The Korean Journal of Physiology and Pharmacology

The Korean Society of Pharmacology (대한약리학회)
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Silymarin Inhibits Morphological Changes in LPS-Stimulated Macrophages by Blocking NF-${\kappa}B$ Pathway

  • Kim, Eun Jeong (Department of Pharmacology, School of Medicine, Chosun University) ;
  • Lee, Min Young (Department of Pharmacology, School of Medicine, Chosun University) ;
  • Jeon, Young Jin (Department of Pharmacology, School of Medicine, Chosun University)
  • Received : 2014.10.20
  • Accepted : 2015.01.21
  • Published : 2015.05.01
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Abstract

The present study showed that silymarin, a polyphenolic flavonoid isolated from milk thistle (Silybum marianum), inhibited lipopolysaccharide (LPS)-induced morphological changes in the mouse RAW264.7 macrophage cell line. We also showed that silymarin inhibited the nuclear translocation and transactivation activities of nuclear factor-kappa B (NF-${\kappa}B$), which is important for macrophage activation-associated changes in cell morphology and gene expression of inflammatory cytokines. BAY-11-7085, an NF-${\kappa}B$ inhibitor, abrogated LPS-induced morphological changes and NO production, similar to silymarin. Treatment of RAW264.7 cells with silymarin also inhibited LPS-stimulated activation of mitogen-activated protein kinases (MAPKs). Collectively, these experiments demonstrated that silymarin inhibited LPS-induced morphological changes in the RAW264.7 mouse macrophage cell line. Our findings indicated that the most likely mechanism underlying this biological effect involved inhibition of the MAPK pathway and NF-${\kappa}B$ activity. Inhibition of these activities by silymarin is a potentially useful strategy for the treatment of inflammation because of the critical roles played by MAPK and NF-${\kappa}B$ in mediating inflammatory responses in macrophages.

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References

  1. Radjadian T, Rezazadeh SH, Huseini HF. Analysis of silymarin components in the seed extracts of some milk thistle ecotypes from Iran by Hplc. Iranian Journal of Science and Technology Transaction ${\alpha}$-Science. 2008;32:141-146.
  2. Valenzuela A, Garrido A. Biochemical bases of the pharmacological action of the flavonoid silymarin and of its structural isomer silibinin. Biol Res. 1994;27:105-112.
  3. Zhang W, Hong R, Tian T. Silymarin's Protective effects and possible mechanisms on alcoholic fatty liver for rats. Biomol Ther (Seoul). 2013;21:264-269. https://doi.org/10.4062/biomolther.2013.020
  4. Bektur NE, Sahin E, Baycu C, Unver G. Protective effects of silymarin against acetaminophen-induced hepatotoxicity and nephrotoxicity in mice. Toxicol Ind Health. 2013. [Epub ahead of print]
  5. Mereish KA, Bunner DL, Ragland DR, Creasia DA. Protection against microcystin-LR-induced hepatotoxicity by Silymarin: biochemistry, histopathology, and lethality. Pharm Res. 1991; 8:273-277. https://doi.org/10.1023/A:1015868809990
  6. Kvasnicka F, Biba B, Sevcik R, Voldrich M, Kratka J. Analysis of the active components of silymarin. J Chromatogr A. 2003; 990:239-245. https://doi.org/10.1016/S0021-9673(02)01971-4
  7. Cristofalo R, Bannwart-Castro CF, Magalhaes CG, Borges VT, Peracoli JC, Witkin SS, Peracoli MT. Silibinin attenuates oxidative metabolism and cytokine production by monocytes from preeclamptic women. Free Radic Res. 2013;47:268-275. https://doi.org/10.3109/10715762.2013.765951
  8. Kang JS, Jeon YJ, Kim HM, Han SH, Yang KH. Inhibition of inducible nitric-oxide synthase expression by silymarin in lipopolysaccharide-stimulated macrophages. J Pharmacol Exp Ther. 2002;302:138-144. https://doi.org/10.1124/jpet.302.1.138
