Development and Reproduction (한국발생생물학회지:발생과생식)

The Korean Society of Developmental Biology (한국발생생물학회)
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Metformin Induces Lipogenesis and Apoptosis in H4IIE Hepatocellular Carcinoma Cells

  • Deokbae Park (Department of Histology, Jeju National University College of Medicine) ;
  • Sookyoung Lee (Department of Histology, Jeju National University College of Medicine) ;
  • Hyejin Boo (Department of Histology, Jeju National University College of Medicine)
  • Received : 2023.04.11
  • Accepted : 2023.06.15
  • Published : 2023.06.30
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Abstract

Metformin is the most widely used anti-diabetic drug that helps maintain normal blood glucose levels primarily by suppressing hepatic gluconeogenesis in type II diabetic patients. We previously found that metformin induces apoptotic death in H4IIE rat hepatocellular carcinoma cells. Despite its anti-diabetic roles, the effect of metformin on hepatic de novo lipogenesis (DNL) remains unclear. We investigated the effect of metformin on hepatic DNL and apoptotic cell death in H4IIE cells. Metformin treatment stimulated glucose consumption, lactate production, intracellular fat accumulation, and the expressions of lipogenic proteins. It also stimulated apoptosis but reduced autophagic responses. These metformin-induced changes were clearly reversed by compound C, an inhibitor of AMP-activated protein kinase (AMPK). Interestingly, metformin massively increased the production of reactive oxygen species (ROS), which was completely blocked by compound C. Metformin also stimulated the phosphorylation of p38 mitogen-activated protein kinase (p38MAPK). Finally, inhibition of p38MAPK mimicked the effects of compound C, and suppressed the metformin-induced fat accumulation and apoptosis. Taken together, metformin stimulates dysregulated glucose metabolism, intracellular fat accumulation, and apoptosis. Our findings suggest that metformin induces excessive glucose-induced DNL, oxidative stress by ROS generation, activation of AMPK and p38MAPK, suppression of autophagy, and ultimately apoptosis.

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Acknowledgement

This work was supported by the 2023 education, research and student guidance grant funded by Jeju National University.

