European Journal of Nutrition

, Volume 56, Issue 2, pp 635–647 | Cite as

AMPK activation by liquiritigenin inhibited oxidative hepatic injury and mitochondrial dysfunction induced by nutrition deprivation as mediated with induction of farnesoid X receptor

  • Eun Hye Jung
  • Ju-Hee Lee
  • Sang Chan Kim
  • Young Woo Kim
Original Contribution



Nutrition is indispensable for cell survival and proliferation. Thus, loss of nutrition caused by serum starvation in cells could induce formation of reactive oxygen species (ROS), resulting in cell death. Liquiritigenin (LQ) is an active flavonoid in licorice and plays a role in the liver as a hepatic protectant.


This study investigated the effect of LQ, metformin [an activator of activated AMP-activated protein kinase (AMPK)] and GW4064 [a ligand of farnesoid X receptor (FXR)] on mitochondrial dysfunction and oxidative stress induced by serum deprivation as well as its molecular mechanism, as assessed by immunoblot and flow cytometer assays.


Serum deprivation in HepG2, H4IIE and AML12 cells successfully induced oxidative stress and apoptosis, as indicated by depletion of glutathione, formation of ROS, and altered expression of apoptosis-related proteins such as procaspase-3, poly(ADP-ribose) polymerase, and Bcl-2. However, LQ pretreatment significantly blocked these pathological changes and mitochondrial dysfunction caused by serum deprivation. Moreover, LQ activated AMPK in HepG2 cells and mice liver, as shown by phosphorylation of AMPK and ACC, and this activation was mediated by its upstream kinase (i.e., LKB1). Experiments using a chemical inhibitor of AMPK with LKB1-deficient Hela cells revealed the role of the LKB1–AMPK pathway in cellular protection conferred by LQ. LQ also induced protein and mRNA expression of both FXR as well as small heterodimer partner, which is important since treatment with FXR ligand GW4064 protected hepatocytes against cell death and mitochondrial damage induced by serum deprivation.


AMPK activators such as LQ can protect hepatocytes against oxidative hepatic injury and mitochondrial dysfunction induced by serum deprivation, and the beneficial effect might be mediated through the LKB1 pathway as well as FXR induction.


AMPK FXR Liquiritigenin Mitochondria Nutrition deprivation 



Acetyl-CoA carboxylase


AMP-activated protein kinase


Fluorescence-activated cell sorter


Farnesoid X receptor




2′,7′-Dihydrodichlorofluorescein diacetate




Mitochondrial membrane permeability




Poly(ADP-ribose) polymerase


Rhodamine 123


Reactive oxygen species


Serum deprivation


Small heterodimer partner



This work was supported by the National Research Foundation of Korea (NRF) Grant funded by the Korea government [MSIP] (No. 2014R1A2A2A01007375) and also supported by NRF Grant (No. 2012R1A5A2A42671316).

Compliance with ethical standards

Conflict of interest

The authors declare that we have no conflict of interests.