  9. Mokhtari MJ, Motamed N, Shokrgozar MA. Evaluation of silibinin on the viability, migration and adhesion of the human prostate adenocarcinoma (PC-3) cell line. Cell Biol Int. 2008;32: 888-892. https://doi.org/10.1016/j.cellbi.2008.03.019
  10. Hogan FS, Krishnegowda NK, Mikhailova M, Kahlenberg MS. Flavonoid, silibinin, inhibits proliferation and promotes cellcycle arrest of human colon cancer. J Surg Res. 2007;143:58-65. https://doi.org/10.1016/j.jss.2007.03.080
  11. Ahmed-Belkacem A, Ahnou N, Barbotte L, Wychowski C, Pallier C, Brillet R, Pohl RT, Pawlotsky JM. Silibinin and related compounds are direct inhibitors of hepatitis C virus RNAdependent RNA polymerase. Gastroenterology. 2010;138:1112-1122. https://doi.org/10.1053/j.gastro.2009.11.053
  12. Pferschy-Wenzig EM, Atanasov AG, Malainer C, Noha SM, Kunert O, Schuster D, Heiss EH, Oberlies NH, Wagner H, Bauer R, Dirsch VM. Identification of isosilybin a from milk thistle seeds as an agonist of peroxisome proliferator-activated receptor gamma. J Nat Prod. 2014;77:842-847. https://doi.org/10.1021/np400943b
  13. Letteron P, Labbe G, Degott C, Berson A, Fromenty B, Delaforge M, Larrey D, Pessayre D. Mechanism for the protective effects of silymarin against carbon tetrachloride-induced lipid peroxidation and hepatotoxicity in mice. Evidence that silymarin acts both as an inhibitor of metabolic activation and as a chain-breaking antioxidant. Biochem Pharmacol. 1990;39: 2027-2034. https://doi.org/10.1016/0006-2952(90)90625-U
  14. Zhao J, Sharma Y, Agarwal R. Significant inhibition by the flavonoid antioxidant silymarin against 12-O-tetradecanoylphorbol 13-acetate-caused modulation of antioxidant and inflammatory enzymes, and cyclooxygenase 2 and interleukin-1alpha expression in SENCAR mouse epidermis: implications in the prevention of stage I tumor promotion. Mol Carcinog. 1999; 26:321-333. https://doi.org/10.1002/(SICI)1098-2744(199912)26:4<321::AID-MC11>3.0.CO;2-9
  15. Singh RP, Tyagi AK, Zhao J, Agarwal R. Silymarin inhibits growth and causes regression of established skin tumors in SENCAR mice via modulation of mitogen-activated protein kinases and induction of apoptosis. Carcinogenesis. 2002;23: 499-510. https://doi.org/10.1093/carcin/23.3.499
  16. Kim S, Choi MG, Lee HS, Lee SK, Kim SH, Kim WW, Hur SM, Kim JH, Choe JH, Nam SJ, Yang JH, Kim S, Lee JE, Kim JS. Silibinin suppresses TNF-alpha-induced MMP-9 expression in gastric cancer cells through inhibition of the MAPK pathway. Molecules. 2009;14:4300-4311. https://doi.org/10.3390/molecules14114300
  17. Kim JH, Kim K, Jin HM, Song I, Youn BU, Lee J, Kim N. Silibinin inhibits osteoclast differentiation mediated by TNF family members. Mol Cells. 2009;28:201-207. https://doi.org/10.1007/s10059-009-0123-y
  18. Pi J, Li T, Liu J, Su X, Wang R, Yang F, Bai H, Jin H, Cai J. Detection of lipopolysaccharide induced inflammatory responses in RAW264.7 macrophages using atomic force microscope. Micron. 2014;65:1-9. https://doi.org/10.1016/j.micron.2014.03.012
  19. Machesky LM. Lamellipodia and filopodia in metastasis and invasion. FEBS Lett. 2008;582:2102-2111. https://doi.org/10.1016/j.febslet.2008.03.039
  20. Bhavsar PJ, Vigorito E, Turner M, Ridley AJ. Vav GEFs regulate macrophage morphology and adhesion-induced Rac and Rho activation. Exp Cell Res. 2009;315:3345-3358. https://doi.org/10.1016/j.yexcr.2009.08.010
  21. Guo AK, Hou YY, Hirata H, Yamauchi S, Yip AK, Chiam KH, Tanaka N, Sawada Y, Kawauchi K. Loss of p53 enhances NF-κ B-dependent lamellipodia formation. J Cell Physiol. 2014;229: 696-704. https://doi.org/10.1002/jcp.24505
