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Clinical Science (2010) 118, (727–736) (Printed in Great Britain)

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Myriocin-mediated up-regulation of hepatocyte apoA-I synthesis is associated with ERK inhibition

Elias N. Glaros^*†, Woojin S. Kim^*† and Brett Garner^*†‡

*Prince of Wales Medical Research Institute, Sydney, NSW 2031, Australia, †School of Medical Sciences, Faculty of Medicine, University of New South Wales, Sydney, NSW 2052, Australia, and ‡School of Biological Sciences, University of Wollongong, Wollongong, NSW 2522, Australia

Key words: apolipoprotein A-I (apoA-I), atherosclerosis, extracellular-signal regulated kinase (ERK), hepatocyte, myriocin, sphingolipid.

Abbreviations: 4-AAP, 4-aminoantipyrine; apoA-I, apolipoprotein A-I; apoE, apolipoprotein-E; DAOS, N-ethyl-N-(2-hydroxy-3-sulfopropyl)-3,5-dimethoxyaniline sodium salt; DMEM, Dulbecco's modified Eagle's medium; ERK, extracellular-signal-related kinase; FCS, fetal calf serum; GCS, glucosylceramide synthase; GSL, glycosphingolipid; HDL, high-density lipoprotein; HRP, horseradish peroxidase; i.p., intraperitoneal(ly); LDL, low-density lipoprotein; MAPK, mitogen-activated protein kinase; MEK, MAPK/ERK kinase; MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide; qPCR, quantitative PCR; S1P, sphingosine 1-phosphate; SM, sphingomyelin; SMase, sphingomyelinase; SPT, serine palmitoyltransferase.

Correspondence: Professor Brett Garner (email b.garner@powmri.edu.au).

Sphingolipids including sphingomyelin have been implicated as potential atherogenic lipids. Studies in apoE (apolipoprotein E)-null mice have revealed that the serine palmitoyltransferase inhibitor myriocin reduces plasma levels of sphingomyelin, ceramide, sphingosine-1-phosphate and glycosphingolipids and that this is associated with potent inhibition of atherosclerosis. Interestingly, hepatic apoA-I (apolipoprotein A-I) synthesis and plasma HDL (high-density lipoprotein)-cholesterol levels were also increased in apoE-null mice treated with myriocin. Since myriocin is a known inhibitor of ERK (extracellular-signal-related kinase) phosphorylation, we assessed the possibility that myriocin may be acting to increase hepatic apoA-I production via this pathway. To address this, HepG2 cells and primary mouse hepatocytes were treated with 200 μM myriocin for up to 48 h. Myriocin increased apoA-I mRNA and protein levels by approx. 3- and 2-fold respectively. Myriocin also increased apoA-I secretion up to 3.5-fold and decreased ERK phosphorylation by approx. 70%. Similar findings were obtained when primary hepatocytes were isolated from apoE-null mice that were treated with myriocin (intraperitoneal injection at a dose of 0.3 mg/kg body weight). Further experiments revealed that the MEK (mitogen-activated protein kinase/ERK kinase) inhibitor PD98059 potently inhibited ERK phosphorylation, as expected, and increased primary hepatocyte apoA-I production by 3-fold. These results indicate that ERK phosphorylation plays a role in regulating hepatic apoA-I expression and suggest that the anti-atherogenic mechanism of action for myriocin may be linked to this pathway.

Received 3 September 2009/14 December 2009; accepted 26 January 2010

Published as Immediate Publication 26 January 2010, doi:10.1042/CS20090452

© 2010 The Author(s)
The author(s) has paid for this article to be freely available under the terms of the Creative Commons Attribution Non-Commercial Licence (http://creativecommons.org/licenses/by-nc/2.5/) which permits unrestricted non-commercial use, distribution and reproduction in any medium, provided the original work is properly cited.