Treatment of human skeletal muscle cells with inhibitors of diacylglycerol acyltransferases 1 and 2 to explore isozyme-specific roles on lipid metabolism.
Acetic Acid
/ metabolism
Diacylglycerol O-Acyltransferase
/ antagonists & inhibitors
Enzyme Inhibitors
/ pharmacology
Glucose
/ metabolism
Glycerol
/ metabolism
Humans
Isoenzymes
/ antagonists & inhibitors
Lipid Metabolism
/ drug effects
Muscle Fibers, Skeletal
/ drug effects
Muscle, Skeletal
/ drug effects
Oxidation-Reduction
/ drug effects
Journal
Scientific reports
ISSN: 2045-2322
Titre abrégé: Sci Rep
Pays: England
ID NLM: 101563288
Informations de publication
Date de publication:
14 01 2020
14 01 2020
Historique:
received:
26
09
2019
accepted:
19
12
2019
entrez:
16
1
2020
pubmed:
16
1
2020
medline:
11
11
2020
Statut:
epublish
Résumé
Diacylglycerol acyltransferases (DGAT) 1 and 2 catalyse the final step in triacylglycerol (TAG) synthesis, the esterification of fatty acyl-CoA to diacylglycerol. Despite catalysing the same reaction and being present in the same cell types, they exhibit different functions on lipid metabolism in various tissues. Yet, their roles in skeletal muscle remain poorly defined. In this study, we investigated how selective inhibitors of DGAT1 and DGAT2 affected lipid metabolism in human primary skeletal muscle cells. The results showed that DGAT1 was dominant in human skeletal muscle cells utilizing fatty acids (FAs) derived from various sources, both exogenously supplied FA, de novo synthesised FA, or FA derived from lipolysis, to generate TAG, as well as being involved in de novo synthesis of TAG. On the other hand, DGAT2 seemed to be specialised for de novo synthesis of TAG from glycerol-3-posphate only. Interestingly, DGAT activities were also important for regulating FA oxidation, indicating a key role in balancing FAs between storage in TAG and efficient utilization through oxidation. Finally, we observed that inhibition of DGAT enzymes could potentially alter glucose-FA interactions in skeletal muscle. In summary, treatment with DGAT1 or DGAT2 specific inhibitors resulted in different responses on lipid metabolism in human myotubes, indicating that the two enzymes play distinct roles in TAG metabolism in skeletal muscle.
Identifiants
pubmed: 31937853
doi: 10.1038/s41598-019-57157-5
pii: 10.1038/s41598-019-57157-5
pmc: PMC6959318
doi:
Substances chimiques
Enzyme Inhibitors
0
Isoenzymes
0
Diacylglycerol O-Acyltransferase
EC 2.3.1.20
Glucose
IY9XDZ35W2
Glycerol
PDC6A3C0OX
Acetic Acid
Q40Q9N063P
Types de publication
Address
Journal Article
Research Support, Non-U.S. Gov't
Langues
eng
Sous-ensembles de citation
IM
Pagination
238Références
Sacchetti, M., Saltin, B., Olsen, D. B. & van Hall, G. High triacylglycerol turnover rate in human skeletal muscle. J. Physiol. 561, 883–891, https://doi.org/10.1113/jphysiol.2004.075135 (2004).
doi: 10.1113/jphysiol.2004.075135
pubmed: 15498807
pmcid: 1665384
Yen, C. L., Stone, S. J., Koliwad, S., Harris, C. & Farese, R. V. Jr. Thematic review series: glycerolipids. DGAT enzymes and triacylglycerol biosynthesis. J. Lipid Res. 49, 2283–2301, https://doi.org/10.1194/jlr.R800018-JLR200 (2008).
doi: 10.1194/jlr.R800018-JLR200
pubmed: 18757836
pmcid: 3837458
Cases, S. et al. Identification of a gene encoding an acyl CoA:diacylglycerol acyltransferase, a key enzyme in triacylglycerol synthesis. Proc. Natl Acad. Sci. USA 95, 13018–13023 (1998).
doi: 10.1073/pnas.95.22.13018
Cases, S. et al. Cloning of DGAT2, a second mammalian diacylglycerol acyltransferase, and related family members. J. Biol. Chem. 276, 38870–38876, https://doi.org/10.1074/jbc.M106219200 (2001).