  1. 1.
    Grasl-Kraupp B, Bursch W, Ruttkay-Nedecky B, Wagner A, Lauer B, Schulte-Hermann R (1994) Food restriction eliminates preneoplastic cells through apoptosis and antagonizes carcinogenesis in rat liver. Proc Natl Acad Sci USA 91:9995–9999CrossRefGoogle Scholar
  2. 2.
    Tessitore L, Tomasi C, Greco M (1999) Fasting-induced apoptosis in rat liver is blocked by cycloheximide. Eur J Cell Biol 78:573–579CrossRefGoogle Scholar
  3. 3.
    Zhuge J, Cederbaum AI (2006) Serum deprivation-induced HepG2 cell death is potentiated by CYP2E1. Free Radic Biol Med 40:63–74CrossRefGoogle Scholar
  4. 4.
    Zhang Q, Yang YJ, Wang H, Dong QT, Wang TJ, Qian HY, Xu H (2012) Autophagy activation: a novel mechanism of atorvastatin to protect mesenchymal stem cells from hypoxia and serum deprivation via AMP-activated protein kinase/mammalian target of rapamycin pathway. Stem Cells Dev 21:1321–1332CrossRefGoogle Scholar
  5. 5.
    Valko M, Leibfritz D, Moncol J, Cronin MT, Mazur M, Telser J (2007) Free radicals and antioxidants in normal physiological functions and human disease. Int J Biochem Cell Biol 39:44–84CrossRefGoogle Scholar
  6. 6.
    Medina J, Moreno-Otero R (2005) Pathophysiological basis for antioxidant therapy in chronic liver disease. Drugs 65:2445–2461CrossRefGoogle Scholar
  7. 7.
    Mari M, Colell A, Morales A, von Montfort C, Garcia-Ruiz C, Fernandez-Checa JC (2010) Redox control of liver function in health and disease. Antioxid Redox Signal 12:1295–1331CrossRefGoogle Scholar
  8. 8.
    Muriel P (2009) Role of free radicals in liver diseases. Hepatol Int 3:526–536CrossRefGoogle Scholar
  9. 9.
    Lee JH, Lee JS, Kim YR, Jung WC, Lee KE, Lee SY, Hong EK (2011) Hispidin isolated from Phellinus linteus protects against hydrogen peroxide-induced oxidative stress in pancreatic MIN6N β-cells. J Med Food 14:1431–1438CrossRefGoogle Scholar
  10. 10.
    Spiteller G (2010) Is lipid peroxidation of polyunsaturated acids the only source of free radicals that induce aging and age-related diseases? Rejuvenation Res 13:91–103CrossRefGoogle Scholar
  11. 11.
    Catala A (2010) A synopsis of the process of lipid peroxidation since the discovery of the essential fatty acids. Biochem Biophys Res Commun 399:318–323CrossRefGoogle Scholar
  12. 12.
    Michel S, Wanet A, De Pauw A, Rommelaere G, Arnould T, Renard P (2012) Crosstalk between mitochondrial (dys)function and mitochondrial abundance. J Cell Physiol 227:2297–2310CrossRefGoogle Scholar
  13. 13.
    Hayashi T, Hirshman MF, Fujii N, Habinowski SA, Witters LA, Goodyear LJ (2000) Metabolic stress and altered glucose transport: activation of AMP-activated protein kinase as a unifying coupling mechanism. Diabetes 49:527–531CrossRefGoogle Scholar
  14. 14.
    Terai K, Hiramoto Y, Masaki M, Sugiyama S, Kuroda T, Hori M, Kawase I, Hirota H (2005) AMP-activated protein kinase protects cardiomyocytes against hypoxic injury through attenuation of endoplasmic reticulum stress. Mol Cell Biol 25:9554–9575CrossRefGoogle Scholar
  15. 15.
    Shin SM, Kim SG (2009) Inhibition of arachidonic acid and iron-induced mitochondrial dysfunction and apoptosis by oltipraz and novel 1,2-dithiole-3-thione congeners. Mol Pharmacol 75:242–253CrossRefGoogle Scholar
  16. 16.
    Zhou G, Myers R, Li Y, Chen Y, Shen X, Fenyk-Melody J, Wu M, Ventre J, Doebber T, Fujii N, Musi N, Hirshman MF, Goodyear LJ, Moller DE (2001) Role of AMP-activated protein kinase in mechanism of metformin action. J Clin Invest 108:1167–1174CrossRefGoogle Scholar
