Fatty Acid Regulation of Liver X Receptors (LXR) and Peroxisome Proliferator-activated Receptor α (PPARα) in HEK293 Cells (2002)

Fatty acids bind to and regulate the activity of peroxisome proliferator-activated (PPAR) and liver X receptors (LXR). However, the role lipid metabolism plays in the control of intracellular fatty acid ligands is poorly understood. We have identified two strains of HEK293 cells that display differences in fatty acid regulation of nuclear receptors. Using full-length and Gal4-LBD chimeric receptors in functional assays, 20:4,n6 induced PPARα activity ∼2.2-fold and suppressed LXRα activity by 80% (ED50 ∼25–50 μm) in HEK293-E (early passage) cells but had no effect on PPARα or LXRα receptor activity in HEK293-L (late passage) cells. LXRβ was insensitive to fatty acid regulation in both HEK293 strains. Metabolic labeling studies using [14C]20:4,n6 (at 100 μm) indicated that the uptake of 20:4,n6 and its assimilation into triacylglycerol, diacylglycerol, and polar lipids revealed no difference between the two strains. Such treatment increased total cellular 20:4,n6 (∼11-fold) and its elongation product, 22:4,n6 (∼3.6-fold), within 6 h. Non-esterified 20:4,n6 and 22:4,n6 represented ≤3% of the total cellular 20:4,n6 and 22:4,n6. In HEK293-E cells, non-esterified 20:4,n6 and 22:4,n6 increased 8- and 18-fold, respectively, by 6 h and was sustained at that level for 24 h. In HEK293-L cells, non-esterified 20:4,n6 also increased (5-fold) at 6 h but fell by 70% within 24 h. In contrast to HEK293-E cells, non-esterified 22:4,n6 did not accumulate in HEK293-L cells. Functional assays showed that 22:4,n6 was ∼2-fold more effective than 20:4,n6 at inhibiting oxysterol-induced LXRα activity in HEK293-E cells, but had no effect on LXRα activity in HEK293-L cells. Taken together, these findings demonstrate that the rate of assimilation of exogenously added fatty acids and their metabolites into complex lipids plays an important role in regulating PPARα and LXRα activity. Fatty acids bind to and regulate the activity of peroxisome proliferator-activated (PPAR) and liver X receptors (LXR). However, the role lipid metabolism plays in the control of intracellular fatty acid ligands is poorly understood. We have identified two strains of HEK293 cells that display differences in fatty acid regulation of nuclear receptors. Using full-length and Gal4-LBD chimeric receptors in functional assays, 20:4,n6 induced PPARα activity ∼2.2-fold and suppressed LXRα activity by 80% (ED50 ∼25–50 μm) in HEK293-E (early passage) cells but had no effect on PPARα or LXRα receptor activity in HEK293-L (late passage) cells. LXRβ was insensitive to fatty acid regulation in both HEK293 strains. Metabolic labeling studies using [14C]20:4,n6 (at 100 μm) indicated that the uptake of 20:4,n6 and its assimilation into triacylglycerol, diacylglycerol, and polar lipids revealed no difference between the two strains. Such treatment increased total cellular 20:4,n6 (∼11-fold) and its elongation product, 22:4,n6 (∼3.6-fold), within 6 h. Non-esterified 20:4,n6 and 22:4,n6 represented ≤3% of the total cellular 20:4,n6 and 22:4,n6. In HEK293-E cells, non-esterified 20:4,n6 and 22:4,n6 increased 8- and 18-fold, respectively, by 6 h and was sustained at that level for 24 h. In HEK293-L cells, non-esterified 20:4,n6 also increased (5-fold) at 6 h but fell by 70% within 24 h. In contrast to HEK293-E cells, non-esterified 22:4,n6 did not accumulate in HEK293-L cells. Functional assays showed that 22:4,n6 was ∼2-fold more effective than 20:4,n6 at inhibiting oxysterol-induced LXRα activity in HEK293-E cells, but had no effect on LXRα activity in HEK293-L cells. Taken together, these findings demonstrate that the rate of assimilation of exogenously added fatty acids and their metabolites into complex lipids plays an important role in regulating PPARα and LXRα activity. peroxisome