Naunyn-Schmiedeberg's Archives of Pharmacology

, Volume 381, Issue 1, pp 93–105 | Cite as

Protective effect of verbascoside in activated C6 glioma cells: possible molecular mechanisms

  • Emanuela Esposito
  • Roberto Dal Toso
  • Giovanna Pressi
  • Placido Bramanti
  • Rosaria Meli
  • Salvatore Cuzzocrea
ORIGINAL ARTICLE

Abstract

The glycosylated phenylpropanoid verbascoside (VB), isolated from cultured cells of the medicinal plant Syringa vulgaris (Oleaceae), has previously been characterized as an effective scavenger of biologically active free radicals and an inhibitor of lipid peroxidation. The aim of the present study was to evaluate in a rat glioma cell line (C6) the effect of VB biotechnologically produced by S. vulgaris plant cell cultures in the regulation of the inflammatory response. We used a model of central nervous system inflammation induced by bacterial endotoxin/cytokine (lipopolysaccharide (LPS)/interferon (IFN)-γ, 1 μg/ml and 100 U/ml, respectively). Our results show that the treatment with LPS/IFN-γ for 24 h elicited the induction of inducible nitric oxide synthase (iNOS) activity as determined by NO x accumulation in the culture medium. Preincubation with VB (10–100 μg/ml) abrogated the mixed cytokine-mediated induction of iNOS. The effect was concentration-dependent. Our studies also showed an inhibitory effect of VB on neuronal nitric oxide synthase expression. Moreover, Western blot analysis showed that this glycoside prevents specifically the activation of the proinflammatory enzyme cyclooxygenase (COX)-2 in glioma cells without simultaneous inhibition of COX-1 enzyme. Moreover, we found that VB reduced the expression of proinflammatory enzymes in LPS/IFN-γ through the inhibition of the activation of nuclear factor kappa B and mitogen-activated protein kinase signaling pathway. The mechanisms underlying in vitro the neuroprotective properties of VB involve modulation of transcription factors and consequent altered gene expression, resulting in downregulation of inflammation. These findings provide support that VB may provide a promising approach for the treatment of oxidative-stress-related neurodegenerative diseases.

