title: Dihydromyricetin inhibits migration and invasion of hepatoma cells through regulation of MMP-9 expression.
abstract: OBJECTIVE
  To investigate the effects of dihydromyricetin (DHM) on the migration and invasion of human hepatic cancer cells.
  
  
  METHODS
  The hepatoma cell lines SK-Hep-1 and MHCC97L were used in this study. The cells were cultured in RPIM-1640 medium supplemented with 10% fetal bovine serum at 37 °C in a humidified 5% CO2 incubator. DHM was dissolved in dimethyl sulfoxide and diluted to various concentrations in medium before applying to cells. MTT assays were performed to measure the viability of the cells after DHM treatment. Wound healing and Boyden transwell assays were used to assess cancer cell motility. The invasive capacity of cancer cells was measured using Matrigel-coated transwell chambers. Matrix metalloproteinase (MMP)-2/9 activity was examined by fluorescence analysis. Western blot was carried out to analyze the expression of MMP-2, MMP-9, p-38, JNK, ERK1/2 and PKC-δ proteins. All data were analyzed by Student's t tests in GraphPad prism 5.0 software and are presented as mean ± SD.
  
  
  RESULTS
  DHM was found to strongly inhibit the migration of the hepatoma cell lines SK-Hep-1 (without DHM, 24 h: 120 ± 8 μmol/L vs 100 μmol/L DHM, 24 h: 65 ± 10 μmol/L, P < 0.001) and MHCC97L (without DHM, 24 h: 126 ± 7 μmol/L vs 100 μmol/L DHM, 24 h: 74 ± 6 μmol/L, P < 0.001). The invasive capacity of the cells was reduced by DHM treatment (SK-Hep-1 cells without DHM, 24 h: 67 ± 4 μmol/L vs 100 μmol/L DHM, 24 h: 9 ± 3 μmol/L, P < 0.001; MHCC97L cells without DHM, 24 h: 117 ± 8 μmol/L vs 100 μmol/L DHM, 24 h: 45 ± 2 μmol/L, P < 0.001). MMP2/9 activity was also inhibited by DHM exposure (SK-Hep-1 cells without DHM, 24 h: 600 ± 26 μmol/L vs 100 μmol/L DHM, 24 h: 100 ± 6 μmol/L, P < 0.001; MHCC97L cells without DHM, 24 h: 504 ± 32 μmol/L vs 100 μmol/L DHM 24 h: 156 ± 10 μmol/L, P < 0.001). Western blot analysis showed that DHM decreased the expression level of MMP-9 but had little effect on MMP-2. Further investigation indicated that DHM markedly reduced the phosphorylation levels of p38, ERK1/2 and JNK in a concentration-dependent manner but had no impact on the total protein levels. In addition, PKC-δ protein, a key protein in the regulation of MMP family protein expression, was up-regulated with DHM treatment.
  
  
  CONCLUSIONS
  These findings demonstrate that DHM inhibits the migration and invasion of hepatoma cells and may serve as a potential candidate agent for the prevention of HCC metastasis.
authors.affiliation: Qing-Yu Zhang, Ran Li, Guo-Fang Zeng, Bin Liu, Jie Liu, Yang Shu, Zhong-Kao Liu, Zhi-Dong Qiu, Dong-Jun Wang, Hui-Lai Miao, Ming-Yi Li , Run-Zhi Zhu, Key Laboratory of Hepatic Disease, Affiliated Hospital of Guangdong Medical College, Zhanjiang 524001, Guangdong Province, China.
authors.forename: Qing-Yu
authors.initials: QY
authors.lastname: Zhang
authors.affiliation: Qing-Yu Zhang, Ran Li, Guo-Fang Zeng, Bin Liu, Jie Liu, Yang Shu, Zhong-Kao Liu, Zhi-Dong Qiu, Dong-Jun Wang, Hui-Lai Miao, Ming-Yi Li , Run-Zhi Zhu, Key Laboratory of Hepatic Disease, Affiliated Hospital of Guangdong Medical College, Zhanjiang 524001, Guangdong Province, China.