proliferator activated receptor liver X receptor retinoid X receptor polyunsaturated fatty acids non-esterified fatty acids butylated hydroxytoluene Dulbecco's modified Eagle's medium bovine serum albumin ligand binding domain triacylglycerol, DAG, Diacylglycerol cholesterol esters reverse phase-high pressure liquid chromatography Several nuclear receptors have been identified as targets for regulation by fatty acids or their metabolites. These include the peroxisome proliferator-activated receptor family (PPARα1 (NR1C1), PPARδ (NR1C2), PPARγ (NR1C3)), liver X receptors (LXRα (NR1H3) and LXRβ (NR1H2)), RXRα, and HNF-4α (1Jump D.B. J. Biol. Chem. 2002; 277: 8755-8758Abstract Full Text Full Text PDF PubMed Scopus (517) Google Scholar). The best characterized are members of the PPAR family. All PPARs bind 20-carbon polyunsaturated fatty acids, e.g. 20:4,n6 and 20:5,n3, with an apparent K d of ∼1–4 μm (2Desvergne B. Wahli W. Endocr. Rev. 1999; 20: 649-688Crossref PubMed Scopus (2746) Google Scholar). As class II nuclear receptors, PPARs heterodimerize with RXR and bind direct repeats (PPREs, a DR1, rat AOX PPRE: CCGAACGTGACCTNTGTCCT) in promoters of regulated genes. PPARs play a major role in whole body lipid metabolism, inflammatory, and immune responses (3Xu H.E. Lambert M.H. Montana V.G. Parks D.J. Blanchard S.G. Brown P.J. Sernbach D.D. Lehmann J.M. Wisely G.B. Willson T.M. Kliewer S.A. Milburn M.V. Mol. Cell. 1999; 3: 397-403Abstract Full Text Full Text PDF PubMed Scopus (970) Google Scholar). Liver X receptors (LXRα and LXRβ) are also class II nuclear receptors that bind direct repeats (LXRE, a DR-4, murine Cyp7A1: TGGTCActcaAGTTCA) as a heterodimer with RXRα (4Lu T.T. Repa J.J. Mangelsdorf D.J. J. Biol. Chem. 2001; 276: 37735-37738Abstract Full Text Full Text PDF PubMed Scopus (327) Google Scholar). Oxysterols, like 22(R)-hydroxycholesterol or 24,25-epoxycholesterol, bind and activate LXRs (5Joseph S.B. Laffitte B.A. Patel P.H. Watson M.A. Matsukuma K.E. Walczak R. Collins J. Osborne T.F. Tontonoz P. J. Biol. Chem. 2002; 277: 11019-11025Abstract Full Text Full Text PDF PubMed Scopus (626) Google Scholar, 6Ou J., Tu, H. Shan B. Luk A. DeBose-Boyd R.A. Bashmakov Y. Goldstein J.L. Brown M.S. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 6027-6032Crossref PubMed Scopus (409) Google Scholar, 7Schultz J.R., Tu, H. Luk A. Repa J.J. Medina J.C., Li, L. Schwendner S. Wang S. Thoolen M. Mangelsdorf D.J. Lustig K.D. Shan B. Genes Dev. 2000; 14: 2831-2838Crossref PubMed Scopus (1404) Google Scholar). LXR/RXR heterodimers bind LXREs in promoters of enzymes involved in hepatic bile acid synthesis, e.g.7α-hydroxylase (Cyp7A), the main route for cholesterol elimination from the body. LXRs also regulate lipogenic gene expression through two mechanisms either by controlling the expression of SREBP-1c or by direct binding to promoters of certain lipogenic genes, e.g.fatty acid synthase (5Joseph S.B. Laffitte B.A. Patel P.H. Watson M.A. Matsukuma K.E. Walczak R. Collins J. Osborne T.F. Tontonoz P. J. Biol. Chem. 2002; 277: 11019-11025Abstract Full Text Full Text PDF PubMed Scopus (626) Google Scholar, 7Schultz J.R., Tu, H. Luk A. Repa J.J. Medina J.C., Li, L. Schwendner S. Wang S. Thoolen M. Mangelsdorf D.J. Lustig K.D. Shan B. Genes Dev. 2000; 14: 2831-2838Crossref PubMed Scopus (1404) Google Scholar). In contrast to PPARs, unsaturated fatty acids act as antagonist to oxysterol activation of LXRα in HEK293 and hepatoma cell lines (6Ou J., Tu, H. Shan B. Luk A. DeBose-Boyd R.A. Bashmakov Y. Goldstein J.L. Brown M.S. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 6027-6032Crossref PubMed Scopus (409) Google Scholar). The hierarchy for the fatty acid effect on LXR is 20:4,n6 > 18:2,n6 > 18:1,n9; saturated fatty acids have no effect. The half-maximal effect for 20:4,n6 antagonism of oxysterol binding to the LXR-ligand binding domain is ∼1.5 μm. This level of fatty acid is comparable to the K d for 20-carbon PUFA binding to PPARs (3Xu H.E. Lambert M.H. Montana V.G. Parks D.J. Blanchard S.G. Brown P.J. Sernbach D.D. Lehmann J.M. Wisely G.B. Willson T.M. Kliewer S.A. Milburn M.V. Mol. Cell. 1999; 3: 397-403Abstract Full Text Full Text PDF PubMed Scopus (970) Google Scholar). Based on in vitro binding and cell culture studies both PPARs and LXR have emerged as prospective monitors of intracellular NEFA levels, and in liver these receptors would respond accordingly by altering metabolism to prevent lipid and cholesterol overload. However, there remains little information that documents changes in intracellular NEFA levels in cells or how lipid metabolism might contribute to the regulation of NEFA levels and influence nuclear receptor activity. During the course of our studies, we observed that the fatty acid regulation of PPARα and LXRα in HEK293 cells was strain-dependent. This difference in fatty acid responsiveness provided us with an opportunity to identify prospective metabolic pathways that impact intracellular NEFA levels and affect nuclear receptor activity. Our studies show that while both cell types share the metabolic pathways for 20:4,n6 assimilation complex there are These differences contribute to the level of non-esterified PUFA in cells in LXR and PPAR activity. Using the HEK293 cell provided for the that both LXRα and PPARα are of intracellular strains of HEK293 identified as HEK293-E (early passage) and HEK293-L (late passage) from and at the of at in with serum in culture or culture or to and in fatty acid and J.M. D.B. J. 2000; Full Text Full Text PDF PubMed Google Scholar, B.A. S.A. Wisely G.B. Kliewer S.A. Mangelsdorf D.J. Proc. Natl. Acad. Sci. U. S. A. 1999; PubMed Scopus Google Scholar). was using the the to the The for and was from and as an control for All to in culture in of and of receptor expression using in an cells with fatty acids or for 24 h in bovine serum albumin The was to a of fatty acid to LXRs activated with 22(R)-hydroxycholesterol at μm. The for 22(R)-hydroxycholesterol is cells and for and activity using the with a was using the with as All in and at to in in to fatty acid treatment cells in with [14C]20:4,n6 of 100 μm fatty acids from and fatty acids from Fatty is added to μm. with fatty acids for and 24 h. was cells with μm with and in of The was with to and lipids with butylated hydroxytoluene A. J. Biol. Chem. Full Text PDF PubMed Google Scholar). and with The in and at are by chromatography in acid and a of lipids was with for cholesterol esters fatty acids, fatty acid esters and and of into the was by liquid of fatty acids from was by and chromatography by as HEK293 cells in with 100 μm fatty acid as cells in and was added as a lipid was as lipids in 80% h at in and in for to with a of acid to acid was to unsaturated fatty and of unsaturated fatty acids by fatty acid and for from The of fatty acids was by at the at The NEFA in total cellular lipids was on J. Full Text PDF PubMed Google Scholar). in to a and with to NEFA with on the The acid was in and for and of unsaturated fatty The of NEFA from whole cells was PPARα is a for fatty acid regulation (1Jump D.B. J. Biol. Chem. 2002; 277: 8755-8758Abstract Full Text Full Text PDF PubMed Scopus (517) Google Scholar, B. Wahli W. Endocr. Rev. 1999; 20: 649-688Crossref PubMed Scopus (2746) Google Scholar, H.E. Lambert M.H. Montana V.G. Parks D.J. Blanchard S.G. Brown P.J. Sernbach D.D. Lehmann J.M. Wisely G.B. Willson T.M. Kliewer S.A. Milburn M.V. Mol. Cell. 1999; 3: 397-403Abstract Full Text Full Text PDF PubMed Scopus (970) Google Scholar). However, LXRα been identified as a for fatty acid regulation (6Ou J., Tu, H. Shan B. Luk A. DeBose-Boyd R.A. Bashmakov Y. Goldstein J.L. Brown M.S. