Hepatic FoxOs induce apolipoprotein M and are required for sphingosine-1-phosphate to bind high density lipoproteins

Abstract

The authors declare no conflicts of interest.Abbreviations: AAV=adeno-associated virus, ApoM=apolipoprotein M, DIO=diet induced obese, FPLC=fast protein liquid chromatography, HDL=high density lipoprotein, HFD=high fat diet, LC-MS/MS=liquid chromatography-mass spectrometry, LDL=low density lipoprotein, LPD=lipoprotein depleted, S1P=sphingosine-1-phosphate, Tbg=thyroxin binding globulin, VLDL=very low density lipoprotein, WTD=western type diet ABSTRACTThe FoxO family of transcription factors play an important role in mediating insulin action on glucose, lipid, and lipoprotein metabolism. Liver-specific triple FoxO knockout mice (L-FoxO1,3,4) have defects in expression of genes related to glucose production, bile acid synthesis, and high density lipoprotein (HDL)-cholesterol uptake. We have now identified Apolipoprotein M (Apom) as a novel transcriptional target of liver FoxO. ApoM is a liver-secreted apolipoprotein that is bound to HDL in the circulation, and it serves as a chaperone for the bioactive lipid, sphingosine-1-phosphate (S1P). Several recent studies have demonstrated that S1P bound to ApoM induces unique effects, compared to S1P bound to albumin. We now show that liver FoxOs are required for ApoM mRNA and protein expression, and that ApoM is a transcriptional target of FoxOs. Moreover, while total plasma S1P levels are similar between control and L-FoxO1,3,4 mice, S1P is nearly absent from HDL in L-FoxO1,3,4 mice, and is instead increased in the lipoprotein depleted fraction. We also observed that leptin receptor deficient db/db mice have low hepatic Apom mRNA, and low levels of ApoM and S1P in HDL, without changes in total plasma S1P. These data demonstrate that FoxO transcription factors are novel regulators of the ApoM-S1P pathway, and indicate a potential link between hepatic insulin action and HDL function. METHODS Mice and diets.All experiments were approved by the Columbia University Institutional Animal Care and Use Committee. L-FoxO1,3,4 mice have been previously described 5,9 . Males at least 12 weeks old were studied, except in studies of 2-day-old pups, which were of mixed sex. For the acute FoxO depletion experiments, adult Foxo1 fl/fl , Foxo3 fl/fl , and Foxo4 fl/Y control mice were transduced with adeno-associated virus (serotype 8) expressing Cre recombinase driven by the hepatocyte-specific Tbg promoter (AAV8.Tbg. Cre) or control virus (AAV.GFP). Mice were injected intravenously with 1×10 11 virus particles/mouse, 4 weeks prior to euthanasia. AAV8.Tbg.Cre was a gift ). For the adenovirus experiments, adult male mice were injected intravenously with murine ApoM adenovirus (Welgen) 0.5 × 10 9 virus particles/gram of body weight, 8 days prior to euthanasia. For the db/db studies, male db/db and db/+ mice were purchased from Jackson Laboratory and were studied when they were 12, 13 or 25 weeks old. For the diet induced obesity studies, male C57BL/6J mice were purchased from Jackson Laboratory when they were 6 weeks old. Mice were fed either a standard chow diet (Purina) or a high fat diet ...