title: [The study of the protective effect and its mechanism of Edaravone to neurons with hydrogen peroxide stimulated].
abstract: OBJECTIVE
  To prove the protective effect of Edaravone to neurons and to study the particular mechanism.
  
  
  METHODS
  Neurons were collected from 18-day fetal rat brains and a culture of almost pure neurons was obtained after 14-day culture, then the cells were randomly assigned to one of the three groups: control group, hydrogen peroxide (H₂O₂)-treated group, and Edaravone-treated group. In H₂O₂-treated group, 300 µmol/L H₂O₂ was added to the medium, followed by returning to the normal culture for the presupposition of time. In Edaravone-treated group, 500 µmol/L Edaravone was prophylactically added to the medium for 30 minutes before the insult. Morphology of mitochondria was visualized by transmission electron microscopy. The rate of apoptotic cells was detected by flow cytometry analysis. The relationships between the proteins and the key proteins expressions were observed by immunoprecipitation and immunoblotting.
  
  
  RESULTS
  Compared to the Edaravone-treated group, mitochondria in H₂O₂-treated group displayed more vesicular matrix compartments at the same time. Percentage of apoptotic cells in H₂O₂-treated group after 0.5, 2, 6 and 12 h were 14.40% ± 1.23%, 45.50% ± 2.81%, 56.40% ± 3.53%, 62.50% ± 4.23%, which were higher than control group (F = 274.8, P < 0.01). Edaravone-treated group were 0.90% ± 0.07%, 1.10% ± 0.08%, 3.50% ± 1.90%, 12.60% ± 1.10%, which were lower than H₂O₂-treated group (F = 362.7, P < 0.01). After H₂O₂ stimulation for 0.5 h in H₂O₂-treated group, the levels of p-JNK (Thr183/Tyr185) and cytochrome c in cytosol and BAX in heavy membrane were increased significantly at 0.5 h, reaching a peak at 12 h after stimulation, In addition, the expressions of p-BAD, BAX, BAD and 14-3-3 of cytoplasm decreased, however, these changes were inhibited in the Edaravone-treated group.
  
  
  CONCLUSIONS
  As a free radical scavenger, the Edaravone could protect neurons by inhibiting the activity of JNK, the disassociation of BAD from 14-3-3 and the translocation of BAX from the cytosol to mitochondria.
authors.affiliation: Department of Spine Surgery, the First Affiliated Hospital of Nanjing Medical University, Nanjing 210029, China.
authors.forename: Hao
authors.initials: H
authors.lastname: Long
authors.forename: Ning
authors.initials: N
authors.lastname: Zhang
authors.forename: Jin
authors.initials: J
authors.lastname: Fan
authors.forename: Qing-qing
authors.initials: QQ
authors.lastname: Li
authors.forename: Yi-ming
authors.initials: YM
authors.lastname: Li
authors.forename: Jian
authors.initials: J
authors.lastname: Tang
authors.forename: Gang
authors.initials: G
authors.lastname: Cheng
authors.forename: Guo-yong
authors.initials: GY
authors.lastname: Yin
authors.forename: Wei-hua
authors.initials: WH
authors.lastname: Cai
mesh_terms: D048948:14-3-3 Proteins / Q000378:metabolism; D000818:Animals; D000983:Antipyrine / Q000031:analogs & derivatives* / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D002478:Cells, Cultured; D000077553:Edaravone; D016166:Free Radical Scavengers / Q000494:pharmacology*; D006861:Hydrogen Peroxide / Q000378:metabolism; D020935:MAP Kinase Signaling System; D008928:Mitochondria / Q000187:drug effects; D009474:Neurons / Q000187:drug effects*; D018696:Neuroprotective Agents / Q000494:pharmacology*; D061251:Primary Cell Culture; D051381:Rats; D017207:Rats, Sprague-Dawley; D051028:bcl-2-Associated X Protein / Q000378:metabolism; D051025:bcl-Associated Death Protein / Q000378:metabolism
chemical_list: D048948:14-3-3 Proteins; C494821:Bad protein, rat; C494828:Bax protein, rat; D016166:Free Radical Scavengers; D018696:Neuroprotective Agents; D051028:bcl-2-Associated X Protein; D051025:bcl-Associated Death Protein; D006861:Hydrogen Peroxide; D000077553:Edaravone; D000983:Antipyrine