Neurochemical Research

, Volume 33, Issue 9, pp 1795–1803 | Cite as

Artepillin C Derived from Propolis Induces Neurite Outgrowth in PC12m3 Cells via ERK and p38 MAPK Pathways

  • Yoshio Kano
  • Noboru Horie
  • Shima Doi
  • Fumika Aramaki
  • Hidefumi Maeda
  • Fukumi Hiragami
  • Kenji Kawamura
  • Hirotoshi Motoda
  • Yoshihisa Koike
  • Junichi Akiyama
  • Sueo Eguchi
  • Ken Hashimoto
Original Paper


We investigated whether artepillin C, a major component of Brazilian propolis, acts as a neurotrophic-like factor in rat PC12m3 cells, in which nerve growth factor (NGF)-induced neurite outgrowth is impaired. When cultures of PC12m3 cells were treated with artepillin C at a concentration of 20 μM, the frequency of neurite outgrowth induced by artepillin C was approximately 7-fold greater than that induced by NGF alone. Artepillin C induced-neurite outgrowth of PC12m3 cells was inhibited by the ERK inhibitor U0126 and by the p38 MAPK inhibitor SB203580. Although artepillin C-induced p38 MAPK activity was detected in PC12m3 cells, phosphorylation of ERK induced by artepillin C was not observed. On the other hand, artepillin C caused rapid activation of ERK and the time course of the activation was similar to that induced by NGF treatment in PC12 parental cells. However, NGF-induced neurite outgrowth was inhibited by artepillin C treatment. Interestingly, inhibition of ERK by U0126 completely prevented artepillin C-induced p38 MAPK phosphorylation of PC12m3 cells. These findings suggest that artepillin C-induced activation of p38 MAPK through the ERK signaling pathway is responsible for the neurite outgrowth of PC12m3 cells.


