Abstract
Many metabolic diseases in fish are often associated with lowered mitochondrial fatty acid β-oxidation (FAO). However, the physiological role of mitochondrial FAO in lipid metabolism has not been verified in many carnivorous fish species, for example in largemouth bass (Micropterus salmonids). In the present study, a specific mitochondrial FAO inhibitor, mildronate (MD), was used to investigate the effects of impaired mitochondrial FAO on growth performance, health status, and lipid metabolism of largemouth bass. The results showed that the dietary MD treatment significantly suppressed growth performance and caused heavy lipid accumulation, especially neutral lipid, in the liver. The MD-treated fish exhibited lower monounsaturated fatty acid and higher long-chain polyunsaturated fatty acids in the muscle. The MD treatment downregulated the gene expressions in lipolysis and lipogenesis, as well as the expressions of the genes and some key proteins in FAO without enhancing peroxisomal FAO. Additionally, the MD-treated fish had lower serum aspartate aminotransferase activity and lower pro-inflammation- and apoptosis-related genes in the liver. Taken together, MD treatment markedly induced lipid accumulation via depressing lipid catabolism. Our findings reveal the pivotal roles of mitochondrial FAO in maintaining health and lipid homeostasis in largemouth bass and could be hopeful in understanding metabolic diseases in farmed carnivorous fish.
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References
Amoah A, Coyle SD, Webster CD et al (2008) Effects of graded levels of carbohydrate on growth and survival of largemouth bass, Micropterus salmoides. J World Aquac Soc 39:397–405. https://doi.org/10.1111/j.1749-7345.2008.00168.x
Anderson FH, Zeng L, Rock NR, Yoshida EM (2000) An assessment of the clinical utility of serum ALT and AST in chronic hepatitis C. Hepatol Res 18:63–71. https://doi.org/10.1016/S1386-6346(99)00085-6.10.1016/S1386-6346(99)00085-6
Aurousseau B, Bauchart D, Calichon E et al (2004) Effect of grass or concentrate feeding systems and rate of growth on triglyceride and phospholipid and their fatty acids in the M. longissimus thoracis of lambs. Meat Sci 66:531–541. https://doi.org/10.1016/S0309-1740(03)00156-6
Bright LA, Coyle SD, Tidwell JH (2005) Effect of dietary lipid level and protein energy ratio on growth and body composition of largemouth bass Micropterus salmoides. J World Aquac Soc 36:129–134. https://doi.org/10.1111/j.1749-7345.2005.tb00139.x
Cejas JR, Almansa E, Villamandos JE et al (2003) Lipid and fatty acid composition of ovaries from wild fish and ovaries and eggs from captive fish of white sea bream (Diplodus sargus). Aquaculture 216:299–313. https://doi.org/10.1016/S0044-8486(02)00525-2
Chen Y, Sun Z, Liang Z et al (2020) Addition of L-carnitine to formulated feed improved growth performance, antioxidant status and lipid metabolism of juvenile largemouth bass, Micropterus salmoides. Aquaculture 518:734434. https://doi.org/10.1016/j.aquaculture.2019.734434
Craig S, Helfrich LA, Kuhn D, Schwarz MH (2017) Understanding fish nutrition, feeds, and feeding. Fisheries. 420–256, Virginia Cooperative Extension 4 pp
Crockett EL, Sidell BD (1993a) Substrate selectivities differ for hepatic mitochondrial and peroxisomal β-oxidation in an Antarctic fish, Notothenia gibberifrons. Biochem J 289:427–433. https://doi.org/10.1042/bj2890427
Crockett EL, Sidell BD (1993b) Peroxisomal beta-oxidation is a significant pathway for catabolism of fatty acids in a marine teleost. Am J Physiol Integr Comp Physiol 264:R1004–R1009. https://doi.org/10.1152/ajpregu.1993.264.5.R1004
da Costa RM, Fais RS, Dechandt CRP et al (2017) Increased mitochondrial ROS generation mediates the loss of the anti-contractile effects of perivascular adipose tissue in high-fat diet obese mice. Br J Pharmacol 174:3527–3541. https://doi.org/10.1111/bph.13687