doi: 10.1074/jbc.M106219200
pubmed: 11481335
Stone, S. J. et al. The endoplasmic reticulum enzyme DGAT2 is found in mitochondria-associated membranes and has a mitochondrial targeting signal that promotes its association with mitochondria. J. Biol. Chem. 284, 5352–5361, https://doi.org/10.1074/jbc.M805768200 (2009).
doi: 10.1074/jbc.M805768200
pubmed: 19049983
pmcid: 2643492
Kuerschner, L., Moessinger, C. & Thiele, C. Imaging of lipid biosynthesis: how a neutral lipid enters lipid droplets. Traffic (Copenhagen, Den.) 9, 338–352, https://doi.org/10.1111/j.1600-0854.2007.00689.x (2008).
doi: 10.1111/j.1600-0854.2007.00689.x
Wilfling, F. et al. Triacylglycerol Synthesis Enzymes Mediate Lipid Droplet Growth by Relocalizing from the ER to Lipid Droplets. Developmental Cell 24, 384–399, https://doi.org/10.1016/j.devcel.2013.01.013 (2013).
doi: 10.1016/j.devcel.2013.01.013
pubmed: 23415954
pmcid: 3727400
Chen, H. C. et al. Increased insulin and leptin sensitivity in mice lacking acyl CoA:diacylglycerol acyltransferase 1. The J. Clin. investigation 109, 1049–1055, https://doi.org/10.1172/JCI14672 (2002).
doi: 10.1172/JCI14672
Stone, S. J. et al. Lipopenia and skin barrier abnormalities in DGAT2-deficient mice. J. Biol. Chem. 279, 11767–11776, https://doi.org/10.1074/jbc.M311000200 (2004).
doi: 10.1074/jbc.M311000200
pubmed: 14668353
Smith, S. J. et al. Obesity resistance and multiple mechanisms of triglyceride synthesis in mice lacking Dgat. Nat. Genet. 25, 87–90, https://doi.org/10.1038/75651 (2000).
doi: 10.1038/75651
pubmed: 10802663
Irshad, Z., Dimitri, F., Christian, M. & Zammit, V. A. Diacylglycerol acyltransferase 2 links glucose utilization to fatty acid oxidation in the brown adipocytes. J. Lipid Res. 58, 15–30, https://doi.org/10.1194/jlr.M068197 (2017).
doi: 10.1194/jlr.M068197
pubmed: 27836993
Zammit, V. A. Hepatic triacylglycerol synthesis and secretion: DGAT2 as the link between glycaemia and triglyceridaemia. Biochem. J. 451, 1–12, https://doi.org/10.1042/bj20121689 (2013).
doi: 10.1042/bj20121689
pubmed: 23489367
Han, R. H., Wang, M., Fang, X. & Han, X. Simulation of triacylglycerol ion profiles: bioinformatics for interpretation of triacylglycerol biosynthesis. J. Lipid Res. 54, 1023–1032, https://doi.org/10.1194/jlr.M033837 (2013).
doi: 10.1194/jlr.M033837
pubmed: 23365150
pmcid: 3605979
Hung, Y.-H., Carreiro, A. L. & Buhman, K. K. Dgat1 and Dgat2 regulate enterocyte triacylglycerol distribution and alter proteins associated with cytoplasmic lipid droplets in response to dietary fat. Biochimica et. Biophysica Acta (BBA) - Mol. Cell Biol. Lipids 1862, 600–614, https://doi.org/10.1016/j.bbalip.2017.02.014 (2017).
doi: 10.1016/j.bbalip.2017.02.014
Wurie, H. R., Buckett, L. & Zammit, V. A. Diacylglycerol acyltransferase 2 acts upstream of diacylglycerol acyltransferase 1 and utilizes nascent diglycerides and de novo synthesized fatty acids in HepG2 cells. FEBS J. 279, 3033–3047, https://doi.org/10.1111/j.1742-4658.2012.08684.x (2012).
doi: 10.1111/j.1742-4658.2012.08684.x
pubmed: 22748069
Qi, J. et al. The use of stable isotope-labeled glycerol and oleic acid to differentiate the hepatic functions of DGAT1 and −2. J. Lipid Res. 53, 1106–1116, https://doi.org/10.1194/jlr.M020156 (2012).