Artepillin C NGF MAP kinase PC12m3 cells 


  1. 1.
    Shimazawa M, Chikamatsu S, Morimoto N, Mishima S, Nagai H, Hara H (2005) Neuroprotection by Brazilian green propolis against in vitro and in vivo ischemic neuronal damage. eCAM 2:201–207PubMedCrossRefGoogle Scholar
  2. 2.
    Isla MI, Nieva Moreno MI, Sampietro AR, Vattuone MA (2001) Antioxidant activity of Argentine propolis extracts. J Ethnopharmacol 76:165–170PubMedCrossRefGoogle Scholar
  3. 3.
    Natarajan K, Singh S, Burker TR Jr, Grunberger D, Aggarwal BB (1996) Caffeic acid phenethyl ester is a potent and specific inhibitor of activation of nuclear transcription factor NF-KB. Proc Natl Acad Sci USA 93:9090–9095PubMedCrossRefGoogle Scholar
  4. 4.
    Amodio R, De Ruvo C, Di Matteo V, Poggi A, Di Santo A, Martelli N, Di Matteo V, Lorenzet R, Poqqi A, Rotilio D, Cacchio M, Esposito E (2003) Caffeic acid phenethyl ester blocks apoptosis by low potassium in cerebellar granule cells. Int J Dev Neurosci 21:379–389PubMedCrossRefGoogle Scholar
  5. 5.
    Wei X, Zhao L, Ma Z, Holtzman DM, Yan C, Dodel RC, Hampel H, Oertel W, Farlow MO, Du Y (2004) Caffeic acid phenethl ester prevents neonatal hypoxic-ischaemic brain injury. Brain 127:2629–2635PubMedCrossRefGoogle Scholar
  6. 6.
    Ilhan A, Iraz M, Gurel A, Armutcu F, Akyol O (2004) Caffeic acid phenethyl ester exerts a neuroprotective effect on CNS against pentylnetetrazol-induced seizures in mice. Neurochem Res 29:2287–2292PubMedCrossRefGoogle Scholar
  7. 7.
    Kimoto T, Koya-Miyata S, Hino K, Micallet MJ, Hanaya T, Arai S, Ikeda M, Kurimoto M (2001) Pulmonary carcinogenesis induced by ferric nitrilotriacetate in mice and protection from it by Brazilian propolis and artepillin C. Virchows Arch 438:259–270PubMedCrossRefGoogle Scholar
  8. 8.
    Matsuno T, Jung S-K, Matsumoto Y, Saito M, Morikawa J (1997) Preferential cytotoxcity to tumor cells of 3,5-diprenyl-4-hydroxycinnamic acid (Artepillin C) isolated from propolis. Anticancer Res 17:3565–3568PubMedGoogle Scholar
  9. 9.
    Balasubramanian S, Efimova T, Eckert RL (2002) Green tea polyphenol stimulates a Ras, MEKK1, MEK3, and p38 cascade to increase activator protein 1 factor-dependent involucrin gene expression in normal human keratinocytes. J Biol Chem 277:1828–1836PubMedCrossRefGoogle Scholar
  10. 10.
    Greene LA, Tischler AS (1976) Establishment of a noradrenergic clonal line of rat adrenal pheochromocytoma cells which respond to nerve growth factor. Proc Natl Acad Sci USA 73:2424–2428PubMedCrossRefGoogle Scholar
  11. 11.
    Qui MS, Green SH (1991) NGF and EGF rapidly activate p21 ras in PC12 cells by distinct convergent pathways including tyrosin phoshorylation. Neuron 7:937–946CrossRefGoogle Scholar
  12. 12.
    Fukuda M, Gotoh Y,Tachibana T, Dell K, Hattori S, Yoneda Y, Nishida E (1995) Induction of neurite outgrowth by MAP kinase in PC12 cells. Oncogene 11:239–244PubMedGoogle Scholar
  13. 13.
    Thomas V, Hansen O, Rehfeld JF, Nielsen FC (2000) Cyclic AMP-induced neuronal differentiation via activation of p38 mitogen-activated protein kinase. J Neurochem 75:1870–1877CrossRefGoogle Scholar
  14. 14.
    Ichijo H (1999) From receptors to stress-activated MAP kinases. Oncogene 18:6087–6093PubMedCrossRefGoogle Scholar
  15. 15.
    Yao R, Osada H (1997) Induction of neurite outgrowth in PC12 cells by γ- lactam-related compounds via Ras-MAP kinase signaling pathway independent mechanism. Exp Cell Res 234:233–239PubMedCrossRefGoogle Scholar
  16. 16.
    Sakai T, Furuyama T, Ohoka Y, Miyazaki N, Fujioka S, Sugimoto H, Amasaki M, Hattori S, Matsuya T, Inagaki S (1999) Mouse semaphorin H induces PC12 cell neurite outgrowth activating ras-mitogen-activated protein kinase signaling pathway via Ca2+ influx. J Biol Chem 274:29666–29671PubMedCrossRefGoogle Scholar
  17. 17.
    Hirao T, Tsuge N, Imai S, Shin-ya K, Seto H (1995) Nerfilin I, a novel microbial metabolite inducing neurite outgrowth of PC12 cells. J Antibiotics 48:1494–1496Google Scholar