Degrace P, Demizieux L, Du ZY et al (2007) Regulation of lipid flux between liver and adipose tissue during transient hepatic steatosis in carnitine-depleted rats. J Biol Chem 282:20816–20826. https://doi.org/10.1074/jbc.M611391200
Di Cristo F, Finicelli M, Digilio FA et al (2019) Meldonium improves Huntington’s disease mitochondrial dysfunction by restoring peroxisome proliferator-activated receptor γ coactivator 1α expression. J Cell Physiol 234:9233–9246. https://doi.org/10.1002/jcp.27602
Du Z-Y, Ma T, Liaset B et al (2013) Dietary eicosapentaenoic acid supplementation accentuates hepatic triglyceride accumulation in mice with impaired fatty acid oxidation capacity. Biochim Biophys Acta (BBA)-Molecular Cell Biol Lipids. 1831:291–299. https://doi.org/10.1016/j.bbalip.2012.10.002
Du ZY, Clouet P, Degrace P et al (2008a) Hypolipidaemic effects of fenofibrate and fasting in the herbivorous grass carp (Ctenopharyngodon idella) fed a high-fat diet. Br J Nutr 100:1200–1212. https://doi.org/10.1017/S0007114508986840
Du ZY, Clouet P, Huang LM et al (2008b) Utilization of different dietary lipid sources at high level in herbivorous grass carp (Ctenopharyngodon idella): mechanism related to hepatic fatty acid oxidation. Aquac Nutr 14:77–92. https://doi.org/10.1111/j.1365-2095.2007.00507.x
Dutta A, Ray K, Singh VK et al (2008) L-carnitine supplementation attenuates intermittent hypoxia-induced oxidative stress and delays muscle fatigue in rats. Exp Physiol 93:1139–1146. https://doi.org/10.1113/expphysiol.2008.042465
Elmore S (2007) Apoptosis: a review of programmed cell death. Toxicol Pathol 35:495–516. https://doi.org/10.1080/01926230701320337
Fogarty CE, Bergmann A (2017) Killers creating new life: caspases drive apoptosis-induced proliferation in tissue repair and disease. Cell Death Differ 24:1390–1400. https://doi.org/10.1038/cdd.2017.47
Goodwin AE, Lochmann RT, Tieman DM, Mitchell AJ (2002) Massive hepatic necrosis and nodular regeneration in largemouth bass fed diets high in available carbohydrate. J World Aquac Soc 33:466–477. https://doi.org/10.1111/j.1749-7345.2002.tb00026.x
Harpaz S (2005) L-Carnitine and its attributed functions in fish culture and nutrition - A review. Aquaculture 249:3–21. https://doi.org/10.1016/j.aquaculture.2005.04.007
Hayashi Y, Muranaka Y, Kirimoto T et al (2000) Effects of MET-88, a γ-butyrobetaine hydroxylase inhibitor, on tissue carnitine and lipid levels in rats. Biol Pharm Bull 23:770–773. https://doi.org/10.1248/bpb.23.770
Hemre G, Mommsen TP, Krogdahl Å (2002) Carbohydrates in fish nutrition: effects on growth, glucose metabolism and hepatic enzymes. Aquac Nutr 8:175–194. https://doi.org/10.1046/j.1365-2095.2002.00200.x
Henderson RJ, Sargent JR (1985) Chain-length specificities of mitochondrial and peroxisomal beta-oxidation of fatty acids in livers of rainbow trout (Salmo gairdneri). Comp Biochem Physiol B 82:79–85. https://doi.org/10.1016/0305-0491(85)90131-2
Horiuchi M, Kobayashi K, Yamaguchi S et al (1994) Primary defect of juvenile visceral steatosis (jvs) mouse with systemic carnitine deficiency is probably in renal carnitine transport system. Biochim Biophys Acta (BBA)-Molecular Basis Dis 1226:25–30. https://doi.org/10.1016/0925-4439(94)90054-X
Hovik R, Osmundsen H (1987) Peroxisomal β-oxidation of long-chain fatty acids possessing different extents of unsaturation. Biochem J 247:531–535. https://doi.org/10.1042/bj2470531
Jain S, Singh SN (2015) Effect of L-carnitine supplementation on nutritional status and physical performance under calorie restriction. Indian J Clin Biochem 30:187–193. https://doi.org/10.1007/s12291-014-0437-1
Jänicke RU, Sprengart ML, Wati MR, Porter AG (1998) Caspase-3 is required for DNA fragmentation and morphological changes associated with apoptosis. J Biol Chem 273:9357–9360. https://doi.org/10.1074/jbc.273.16.9357
Jin M, Pan T, Tocher DR, et al (2019) Dietary choline supplementation attenuated high-fat diet-induced inflammation through regulation of lipid metabolism and suppression of NFκB activation in juvenile black seabream (Acanthopagrus schlegelii). J Nutr Sci 8:. https://doi.org/10.1017/jns.2019.34
