title: Engagement of ubiquitination and de-ubiquitination at rostral ventrolateral medulla in experimental brain death.
abstract: BACKGROUND
  Whereas brain death is a vitally important clinical phenomenon, our contemporary understanding on its underlying cellular mechanisms remains elusive. This study evaluated whether the ubiquitin-proteasome system (UPS) in the rostral ventrolateral medulla (RVLM), a neural substrate that our laboratory identified previously to be intimately related to brain death, is engaged in this fatal process.
  
  
  METHODS
  We performed proteomics, Western Blot, real-time PCR, ELISA and pharmacological experiments in conjunction with a clinically relevant experimental endotoxemia model of brain death based on intravenous administration of Escherichia coli lipopolysaccharide in adult male Sprague-Dawley rats.
  
  
  RESULTS
  Proteomics, Western blot and enzyme activity analyses demonstrated that polyubiquitination was preserved and de-ubiquitination by ubiquitin C-terminal hydrolase isozyme-L1 (UCH-L1) was sustained, alongside increased monoubiquitin availability or proteasome activity in RVLM over the course of experimental endotoxemia. However, real-time PCR revealed no significant alteration in proteasome subunit alpha type-1, ubiquitin or UCH-L1 at mRNA level. Functionally, whereas microinjection into the bilateral RVLM of proteasome inhibitors (lactacystin or proteasome inhibitor II) potentiated survival, an inhibitor of ubiquitin-recycling (ubiquitin aldehyde) or an UCH-L1 inhibitor exacerbated mortality.
  
  
  CONCLUSIONS
  We proposed previously that the progression towards brain death entails a tug-of-war between pro-death and pro-life programs in RVLM. It is conceivable that ubiquitination or de-ubiquitination in RVLM participate in brain death by regulating the degradation of the proteins involved in those programs.
authors.affiliation: Center for Translational Research in Biomedical Sciences, Kaohsiung Chang Gung Memorial Hospital, Kaohsiung, 83301, Taiwan, Republic of China.
authors.forename: Carol H Y
authors.initials: CH
authors.lastname: Wu
authors.forename: Julie Y H
authors.initials: JY
authors.lastname: Chan
authors.forename: Jimmy Li-Jer
authors.initials: JL
authors.lastname: Chou
authors.forename: Samuel H H
authors.initials: SH
authors.lastname: Chan
authors.forename: Alice Y W
authors.initials: AY
authors.lastname: Chang
mesh_terms: D000818:Animals; D001926:Brain Death / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000633:toxicity; D008297:Male; D008526:Medulla Oblongata / Q000201:enzymology / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D025801:Ubiquitin / Q000378:metabolism; D043222:Ubiquitin Thiolesterase / Q000378:metabolism*; D054875:Ubiquitination
chemical_list: D008070:Lipopolysaccharides; D012333:RNA, Messenger; D025801:Ubiquitin; C489034:UCHL1 protein, rat; D043222:Ubiquitin Thiolesterase; D046988:Proteasome Endopeptidase Complex