title: Paeoniflorin ameliorates ischemic neuronal damage in vitro via adenosine A1 receptor-mediated transactivation of epidermal growth factor receptor.
abstract: OBJECTIVE
  Paeoniflorin from Chinese herb Paeoniae Radix has been shown to ameliorate middle cerebral artery occlusion-induced ischemia in rats. The aim of this study was to investigate the mechanisms underlying the neuroprotective action of PF in cultured rat cortical neurons.
  
  
  METHODS
  Primary cultured cortical neurons of rats were subjected to oxygen-glucose deprivation and reoxygenation (OGD/R) insult. Cell survival was determined using MTT assay. HEK293 cells stably transfected with A1R (HEK293/A1R) were used for detailed analysis. Phosphorylation of the signaling proteins was evaluated by Western blot or immunoprecipitation. Receptor interactions were identified using co-immunoprecipitation and immunofluorescence staining.
  
  
  RESULTS
  Paeoniflorin (10 nmol/L to 1 μmol/L) increased the survival of neurons subjected to OGD/R. Furthermore, paeoniflorin increased the phosphorylation of Akt and ERK1/2 in these neurons. These effects were blocked by PI3K inhibitor wortmannin or MEK inhibitor U0126. Paeoniflorin also increased the phosphorylation of Akt and ERK1/2 in HEK293/A1R cells. Both A1R antagonist DPCPX and EGFR inhibitor AG1478 not only blocked paeoniflorin-induced phosphorylation of ERK1/2 and Akt in HEK293/A1R cells, but also paeoniflorin-increased survival of neurons subjected to OGD/R. In addition, paeoniflorin increased the phosphorylation of Src kinase and activation of MMP-2 in HEK293/A1R cells. Both Src inhibitor PP2 and MMP-2/MMP-9 inhibitor BiPs not only blocked paeoniflorin-induced phosphorylation of ERK1/2 (and Akt) in HEK293/A1R cells, but also paeoniflorin-increased survival of neurons subjected to OGD/R.
  
  
  CONCLUSIONS
  Paeoniflorin promotes the survival of cultured cortical neurons by increasing Akt and ERK1/2 phosphorylation via A1R-mediated transactivation of EGFR.
authors: CAS Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China.
authors: Min
authors: M
authors: Zhong
authors: CAS Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China.
authors: Wan-ling
authors: WL
authors: Song
authors: CAS Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China.
authors: Ye-chun
authors: YC
authors: Xu
authors: CAS Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China.
authors: Yang
authors: Y
authors: Ye
authors: CAS Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China.
authors: Lin-yin
authors: LY
authors: Feng
mesh_terms: D000818:Animals; D002545:Brain Ischemia / Q000188:drug therapy* / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D015687:Cell Hypoxia; D002470:Cell Survival / Q000187:drug effects; D002540:Cerebral Cortex / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D019610:Cytoprotection; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D066246:ErbB Receptors / Q000187:drug effects* / Q000378:metabolism; D005865:Gestational Age; D005960:Glucosides / Q000494:pharmacology*; D057809:HEK293 Cells; D006801:Humans; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D039821:Monoterpenes / Q000494:pharmacology*; D009410:Nerve Degeneration; D009474:Neurons / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D018696:Neuroprotective Agents / Q000494:pharmacology*; D010766:Phosphorylation; D061251:Primary Cell Culture; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D017207:Rats, Sprague-Dawley; D020239:Receptor Cross-Talk / Q000187:drug effects*; D043682:Receptor, Adenosine A1 / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D013997:Time Factors; D014162:Transfection; D019061:src-Family Kinases / Q000378:metabolism; D006027:Glycosides
chemical_list: D066246:ErbB Receptors; D005960:Glucosides; D020778:Matrix Metalloproteinase 2; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D039821:Monoterpenes; D018696:Neuroprotective Agents; D051057:Proto-Oncogene Proteins c-akt; D043682:Receptor, Adenosine A1; D019061:src-Family Kinases; C015423:peoniflorin; D006027:Glycosides; C512478:EGFR protein, human; C483803:Egfr protein, rat; C535150:MAPK1 protein, human; C535149:Mapk1 protein, rat; C522361:MMP2 protein, human; C522363:Mmp2 protein, rat