title: Shikonin suppresses ERK 1/2 phosphorylation during the early stages of adipocyte differentiation in 3T3-L1 cells.
abstract: BACKGROUND
  The naphthoquinone pigment, shikonin, is a major component of Lithospermum erythrorhizon and has been shown to have various biological functions, including antimicrobial, anti-inflammatory, and antitumor effects. In this study, we investigated the effect of shikonin on adipocyte differentiation and its mechanism of action in 3T3-L1 cells.
  
  
  METHODS
  To investigate the effects of shikonin on adipocyte differentiation, 3T3-L1 cells were induced to differentiate using 3-isobutyl-1-methylzanthine, dexamethasone, and insulin (MDI) for 8 days in the presence of 0-2 μM shikonin. Oil Red O staining was performed to determine the lipid accumulation in 3T3-L1 cells. To elucidate the anti-adipogenic mechanism of shikonin, adipogenic transcription factors, the phosphorylation levels of ERK, and adipogenic gene expression were analyzed by Western blotting and quantitative real-time PCR. To further confirm that shikonin inhibits adipogenic differentiation through downregulation of ERK 1/2 activity, 3T3-L1 cells were treated with shikonin in the presence of FGF-2, an activator, or PD98059, an inhibitor, of the ERK1/2 signaling pathway.
  
  
  RESULTS
  Shikonin effectively suppressed adipogenesis and downregulated the protein levels of 2 major transcription factors, PPARγ and C/EBPα, as well as the adipocyte specific gene aP2 in a dose-dependent manner. qRT-PCR analysis revealed that shikonin inhibited mRNA expression of adipogenesis-related genes, such as PPARγ, C/EBPα, and aP2. Adipocyte differentiation was mediated by ERK 1/2 phosphorylation, which was confirmed by pretreatment with PD98059 (an ERK 1/2 inhibitor) or FGF-2 (an ERK 1/2 activator). The phosphorylation of ERK1/2 during the early stages of adipogenesis in 3T3-L1 cells was inhibited by shikonin. We also confirmed that FGF-2-stimulated ERK 1/2 activity was attenuated by shikonin.
  
  
  CONCLUSIONS
  These results demonstrate that shikonin inhibits adipogenic differentiation via suppression of the ERK signaling pathway during the early stages of adipogenesis.
authors.affiliation: Functional Food Technology Research Group, Korea Food Research Institute, 516, Baekhyn-dong, Seongnam-si, Gyeonggi-do 463-746, Republic of Korea.
authors.forename: So Young
authors.initials: SY
authors.lastname: Gwon
authors.forename: Ji Yun
authors.initials: JY
authors.lastname: Ahn
authors.forename: Chang Hwa
authors.initials: CH
authors.lastname: Jung
authors.forename: Bo Kyung
authors.initials: BK
authors.lastname: Moon
authors.forename: Tae Youl
authors.initials: TY
authors.lastname: Ha
mesh_terms: D041721:3T3-L1 Cells; D017667:Adipocytes / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D050156:Adipogenesis / Q000187:drug effects*; D000818:Animals; D015536:Down-Regulation / Q000187:drug effects; D029684:Lithospermum / Q000737:chemistry*; D020935:MAP Kinase Signaling System / Q000187:drug effects; D051379:Mice; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism*; D009285:Naphthoquinones / Q000494:pharmacology*; D010766:Phosphorylation / Q000187:drug effects; D010936:Plant Extracts / Q000494:pharmacology*
chemical_list: D009285:Naphthoquinones; D010936:Plant Extracts; C016101:shikonin; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3
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