15(S)-Lipoxygenase-1 associates with neutral lipid droplets in macrophage foam cells: evidence of lipid droplet metabolism

Ginny L. Weibel¹^,*, Michelle R. Joshi^*, Cong Wei, Sandra R. Bates, Ian A. Blair and George H. Rothblat^*

^* Division of Gastroenterology and Nutrition, The Children's Hospital of Philadelphia, Philadelphia, PA
Centers for Cancer Pharmacology and Excellence in Environmental Toxicology, University of Pennsylvania School of Medicine Philadelphia PA
Institute for Environmental Medicine and Department of Physiology, University of Pennsylvania School of Medicine, Philadelphia, PA

¹ To whom correspondence should be addressed. e-mail: weibel{at}email.chop.edu

15(S)-lipoxygenase-1 (15-LO-1) was present in the whole-cellhomogenate of an acute human monocytic leukemia cell line (THP-1).Additionally, 15-LO-1 was detected on neutral lipid dropletsisolated from THP-1 foam cells. To investigate if 15-LO-1 isactive on lipid droplets, we used the mouse leukemic monocyticmacrophage cell line (RAW 264.7), which are stably transfectedwith human 15-LO-1. The RAW 15-LO-1 cells were incubated withacetylated low density lipoprotein to generate foam cells. 15(S)-hydroxyeicosatetraenoicacid [15(S)-HETE], the major 15-LO-1 metabolite of arachidonicacid, was produced in the 15-LO-1 RAW but not in the mock transfectedcells when incubated with arachidonic acid. Lipid droplets wereisolated from the cells and incubated with arachidonic acid,and production of 15(S)-HETE was measured over 2 h. 15(S)-HETEwas produced in the incubations with the lipid droplets, andthis production was attenuated when the lipid droplet fractionwas subjected to enzyme inactivation through heating. Effluxof 15(S)-HETE from cholesteryl ester-enriched 15-LO RAW cells,when lipid droplets are present, was significantly reduced comparedwith that from cells enriched with free cholesterol (lipid dropletsare absent). We propose that 15-LO-1 is present and functionalon cytoplasmic neutral lipid droplets in macrophage foam cells,and these droplets may act to accumulate the anti-inflammatorylipid mediator 15(S)-HETE.

Supplementary key words lipoxygenases • cholesterol • cholesteryl ester

Abbreviations: 15-LO-1, 15(S)-lipoxygenase-1; acLDL, acetylated low density lipoprotein; AU, arbitrary unit; CE, cholesteryl ester; ECAPCI, electron capture atmospheric pressure chemical ionization; FC, free cholesterol; 15(S)-HETE, 15(S)-hydroxyeicosatetraenoic acid; HODE, hydroxyoctadecadienoic acid; MRM, multiple reaction monitoring

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