doi: 10.1194/jlr.M020156
pubmed: 22493088
pmcid: 3351817
Chitraju, C., Walther, T. C. & Farese, R. V. The triglyceride synthesis enzymes DGAT1 and DGAT2 have distinct and overlapping functions in adipocytes. J. Lipid Res. https://doi.org/10.1194/jlr.M093112 (2019).
doi: 10.1194/jlr.M093112
pubmed: 30936184
pmcid: 6547635
Chitraju, C. et al. Triglyceride Synthesis by DGAT1 Protects Adipocytes from Lipid-Induced ER Stress during Lipolysis. Cell Metab. 26, 407–418.e403, https://doi.org/10.1016/j.cmet.2017.07.012 (2017).
doi: 10.1016/j.cmet.2017.07.012
pubmed: 28768178
pmcid: 6195226
Liu, L. et al. DGAT1 deficiency decreases PPAR expression and does not lead to lipotoxicity in cardiac and skeletal muscle. J. Lipid Res. 52, 732–744, https://doi.org/10.1194/jlr.M011395 (2011).
doi: 10.1194/jlr.M011395
pubmed: 21205704
pmcid: 3284165
Roe, N. D., Handzlik, M. K., Li, T. & Tian, R. The Role of Diacylglycerol Acyltransferase (DGAT) 1 and 2 in Cardiac Metabolism and Function. Sci. Rep. 8, 4983, https://doi.org/10.1038/s41598-018-23223-7 (2018).
doi: 10.1038/s41598-018-23223-7
pubmed: 29563512
pmcid: 5862879
Liu, L. et al. Upregulation of myocellular DGAT1 augments triglyceride synthesis in skeletal muscle and protects against fat-induced insulin resistance. J. Clin. Invest. 117, 1679–1689, https://doi.org/10.1172/jci30565 (2007).
doi: 10.1172/jci30565
pubmed: 17510710
pmcid: 1866250
Levin, M. C. et al. Increased lipid accumulation and insulin resistance in transgenic mice expressing DGAT2 in glycolytic (type II) muscle. Am. J. Physiol. Endocrinol. Metab. 293, E1772–1781, https://doi.org/10.1152/ajpendo.00158.2007 (2007).
doi: 10.1152/ajpendo.00158.2007
pubmed: 17940217
Ehrenborg, E. & Krook, A. Regulation of skeletal muscle physiology and metabolism by peroxisome proliferator-activated receptor delta. Pharmacol. Rev. 61, 373–393, https://doi.org/10.1124/pr.109.001560 (2009).
doi: 10.1124/pr.109.001560
pubmed: 19805479
van Loon, L. J. & Goodpaster, B. H. Increased intramuscular lipid storage in the insulin-resistant and endurance-trained state. Pflug. Arch. 451, 606–616, https://doi.org/10.1007/s00424-005-1509-0 (2006).
doi: 10.1007/s00424-005-1509-0
Walther, T. C. & Farese, R. V. Jr. Lipid droplets and cellular lipid metabolism. Annu. Rev. Biochem. 81, 687–714, https://doi.org/10.1146/annurev-biochem-061009-102430 (2012).
doi: 10.1146/annurev-biochem-061009-102430
pubmed: 22524315
pmcid: 3767414
Zhao, G. et al. Validation of diacyl glycerolacyltransferase I as a novel target for the treatment of obesity and dyslipidemia using a potent and selective small molecule inhibitor. J. Med. Chem. 51, 380–383, https://doi.org/10.1021/jm7013887 (2008).
doi: 10.1021/jm7013887
pubmed: 18183944
Gaster, M., Kristensen, S. R., Beck-Nielsen, H. & Schroder, H. D. A cellular model system of differentiated human myotubes. Apmis 109, 735–744 (2001).
doi: 10.1034/j.1600-0463.2001.d01-140.x
Lund, J. et al. Glucose metabolism and metabolic flexibility in cultured skeletal muscle cells is related to exercise status in young male subjects. Arch. Physiol. Biochem. 124, 119–130, https://doi.org/10.1080/13813455.2017.1369547 (2018).
doi: 10.1080/13813455.2017.1369547
pubmed: 28862046
Lund, J. et al. Higher lipid turnover and oxidation in cultured human myotubes from athletic versus sedentary young male subjects. Sci. Rep. 8, 17549, https://doi.org/10.1038/s41598-018-35715-7 (2018).