Keywords

Verbascoside NF-κB pathway Glioma cell line iNOS COX-2 

Abbreviations

PPGs

Phenylpropanoid glycosides

PPs

Phenylpropanoids

VB

Verbascoside

NO

Nitric oxide

NOS

Nitric oxide synthase

CNS

Central nervous system

NF-κB

Nuclear factor kappa B

MAPKs

Mitogen-activated protein kinases

DMEM

Dulbecco’s modified essential medium

LPS

Lipopolysaccharide

IFN

Interferon

MDA

Malondialdehyde

COX

Cyclooxygenase

References

  1. Aggarwal BB (2004) Nuclear factor-kappa B: the enemy within. Cancer Cell 6:203–208CrossRefPubMedGoogle Scholar
  2. Ames BN, Shigenaga MK, Hagen TM (1993) Oxidants, antioxidants, and the degenerative diseases of aging. Proc Natl Acad Sci U S A 90:7915–7922CrossRefPubMedGoogle Scholar
  3. Aoki E, Semba R, Mikoshiba K, Kashiwamata S (1991) Predominant localization in glial cells of free l-arginine. Immunocytochemical evidence. Brain Res 547:190–192CrossRefPubMedGoogle Scholar
  4. Beal MF (2003) Bioenergetic approaches for neuroprotection in Parkinson’s disease. Ann Neurol 53(Suppl 3):S39–S47 discussion S47–38CrossRefPubMedGoogle Scholar
  5. Bissell MG, Rubinstein LJ, Bignami A, Herman MM (1974) Characteristics of the rat C-6 glioma maintained in organ culture systems. Production of glial fibrillary acidic protein in the absence of gliofibrillogenesis. Brain Res 82:77–89CrossRefPubMedGoogle Scholar
  6. Bolanos JP, Almeida A, Stewart V, Peuchen S, Land JM, Clark JB, Heales SJ (1997) Nitric oxide-mediated mitochondrial damage in the brain: mechanisms and implications for neurodegenerative diseases. J Neurochem 68:2227–2240PubMedGoogle Scholar
  7. Brambilla R, Neary JT, Cattabeni F, Cottini L, D'Ippolito G, Schiller PC, Abbracchio MP (2002) Induction of COX-2 and reactive gliosis by P2Y receptors in rat cortical astrocytes is dependent on ERK1/2 but independent of calcium signalling. J Neurochem 83:1285–1296CrossRefPubMedGoogle Scholar
  8. Bredt DS, Snyder SH (1990) Isolation of nitric oxide synthetase, a calmodulin-requiring enzyme. Proc Natl Acad Sci U S A 87:682–685CrossRefPubMedGoogle Scholar
  9. Calabrese V, Scapagnini G, Giuffrida Stella AM, Bates TE, Clark JB (2001) Mitochondrial involvement in brain function and dysfunction: relevance to aging, neurodegenerative disorders and longevity. Neurochem Res 26:739–764CrossRefPubMedGoogle Scholar
  10. Calabrese V, Boyd-Kimball D, Scapagnini G, Butterfield DA (2004) Nitric oxide and cellular stress response in brain aging and neurodegenerative disorders: the role of vitagenes. In Vivo 18:245–267PubMedGoogle Scholar
  11. Cieslik K, Zhu Y, Wu KK (2002) Salicylate suppresses macrophage nitric-oxide synthase-2 and cyclo-oxygenase-2 expression by inhibiting CCAAT/enhancer-binding protein-beta binding via a common signaling pathway. J Biol Chem 277:49304–49310CrossRefPubMedGoogle Scholar
  12. Dawson VL, Dawson TM (1996) Nitric oxide neurotoxicity. J Chem Neuroanat 10:179–190CrossRefPubMedGoogle Scholar
  13. de Vera ME, Shapiro RA, Nussler AK, Mudgett JS, Simmons RL, Morris SM Jr, Billiar TR, Geller DA (1996) Transcriptional regulation of human inducible nitric oxide synthase (NOS2) gene by cytokines: initial analysis of the human NOS2 promoter. Proc Natl Acad Sci U S A 93:1054–1059CrossRefPubMedGoogle Scholar
  14. Delacourte A (1990) General and dramatic glial reaction in Alzheimer brains. Neurology 40:33–37PubMedGoogle Scholar
  15. Dexter DT, Carter CJ, Wells FR, Javoy-Agid F, Agid Y, Lees A, Jenner P, Marsden CD (1989) Basal lipid peroxidation in substantia nigra is increased in Parkinson’s disease. J Neurochem 52:381–389CrossRefPubMedGoogle Scholar