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 6027-6032Crossref PubMed Scopus (409) Google Scholar). In we characterized the fatty acid regulation of oxysterol activation of LXRα and LXRβ and fatty acid regulation of PPARα in two strains of HEK293 cells. These studies show that these two strains of HEK293 cells for fatty acid on LXR and PPAR activity. The two strains of HEK293 cells in HEK293-E are cells and HEK293-L are cells. HEK293-L cells than HEK293-E with a of HEK293-L cells to a than HEK293-E However, and fatty acid metabolism studies at in cell was of 20:4,n6 and on PPAR gene expression in HEK293-E and are with and the are with 20:4,n6 or at 100 μm activity and was J.M. D.B. J. 2000; Full Text Full Text PDF PubMed Google and as of cells with The was > are as in or activity. These are of two acid on oxysterol of LXR activity in HEK293-L cells. HEK293-L cells with expression full-length and as in A. The was and the control for was > These are of acid on oxysterol of LXR activity in HEK293-E cells. HEK293-E cells with expression full-length and The was and the control for was > HEK293-E cells with expression to the The was of the binding an cells or with 22(R)-hydroxycholesterol and and with fatty acids at 100 μm for 24 h. and for and activity and cells as in and an cells or with μm 22(R)-hydroxycholesterol and and with fatty acids 20:5,n3, and at 100 24 h. > These are of at two Fatty acid regulation of nuclear receptors was by using either full-length receptors or Gal4-LBD chimeric receptors. cells with the and expression full-length LXRα or LXRβ The LXR 22(R)-hydroxycholesterol induced a and in activity in HEK293-E cells with full-length LXRα and 22(R)-hydroxycholesterol had no effect on activity LXR expression from the of cells with 100 μm had no effect on either or of activity. of cells with or 20:4,n6 the effect of 22(R)-hydroxycholesterol on by and LXRβ was induced by 22(R)-hydroxycholesterol The oxysterol of LXRβ was not by fatty acid the effect of fatty acids on LXRs is for of the fatty acid on LXR receptors the LXR to the binding domain of HEK293-E cells with to a comparable level of of activity 22(R)-hydroxycholesterol treatment as with the full-length the effect of fatty acids on the oxysterol regulation of was the as that with the full-length but not was to unsaturated fatty acid (at a LXR B.A. S.A. Wisely G.B. Kliewer S.A. Mangelsdorf D.J. Proc. Natl. Acad. Sci. U. S. A. 1999; PubMed Scopus Google for 22(R)-hydroxycholesterol the for both the LXRα and LXRβ as in and B. for their on activation was as effective as 20:4,n6 at while was the effective at of These on the effect of unsaturated fatty acids on the oxysterol regulation of LXRα (6Ou J., Tu, H. Shan B. Luk A. DeBose-Boyd R.A. Bashmakov Y. Goldstein J.L. Brown M.S. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 6027-6032Crossref PubMed Scopus (409) Google Scholar). We have these to include and as effective of oxysterol activation of However, PUFA oxysterol activation of in the HEK293-L cells a to fatty 22(R)-hydroxycholesterol induced activity by 8- and in cells with the LXRα and LXRβ expression This is comparable to that in the HEK293-E cells In contrast to the HEK293-E cells, oxysterol activation of LXRα and LXRβ in HEK293-L cells was insensitive to fatty acid In both the LXRα and chimeric Gal4-LBD receptors insensitive to fatty acid regulation in HEK293-L cells. levels of fatty acids μm) had no effect on activity fatty acid regulation of LXRα was we the fatty acid regulation of a fatty nuclear in the two HEK293 strains. As the fatty acid regulation of PPARα is (2Desvergne B. Wahli W. Endocr. Rev. 1999; 20: 649-688Crossref PubMed Scopus (2746) Google Scholar, H.E. Lambert M.H. Montana V.G. Parks D.J. Blanchard S.G. Brown P.J. Sernbach D.D. Lehmann J.M. Wisely G.B. Willson T.M. Kliewer S.A. Milburn M.V. Mol. Cell. 1999; 3: 397-403Abstract Full Text Full Text PDF PubMed Scopus (970) Google Scholar). The two HEK293 cell strains with a the from the and the