Johnston IA, Bower NI, Macqueen DJ (2011) Growth and the regulation of myotomal muscle mass in teleost fish. J Exp Biol 214:1617–1628. https://doi.org/10.1242/jeb.038620
Karin M, Clevers H (2016) Reparative inflammation takes charge of tissue regeneration. Nature 529:307–315. https://doi.org/10.1038/nature17039
Kim S, Sohn I, Ahn J-I et al (2004) Hepatic gene expression profiles in a long-term high-fat diet-induced obesity mouse model. Gene 340:99–109. https://doi.org/10.1016/j.gene.2004.06.015
Kreeft AJ, Moen CJA, Porter G et al (2005) Genomic analysis of the response of mouse models to high-fat feeding shows a major role of nuclear receptors in the simultaneous regulation of lipid and inflammatory genes. Atherosclerosis 182:249–257. https://doi.org/10.1016/j.atherosclerosis.2005.01.049
Lazarow PB (1978) Rat liver peroxisomes catalyze the beta oxidation of fatty acids. J Biol Chem 253:1522–1528. https://doi.org/10.1016/S0021-9258(17)34897-4
Lazarow PB, De Duve C (1976) A fatty acyl-CoA oxidizing system in rat liver peroxisomes; enhancement by clofibrate, a hypolipidemic drug. Proc Natl Acad Sci 73:2043–2046. https://doi.org/10.1073/pnas.73.6.2043
Li J-M, Li L-Y, Qin X et al (2018) Inhibited carnitine synthesis causes systemic alteration of nutrient metabolism in zebrafish. Front Physiol 9:509. https://doi.org/10.3389/fphys.2018.00509
Li L-Y, Li J-M, Ning L-J, et al (2020a) Mitochondrial fatty acid β-oxidation inhibition promotes glucose utilization and protein deposition through energy homeostasis remodeling in fish. J Nutr 150:2322–2335. Inhibited carnitine synthesis impairs adaptation to high-fat diet in Nile tilapia (Oreochromis niloticus)
Li L-Y, Li J-M, Qiao F et al (2020b) Inhibited carnitine synthesis impairs adaptation to high-fat diet in Nile tilapia (Oreochromis niloticus). Aquac Reports 16:100249. https://doi.org/10.1016/j.aqrep.2019.100249
Li LY, Limbu SM, Ma Q et al (2019) The metabolic regulation of dietary L-carnitine in aquaculture nutrition: present status and future research strategies. Rev Aquac 11:1228–1257. https://doi.org/10.1111/raq.12289
Li X, Jiang Y, Liu W, Ge X (2012) Protein-sparing effect of dietary lipid in practical diets for blunt snout bream (Megalobrama amblycephala) fingerlings: effects on digestive and metabolic responses. Fish Physiol Biochem 38:529–541. https://doi.org/10.1007/s10695-011-9533-9
Liepinsh E, Makrecka-Kuka M, Kuka J et al (2015) Inhibition of L-carnitine biosynthesis and transport by methyl-γ-butyrobetaine decreases fatty acid oxidation and protects against myocardial infarction. Br J Pharmacol 172:1319–1332. https://doi.org/10.1111/bph.13004
Liepinsh E, Vilskersts R, Zvejniece L et al (2009) Protective effects of mildronate in an experimental model of type 2 diabetes in Goto-Kakizaki rats. Br J Pharmacol 157:1549–1556. https://doi.org/10.1111/j.1476-5381.2009.00319.x
Liu G, Yu H, Wang C et al (2021) Nano-selenium supplements in high-fat diets relieve hepatopancreas injury and improve survival of grass carp Ctenopharyngodon Idella by reducing lipid deposition. Aquaculture 538:736580. https://doi.org/10.1016/j.aquaculture.2021.736580
Liu Y, Han SL, Luo Y et al (2020) Impaired peroxisomal fat oxidation induces hepatic lipid accumulation and oxidative damage in Nile tilapia. Fish Physiol Biochem 46:1229–1242. https://doi.org/10.1007/s10695-020-00785-w
Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative pcr and the 2−ΔΔCT Method. Methods 25:402–408. https://doi.org/10.1006/meth.2001.1262
Luo H, Jiang M, Lian G et al (2018) AIDA Selectively mediates downregulation of fat synthesis enzymes by ERAD to retard intestinal fat absorption and prevent obesity. Cell Metab 27:843-853.e6. https://doi.org/10.1016/j.cmet.2018.02.021
Madureira TV, Castro LFC, Rocha E (2016) Acyl-coenzyme A oxidases 1 and 3 in brown trout (Salmo trutta f. fario): can peroxisomal fatty acid β-oxidation be regulated by estrogen signaling? Fish Physiol Biochem 42:389–401. https://doi.org/10.1007/s10695-015-0146-6