References

  1. Barzilai N, Crandall JP, Kritchevsky SB, Espeland MA (2016) Metformin as a tool to target aging. Cell Metab 23:1060-1065. https://doi.org/10.1016/j.cmet.2016.05.011
  2. Bhalla K, Hwang BJ, Dewi RE, Twaddel W, Goloubeva OG, Wong KK, Saxena NK, Biswal S, Girnun GD (2012) Metformin prevents liver tumorigenesis by inhibiting pathways driving hepatic lipogenesis. Cancer Prev Res 5:544-552. https://doi.org/10.1158/1940-6207.CAPR-11-0228
  3. Bray F, Ferlay J, Soerjomataram I, Siegel RL, Torre LA, Jemal A (2018) Global cancer statistics 2018: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA: A Cancer J Clin 68:394-424. https://doi.org/10.3322/caac.21492
  4. Das S, Shukla N, Singh SS, Kushwaha S, Shrivastava R (2021) Mechanism of interaction between autophagy and apoptosis in cancer. Apoptosis 26:512-533. https://doi.org/10.1007/s10495-021-01687-9
  5. DePeralta DK, Wei L, Ghoshal S, Schmidt B, Lauwers GY, Lanuti M, Chung RT, Tanabe KK, Fuchs BC (2016). Metformin prevents hepatocellular carcinoma development by suppressing hepatic progenitor cell activation in a rat model of cirrhosis. Cancer 122:1216-1227. https://doi.org/10.1002/cncr.29912
  6. DeWaal D, Nogueira V, Terry AR, Patra KC, Jeon SM, Guzman G, Au J, Long CP, Antoniewicz MR, Hay N (2018) Hexokinase-2 depletion inhibits glycolysis and induces oxidative phosphorylation in hepatocellular carcinoma and sensitizes to metformin. Nat Commun 9:446.
  7. Forslund K, Hildebrand F, Nielsen T, Falony G, Le Chatelier E, Sunagawa S, et al. (2015) Disentangling type 2 diabetes and metformin treatment signatures in the human gut microbiota. Nature 528:262-266. https://doi.org/10.1038/nature15766
  8. Halliwell B, Whiteman M (2004) Measuring reactive species and oxidative damage in vivo and in cell culture: How should you do it and what do the results mean? Br J Pharmacol 142:231-255. https://doi.org/10.1038/sj.bjp.0705776
  9. Han Y, Hu Z, Cui A, Liu Z, Ma F, Xue Y, Liu Y, Zhang F, Zhao Z, Yu Y, Gao J, Wei C, Li J, Fang J, Li J, Fan JG, Song BL, Li Y (2019) Post-translational regulation of lipogenesis via AMPK-dependent phosphorylation of insulin-induced gene. Nat Commun 10:623.
  10. Hardie DG, Pan DA (2002) Regulation of fatty acid synthesis and oxidation by the AMP-activated protein kinase. Biochem Soc Trans 30:1064-1070. https://doi.org/10.1042/bst0301064
  11. Hay N (2016) Reprogramming glucose metabolism in cancer: Can it be exploited for cancer therapy? Nat Rev Cancer 16:635-649. https://doi.org/10.1038/nrc.2016.77
  12. Jones RG, Plas DR, Kubek S, Buzzai M, Mu J, Xu Y, Birnbaum MJ, Thompson CB (2005) AMP-activated protein kinase induces a p53-dependent metabolic checkpoint. Mol Cell 18:283-293. https://doi.org/10.1016/j.molcel.2005.03.027
  13. Kim J, Yang G, Kim Y, Kim J, Ha J (2016) AMPK activators: Mechanisms of action and physiological activities. Exp Mol Med 48:e224.
  14. Kim YD, Park KG, Lee YS, Park YY, Kim DK, Nedumaran B, Jang WG, Cho WJ, Ha J, Lee IK, Lee CH, Choi HS (2008) Metformin inhibits hepatic gluconeogenesis through AMP-activated protein kinase-dependent regulation of the orphan nuclear receptor SHP. Diabetes 57:306-314. https://doi.org/10.2337/db07-0381
  15. Ko E, Baek S, Kim J, Park D, Lee Y (2020) Antitumor activity of combination therapy with metformin and trametinib in non-small cell lung cancer cells. Dev Reprod 24:113-123. https://doi.org/10.12717/DR.2020.24.2.113
  16. Krishnan UA, Viswanathan P, Venkataraman AC (2023) AMPK activation by AICAR reduces diet induced fatty liver in C57BL/6 mice. Tissue Cell 82:102054.
  17. Lee J, Park D, Lee Y (2017) Metformin synergistically potentiates the antitumor effects of imatinib in colorectal cancer cells. Dev Reprod 21:139-150. https://doi.org/10.12717/DR.2017.21.2.139
  18. Liu W, Sun C, Yan Y, Cao H, Niu Z, Shen S, Liu S, Wu Y, Li Y, Hui L, Li Y, Zhao L, Hu C, Ding Q, Jiang J, Ying H (2022) Hepatic P38 activation modulates systemic metabolism through Fgf21-mediated interorgan communication. Diabetes 71:60-72. https://doi.org/10.2337/db21-0240
  19. Ma X, Qiu Y, Sun Y, Zhu L, Zhao Y, Li T, Lin Y, Ma D, Qin Z, Sun C, Han L (2020) NOD2 inhibits tumorigenesis and increases chemosensitivity of hepatocellular carcinoma by targeting AMPK pathway. Cell Death Dis 11:174.