  22. Kucik DF, Wu C. Cell-adhesion assays. Methods Mol Biol. 2005; 294:43-54.
  23. Green LC, Wagner DA, Glogowski J, Skipper PL, Wishnok JS, Tannenbaum SR. Analysis of nitrate, nitrite, and [15N]nitrate in biological fluids. Anal Biochem. 1982;126:131-138. https://doi.org/10.1016/0003-2697(82)90118-X
  24. Huong PT, Lee MY, Lee KY, Chang IY, Lee SK, Yoon SP, Lee DC, Jeon YJ. Synergistic Induction of iNOS by IFN-γ and Glycoprotein Isolated from Dioscorea batatas. Korean J Physiol Pharmacol. 2012;16:431-436. https://doi.org/10.4196/kjpp.2012.16.6.431
  25. Kim EJ, Kim J, Lee MY, Sudhanva MS, Devakumar S, Jeon YJ. Silymarin inhibits cytokine-stimulated pancreatic beta cells by blocking the ERK1/2 pathway. Biomol Ther (Seoul). 2014;22: 282-287. https://doi.org/10.4062/biomolther.2014.072
  26. Hibbs JB Jr, Taintor RR, Vavrin Z. Macrophage cytotoxicity: role for L-arginine deiminase and imino nitrogen oxidation to nitrite. Science. 1987;235:473-476. https://doi.org/10.1126/science.2432665
  27. Palmer RM, Ashton DS, Moncada S. Vascular endothelial cells synthesize nitric oxide from L-arginine. Nature. 1988;333:664-666. https://doi.org/10.1038/333664a0
  28. Matsuda T, Ferreri K, Todorov I, Kuroda Y, Smith CV, Kandeel F, Mullen Y. Silymarin protects pancreatic beta-cells against cytokine-mediated toxicity: implication of c-Jun NH2-terminal kinase and janus kinase/signal transducer and activator of transcription pathways. Endocrinology. 2005;146:175-185. https://doi.org/10.1210/en.2004-0850
  29. Cetkovic-Cvrlje M, Eizirik DL. TNF-alpha and IFN-gamma potentiate the deleterious effects of IL-1 beta on mouse pancreatic islets mainly via generation of nitric oxide. Cytokine. 1994;6:399-406. https://doi.org/10.1016/1043-4666(94)90064-7
  30. Darville MI, Eizirik DL. Regulation by cytokines of the inducible nitric oxide synthase promoter in insulin-producing cells. Diabetologia. 1998;41:1101-1108. https://doi.org/10.1007/s001250051036
  31. Soto C, Raya L, Juarez J, Perez J, Gonzalez I. Effect of Silymarin in Pdx-1 expression and the proliferation of pancreatic ${\beta}$-cells in a pancreatectomy model. Phytomedicine. 2014;21: 233-239. https://doi.org/10.1016/j.phymed.2013.09.008
  32. Agarwal R, Agarwal C, Ichikawa H, Singh RP, Aggarwal BB. Anticancer potential of silymarin: from bench to bed side. Anticancer Res. 2006;26:4457-4498.
  33. Iademarco MF, McQuillan JJ, Rosen GD, Dean DC. Characterization of the promoter for vascular cell adhesion molecule-1 (VCAM-1). J Biol Chem. 1992;267:16323-16329.