Makrecka M, Kuka J, Liepinsh E, Dambrova M (2011) 04 The regulation of mitochondrial energy metabolism by L-carnitine lowering agents in ischaemia-reperfusion injury. Heart 97:e8–e8. https://doi.org/10.1136/heartjnl-2011-301156.4
Mannaerts GP, Debeer LJ, Thomas J, De Schepper PJ (1979) Mitochondrial and peroxisomal fatty acid oxidation in liver homogenates and isolated hepatocytes from control and clofibrate-treated rats. J Biol Chem 254:4585–4595. https://doi.org/10.1016/S0021-9258(17)30051-0
Ning L-J, He A-Y, Li J-M et al (2016) Mechanisms and metabolic regulation of PPARα activation in Nile tilapia (Oreochromis niloticus). Biochim Biophys Acta (BBA)-Molecular Cell Biol Lipids 1861:1036–1048. https://doi.org/10.1016/j.bbalip.2016.06.005
Ning L-J, He A-Y, Lu D-L et al (2017) Nutritional background changes the hypolipidemic effects of fenofibrate in Nile tilapia (Oreochromis niloticus). Sci Rep 7:1–11. https://doi.org/10.1038/srep41706
Ning LJ, He AY, Li JM et al (2016b) Mechanisms and metabolic regulation of PPARα activation in Nile tilapia (Oreochromis niloticus). Biochim Biophys Acta - Mol Cell Biol Lipids 1861:1036–1048. https://doi.org/10.1016/j.bbalip.2016.06.005
Pan H, Li LY, Li JM et al (2017) Inhibited fatty acid β-oxidation impairs stress resistance ability in Nile tilapia (Oreochromis niloticus). Fish Shellfish Immunol 68:500–508. https://doi.org/10.1016/j.fsi.2017.07.058
Pérez-Garijo A, Martín FA, Morata G (2004) Caspase inhibition during apoptosis causes abnormal signalling and developmental aberrations in Drosophila. Development 131:5591–5598. https://doi.org/10.1242/dev.01432
Peschechera A, Scalibastri M, Russo F et al (2005) Carnitine depletion in rat pups from mothers given mildronate: a model of carnitine deficiency in late fetal and neonatal life. Life Sci 77:3078–3091. https://doi.org/10.1016/j.lfs.2005.03.029
Pupure J, Isajevs S, Skapare E et al (2010) Neuroprotective properties of mildronate, a mitochondria-targeted small molecule. Neurosci Lett 470:100–105. https://doi.org/10.1016/j.neulet.2009.12.055
Ruggiero C, Ehrenshaft M, Cleland E, Stadler K (2011) High-fat diet induces an initial adaptation of mitochondrial bioenergetics in the kidney despite evident oxidative stress and mitochondrial ROS production. Am J Physiol Metab 300:E1047–E1058. https://doi.org/10.1152/ajpendo.00666.2010
Schulz H (1991) Beta oxidation of fatty acids. Biochim Biophys Acta (BBA)-Lipids Lipid Metab 1081:109–120. https://doi.org/10.1016/0005-2760(91)90015-A
Shi X, Jin A, Sun J et al (2018) The protein-sparing effect of α-lipoic acid in juvenile grass carp, Ctenopharyngodon idellus: effects on lipolysis, fatty acid β-oxidation and protein synthesis. Br J Nutr 120:977–987. https://doi.org/10.1017/S000711451800226X
Simkhovich BZ, Shutenko ZV, Meirēna DV et al (1988) 3-(2, 2, 2-Trimethylhydrazinium) propionate (thp)-a novel γ-butyrobetaine hydroxylase inhibitor with cardioprotective properties. Biochem Pharmacol 37:195–202. https://doi.org/10.1016/0006-2952(88)90717-4
Stephens FB, Constantin-Teodosiu D, Greenhaff PL (2007) New insights concerning the role of carnitine in the regulation of fuel metabolism in skeletal muscle. J Physiol 581:431–444. https://doi.org/10.1113/jphysiol.2006.125799
Stephens FB, Wall BT, Marimuthu K et al (2013) Skeletal muscle carnitine loading increases energy expenditure, modulates fuel metabolism gene networks and prevents body fat accumulation in humans. J Physiol 591:4655–4666. https://doi.org/10.1113/jphysiol.2013.255364
Stubhaug I, Lie Ø, Torstensen BE (2007) Fatty acid productive value and β-oxidation capacity in Atlantic salmon (Salmo salar L.) fed on different lipid sources along the whole growth period. Aquac Nutr 13:145–155. https://doi.org/10.1111/j.1365-2095.2007.00462.x