  17. 17.
    Shaw RJ, Lamia KA, Vasquez D, Koo SH, Bardeesy N, Depinho RA, Montminy M, Cantley LC (2005) The kinase LKB1 mediates glucose homeostasis in liver and therapeutic effects of metformin. Science 310:1642–1646CrossRefGoogle Scholar
  18. 18.
    Imamura K, Oqura T, Kishimoto A, Kaminishi M, Esumi H (2001) Cell cycle regulation via p53 phosphorylation by a 5′-AMP activated protein kinase activator, 5-aminomidiazole-4-carboxamide-1-beta-D-ribofuranoside, in a human hepatocellular carcinoma cell line. Biochem Biophys Res Commun 287:562–567CrossRefGoogle Scholar
  19. 19.
    Wang HM, Mehta S, Bansode R, Huang W, Mehta KD (2008) AICAR positively regulate glycogen synthase activity and LDL receptor expression through Raf-1/MEK/p42/44MAPK/p90RSK/GSK-3 signaling cascade. Biochem Pharmacol 75:457–467CrossRefGoogle Scholar
  20. 20.
    Kamei J, Nakamura R, Ichiki H, Kudo M (2003) Antitussive principles of Glycyrrhizae radix, a main component of the Kampo preparations Bakumondo-to (Mai-men-dong-tang). Eur J Pharmacol 469:159–163CrossRefGoogle Scholar
  21. 21.
    Choi EM, Suh KS, Lee YS (2014) Liquiritigenin restores osteoblast damage through regulating oxidative stress and mitochondrial dysfunction. Phytother Res 28:880–886CrossRefGoogle Scholar
  22. 22.
    Kim YW, Kim YM, Yang YM, Kay HY, Kim WD, Lee JW, Hwang SJ, Kim SG (2011) Inhibition of LXRα-dependent steatosis and oxidative injury by liquiritigenin, a licorice flavonoid, as mediated with Nrf2 activation. Antioxid Redox Signal 14:733–745CrossRefGoogle Scholar
  23. 23.
    Kim YW, Zhao RJ, Park SJ, Lee JR, Cho IJ, Yang CH, Kim SG, Kim SC (2008) Anti-inflammatory effects of liquiritigenin as a consequence of the inhibition of NF-kappaB-dependent iNOS and proinflammatory cytokines production. Br J Pharmacol 154:165–173CrossRefGoogle Scholar
  24. 24.
    Liu Y, Xie S, Wang Y, Luo K, Wang Y, Cai Y (2012) Liquiritigenin inhibits tumor growth and vascularization in a mouse model of HeLa cells. Molecules 17:7206–7216CrossRefGoogle Scholar
  25. 25.
    Zhou M, Higo H, Cai Y (2010) Inhibition of hepatoma 22 tumor by Liquiritigenin. Phytother Res 24:827–833CrossRefGoogle Scholar
  26. 26.
    Gaur R, Yadav KS, Verma RK, Yadav NP, Bhakuni RS (2014) In vivo anti-diabetic activity of derivatives of isoliquiritigenin and liquiritigenin. Phytomedicine 21:415–422CrossRefGoogle Scholar
  27. 27.
    Kim SC, Byun SH, Yang CH, Kim CY, Kim JW, Kim SG (2004) Cytoprotective effects of Glycyrrhizae radix extract and its active component liquiritigenin against cadmium-induced toxicity (effects on bad translocation and cytochrome c-mediated PARP cleavage). Toxicology 197:239–251CrossRefGoogle Scholar
  28. 28.
    Kim YW, Ki SH, Lee JR, Lee SJ, Kim CW, Kim SC, Kim SG (2006) Liquiritigenin, an aglycone of liquiritin in Glycyrrhizae radix, prevents acute liver injuries in rats induced by acetaminophen with or without buthionine sulfoximine. Chem Biol Interact 161:125–138CrossRefGoogle Scholar
  29. 29.
    Dong S, Inoue A, Zhu Y, Tanji M, Kiyama R (2007) Activation of rapid signaling pathways and the subsequent transcriptional regulation for the proliferation of breast cancer MCF-7 cells by the treatment with an extract of Glycyrrhiza glabra root. Food Chem Toxicol 45:2470–2478CrossRefGoogle Scholar
  30. 30.
    Choi SH, Kim YW, Kim SG (2010) AMPK-mediated GSK3beta inhibition by isoliquiritigenin contributes to protecting mitochondria against iron-catalyzed oxidative stress. Biochem Pharmacol 79:1352–1362CrossRefGoogle Scholar
