title: Fingolimod induces neuroprotective factors in human astrocytes.
abstract: BACKGROUND
  Fingolimod (FTY720) is the first sphingosine-1-phosphate (S1P) receptor modulator approved for the treatment of multiple sclerosis. The phosphorylated active metabolite FTY720-phosphate (FTY-P) interferes with lymphocyte trafficking. In addition, it accumulates in the CNS and reduces brain atrophy in multiple sclerosis (MS), and neuroprotective effects are hypothesized.
  
  
  METHODS
  Human primary astrocytes as well as human astrocytoma cells were stimulated with FTY-P or S1P. We analyzed gene expression by a genome-wide microarray and validated induced candidate genes by quantitative PCR (qPCR) and ELISA. To identify the S1P-receptor subtypes involved, we applied a membrane-impermeable S1P analog (dihydro-S1P), receptor subtype specific agonists and antagonists, as well as RNAi silencing.
  
  
  RESULTS
  FTY-P induced leukemia inhibitory factor (LIF), interleukin 11 (IL11), and heparin-binding EGF-like growth factor (HBEGF) mRNA, as well as secretion of LIF and IL11 protein. In order to mimic an inflammatory milieu as observed in active MS lesions, we combined FTY-P application with tumor necrosis factor (TNF). In the presence of this key inflammatory cytokine, FTY-P synergistically induced LIF, HBEGF, and IL11 mRNA, as well as secretion of LIF and IL11 protein. TNF itself induced inflammatory, B-cell promoting, and antiviral factors (CXCL10, BAFF, MX1, and OAS2). Their induction was blocked by FTY-P. After continuous exposure of cells to FTY-P or S1P for up to 7 days, the extent of induction of neurotrophic factors and the suppression of TNF-induced inflammatory genes declined but was still detectable. The induction of neurotrophic factors was mediated via surface S1P receptors 1 (S1PR1) and 3 (S1PR3).
  
  
  CONCLUSIONS
  We identified effects of FTY-P on astrocytes, namely induction of neurotrophic mediators (LIF, HBEGF, and IL11) and inhibition of TNF-induced inflammatory genes (CXCL10, BAFF, MX1, and OAS2). This supports the view that a part of the effects of fingolimod may be mediated via astrocytes.
authors.affiliation: Institute of Clinical Neuroimmunology, Ludwig Maximilian University, 81377, Munich, Germany. franziska.hoffmann@med.uni-muenchen.de.
authors.forename: Franziska S
authors.initials: FS
authors.lastname: Hoffmann
authors.affiliation: Institute of Clinical Neuroimmunology, Ludwig Maximilian University, 81377, Munich, Germany. johann.hofereiter@med.uni-muenchen.de.
authors.forename: Johann
authors.initials: J
authors.lastname: Hofereiter
authors.affiliation: Institute of Clinical Neuroimmunology, Ludwig Maximilian University, 81377, Munich, Germany. heike.ruebsamen@med.uni-muenchen.de.
authors.forename: Heike
authors.initials: H
authors.lastname: Rübsamen
authors.affiliation: German Center for Neurodegenerative Diseases (DZNE) and Technical University, 81377, Munich, Germany. johannes.melms@dzne.de.
authors.forename: Johannes
authors.initials: J
authors.lastname: Melms
authors.affiliation: German Center for Neurodegenerative Diseases (DZNE) and Technical University, 81377, Munich, Germany. sigrid.schwarz@dzne.de.
authors.forename: Sigrid
authors.initials: S
authors.lastname: Schwarz
authors.affiliation: Max Planck Institute of Psychiatry, 80804, Munich, Germany. faber@psych.mpg.de.
authors.forename: Hans
authors.initials: H
authors.lastname: Faber
authors.affiliation: Max Planck Institute of Psychiatry, 80804, Munich, Germany. pweber@psych.mpg.de.
authors.forename: Peter
authors.initials: P
authors.lastname: Weber
authors.affiliation: Max Planck Institute of Psychiatry, 80804, Munich, Germany. puetz@psych.mpg.de.
authors.forename: Benno
authors.initials: B
authors.lastname: Pütz
authors.affiliation: Institute of Clinical Neuroimmunology, Ludwig Maximilian University, 81377, Munich, Germany. verena.loleit@gmx.de.
authors.forename: Verena
authors.initials: V
authors.lastname: Loleit
authors.affiliation: Max Planck Institute of Psychiatry, 80804, Munich, Germany. fweber@psych.mpg.de.
authors.forename: Frank
authors.initials: F
authors.lastname: Weber
authors.affiliation: Institute of Clinical Neuroimmunology, Ludwig Maximilian University, 81377, Munich, Germany. reinhard.hohlfeld@med.uni-muenchen.de.|Munich Cluster for Systems Neurology (SyNergy), Munich, Germany. reinhard.hohlfeld@med.uni-muenchen.de.
authors.forename: Reinhard
authors.initials: R
authors.lastname: Hohlfeld
authors.affiliation: Institute of Clinical Neuroimmunology, Ludwig Maximilian University, 81377, Munich, Germany. edgar.meinl@med.uni-muenchen.de.
authors.forename: Edgar
authors.initials: E
authors.lastname: Meinl
authors.affiliation: Institute of Clinical Neuroimmunology, Ludwig Maximilian University, 81377, Munich, Germany. markus.krumbholz@uni-tuebingen.de.|Center of Neurology and Hertie Institute for Clinical Brain Research, University of Tübingen, Tübingen, Germany. markus.krumbholz@uni-tuebingen.de.
authors.forename: Markus
authors.initials: M
authors.lastname: Krumbholz
mesh_terms: D001253:Astrocytes / Q000187:drug effects*; D054357:Chemokine CXCL10 / Q000235:genetics / Q000378:metabolism; D003342:Corpus Striatum / Q000166:cytology; D004305:Dose-Response Relationship, Drug; D004797:Enzyme-Linked Immunosorbent Assay; D005333:Fetus / Q000166:cytology; D000068876:Fingolimod Hydrochloride / Q000494:pharmacology*; D066257:Heparin-binding EGF-like Growth Factor / Q000235:genetics / Q000378:metabolism; D006801:Humans; D017370:Interleukin-11 / Q000235:genetics / Q000378:metabolism; D008246:Lysophospholipids / Q000494:pharmacology; D046228:Microarray Analysis; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism; D058953:Neural Stem Cells / Q000187:drug effects*; D018696:Neuroprotective Agents / Q000494:pharmacology*; D012333:RNA, Messenger; D034741:RNA, Small Interfering / Q000494:pharmacology; D013110:Sphingosine / Q000031:analogs & derivatives / Q000494:pharmacology; D013997:Time Factors
chemical_list: D054357:Chemokine CXCL10; C587883:HBEGF protein, human; D066257:Heparin-binding EGF-like Growth Factor; D017370:Interleukin-11; D008246:Lysophospholipids; D016328:NF-kappa B; D018696:Neuroprotective Agents; D012333:RNA, Messenger; D034741:RNA, Small Interfering; C060506:sphingosine 1-phosphate; D000068876:Fingolimod Hydrochloride; D013110:Sphingosine