  16. Dignam JD, Lebovitz RM, Roeder RG (1983) Accurate transcription initiation by RNA polymerase II in a soluble extract from isolated mammalian nuclei. Nucleic Acids Res 11:1475–1489CrossRefPubMedGoogle Scholar
  17. Eddleston M, Mucke L (1993) Molecular profile of reactive astrocytes—implications for their role in neurologic disease. Neuroscience 54:15–36CrossRefPubMedGoogle Scholar
  18. Emerit J, Edeas M, Bricaire F (2004) Neurodegenerative diseases and oxidative stress. Biomed Pharmacother 58:39–46CrossRefPubMedGoogle Scholar
  19. Endoh M, Maiese K, Wagner J (1994) Expression of the inducible form of nitric oxide synthase by reactive astrocytes after transient global ischemia. Brain Res 651:92–100CrossRefPubMedGoogle Scholar
  20. Ennamany R, Marzetto S, Saboureau D, Creppy EE (1995) Lipid peroxidation induced by bolesatine, a toxin of Boletus satanas: implication in m5dC variation in Vero cells related to inhibition of cell growth. Cell Biol Toxicol 11:347–354CrossRefPubMedGoogle Scholar
  21. Galea E, Reis DJ, Feinstein DL (1994) Cloning and expression of inducible nitric oxide synthase from rat astrocytes. J Neurosci Res 37:406–414CrossRefPubMedGoogle Scholar
  22. Gegg ME, Beltran B, Salas-Pino S, Bolanos JP, Clark JB, Moncada S, Heales SJ (2003) Differential effect of nitric oxide on glutathione metabolism and mitochondrial function in astrocytes and neurones: implications for neuroprotection/neurodegeneration? J Neurochem 86:228–237CrossRefPubMedGoogle Scholar
  23. Gilmore TD (1999) The Rel/NF-kappa B signal transduction pathway: introduction. Oncogene 18:6842–6844CrossRefPubMedGoogle Scholar
  24. Haddad JJ (2004) Oxygen sensing and oxidant/redox-related pathways. Biochem Biophys Res Commun 316:969–977CrossRefPubMedGoogle Scholar
  25. Hsu S (2005) Green tea and the skin. J Am Acad Dermatol 52:1049–1059CrossRefPubMedGoogle Scholar
  26. Ito CY, Kazantsev AG, Baldwin AS Jr (1994) Three NF-kappa B sites in the I kappa B-alpha promoter are required for induction of gene expression by TNF alpha. Nucleic Acids Res 22:3787–3792CrossRefPubMedGoogle Scholar
  27. Kelleher ZT, Matsumoto A, Stamler JS, Marshall HE (2007) NOS2 regulation of NF-kappaB by S-nitrosylation of p65. J Biol Chem 282:30667–30672CrossRefPubMedGoogle Scholar
  28. Korkina LG (2007) Phenylpropanoids as naturally occurring antioxidants: from plant defense to human health. Cell Mol Biol 53:15–25PubMedGoogle Scholar
  29. Korkina LG, Mikhal’chik E, Suprun MV, Pastore S, Dal Toso R (2007) Molecular mechanisms underlying wound healing and anti-inflammatory properties of naturally occurring biotechnologically produced phenylpropanoid glycosides. Cell Mol Biol 53:84–91PubMedGoogle Scholar
  30. Kris-Etherton PM, Hecker KD, Bonanome A, Coval SM, Binkoski AE, Hilpert KF, Griel AE, Etherton TD (2002) Bioactive compounds in foods: their role in the prevention of cardiovascular disease and cancer. Am J Med 113(Suppl 9B):71S–88SCrossRefPubMedGoogle Scholar
  31. Lee JK, Choi SS, Won JS, Suh HW (2003) The regulation of inducible nitric oxide synthase gene expression induced by lipopolysaccharide and tumor necrosis factor-alpha in C6 cells: involvement of AP-1 and NFkappaB. Life Sci 73:595–609PubMedGoogle Scholar
  32. Lee JY, Woo ER, Kang KW (2005) Inhibition of lipopolysaccharide-inducible nitric oxide synthase expression by acteoside through blocking of AP-1 activation. J Ethnopharmacol 97:561–566CrossRefPubMedGoogle Scholar