authors.forename: Ran
authors.initials: R
authors.lastname: Li
authors.affiliation: Qing-Yu Zhang, Ran Li, Guo-Fang Zeng, Bin Liu, Jie Liu, Yang Shu, Zhong-Kao Liu, Zhi-Dong Qiu, Dong-Jun Wang, Hui-Lai Miao, Ming-Yi Li , Run-Zhi Zhu, Key Laboratory of Hepatic Disease, Affiliated Hospital of Guangdong Medical College, Zhanjiang 524001, Guangdong Province, China.
authors.forename: Guo-Fang
authors.initials: GF
authors.lastname: Zeng
authors.affiliation: Qing-Yu Zhang, Ran Li, Guo-Fang Zeng, Bin Liu, Jie Liu, Yang Shu, Zhong-Kao Liu, Zhi-Dong Qiu, Dong-Jun Wang, Hui-Lai Miao, Ming-Yi Li , Run-Zhi Zhu, Key Laboratory of Hepatic Disease, Affiliated Hospital of Guangdong Medical College, Zhanjiang 524001, Guangdong Province, China.
authors.forename: Bin
authors.initials: B
authors.lastname: Liu
authors.affiliation: Qing-Yu Zhang, Ran Li, Guo-Fang Zeng, Bin Liu, Jie Liu, Yang Shu, Zhong-Kao Liu, Zhi-Dong Qiu, Dong-Jun Wang, Hui-Lai Miao, Ming-Yi Li , Run-Zhi Zhu, Key Laboratory of Hepatic Disease, Affiliated Hospital of Guangdong Medical College, Zhanjiang 524001, Guangdong Province, China.
authors.forename: Jie
authors.initials: J
authors.lastname: Liu
authors.affiliation: Qing-Yu Zhang, Ran Li, Guo-Fang Zeng, Bin Liu, Jie Liu, Yang Shu, Zhong-Kao Liu, Zhi-Dong Qiu, Dong-Jun Wang, Hui-Lai Miao, Ming-Yi Li , Run-Zhi Zhu, Key Laboratory of Hepatic Disease, Affiliated Hospital of Guangdong Medical College, Zhanjiang 524001, Guangdong Province, China.
authors.forename: Yang
authors.initials: Y
authors.lastname: Shu
authors.affiliation: Qing-Yu Zhang, Ran Li, Guo-Fang Zeng, Bin Liu, Jie Liu, Yang Shu, Zhong-Kao Liu, Zhi-Dong Qiu, Dong-Jun Wang, Hui-Lai Miao, Ming-Yi Li , Run-Zhi Zhu, Key Laboratory of Hepatic Disease, Affiliated Hospital of Guangdong Medical College, Zhanjiang 524001, Guangdong Province, China.
authors.forename: Zhong-Kao
authors.initials: ZK
authors.lastname: Liu
authors.affiliation: Qing-Yu Zhang, Ran Li, Guo-Fang Zeng, Bin Liu, Jie Liu, Yang Shu, Zhong-Kao Liu, Zhi-Dong Qiu, Dong-Jun Wang, Hui-Lai Miao, Ming-Yi Li , Run-Zhi Zhu, Key Laboratory of Hepatic Disease, Affiliated Hospital of Guangdong Medical College, Zhanjiang 524001, Guangdong Province, China.
authors.forename: Zhi-Dong
authors.initials: ZD
authors.lastname: Qiu
authors.affiliation: Qing-Yu Zhang, Ran Li, Guo-Fang Zeng, Bin Liu, Jie Liu, Yang Shu, Zhong-Kao Liu, Zhi-Dong Qiu, Dong-Jun Wang, Hui-Lai Miao, Ming-Yi Li , Run-Zhi Zhu, Key Laboratory of Hepatic Disease, Affiliated Hospital of Guangdong Medical College, Zhanjiang 524001, Guangdong Province, China.
