DOI QR코드

DOI QR Code

The Effect of L-Ornithine on the Phosphorylation of mTORC1 Downstream Targets in Rat Liver

  • Kokubo, Takeshi (Research and Development Division, Kirin Company, Limited) ;
  • Maeda, Shyuichi (Research Laboratories for Health Science and Food Technologies, Kirin Company, Limited) ;
  • Tazumi, Kyoko (Research Laboratories for Health Science and Food Technologies, Kirin Company, Limited) ;
  • Nozawa, Hajime (Research Laboratories for Key Technologies, Kirin Company, Limited) ;
  • Miura, Yutaka (Research Laboratories for Health Science and Food Technologies, Kirin Company, Limited) ;
  • Kirisako, Takayoshi (Research Laboratories for Key Technologies, Kirin Company, Limited)
  • Received : 2015.09.24
  • Accepted : 2015.12.08
  • Published : 2015.12.31

Abstract

A non-protein amino acid, L-ornithine (Orn), has been shown to stimulate the urea cycle and tissue protein synthesis in the liver. The purpose of the current study was to assess whether Orn affects the mammalian target of rapamycin (mTOR) complex 1 (mTORC1) pathway, which is involved in protein synthesis. Primary cultured cells isolated from Wistar rat liver were incubated in an amino acid-free medium, followed by addition of Orn for 3 h. The cell lysate was subjected to immunoblotting to evaluate the phosphorylation of downstream targets of mTORC1, including p70S6K, S6, and 4EBP1. To assess the involvement of mTORC1 for the effect of Orn, the cells were pretreated with the mTOR inhibitor rapamycin before the addition of Orn and the cell lysate was subjected to immunoblotting. We next examined whether the effects of Orn were exerted in vivo. Orn was orally administered to 18 h food-deprived rats, the blood and the livers were collected at 1 and 3 h after administration for immunoblotting. Orn treatment for primary cultured cells for 3 h enhanced the phosphorylation of p70S6K, S6, and 4EBP1. In addition, rapamycin blocked the effects of Orn completely (p70S6K and S6) or partially (4EBP1). The oral administration of Orn to the rat also augmented the phosphorylation of mTORC1 downstream targets notably in S6 at 1 h. Our findings demonstrate that Orn has the potential to induce the phosphorylation of downstream targets of mTORC1 in the rat liver. This may be mediated by the augmentation of mTORC1 activity.