gene with the expression with 100 μm 20:4,n6 or PPARα for 24 h. In HEK293-E with activity was induced ∼2.2-fold by 20:4,n6 or induced activity in HEK293-L cells, 20:4,n6 had no effect on activity. In the of activity was not by either 20:4,n6 or like PPARα is not to fatty acid regulation in HEK293-L cells. We the fatty acid of LXRα and PPARα in HEK293-E cells by a In the chimeric receptor was to the of LXRα to fatty acid The effect of 20:4,n6 on activity was an 80% at μm of of LXRα by 20:4,n6 was at μm. of fatty acid activation of PPARα both the full-length expression and the to activate the or genes, 20:4,n6 induced gene activity This level was comparable to the effect of μm) on gene activity. The for the effect of 20:4,n6 was μm. These that in HEK293-E cells, both LXRα and PPARα display a to exogenously added The fatty acid regulation of LXRα and PPARα is in the two HEK293 strains. As there are that for difference as cell and However, the of cells at and fatty acid in these on metabolic differences in the cell and to how these two cell types fatty both receptors bind NEFA (3Xu H.E. Lambert M.H. Montana V.G. Parks D.J. Blanchard S.G. Brown P.J. Sernbach D.D. Lehmann J.M. Wisely G.B. Willson T.M. Kliewer S.A. Milburn M.V. Mol. Cell. 1999; 3: 397-403Abstract Full Text Full Text PDF PubMed Scopus (970) Google Scholar, 6Ou J., Tu, H. Shan B. Luk A. DeBose-Boyd R.A. Bashmakov Y. Goldstein J.L. Brown M.S. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 6027-6032Crossref PubMed Scopus (409) Google we that difference in to levels of non-esterified 20:4,n6 in HEK293-E cells than in HEK293-L cells. two to 20:4,n6 metabolism in these cells. The a of [14C]20:4,n6 uptake and The changes of fatty acids in cells treatment with fatty HEK293 cells with [14C]20:4,n6 at 100 μm the as for the the assimilation of [14C]20:4,n6 into the of cells and the of into lipid difference in fatty acid uptake or its assimilation into the was of lipids for the of the of [14C]20:4,n6 into DAG, and polar lipids there was no differences in the of [14C]20:4,n6 between DAG, or polar lipids between the two cells. the is between the two cell types at This is of cholesterol and NEFA are differences in the of fatty acids in these the fatty acid ≤3% of the fatty acid into the of these cells. HEK293-E cells a of the fatty acid into cholesterol while HEK293-L have a of fatty acid into a of In the level of 20:4,n6 assimilation into the level of fatty acid assimilation into these studies differences in the level of acid in the NEFA and its assimilation into two lipid cholesterol and The of fatty acids important as to how fatty acids are to lipid However, to how of fatty acids affect levels of fatty acids as as the of fatty acid we the fatty acid in the total as as the NEFA cells with 20:4,n6 at 100 μm and total lipid was from cells at and 24 h lipids in the and by is to unsaturated fatty acids, fatty acid using and is a major unsaturated fatty acid in both cell types The indicated that both and are lipids in HEK293 cells and are PUFA at 100 of cells with 100 μm 20:4,n6 to a and in total 20:4,n6 in HEK293-E cells and a and in HEK293-L cells and 24 acid increased in cell 20:4,n6 the of 20:4,n6 to 22:4,n6 by an The changes in 22:4,n6 and for HEK293-E cells and for HEK293-L cells. the of 20:4,n6 to major changes in cellular 20:4,n6 and 22:4,n6 levels in both cell of 20:4,n6 to cells did not major changes in cellular levels of or the course of the NEFA of the total lipid the was on and by 6 The of are represented as a of total fatty acid from the cell and The of and 22:4,n6 in the NEFA the of these fatty acids in the total cell In HEK293-E cells, these fatty acids at and and in HEK293-L cells the fatty acids and of these non-esterified fatty acids of total or in the 20:4,n6 remains between and of the total cellular 20:4,n6 the treatment in HEK293-E cells In non-esterified 20:4,n6 in HEK293-L