Endogenous 12(S)-HETE production by tumor cells and its role in metastasis.
12(S)-Hydroxyeicosatetraenoic acid [12(S)-HETE] is the 12-lipoxygenase metabolite of arachidonic acid. Previously, we have demonstrated that exogenous 12(S)-HETE can activate protein kinase C, increase cell surface expression of integrins, enhance adhesion, induce endothelial cell retraction, and increase experimental metastasis of tumor cells. Because of these prominent effects of exogenous 12(S)-HETE on tumor cell metastatic potential, it is important to determine whether there is endogenous 12(S)-HETE production by tumor cells. In the present study, mRNAs from human, rat, and mouse platelets as well as human colon carcinoma (Clone A), rat Walker carcinoma (W256), and mouse melanoma (B16a) and lung carcinoma (3LL) were reverse transcribed and amplified by polymerase chain reaction with platelet 12-lipoxygenase specific primers. Identity of the polymerase chain reaction fragments was confirmed by sequencing. 12-Lipoxygenase protein was detected by Western blotting. Tumor cell-derived 12-HETE was determined by reverse phase-high performance liquid chromatography analysis. In addition, the effect of endogenous 12(S)-HETE on tumor cells was studied by using a platelet-type 12-lipoxygenase selective inhibitor (N-benzyl-N-hydroxy-5-phenylpentanamide). Our results suggest that some tumor cells express platelet-type 12-lipoxygenase mRNA, protein and metabolize arachidonic acid to 12(S)-HETE and that endogenous 12(S)-HETE, like the exogenous 12(S)-HETE, may play an important role in tumor cell adhesion to matrix in vitro and lung colonization in vivo.
Chen
Y Q
YQ
Department of Radiation Oncology, Wayne State University, Detroit, Michigan 48202.
Duniec
Z M
ZM
Liu
B
B
Hagmann
W
W
Gao
X
X
Shimoji
K
K
Marnett
L J
LJ
Johnson
C R
CR
Honn
K V
KV
D019377:12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; D000595:Amino Acid Sequence; D000818:Animals; D001092:Arachidonate 12-Lipoxygenase / Q000032:analysis* / Q000502:physiology; D001483:Base Sequence; D015153:Blotting, Western; D002294:Carcinoma, Squamous Cell / Q000201:enzymology / Q000378:metabolism / Q000473:pathology; D002448:Cell Adhesion / Q000502:physiology; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D005109:Extracellular Matrix / Q000502:physiology; D006801:Humans; D006893:Hydroxyeicosatetraenoic Acids / Q000096:biosynthesis* / Q000502:physiology*; D016859:Lipoxygenase Inhibitors; D008175:Lung Neoplasms / Q000556:secondary; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D009362:Neoplasm Metastasis / Q000503:physiopathology*; D009374:Neoplasms, Experimental / Q000201:enzymology / Q000378:metabolism* / Q000473:pathology*; D016133:Polymerase Chain Reaction / Q000379:methods; D012194:RNA-Directed DNA Polymerase / Q000378:metabolism; D051381:Rats
D006893:Hydroxyeicosatetraenoic Acids; D016859:Lipoxygenase Inhibitors; D019377:12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; D001092:Arachidonate 12-Lipoxygenase; D012194:RNA-Directed DNA Polymerase