  18. 18.
    Saito Y, Kawashima S (1988) Enhancement of neurite outgrowth in PC12h cells by a protease inhibitor. Neurosci Lett 89:102–107PubMedCrossRefGoogle Scholar
  19. 19.
    Vossler MR, Yao H, York RD, Pan MG, Rim CS, Stork PJ (1997) cAMP activates MAP kinase and Elk-1 through a B-Raf- and Rap1-dependent pathway. Cell 89:73–82PubMedCrossRefGoogle Scholar
  20. 20.
    Rusanescu G, Qi H, Thomas SM, Brugge JS, Halegoua S (1995) Calcium influx induces neurite outgrowth through a src-ras signaling cassette. Neuron 15:1415–1425PubMedCrossRefGoogle Scholar
  21. 21.
    Kano Y, Nohno T, Hasegawa T,Takahashi R, Hiragami F, Kawamura K, Iwama MK, Motoda H, Miyamoto K (2002) Immunosuppressant FK506 induce neurite outgrowth in PC12 mutant cells with impaired NGF-promoted neuritogenesis via a novel MAP kinase signaling pathway. Neurochem Res 27:1655–1661PubMedCrossRefGoogle Scholar
  22. 22.
    Riese U, Ziegler E, Hamburger M (2004) Militarinone A induces differentiation in PC12 cells via MAP and Akt kinase signal transduction pathways. FEBS Letters 577:455–459PubMedCrossRefGoogle Scholar
  23. 23.
    Krek W, DeCaprio JA (1995) Cell synchronization. Methods Enzymol 254:114–124PubMedCrossRefGoogle Scholar
  24. 24.
    Jang JH, Surh YJ (2004) Bcl-2 attenuation of oxidative cell death is associated with up-regulation of γ-glutamylcysteine ligase via constitutive NF-κB activation. J Biol Chem 279:38779–38786PubMedCrossRefGoogle Scholar
  25. 25.
    Morooka T, Nishida E (1998) Requirement of p38 mitogen-activated protein kinase for neuronal differentiation in PC12 cells. J Biol Chem 273:24285–24288PubMedCrossRefGoogle Scholar
  26. 26.
    Kimoto T, Arai S, Kohguchi M, Aga M, Nomura Y, Micallef MJ, Kurimoto M, Mito K (1998) Apoptosis and suppression of tumor growth by artepillin C extracted from Brazilian propolis. Cancer Detect Prev 22:506–515PubMedCrossRefGoogle Scholar
  27. 27.
    Kimoto T, Koya S, Hino K, Yamamoto Y, Nomura Y, Micallet MJ, Hanaya T, Arai S, Ikeda M, Kurimoto M (2000) Renal carcinogenesis induced by ferric nitrilotriacetate in mice, and protection from it by Brazilian propolis and artepillin C. Pathol Int 50:679–689PubMedCrossRefGoogle Scholar
  28. 28.
    Kawasaki H, Morooka T, Shimohama S, Kimura J, Hirano T, Gotoh Y, Nishida E (1997) Activation and involvement of p38 mitogen-activated protein kinase in glutamate-induced apoptosis in rat cerebellar granule cells. J Biol Chem 272:18518–18521PubMedCrossRefGoogle Scholar
  29. 29.
    Yilmaz A, Kliche S, Mayr-Beyrle U, Fellbric G, Waltenberger J (2003) p38 MAPK inhibition is critically involved in VEGFR-2-mediated endothelial cell survival. Biochem Biophys Res Commun 306:730–736PubMedCrossRefGoogle Scholar
  30. 30.
    Kano Y, Nakagiri S, Nohno T, Hiragami F, Kawamura K, Kadota M, Numata K, Koike T, Furuta T (2004) Heat shock induces neurite outgrowth in PC12m3 cells via the p38 mitogen-activated protein kinase pathway. Brain Res 1026:302–306PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2008

Authors and Affiliations

  • Yoshio Kano
    • 1
  • Noboru Horie
    • 2
  • Shima Doi
    • 3
  • Fumika Aramaki
    • 3
  • Hidefumi Maeda
    • 4
  • Fukumi Hiragami
    • 5
  • Kenji Kawamura
    • 5
  • Hirotoshi Motoda
    • 5
  • Yoshihisa Koike
    • 1
  • Junichi Akiyama
    • 5
  • Sueo Eguchi
    • 5
  • Ken Hashimoto
    • 3
  1. 1.Department of Occupational Therapy, School of Health ScienceKibi International UniversityTakahashi-cityJapan
  2. 2.Department of Food Science and Nutrition, School of Human Environment ScienceMukogawa Women’s UniversityNishinomiyaJapan
  3. 3.Department of Research and DevelopmentYamada Apiculture Center, Inc.Tomata-gunJapan
  4. 4.Department of Endodontology and Operative Dentistry, Division of Oral Rehabilitation, Faculty of Dental ScienceKyushu University Higashi-kuJapan
  5. 5.Department of Physical Therapy, School of Health ScienceKibi International UniversityTakahashiJapan

Personalised recommendations