  31. 31.
    Wang YD, Yang F, Chem WD, Huang X, Lai L, Forman BM, Huang W (2008) Farnesoid X receptor protects liver cells from apoptosis induced by serum deprivation in vitro and fasting in vivo. Mol Endocrinol 22:1622–1632CrossRefGoogle Scholar
  32. 32.
    Dong GZ, Jang EJ, Kang SH, Cho IJ, Park SD, Kim SC, Kim YW (2013) Red ginseng abrogates oxidative stress via mitochondria protection mediated by LKB1-AMPK pathway. BMC Complement Altern Med 13:64CrossRefGoogle Scholar
  33. 33.
    Lee JS, Kim YR, Park JM, Ha SJ, Kim YE, Baek NI, Hong EK (2014) Mulberry fruit extract protects pancreatic β-cells against hydrogen peroxide-induced apoptosis via antioxidative activity. Molecules 19:8904–8915CrossRefGoogle Scholar
  34. 34.
    Ye S, Chen M, Jiang Y, Chen M, Zhou T, Wang Y, Hou Z, Ren L (2014) Polyhydroxylated fullerene attenuates oxidative stress-induced apoptosis via a fortifying Nrf2-regulated cellular antioxidant defence system. Int J Nanomedicine 9:2073–2087CrossRefGoogle Scholar
  35. 35.
    Deevska G, Sunkara M, Karakashian C, Peppers B, Morris AJ, Nikolova-Karakashian MN (2014) Effect of pro-cysteine on aging-associated changes in hepatic GSH and Sphingomyelinase: evidence for transcriptional regulation of Smpd3. J Lipid Res 55:2041–2052CrossRefGoogle Scholar
  36. 36.
    Netto AS, Zanetti MA, Claro GR, de Melo MP, Vilela FG, Correa LB (2014) Effects of copper and selenium supplementation on performance and lipid metabolism in confined brangus bulls. Asian-Australas J Anim Sci 27:488–494CrossRefGoogle Scholar
  37. 37.
    Zhao J, Ming Y, Wan Q, Ye S, Xie S, Zhu Y, Wang Y, Zhong Z, Li L, Ye Q (2014) Gypenoside attenuates hepatic ischemia/reperfusion injury in mice via anti-oxidative and anti-apoptotic bioactivities. Exp Ther Med 7:1388–1392Google Scholar
  38. 38.
    Dong GZ, Lee JH, Ki SH, Yang JH, Cho IJ, Kang SH, Zhao RJ, Kim SC, Kim YW (2014) AMPK activation by isorhamnetin protects hepatocytes against oxidative stress and mitochondrial dysfunction. Eur J Pharm 740:634-640CrossRefGoogle Scholar
  39. 39.
    Browning JD, Horton JD (2004) Molecular mediators of hepatic steatosis and liver injury. J Clin Invest 114:L147–L152CrossRefGoogle Scholar
  40. 40.
    Wu SB, Wu YT, Wu TP, Wei YH (2014) Role of AMPK-mediated adaptive responses in human cells with mitochondrial dysfunction to oxidative stress. Biochim Biophys Acta 1840:1331–1344CrossRefGoogle Scholar
  41. 41.
    Cardin R, Piciocchi M, Bortolami M, Kotsafti A, Barzon L, Lavezzo E, Sinigaglia A, Rodriguez-Castro KI, Rugge M, Farinati F (2014) Oxidative damage in the progression of chronic liver disease to hepatocellular carcinoma: an intricate pathway. World J Gastroenterol 20:3078–3086CrossRefGoogle Scholar
  42. 42.
    Lemasters JJ, Nieminen AL (1997) Mitochondrial oxygen radical formation during reductive and oxidative stress to intact hepatocytes. Biosci Rep 17:281–291CrossRefGoogle Scholar
  43. 43.
    Kim YW, Lee SM, Shin SM, Hwang SJ, Brooks JS, Kang HE, Lee MG, Kim SC, Kim SG (2009) Efficacy of sauchinone as a novel AMPK-activating lignan for preventing iron-induced oxidative stress and liver injury. Free Radic Biol Med 47:1082–1092CrossRefGoogle Scholar
  44. 44.
    Kwon YN, Shin SM, Cho IJ, Kim SG (2009) Oxidized metabolites of oltipraz exert cytoprotective effects against arachidonic acid through AMP-activated protein kinase-dependent cellular antioxidant effect and mitochondrial protection. Drug Metab Dispos 37:1187–1197CrossRefGoogle Scholar
