ORIGINAL ARTICLE
The Antiproliferative Effect of EPA in HL60 Cells is Mediated
by Alterations in Calcium Homeostasis
Jens Erik Slagsvold Æ Caroline Hild Hakva
˚
g Pettersen Æ
Turid Follestad Æ Hans Einar Krokan Æ
Svanhild Arentz Schønberg
Received: 7 August 2008 / Accepted: 27 October 2008 / Published online: 20 November 2008
Ó AOCS 2008
Abstract Studies show that n-3 polyunsaturated fatty
acids (PUFA) inhibit proliferation and induce apoptosis in
cancer cells. Recent reports indicate that this effect is due
to activation of the unfolded protein response (UPR).
However, what causes this activation has been unclear. We
examined the effects of eicosapentaenoic acid (EPA) on the
human leukemia cell line HL60 and the econazole (Ec)
resistant HL60 clone E2R2. Ec depletes Ca
2?
from the ER
and blocks Ca
2?
influx in mammalian cells, leading to
activation of the UPR and apoptosis. EPA inhibited growth
of HL60 cells strongly, while E2R2 cells were much less
affected. Gene expression analysis of HL60 cells revealed
extensive changes in transcripts related to the ER homeo-
stasis, Ca
2?
-homeostasis and cell cycle/apoptosis. Protein
levels of phosphorylated eIF2a, a selective translation
inhibitor and UPR hallmark, activating transcription factor
4 (ATF4) and sequestosome-1 were moderately increased,
whereas the cell cycle/progression protein cyclin D1 was
decreased in HL60. In contrast, EPA concentrations that
strongly inhibited and caused activation of the UPR in
HL60 cells had no effect on the expression level of these
UPR markers in E2R2 cells. Given that the only known
difference between these cells is Ec-resistance, our results
strongly suggest that the inhibitory effect of EPA on HL60
cells is initially meditated through alterations of the Ca
2?
-
homeostasis followed by activation of the UPR.
Keywords Cancer Á E2R2 Á Econazole-resistant Á
EPA Á HL60 Á UPR Á PUFA
Abbreviations
ATF3, ATF4
and ATF6
Activating transcription factor
3, 4 and 6
CHOP Growth arrest- and DNA damage-
inducible gene 153/C/EBP-homologous
protein (CHOP/Gadd153)
Ec Econazole
EPA Eicosapentaenoic acid (20:5n-3)
ER Endoplasmic reticulum
ERAD ER-associated degradation
eIF2a Eukaryote translation initiation
factor 2a
EIF2AK3/PERK eIF2a kinase 3
HMOX-1 Heme oxygenase (decycling) 1
IP3 Inositol 1,4,5-triphosphate
PUFA Polyunsaturated fatty acids
SQSTM1 Sequestosome-1
SOC Store-operated Ca
2?
channels
UPR Unfolded protein response
J. E. Slagsvold Á C. H. H. Pettersen Á S. A. Schønberg
Department of Laboratory Medicine,
Children’s and Women’s Health,
Norwegian University of Science and Technology (NTNU),
Erling Skjalgssons gate 1, 7006 Trondheim, Norway
T. Follestad
Department of Mathematical Sciences,
Norwegian University of Science and Technology (NTNU),
7491 Trondheim, Norway
H. E. Krokan
Department of Cancer Research and Molecular Medicine,
Norwegian University of Science and Technology (NTNU),
Erling Skjalgssons gate 1, 7006 Trondheim, Norway
S. A. Schønberg (&)
Department of Laboratory Medicine,
Children’s and Women’s Health,
Norwegian University of Science and Technology,
St Olav’s Hospital, 7489 Trondheim, Norway
e-mail: svanhild.schonberg@ntnu.no
123
Lipids (2009) 44:103–113
DOI 10.1007/s11745-008-3263-5