  34. Ledebur HC, Parks TP. Transcriptional regulation of the intercellular adhesion molecule-1 gene by inflammatory cytokines in human endothelial cells. Essential roles of a variant NFkappa B site and p65 homodimers. J Biol Chem. 1995;270: 933-943. https://doi.org/10.1074/jbc.270.2.933
  35. Whelan J, Ghersa P, Hooft van Huijsduijnen R, Gray J, Chandra G, Talabot F, DeLamarter JF. An NF kappa B-like factor is essential but not sufficient for cytokine induction of endothelial leukocyte adhesion molecule 1 (ELAM-1) gene transcription. Nucleic Acids Res. 1991;19:2645-2653. https://doi.org/10.1093/nar/19.10.2645
  36. Kang JS, Park SK, Yang KH, Kim HM. Silymarin inhibits TNF-alpha-induced expression of adhesion molecules in human umbilical vein endothelial cells. FEBS Lett. 2003;550:89-93. https://doi.org/10.1016/S0014-5793(03)00827-5
  37. Su B, Karin M. Mitogen-activated protein kinase cascades and regulation of gene expression. Curr Opin Immunol. 1996;8: 402-411. https://doi.org/10.1016/S0952-7915(96)80131-2
  38. Raingeaud J, Gupta S, Rogers JS, Dickens M, Han J, Ulevitch RJ, Davis RJ. Pro-inflammatory cytokines and environmental stress cause p38 mitogen-activated protein kinase activation by dual phosphorylation on tyrosine and threonine. J Biol Chem. 1995;270:7420-7426. https://doi.org/10.1074/jbc.270.13.7420
  39. Oh SJ, Jung SP, Han J, Kim S, Kim JS, Nam SJ, Lee JE, Kim JH. Silibinin inhibits TPA-induced cell migration and MMP-9 expression in thyroid and breast cancer cells. Oncol Rep. 2013; 29:1343-1348. https://doi.org/10.3892/or.2013.2252
  40. Kim S, Choi JH, Lim HI, Lee SK, Kim WW, Kim JS, Kim JH, Choe JH, Yang JH, Nam SJ, Lee JE. Silibinin prevents TPAinduced MMP-9 expression and VEGF secretion by inactivation of the Raf/MEK/ERK pathway in MCF-7 human breast cancer cells. Phytomedicine. 2009;16:573-580. https://doi.org/10.1016/j.phymed.2008.11.006
  41. Larsen L, Størling J, Darville M, Eizirik DL, Bonny C, Billestrup N, Mandrup-Poulsen T. Extracellular signal-regulated kinase is essential for interleukin-1-induced and nuclear factor kappaB-mediated gene expression in insulin-producing INS-1E cells. Diabetologia. 2005;48:2582-2590. https://doi.org/10.1007/s00125-005-0039-9
  42. Youn CK, Park SJ, Li MH, Lee MY, Lee KY, Cha MJ, Kim OH, You HJ, Chang IY, Yoon SP, Jeon YJ. Radicicol inhibits inos expression in cytokine-stimulated pancreatic beta cells. Korean J Physiol Pharmacol. 2013;17:315-320. https://doi.org/10.4196/kjpp.2013.17.4.315
  43. Svitkina TM, Bulanova EA, Chaga OY, Vignjevic DM, Kojima S, Vasiliev JM, Borisy GG. Mechanism of filopodia initiation by reorganization of a dendritic network. J Cell Biol. 2003; 160:409-421. https://doi.org/10.1083/jcb.200210174
  44. Adams JC, Schwartz MA. Stimulation of fascin spikes by thrombospondin-1 is mediated by the GTPases Rac and Cdc42. J Cell Biol. 2000;150:807-822. https://doi.org/10.1083/jcb.150.4.807
  45. Hashimoto Y, Parsons M, Adams JC. Dual actin-bundling and protein kinase C-binding activities of fascin regulate carcinoma cell migration downstream of Rac and contribute to metastasis. Mol Biol Cell. 2007;18:4591-4602. https://doi.org/10.1091/mbc.E07-02-0157
  46. Faix J, Grosse R. Staying in shape with formins. Dev Cell. 2006; 10:693-706.
  47. Applewhite DA, Barzik M, Kojima S, Svitkina TM, Gertler FB, Borisy GG. Ena/VASP proteins have an anti-capping independent function in filopodia formation. Mol Biol Cell. 2007;18:2579-2591. https://doi.org/10.1091/mbc.E06-11-0990

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