Keywords

References

  1. Uchisawa H, Sato A, Ichita J, Matsue H, Ono T. 2004. Influence of low-temperature processing of the brackish-water bivalve, Corbicula japonica, on the ornithine content of its extract. Biosci Biotechnol Biochem 68: 1228-1234. https://doi.org/10.1271/bbb.68.1228
  2. Antoine FR, Wei CI, Littell RC, Quinn BP, Hogle AD, Marshall MR. 2001. Free amino acids in dark-and white-muscle fish as determined by O-phthaldialdehyde precolumn derivatization. J Food Sci 66: 72-77. https://doi.org/10.1111/j.1365-2621.2001.tb15584.x
  3. Frau M, Massanet J, Rossello C, Simal S, Canellas J. 1997. Evolution of free amino acid content during ripening of Mahon cheese. Food Chem 60: 651-657. https://doi.org/10.1016/S0308-8146(97)00051-4
  4. Morris SM Jr. 2002. Regulation of enzymes of the urea cycle and arginine metabolism. Annu Rev Nutr 22: 87-105. https://doi.org/10.1146/annurev.nutr.22.110801.140547
  5. Krebs HA, Hems R, Lund P. 1973. Accumulation of amino acids by the perfused rat liver in the presence of ethanol. Biochem J 134: 697-705. https://doi.org/10.1042/bj1340697
  6. Staedt U, Leweling H, Gladisch R, Kortsik C, Hagmuller E, Holm E. 1993. Effects of ornithine aspartate on plasma ammonia and plasma amino acids in patients with cirrhosis. A double-blind, randomized study using a four-fold crossover design. J Hepatol 19: 424-430. https://doi.org/10.1016/S0168-8278(05)80553-7
  7. Sugino T, Shirai T, Kajimoto Y, Kajimoto O. 2008. L-ornithine supplementation attenuates physical fatigue in healthy volunteers by modulating lipid and amino acid metabolism. Nutr Res 28: 738-743. https://doi.org/10.1016/j.nutres.2008.08.008
  8. Kokubo T, Ikeshima E, Kirisako T, Miura Y, Horiuchi M, Tsuda A. 2013. A randomized, double-masked, placebo-controlled crossover trial on the effects of L-ornithine on salivary cortisol and feelings of fatigue of flushers the morning after alcohol consumption. Biopsychosoc Med 7: 6. https://doi.org/10.1186/1751-0759-7-6
  9. Demura S, Yamada T, Yamaji S, Komatsu M, Morishita K. 2010. The effect of L-ornithine hydrochloride ingestion on human growth hormone secretion after strength training. Adv Biosci Biotechnol 1: 7-11. https://doi.org/10.4236/abb.2010.11002
  10. Horiuchi M, Kanesada H, Miyata T, Watanabe K, Nishimura A, Kokubo T, Kirisako T. 2013. Ornithine ingestion improved sleep disturbances but was not associated with correction of blood tryptophan ratio in Japanese Antarctica expedition members during summer. Nutr Res 33: 557-564. https://doi.org/10.1016/j.nutres.2013.05.001
  11. Miyake M, Kirisako T, Kokubo T, Miura Y, Morishita K, Okamura H, Tsuda A. 2014. Randomised controlled trial of the effects of L-ornithine on stress markers and sleep quality in healthy workers. Nutr J 13: 53. https://doi.org/10.1186/1475-2891-13-53
  12. Tujioka K, Yamada T, Aoki M, Morishita K, Hayase K, Yokogoshi H. 2012. Dietary ornithine affects the tissue protein synthesis rate in young rats. J Nutr Sci Vitaminol (Tokyo) 58: 297-302. https://doi.org/10.3177/jnsv.58.297
  13. Marshall S. 2006. Role of insulin, adipocyte hormones, and nutrient-sensing pathways in regulating fuel metabolism and energy homeostasis: a nutritional perspective of diabetes, obesity, and cancer. Sci STKE 2006: re7.
  14. Patti ME, Kahn BB. 2004. Nutrient sensor links obesity with diabetes risk. Nat Med 10: 1049-1050. https://doi.org/10.1038/nm1004-1049
  15. Betz C, Hall MN. 2013. Where is mTOR and what is it doing there?. J Cell Biol 203: 563-574. https://doi.org/10.1083/jcb.201306041
  16. Proud CG. 2004. mTOR-mediated regulation of translation factors by amino acids. Biochem Biophys Res Commun 313: 429-436. https://doi.org/10.1016/j.bbrc.2003.07.015
  17. Reiter AK, Anthony TG, Anthony JC, Jefferson LS, Kimball SR. 2004. The mTOR signaling pathway mediates control of ribosomal protein mRNA translation in rat liver. Int J Biochem Cell Biol 36: 2169-2179. https://doi.org/10.1016/j.biocel.2004.04.004
  18. Kwon G, Marshall CA, Pappan KL, Remedi MS, McDaniel ML. 2004. Signaling elements involved in the metabolic regulation of mTOR by nutrients, incretins, and growth factors in islets. Diabetes 53: S225-S232. https://doi.org/10.2337/diabetes.53.suppl_3.S225
  19. Le Plenier S, Walrand S, Noirt R, Cynober L, Moinard C. 2012. Effects of leucine and citrulline versus non-essential amino acids on muscle protein synthesis in fasted rat: a common activation pathway?. Amino Acids 43: 1171-1178. https://doi.org/10.1007/s00726-011-1172-z
  20. Schanne FA, Kane AB, Young EE, Farber JL. 1979. Calcium dependence of toxic cell death: a final common pathway. Science 206: 700-702. https://doi.org/10.1126/science.386513
  21. Dutcher JP. 2004. Mammalian target of rapamycin (mTOR) inhibitors. Curr Oncol Rep 6: 111-115. https://doi.org/10.1007/s11912-004-0022-5
  22. Shigemitsu K, Tsujishita Y, Miyake H, Hidayat S, Tanaka N, Hara K, Yonezawa K. 1999. Structural requirement of leucine for activation of p70 S6 kinase. FEBS Lett 447: 303-306. https://doi.org/10.1016/S0014-5793(99)00304-X
  23. Choo AY, Yoon SO, Kim SG, Roux PP, Blenis J. 2008. Rapamycin differentially inhibits S6Ks and 4E-BP1 to mediate cell-type-specific repression of mRNA translation. Proc Natl Acad Sci USA 105: 17414-17419. https://doi.org/10.1073/pnas.0809136105
  24. Yoshizawa F, Endo M, Ide H, Yagasaki K, Funabiki R. 1995. Translational regulation of protein synthesis in the liver and skeletal muscle of mice in response to refeeding. J Nutr Biochem 6: 130-136. https://doi.org/10.1016/0955-2863(95)00018-U
  25. Yoshizawa F, Kimball SR, Vary TC, Jefferson LS. 1998. Effect of dietary protein on translation initiation in rat skeletal muscle and liver. Am J Physiol 275: E814-E820.
  26. Norton LE, Layman DK, Bunpo P, Anthony TG, Brana DV, Garlick PJ. 2009. The leucine content of a complete meal directs peak activation but not duration of skeletal muscle protein synthesis and mammalian target of rapamycin signaling in rats. J Nutr 139: 1103-1109. https://doi.org/10.3945/jn.108.103853
  27. Sans MD, Tashiro M, Vogel NL, Kimball SR, D'Alecy LG, Williams JA. 2006. Leucine activates pancreatic translational machinery in rats and mice through mTOR independently of CCK and insulin. J Nutr 136: 1792-1799. https://doi.org/10.1093/jn/136.7.1792
  28. Anthony JC, Anthony TG, Layman DK. 1999. Leucine supplementation enhances skeletal muscle recovery in rats following exercise. J Nutr 129: 1102-1106. https://doi.org/10.1093/jn/129.6.1102
  29. D'Agata R, Vigneri R. 1971. Circadian variations of human growth hormone serum levels. Longitudinal study. Ann Endocrinol (Paris) 32: 383-387.
  30. Hayashi AA, Proud CG. 2007. The rapid activation of protein synthesis by growth hormone requires signaling through mTOR. Am J Physiol Endocrinol Metab 292: E1647-E1655. https://doi.org/10.1152/ajpendo.00674.2006

Cited by

  1. Liver metabolomics study reveals protective function of Phyllanthus urinaria against CCl 4 -induced liver injury vol.15, pp.7, 2017, https://doi.org/10.1016/S1875-5364(17)30078-X
  2. Supplements with purported effects on muscle mass and strength pp.1436-6215, 2019, https://doi.org/10.1007/s00394-018-1882-z