cells from in cells to 24 h of This difference more assimilation of 20:4,n6 into complex lipids in HEK293-L cells than HEK293-E cells. of 22:4,n6 that fatty acid from at the of the to of the total 22:4,n6 within 6 h in HEK293-E cells, but remains between and in HEK293-L cells. As with 22:4,n6 is more from the NEFA and into complex lipids in HEK293-L cells than HEK293-E cells. unsaturated fatty acids in the NEFA the treatment that acid is in cells 20:4,n6 treatment activity in both cell the effect of acid on LXRα a was acid μm) was ∼2-fold more than 20:4,n6 (ED50 μm) at inhibiting LXRα activity in HEK293-E cells. fatty acid LXRα activity in HEK293-L cells, to μm. Based on these metabolism studies, we that non-esterified 20:4,n6 and its are more into complex lipids in HEK293-L cells than in HEK293-E cells. This difference in of fatty acid and its from the NEFA for the differences in fatty acid regulation of LXRα and PPARα in these two cell This was to the fatty acid regulation of these studies we that cell the of these PPARα and LXRα bind NEFA with an d of μm (3Xu H.E. Lambert M.H. Montana V.G. Parks D.J. Blanchard S.G. Brown P.J. Sernbach D.D. Lehmann J.M. Wisely G.B. Willson T.M. Kliewer S.A. Milburn M.V. Mol. Cell. 1999; 3: 397-403Abstract Full Text Full Text PDF PubMed Scopus (970) Google Scholar, 6Ou J., Tu, H. Shan B. Luk A. DeBose-Boyd R.A. Bashmakov Y. Goldstein J.L. Brown M.S. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 6027-6032Crossref PubMed Scopus (409) Google both receptors respond to a in intracellular NEFA of fatty acids to cells. However, our studies with two strains of HEK293 cells responses to fatty acid The that lipid metabolism to nuclear receptor regulation been in the of regulation of PPARs S.A. S.A. Brown P.J. Wisely G.B. P. Wahli W. Willson T.M. J.M. Lehmann J.M. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar, W. Brown M. M.A. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google or the on and polyunsaturated fatty acids on lipogenic gene expression (1Jump D.B. J. Biol. Chem. 2002; 277: 8755-8758Abstract Full Text Full Text PDF PubMed Scopus (517) Google Scholar, D.B. Rev. 14: PubMed Scopus Google Scholar, D.B. Rev. 1999; PubMed Scopus Google Scholar). However, we are of that the rate of in intracellular NEFA or their metabolites of fatty acids to cells and these changes to nuclear receptor Our studies an (6Ou J., Tu, H. Shan B. Luk A. DeBose-Boyd R.A. Bashmakov Y. Goldstein J.L. Brown M.S. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 6027-6032Crossref PubMed Scopus (409) Google by that oxysterol regulation of LXRα is suppressed by unsaturated fatty We have to include the and as of at 100 μm 20:4,n6 > > > 20:4,n6 the LXRα with an d of μm (6Ou J., Tu, H. Shan B. Luk A. DeBose-Boyd R.A. Bashmakov Y. Goldstein J.L. Brown M.S. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 6027-6032Crossref PubMed Scopus (409) Google the for the effect on LXRα activity is μm (6Ou J., Tu, H. Shan B. Luk A. DeBose-Boyd R.A. Bashmakov Y. Goldstein J.L. Brown M.S. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 6027-6032Crossref PubMed Scopus (409) Google the K d for fatty acid binding and the for the activation of PPARα is in (3Xu H.E. Lambert M.H. Montana V.G. Parks D.J. Blanchard S.G. Brown P.J. Sernbach D.D. Lehmann J.M. Wisely G.B. Willson T.M. Kliewer S.A. Milburn M.V. Mol. Cell. 1999; 3: 397-403Abstract Full Text Full Text PDF PubMed Scopus (970) Google at a intracellular level of non-esterified or or both PPARα and LXRα Our studies with that to and LXRβ is to than LXRα to oxysterol activation B.A. S.A. Wisely G.B. Kliewer S.A. Mangelsdorf D.J. Proc. Natl. Acad. Sci. U. S. A. 1999; PubMed Scopus Google we to in contrast to LXRβ was insensitive to fatty acid regulation The acid of the of the two receptors at these differences for the levels of fatty and This also that in both receptors are