authors.forename: Dong-Jun
authors.initials: DJ
authors.lastname: Wang
authors.affiliation: Qing-Yu Zhang, Ran Li, Guo-Fang Zeng, Bin Liu, Jie Liu, Yang Shu, Zhong-Kao Liu, Zhi-Dong Qiu, Dong-Jun Wang, Hui-Lai Miao, Ming-Yi Li , Run-Zhi Zhu, Key Laboratory of Hepatic Disease, Affiliated Hospital of Guangdong Medical College, Zhanjiang 524001, Guangdong Province, China.
authors.forename: Hui-Lai
authors.initials: HL
authors.lastname: Miao
authors.affiliation: Qing-Yu Zhang, Ran Li, Guo-Fang Zeng, Bin Liu, Jie Liu, Yang Shu, Zhong-Kao Liu, Zhi-Dong Qiu, Dong-Jun Wang, Hui-Lai Miao, Ming-Yi Li , Run-Zhi Zhu, Key Laboratory of Hepatic Disease, Affiliated Hospital of Guangdong Medical College, Zhanjiang 524001, Guangdong Province, China.
authors.forename: Ming-Yi
authors.initials: MY
authors.lastname: Li
authors.affiliation: Qing-Yu Zhang, Ran Li, Guo-Fang Zeng, Bin Liu, Jie Liu, Yang Shu, Zhong-Kao Liu, Zhi-Dong Qiu, Dong-Jun Wang, Hui-Lai Miao, Ming-Yi Li , Run-Zhi Zhu, Key Laboratory of Hepatic Disease, Affiliated Hospital of Guangdong Medical College, Zhanjiang 524001, Guangdong Province, China.
authors.forename: Run-Zhi
authors.initials: RZ
authors.lastname: Zhu
mesh_terms: D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology*; D006528:Carcinoma, Hepatocellular / Q000201:enzymology* / Q000473:pathology; D002448:Cell Adhesion / Q000187:drug effects; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000187:drug effects*; D002470:Cell Survival / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation; D044948:Flavonols / Q000494:pharmacology*; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D008113:Liver Neoplasms / Q000201:enzymology* / Q000473:pathology; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism*; D061965:Matrix Metalloproteinase Inhibitors / Q000494:pharmacology*; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D009361:Neoplasm Invasiveness; D010766:Phosphorylation; D051745:Protein Kinase C-delta / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D013997:Time Factors
chemical_list: D000972:Antineoplastic Agents, Phytogenic; D044948:Flavonols; D061965:Matrix Metalloproteinase Inhibitors; C497258:PRKCD protein, human; D051745:Protein Kinase C-delta; D020928:Mitogen-Activated Protein Kinases; C522361:MMP2 protein, human; D020778:Matrix Metalloproteinase 2; C579270:MMP9 protein, human; D020780:Matrix Metalloproteinase 9; C472036:dihydromyricetin
keywords: Dihydromyricetin; Hepatic cancer; Invasion; Matrix metalloproteinase-9; Migration
references.citation: Urology. 2005 May;65(5):1029-34
references.pmid: 15882758
references.citation: PLoS One. 2012;7(6):e38802
references.pmid: 22693649
references.citation: Curr Opin Cell Biol. 2012 Apr;24(2):277-83
references.pmid: 22209238
references.citation: Biochem Pharmacol. 2010 Jun 15;79(12):1714-26
references.pmid: 20152819
references.citation: Exp Mol Pathol. 2012 Feb;92(1):74-81
references.pmid: 22024355
references.citation: Cell Stem Cell. 2011 Jul 8;9(1):50-63
references.pmid: 21726833
references.citation: Curr Pharm Biotechnol. 2012 Jan;13(1):218-28
references.pmid: 21466422
references.citation: J Biol Chem. 2003 Jun 20;278(25):22404-11
references.pmid: 12690120
references.citation: Am J Physiol Cell Physiol. 2005 Apr;288(4):C906-12