  33. Lin LC, Wang YH, Hou YC, Chang S, Liou KT, Chou YC, Wang WY, Shen YC (2006) The inhibitory effect of phenylpropanoid glycosides and iridoid glucosides on free radical production and beta2 integrin expression in human leucocytes. J Pharm Pharmacol 58:129–135CrossRefPubMedGoogle Scholar
  34. Lowenstein CJ, Alley EW, Raval P, Snowman AM, Snyder SH, Russell SW, Murphy WJ (1993) Macrophage nitric oxide synthase gene: two upstream regions mediate induction by interferon gamma and lipopolysaccharide. Proc Natl Acad Sci U S A 90:9730–9734CrossRefPubMedGoogle Scholar
  35. Lyras L, Cairns NJ, Jenner A, Jenner P, Halliwell B (1997) An assessment of oxidative damage to proteins, lipids, and DNA in brain from patients with Alzheimer’s disease. J Neurochem 68:2061–2069PubMedCrossRefGoogle Scholar
  36. Marchetti B (1997) Cross-talk signals in the CNS: role of neurotrophic and hormonal factors, adhesion molecules and intercellular signaling agents in luteinizing hormone-releasing hormone (LHRH)-astroglial interactive network. Front Biosci 2:88–125Google Scholar
  37. Matkowski A (2008) Plant in vitro culture for the production of antioxidants—a review. Biotechnol Adv 26:548–560CrossRefPubMedGoogle Scholar
  38. Mattace Raso G, Esposito E, Iacono A, Pacilio M, Coppola A, Bianco G, Diano S, Di Carlo R, Meli R (2006) Leptin induces nitric oxide synthase type II in C6 glioma cells. Role for nuclear factor-kappaB in hormone effect. Neurosci Lett 396:121–126CrossRefPubMedGoogle Scholar
  39. Mazzon E, Esposito E, Di Paola R, Riccardi L, Caminiti R, Dal Toso R, Pressi G, Cuzzocrea S (2009) Effects of verbascoside biotechnologically produced by Syringa vulgaris plant cell cultures in a rodent model of colitis. Naunyn Schmiedebergs Arch Pharmacol 380:79–94CrossRefPubMedGoogle Scholar
  40. Minagar A, Shapshak P, Fujimura R, Ownby R, Heyes M, Eisdorfer C (2002) The role of macrophage/microglia and astrocytes in the pathogenesis of three neurologic disorders: HIV-associated dementia, Alzheimer disease, and multiple sclerosis. J Neurol Sci 202:13–23CrossRefPubMedGoogle Scholar
  41. Moncada S, Palmer RM, Higgs EA (1991) Nitric oxide: physiology, pathophysiology, and pharmacology. Pharmacol Rev 43:109–142PubMedGoogle Scholar
  42. Mosmann T (1983) Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays. J Immunol Methods 65:55–63CrossRefPubMedGoogle Scholar
  43. Murphy MP (1999) Nitric oxide and cell death. Biochim Biophys Acta 1411:401–414CrossRefPubMedGoogle Scholar
  44. Nathan C (1992) Nitric oxide as a secretory product of mammalian cells. Faseb J 6:3051–3064PubMedGoogle Scholar
  45. Nathan C, Xie QW (1994) Nitric oxide synthases: roles, tolls, and controls. Cell 78:915–918CrossRefPubMedGoogle Scholar
  46. Nomura Y (2001) NF-kappaB activation and IkappaB alpha dynamism involved in iNOS and chemokine induction in astroglial cells. Life Sci 68:1695–1701CrossRefPubMedGoogle Scholar
  47. Perron NR, Brumaghim JL (2009) A review of the antioxidant mechanisms of polyphenol compounds related to iron binding. Cell Biochem Biophys 53:75–100CrossRefPubMedGoogle Scholar
  48. Pfeiffer SE, Herschman HR, Lightbody J, Sato G (1970) Synthesis by a clonal line of rat glial cells of a protein unique to the nervous system. J Cell Physiol 75:329–339CrossRefPubMedGoogle Scholar
  49. Pineda-Molina E, Klatt P, Vazquez J, Marina A, Garcia de Lacoba M, Perez-Sala D, Lamas S (2001) Glutathionylation of the p50 subunit of NF-kappaB: a mechanism for redox-induced inhibition of DNA binding. Biochemistry 40:14134–14142CrossRefPubMedGoogle Scholar