  20. Mosmann T (1983) Rapid colorimetric assay for cellular growth and survival: Application application to proliferation and cytotoxicity assays. J Immunol Methods 65:55-63. https://doi.org/10.1016/0022-1759(83)90303-4
  21. Park D (2015) Metformin promotes apoptosis but suppresses autophagy in glucose-deprived H4IIE hepatocellular carcinoma cells. Diabetes Metab J 39:518-527. https://doi.org/10.4093/dmj.2015.39.6.518
  22. Park D (2019) Metformin induces oxidative stress-mediated apoptosis without the blockade of glycolysis in H4IIE hepatocellular carcinoma cells. Biol Pharm Bull 42:2002-2008. https://doi.org/10.1248/bpb.b19-00474
  23. Raez LE, Papadopoulos K, Ricart AD, Chiorean EG, Dipaola RS, Stein MN, Rocha Lima CM, Schlesselman JJ, Tolba K, Langmuir VK, Kroll S, Jung DT, Kurtoglu M, Rosenblatt J, Lampidis TJ (2013) A phase I dose-escalation trial of 2-deoxy-D-glucose alone or combined with docetaxel in patients with advanced solid tumors. Cancer Chemother Pharmacol 71:523-530. https://doi.org/10.1007/s00280-012-2045-1
  24. Rena G, Hardie DG, Pearson ER (2017) The mechanisms of action of metformin. Diabetologia 60:1577-1585. https://doi.org/10.1007/s00125-017-4342-z
  25. Rudalska R, Harbig J, Snaebjornsson MT, Klotz S, Zwirner S, Taranets L, et al. (2021) LXRα activation and Raf inhibition trigger lethal lipotoxicity in liver cancer. Nat Cancer 2:201-217. https://doi.org/10.1038/s43018-020-00168-3
  26. Schuster S, Penke M, Gorski T, Gebhardt R, Weiss TS, Kiess W, Garten A (2015) FK866-induced NAMPT inhibition activates AMPK and downregulates mTOR signaling in hepatocarcinoma cells. Biochem Biophys Res Commun 458:334-340. https://doi.org/10.1016/j.bbrc.2015.01.111
  27. Stumvoll M, Nurjhan N, Perriello G, Dailey G, Gerich JE (1995) Metabolic effects of metformin in non-insulin-dependent diabetes mellitus. N Engl J Med 333:550-554. https://doi.org/10.1056/NEJM199508313330903
  28. Sun L, Xi S, Zhou Z, Zhang F, Hu P, Cui Y, Wu S, Wang Y, Wu S, Wang Y, Du Y, Zheng J, Yang H, Chen M, Yan Q, Yu D, Shi C, Zhang Y, Xie D, Guan XY, Li Y (2022) Elevated expression of RIT1 hyperactivates RAS/MAPK signal and sensitizes hepatocellular carcinoma to combined treatment with sorafenib and AKT inhibitor. Oncogene 41:732-744. https://doi.org/10.1038/s41388-021-02130-8
  29. Vaupel P, Schmidberger H, Mayer A (2019) The Warburg effect: Essential part of metabolic reprogramming and central contributor to cancer progression. Int J Radiat Biol 95:912-919. https://doi.org/10.1080/09553002.2019.1589653
  30. Viollet B, Foretz M (2013) Revisiting the mechanisms of metformin action in the liver. Ann Endocrinol (Paris) 74:123-129. https://doi.org/10.1016/j.ando.2013.03.006
  31. Wang L, Hu T, Shen Z, Zheng Y, Geng Q, Li L, Sha B, Li M, Sun Y, Guo Y, Xue W, Xuan D, Chen P, Zhao J (2022) Inhibition of USP1 activates ER stress through Ubi-protein aggregation to induce autophagy and apoptosis in HCC. Cell Death Dis 13:951.
  32. Ward PS, Thompson CB (2012) Metabolic reprogramming: A cancer hallmark even warburg did not anticipate. Cancer Cell 21:297-308. https://doi.org/10.1016/j.ccr.2012.02.014
  33. Yang W, Zhu L, Lai S, Ding Q, Xu T, Guo R, Dou X, Chai H, Yu Z, Li S (2022) Cimifugin ameliorates lipotoxicity-induced hepatocyte damage and steatosis through TLR4/p38 MAPK-and SIRT1-involved pathways. Oxid Med Cell Longev 2022:4557532.
  34. Yin J, Hu R, Chen M, Tang J, Li F, Yang Y, Chen J (2002). Effects of berberine on glucose metabolism in vitro. Metabolism 51:1439-1443. https://doi.org/10.1053/meta.2002.34715
  35. Yong HY, Koh MS, Moon A (2009) The p38 MAPK inhibitors for the treatment of inflammatory diseases and cancer. Expert Opin Investig Drugs 18:1893-1905. https://doi.org/10.1517/13543780903321490
  36. Zhang H, Gao C, Fang L, Zhao HC, Yao SK (2013a) Metformin and reduced risk of hepatocellular carcinoma in diabetic patients: A meta-analysis. Scand J Gastroenterol 48:78-87. https://doi.org/10.3109/00365521.2012.719926
  37. Zhang P, Li H, Tan X, Chen L, Wang S (2013b) Association of metformin use with cancer incidence and mortality: A meta-analysis. Cancer Epidemiol 37:207-218. https://doi.org/10.1016/j.canep.2012.12.009
  38. Zhang Y, Wang H, Xiao H (2021) Metformin actions on the liver: Protection mechanisms emerging in hepatocytes and immune cells against NASH-related HCC. Int J Mol Sci 22:5016.