Sun JL, He K, Liu Q, et al (2020) Inhibition of fatty acid oxidation induced by up regulation of miR-124 and miR-205 during exposure of largemouth bass (Micropterus salmoides) to acute hypoxia. Aquaculture 529:735679. https://doi.org/10.1016/j.aquaculture.2020.735679
Wall BT, Stephens FB, Constantin-Teodosiu D et al (2011) Chronic oral ingestion of l-carnitine and carbohydrate increases muscle carnitine content and alters muscle fuel metabolism during exercise in humans. J Physiol 589:963–973. https://doi.org/10.1113/jphysiol.2010.201343
Wang J, Liang D, Yang Q et al (2020) The effect of partial replacement of fish meal by soy protein concentrate on growth performance, immune responses, gut morphology and intestinal inflammation for juvenile hybrid grouper (Epinephelus fuscoguttatus ♀ × Epinephelus lanceolatus ♂). Fish Shellfish Immunol 98:619–631. https://doi.org/10.1016/j.fsi.2019.10.025
Wilson RP (1994) Utilization of dietary carbohydrate by fish. Aquaculture 124:67–80. https://doi.org/10.1016/0044-8486(94)90363-8
Xu H, Meng X, Jia L et al (2020) Tissue distribution of transcription for 29 lipid metabolism-related genes in Takifugu rubripes, a marine teleost storing lipid predominantly in liver. Fish Physiol Biochem 46:1603–1619. https://doi.org/10.1007/s10695-020-00815-7
Yan J, Liao K, Wang T et al (2015) Dietary lipid levels influence lipid deposition in the liver of large yellow croaker (Larimichthys crocea) by regulating lipoprotein receptors, fatty acid uptake and triacylglycerol synthesis and catabolism at the transcriptional level. PLoS ONE 10:e0129937. https://doi.org/10.1371/journal.pone.0129937
Yin P, Xie S, Zhuang Z et al (2021) Dietary supplementation of bile acid attenuate adverse effects of high-fat diet on growth performance, antioxidant ability, lipid accumulation and intestinal health in juvenile largemouth bass (Micropterus salmoides). Aquaculture 531:735864. https://doi.org/10.1016/j.aquaculture.2020.735864
Yu LL, Yu HH, Liang XF et al (2018) Dietary butylated hydroxytoluene improves lipid metabolism, antioxidant and anti-apoptotic response of largemouth bass (Micropterus salmoides). Fish Shellfish Immunol 72:220–229. https://doi.org/10.1016/j.fsi.2017.10.054
Yu H, Zhang L, Chen P et al (2019) Dietary bile acids enhance growth, and alleviate hepatic fibrosis induced by a high starch diet via AKT/FOXO1 and cAMP/AMPK/SREBP1 pathway in Micropterus salmoides. Front Physiol 10:1–15. https://doi.org/10.3389/fphys.2019.01430
Zeng J, Deng S, Wang Y et al (2017) Specific inhibition of Acyl-CoA oxidase-1 by an acetylenic acid improves hepatic lipid and reactive oxygen species (ROS) metabolism in rats fed a high fat diet. J Biol Chem 292:3800–3809. https://doi.org/10.1074/jbc.M116.763532
Zhang D, Yan Y, Tian H et al (2018) Resveratrol supplementation improves lipid and glucose metabolism in high-fat diet-fed blunt snout bream. Fish Physiol Biochem 44:163–173. https://doi.org/10.1007/s10695-017-0421-9
Zhou YL, Guo JL, Tang RJ et al (2020) High dietary lipid level alters the growth, hepatic metabolism enzyme, and anti-oxidative capacity in juvenile largemouth bass Micropterus salmoides. Fish Physiol Biochem 46:125–134. https://doi.org/10.1007/s10695-019-00705-7
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Z.-Y. D. and J.-X. W.: conceived the study and designed the experiments. J.-X. W.: carried out the experiments, analyzed the data, and wrote the manuscript. Y.-Y. Z., J. R., J. W., F. Q., L.-Q. C., and M.-L. Z.: contributed reagents/materials/analysis tools. J.-X. W., S. R., and Z.-Y. D.: wrote and revised the manuscript. All authors read and approved the final manuscript.
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Wang, JX., Rahimnejad, S., Zhang, YY. et al. Mildronate triggers growth suppression and lipid accumulation in largemouth bass (Micropterus salmoides) through disturbing lipid metabolism. Fish Physiol Biochem 48, 145–159 (2022). https://doi.org/10.1007/s10695-021-01040-6
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DOI: https://doi.org/10.1007/s10695-021-01040-6