doi: 10.1038/s41598-018-35715-7
pubmed: 30510272
pmcid: 6277406
Gaster, M., Rustan, A. C., Aas, V. & Beck-Nielsen, H. Reduced Lipid Oxidation in Skeletal Muscle From Type 2 Diabetic Subjects May Be of Genetic Origin. Evid. Cultured Myotubes 53, 542–548, https://doi.org/10.2337/diabetes.53.3.542 (2004).
doi: 10.2337/diabetes.53.3.542
Bradford, M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72, 248–254 (1976).
doi: 10.1016/0003-2697(76)90527-3
Schneider, C. A., Rasband, W. S. & Eliceiri, K. W. NIH Image to ImageJ: 25 years of image analysis. Nat. Methods 9, 671–675, https://doi.org/10.1038/nmeth.2089 (2012).
doi: 10.1038/nmeth.2089
pubmed: 5554542
pmcid: 5554542
Kase, E. T. et al. Liver X receptor antagonist reduces lipid formation and increases glucose metabolism in myotubes from lean, obese and type 2 diabetic individuals. Diabetologia 50, 2171–2180, https://doi.org/10.1007/s00125-007-0760-7 (2007).
doi: 10.1007/s00125-007-0760-7
pubmed: 17661008
Wensaas, A. J. et al. Cell-based multiwell assays for the detection of substrate accumulation and oxidation. J. Lipid Res. 48, 961–967, https://doi.org/10.1194/jlr.D600047-JLR200 (2007).
doi: 10.1194/jlr.D600047-JLR200
pubmed: 17213484
Skrede, S., Bremer, J., Berge, R. K. & Rustan, A. C. Stimulation of fatty acid oxidation by a 3-thia fatty acid reduces triacylglycerol secretion in cultured rat hepatocytes. J. Lipid Res. 35, 1395–1404 (1994).
pubmed: 7989864
Bakke, S. S. et al. Palmitic acid follows a different metabolic pathway than oleic acid in human skeletal muscle cells; lower lipolysis rate despite an increased level of adipose triglyceride lipase. Biochim. Biophys. Acta 1821, 1323–1333, https://doi.org/10.1016/j.bbalip.2012.07.001 (2012).
doi: 10.1016/j.bbalip.2012.07.001
pubmed: 22796147
Rupp, H., Zarain-Herzberg, A. & Maisch, B. The Use of Partial Fatty Acid Oxidation Inhibitors for Metabolic Therapy of Angina Pectoris and Heart Failure. Herz 27, 621–636, https://doi.org/10.1007/s00059-002-2428-x (2002).
doi: 10.1007/s00059-002-2428-x
pubmed: 12439634
Hessvik, N. P. et al. Metabolic switching of human myotubes is improved by n-3 fatty acids. J. Lipid Res. 51, 2090–2104, https://doi.org/10.1194/jlr.M003319 (2010).
doi: 10.1194/jlr.M003319
pubmed: 20363834
pmcid: 2903803
Gaster, M., Rustan, A. C. & Beck-Nielsen, H. Differential utilization of saturated palmitate and unsaturated oleate: evidence from cultured myotubes. Diabetes 54, 648–656 (2005).
doi: 10.2337/diabetes.54.3.648
Glatz, J. F., Luiken, J. J. & Bonen, A. Membrane fatty acid transporters as regulators of lipid metabolism: implications for metabolic disease. Physiol. Rev. 90, 367–417, https://doi.org/10.1152/physrev.00003.2009 (2010).
doi: 10.1152/physrev.00003.2009
pubmed: 20086080
Roorda, B. D. et al. DGAT1 overexpression in muscle by in vivo DNA electroporation increases intramyocellular lipid content. J. Lipid Res. 46, 230–236, https://doi.org/10.1194/jlr.M400416-JLR200 (2005).
doi: 10.1194/jlr.M400416-JLR200
pubmed: 15576838
Yang, F. et al. Upregulation of triglyceride synthesis in skeletal muscle overexpressing DGAT1. Lipids Health Dis. 12, 63–63, https://doi.org/10.1186/1476-511X-12-63 (2013).
doi: 10.1186/1476-511X-12-63
pubmed: 23642106
pmcid: 3671243
Ikeda, S. et al. Up-regulation of SREBP-1c and lipogenic genes in skeletal muscles after exercise training. Biochem. Biophys. Res. Commun. 296, 395–400 (2002).