  45. 45.
    Shin SM, Cho IJ, Kim SG (2009) Resveratrol protects mitochondria against oxidative stress through AMP-activated protein kinase-mediated glycogen synthase kinase-3beta inhibition downstream of poly(ADP-ribose)polymerase-LKB1 pathway. Mol Pharmacol 76:884–895CrossRefGoogle Scholar
  46. 46.
    Shaw RJ, Kosmatka M, Bardeesy N, Hurley RL, Witters LA, DePinho RA, Cantley LC (2004) The tumor suppressor LKB1 kinase directly activates AMP-activated kinase and regulates apoptosis in response to energy stress. Proc Natl Acad Sci USA 101:3329–3335CrossRefGoogle Scholar
  47. 47.
    Hardie DG, Scott JW, Pan DA, Hudson ER (2003) Management of cellular energy by the AMP-activated protein kinase system. FEBS Lett 546:113–120CrossRefGoogle Scholar
  48. 48.
    Bai J, Cederbaum AI (2006) Cycloheximide protects HepG2 cells from serum withdrawal-induced apoptosis by decreasing p53 and phosphorylated p53 levels. J Pharmacol Exp Ther 319:1435–1443CrossRefGoogle Scholar
  49. 49.
    Kim YW, Kang HE, Lee MG, Hwang SJ, Kim SC, Lee CH, Kim SG (2009) Liquiritigenin, a flavonoid aglycone from licorice, has a choleretic effect and the ability to induce hepatic transporters and phase-II enzymes. Am J Physiol Gastrointest Liver Physiol 296:372–381CrossRefGoogle Scholar
  50. 50.
    Yang YM, Han CY, Kim YJ, Kim SG (2010) AMPK-associated signaling to bridge the gap between fuel metabolism and hepatocyte viability. World J Gastroenterol 16:3731–3742CrossRefGoogle Scholar
  51. 51.
    MacIver NJ, Blagih J, Saucillo DC, Tonelli L, Griss T, Rathmell JC, Jones RG (2011) The liver kinase B1 is a central regulator of T cell development, activation, and metabolism. J Immunol 187:4187–4198CrossRefGoogle Scholar
  52. 52.
    Noh K, Kim YM, Kim YW, Kim SG (2011) Farnesoid X receptor activation by chenodeoxycholic acid induces detoxifying enzymes through AMP-activated protein kinase and extracellular signal-regulated kinase 1/2-mediated phosphorylation of CCAAT/enhancer binding protein β. Drug Metab Dispos 39:1451–1459CrossRefGoogle Scholar
  53. 53.
    Lee CG, Kim YW, Kim EH, Meng Z, Huang W, Hwang SJ, Kim SG (2012) Farnesoid X receptor protects hepatocytes from injury by repressing miR-199a-3p, which increases levels of LKB1. Gastroenterology 142:1206–1217CrossRefGoogle Scholar
  54. 54.
    Lee FY, de Aguiar Vallim TQ, Chong HK, Zhang Y, Liu Y, Jones SA, Osborne TF, Edwards PA (2010) Activation of the farnesoid X receptor provides protection against acetaminophen-induced hepatic toxicity. Mol Endocrinol 24:1626–1636CrossRefGoogle Scholar
  55. 55.
    Wang H, Chen J, Hollister K, Sowers LC, Forman BM (1999) Endogenous bile acids are ligands for the nuclear receptor FXR/BAR. Mol Cell 3:543–553CrossRefGoogle Scholar
  56. 56.
    Urquhart BL, Tirona RG, Kim RB (2007) Nuclear receptors and the regulation of drug-metabolizing enzymes and drug transporters: implications for interindividual variability in response to drugs. J Clin Pharmacol 47:566–578CrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2015

Authors and Affiliations

  • Eun Hye Jung
    • 1
  • Ju-Hee Lee
    • 1
    • 2
  • Sang Chan Kim
    • 1
  • Young Woo Kim
    • 1
  1. 1.Department of Herbal Formula, Medical Research Center (MRC-GHF), College of Oriental MedicineDaegu Haany UniversityGyeongsanKorea
  2. 2.College of Oriental MedicineDongguk UniversityGyeongjuKorea

Personalised recommendations