors.initials: AG
authors.lastname: Lundemo
authors.affiliation: Department of Technology, Erling Skjalgssons gt. 1, University College of Sør-Trøndelag, 7006 Trondheim, Norway; Centre of Molecular Inflammation Research, Department of Cancer Research and Molecular Medicine, Norwegian University of Science and Technology, 7491 Trondheim, Norway.
authors.forename: Geir
authors.initials: G
authors.lastname: Bjørkøy
authors.affiliation: Department of Laboratory Medicine, Children's and Women's Health, Faculty of Medicine, Norwegian University of Science and Technology, NTNU, 7006 Trondheim, Norway. Electronic address: svanhild.schonberg@ntnu.no.
authors.forename: Svanhild A
authors.initials: SA
authors.lastname: Schønberg
mesh_terms: D001343:Autophagy / Q000187:drug effects*; D018938:Caco-2 Cells; D003110:Colonic Neoplasms / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D004281:Docosahexaenoic Acids / Q000494:pharmacology*; D006801:Humans; D051267:NF-E2-Related Factor 2 / Q000502:physiology; D018384:Oxidative Stress / Q000187:drug effects*; D017510:Protein Folding
chemical_list: D051267:NF-E2-Related Factor 2; C495635:NFE2L2 protein, human; D004281:Docosahexaenoic Acids
keywords: Autophagy; Colon cancer; DHA; NFE2L2; Oxidative stress; n−3 PUFA