doi: 10.1016/S0006-291X(02)00883-5
Schenk, S. & Horowitz, J. F. Acute exercise increases triglyceride synthesis in skeletal muscle and prevents fatty acid–induced insulin resistance. J. Clin. Investigation 117, 1690–1698, https://doi.org/10.1172/JCI30566 (2007).
doi: 10.1172/JCI30566
Gidda, S. K. et al. Hydrophobic-domain-dependent protein-protein interactions mediate the localization of GPAT enzymes to ER subdomains. Traffic 12, 452–472, https://doi.org/10.1111/j.1600-0854.2011.01160.x (2011).
doi: 10.1111/j.1600-0854.2011.01160.x
pubmed: 21214700
Man, W. C., Miyazaki, M., Chu, K. & Ntambi, J. Colocalization of SCD1 and DGAT2: implying preference for endogenous monounsaturated fatty acids in triglyceride synthesis. J. Lipid Res. 47, 1928–1939, https://doi.org/10.1194/jlr.M600172-JLR200 (2006).
doi: 10.1194/jlr.M600172-JLR200
pubmed: 16751624
Li, C. et al. Roles of Acyl-CoA:Diacylglycerol Acyltransferases 1 and 2 in Triacylglycerol Synthesis and Secretion in Primary Hepatocytes. Arterioscler. Thromb. Vasc. Biol. 35, 1080–1091, https://doi.org/10.1161/atvbaha.114.304584 (2015).
doi: 10.1161/atvbaha.114.304584
pubmed: 25792450
Aas, V., Kase, E. T., Solberg, R., Jensen, J. & Rustan, A. C. Chronic hyperglycaemia promotes lipogenesis and triacylglycerol accumulation in human skeletal muscle cells. Diabetologia 47, 1452–1461, https://doi.org/10.1007/s00125-004-1465-9 (2004).
doi: 10.1007/s00125-004-1465-9
pubmed: 15309295
van Loon, L. J., Greenhaff, P. L., Constantin-Teodosiu, D., Saris, W. H. & Wagenmakers, A. J. The effects of increasing exercise intensity on muscle fuel utilisation in humans. J. Physiol. 536, 295–304 (2001).
doi: 10.1111/j.1469-7793.2001.00295.x
Jordy, A. B. & Kiens, B. Regulation of exercise-induced lipid metabolism in skeletal muscle. Exp. Physiol. 99, 1586–1592, https://doi.org/10.1113/expphysiol.2014.082404 (2014).
doi: 10.1113/expphysiol.2014.082404
pubmed: 25398709
Randle, P. J., Garland, P. B., Hales, C. N. & Newsholme, E. A. The glucose fatty-acid cycle. Its role in insulin sensitivity and the metabolic disturbances of diabetes mellitus. Lancet 1, 785–789 (1963).
doi: 10.1016/S0140-6736(63)91500-9
Hue, L. & Taegtmeyer, H. The Randle cycle revisited: a new head for an old hat. Am. J. Physiol. Endocrinol. Metab. 297, E578–591, https://doi.org/10.1152/ajpendo.00093.2009 (2009).
doi: 10.1152/ajpendo.00093.2009
pubmed: 19531645
pmcid: 2739696
Coleman, R. A. & Mashek, D. G. Mammalian triacylglycerol metabolism: synthesis, lipolysis, and signaling. Chem. Rev. 111, 6359–6386, https://doi.org/10.1021/cr100404w (2011).
doi: 10.1021/cr100404w
pubmed: 21627334
pmcid: 3181269
Tomimoto, D. et al. JTT-553, a novel Acyl CoA:diacylglycerol acyltransferase (DGAT) 1 inhibitor, improves glucose metabolism in diet-induced obesity and genetic T2DM mice. J. Pharmacol. Sci. 129, 51–58, https://doi.org/10.1016/j.jphs.2015.08.005 (2015).
doi: 10.1016/j.jphs.2015.08.005
pubmed: 26354408
Senkal, C. E. et al. Ceramide Is Metabolized to Acylceramide and Stored in Lipid Droplets. Cell Metab. 25, 686–697, https://doi.org/10.1016/j.cmet.2017.02.010 (2017).
doi: 10.1016/j.cmet.2017.02.010
pubmed: 28273483
pmcid: 5472424