LXRα to fatty acid Our studies with have revealed are to PUFA while are J. J. and B. in these two receptors regulate of using LXRα and LXRβ J.J. J. Bashmakov Y. J.M. Shan B. Brown M.S. Goldstein J.L. Mangelsdorf D.J. Genes Dev. 2000; 14: PubMed Scopus Google Scholar). The differences we in nuclear receptor regulation by fatty acids in the two HEK293 strains the of how these cells The of both metabolic labeling and of 20:4,n6 and its elongation product, by into fatty acid metabolism and of of exogenously added fatty acids from the intracellular NEFA for a more assimilation of fatty acid as as its in the HEK293-L cells than in HEK293-E cells. This is on the rate of of [14C]20:4,n6 as as the of 20:4,n6 and 22:4,n6 and in the NEFA We no of of [14C]20:4,n6 in HEK293-L cells. This was on the that levels of fatty acid into cells at the and that the of of was in HEK293-L cells 6 In of culture for of of or the of lipid to major differences between these two cell the difference between these two cell lines is in the assimilation of exogenously added fatty acid into complex This difference levels of expression of have been of fatty using saturated and unsaturated fatty acids of from R.A. T.M. Rev. 2000; 20: PubMed Scopus Google Scholar). However, is on and PUFA R.A. T.M. Rev. 2000; 20: PubMed Scopus Google Scholar). the difference in fatty acid between the HEK293-E and HEK293-L cells is to levels of expression of a remains to differences in fatty acid metabolism are in the assimilation of fatty acid into cholesterol esters and a with the of These differences levels of expression of enzymes involved in the of these complex or for cholesterol or synthase and for 2001; PubMed Scopus Google Scholar, of and Scholar). The of the HEK293-L cells to exogenously added fatty acid into lipids a for the of non-esterified 20:4,n6 and 22:4,n6 from the NEFA of cells. The more of and PUFA from the NEFA these NEFA to a level to affect LXRα or PPARα activity as by that might for the difference in nuclear receptor responsiveness between the two cell types is the assimilation of 22:4,n6 into complex lipids 6 and cell types exogenously added 20:4,n6 to 22:4,n6. However, with HEK293-L cells, HEK293-E cells are to 22:4,n6 into complex that 22:4,n6 is more than the of a more ligand with its assimilation in to complex lipids for the increased of HEK293-E cells to fatty acid regulation of nuclear the remains as to the mechanisms have on PPARα or LXRα regulation in e.g. in liver or of the studies on lipid metabolism have been in added of cells is that we of by PPARα and LXRα than on receptor activation is from these that intracellular NEFA levels are in to fatty acid and as a of of PUFA metabolism in is of for PPARα and LXRα changes in In we have identified two strains of HEK293 cells that display differences in nuclear receptor This difference to of assimilation of fatty acid and its elongation into complex PPARα and but not display comparable to fatty acid This that as PPARα is activated by intracellular LXRα in like the liver that major changes in lipid levels and intracellular NEFA affect both In the is that both and PUFA are PPARα and pathways to fatty and the of fatty acids into pathways (1Jump D.B. J. Biol. Chem. 2002; 277: 8755-8758Abstract Full Text Full Text PDF PubMed Scopus (517) Google Scholar, B. Wahli W. Endocr. Rev. 1999; 20: 649-688Crossref PubMed Scopus (2746) Google Scholar). NEFA as of LXRα and to the of of a controlling PUFA also regulate SREBP-1c levels by J. J. Biol. Chem. 1999; Full Text Full Text PDF PubMed Scopus Google Scholar, D.B. J. Biol. Chem. 1999; Full Text Full Text PDF PubMed Scopus Google Scholar, J. M. J. Biol. Chem. 2001; 276: Full Text Full Text PDF PubMed Scopus Google Scholar). PPARα and LXRα as monitors of intracellular NEFA levels and respond accordingly to prevent lipid A. Repa J.J. Mangelsdorf D.J. 2001; PubMed Scopus Google Scholar).