references.pmid: 15590896
references.citation: Carcinogenesis. 2009 Jul;30(7):1225-33
references.pmid: 19420016
references.citation: CA Cancer J Clin. 2011 Jul-Aug;61(4):212-36
references.pmid: 21685461
references.citation: Biofactors. 2004;21(1-4):175-8
references.pmid: 15630194
references.citation: J Leukoc Biol. 2009 Apr;85(4):656-63
references.pmid: 19164129
references.citation: Int Immunopharmacol. 2012 Jan;12(1):278-87
references.pmid: 22193240
references.citation: J Biol Chem. 1998 Feb 27;273(9):5137-45
references.pmid: 9478967
references.citation: J Biol Chem. 2011 Dec 9;286(49):42072-42081
references.pmid: 22009750
references.citation: J Neurosci. 2012 Jan 4;32(1):390-401
references.pmid: 22219299
references.citation: J Immunol. 2003 Jun 15;170(12):6244-9
references.pmid: 12794156
references.citation: Zhongguo Zhong Yao Za Zhi. 2003 Dec;28(12):1188-90
references.pmid: 15617509
references.citation: J Leukoc Biol. 2006 Jan;79(1):214-22
references.pmid: 16275891
references.citation: J Virol. 2005 Aug;79(15):9714-24
references.pmid: 16014933
references.citation: Zhong Yao Cai. 2006 Feb;29(2):146-50
references.pmid: 16617784
references.citation: J Pharmacol Sci. 2010;112(1):37-45
references.pmid: 20093787
references.citation: Zhongguo Zhong Yao Za Zhi. 2011 Feb;36(4):500-3
references.pmid: 21598552
references.citation: CA Cancer J Clin. 2011 Mar-Apr;61(2):69-90
references.pmid: 21296855
references.citation: Hepatol Res. 2012 Apr;42(4):401-11
references.pmid: 22151918
references.citation: J Agric Food Chem. 2011 Aug 24;59(16):9032-8
references.pmid: 21770460
references.citation: J Asian Nat Prod Res. 2011 Mar;13(3):242-50
references.pmid: 21409686
references.citation: Oncogene. 2004 Apr 15;23(17):3080-8
references.pmid: 15048090
references.citation: Ann Surg Oncol. 2012 Aug;19(8):2733-43
references.pmid: 22207048
references.citation: Hepatology. 2011 Nov;54(5):1808-18
references.pmid: 22045676
references.citation: PLoS One. 2013 Nov 04;8(11):e76886
references.pmid: 24223706
references.citation: Anticancer Drugs. 2012 Jul;23(6):590-6
references.pmid: 22241170
references.citation: Zhongguo Zhong Yao Za Zhi. 2002 Sep;27(9):687-9
references.pmid: 12776572
references.citation: Glia. 2008 Apr 15;56(6):619-32
references.pmid: 18240315
references.citation: Zhong Yao Cai. 2004 Nov;27(11):842-5
references.pmid: 15810594
references.citation: FEBS J. 2011 Jan;278(1):16-27
references.pmid: 21087457
references.citation: J Sep Sci. 2011 Jan;34(2):160-8
references.pmid: 21246721
references.citation: Int J Mol Med. 2012 Feb;29(2):245-51
references.pmid: 22075980
references.citation: Zhong Yao Cai. 2003 May;26(5):338-40
references.pmid: 14535015
references.citation: Zhongguo Zhong Yao Za Zhi. 2003 Oct;28(10):957-61
references.pmid: 15620188
references.citation: Gastroenterology. 2007 Jun;132(7):2557-76
references.pmid: 17570226
references.citation: Zhongguo Zhong Yao Za Zhi. 2009 Mar;34(6):761-5
references.pmid: 19624024
references.citation: Hematol Oncol Clin North Am. 2002 Oct;16(5):1189-227
references.pmid: 12512389
references.citation: Mol Cell Neurosci. 2012 Mar;49(3):282-9
references.pmid: 22197680
references.citation: Anticancer Res. 2006 Sep-Oct;26(5A):3579-83
references.pmid: 17094486