  50. Pu X, Song Z, Li Y, Tu P, Li H (2003) Acteoside from Cistanche salsa inhibits apoptosis by 1-methyl-4-phenylpyridinium ion in cerebellar granule neurons. Planta Med 69:65–66CrossRefPubMedGoogle Scholar
  51. Radi R, Peluffo G, Alvarez MN, Naviliat M, Cayota A (2001) Unraveling peroxynitrite formation in biological systems. Free Radic Biol Med 30:463–488CrossRefPubMedGoogle Scholar
  52. Rudolf E, Andelova H, Cervinka M (2007) Polyphenolic compounds in chemoprevention of colon cancer—targets and signaling pathways. Anticancer Agents Med Chem 7:559–575PubMedGoogle Scholar
  53. Sheng GQ, Zhang JR, Pu XP, Ma J, Li CL (2002) Protective effect of verbascoside on 1-methyl-4-phenylpyridinium ion-induced neurotoxicity in PC12 cells. Eur J Pharmacol 451:119–124CrossRefPubMedGoogle Scholar
  54. Shindo K, Saito E, Sekiya M, Matsui T, Koike Y (2008) Antioxidative activity of the flower of Torenia fournieri. Nat Med 62:247–248CrossRefGoogle Scholar
  55. Singh DK, Lippman SM (1998) Cancer chemoprevention. Part 2: hormones, nonclassic antioxidant natural agents, NSAIDs, and other agents. Oncology (Williston Park) 12:1787–1800; discussion 1802, 1805Google Scholar
  56. Smetanska I (2008) Production of secondary metabolites using plant cell cultures. Adv Biochem Eng Biotechnol 111:187–228PubMedGoogle Scholar
  57. Tognolini M, Barocelli E, Ballabeni V, Bruni R, Bianchi A, Chiavarini M, Impicciatore M (2006) Comparative screening of plant essential oils: phenylpropanoid moiety as basic core for antiplatelet activity. Life Sci 78:1419–1432PubMedGoogle Scholar
  58. Torreilles F, Salman-Tabcheh S, Guerin M, Torreilles J (1999) Neurodegenerative disorders: the role of peroxynitrite. Brain Res Brain Res Rev 30:153–163CrossRefPubMedGoogle Scholar
  59. Vernadakis A (1996) Glia–neuron intercommunications and synaptic plasticity. Prog Neurobiol 49:185–214CrossRefPubMedGoogle Scholar
  60. Xie QW, Whisnant R, Nathan C (1993) Promoter of the mouse gene encoding calcium-independent nitric oxide synthase confers inducibility by interferon gamma and bacterial lipopolysaccharide. J Exp Med 177:1779–1784CrossRefPubMedGoogle Scholar
  61. Xiong Q, Kadota S, Tani T, Namba T (1996) Antioxidative effects of phenylethanoids from Cistanche deserticola. Biol Pharm Bull 19:1580–1585PubMedGoogle Scholar
  62. Xiong Q, Tezuka Y, Kaneko T, Li H, Tran LQ, Hase K, Namba T, Kadota S (2000) Inhibition of nitric oxide by phenylethanoids in activated macrophages. Eur J Pharmacol 400:137–144CrossRefPubMedGoogle Scholar
  63. Xu Z, Wang BR, Wang X, Kuang F, Duan XL, Jiao XY, Ju G (2006) ERK1/2 and p38 mitogen-activated protein kinase mediate iNOS-induced spinal neuron degeneration after acute traumatic spinal cord injury. Life Sci 79:1895–1905CrossRefPubMedGoogle Scholar
  64. Yun HY, Dawson VL, Dawson TM (1997) Nitric oxide in health and disease of the nervous system. Mol Psychiatry 2:300–310CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag 2009

Authors and Affiliations

  • Emanuela Esposito
    • 1
  • Roberto Dal Toso
    • 2
  • Giovanna Pressi
    • 2
  • Placido Bramanti
    • 1
  • Rosaria Meli
    • 3
  • Salvatore Cuzzocrea
    • 1
    • 4
    • 5
  1. 1.IRCCS Centro Neurolesi “Bonino-Pulejo”MessinaItaly
  2. 2.I.R.B. srlAltavilla VicentinaItaly
  3. 3.Department of Experimental PharmacologyUniversity of Naples Federico IINaplesItaly
  4. 4.Department of Clinical and Experimental Medicine and Pharmacology, School of MedicineUniversity of MessinaMessinaItaly
  5. 5.Institute of Pharmacology, School of MedicineUniversity of MessinaMessinaItaly

Personalised recommendations