title: Hypertonic saline alleviates cerebral edema by inhibiting microglia-derived TNF-α and IL-1β-induced Na-K-Cl Cotransporter up-regulation.
abstract: BACKGROUND
  Hypertonic saline (HS) has been successfully used clinically for treatment of various forms of cerebral edema. Up-regulated expression of Na-K-Cl Cotransporter 1 (NKCC1) and inflammatory mediators such as tumor necrosis factor alpha (TNF-α) and interleukin-1 beta (IL-1β) has been demonstrated to be closely associated with the pathogenesis of cerebral edema resulting from a variety of brain injuries. This study aimed to explore if alleviation of cerebral edema by 10% HS might be effected through down-regulation of inflammatory mediator expression in the microglia, and thus result in decreased NKCC1 expression in astrocytes in the cerebral cortex bordering the ischemic core.
  
  
  METHODS
  The Sprague-Dawley (SD) rats that underwent right-sided middle cerebral artery occlusion (MCAO) were used for assessment of NKCC1, TNF-α and IL-1β expression using Western blotting, double immunofluorescence and real time RT-PCR, and the model also was used for evaluation of brain water content (BWC) and infarct size. SB203580 and SP600125, specific inhibitors of the p38 and JNK signaling pathways, were used to treat primary microglia cultures to determine whether the two signaling pathways were required for the inhibition of HS on microglia expressing and secreting TNF-α and IL-1β using Western blotting, double immunofluorescence and enzyme-linked immunosorbent assay (ELISA). The effect of TNF-α and IL-1β on NKCC1 expression in primary astrocyte cultures was determined. In addition, the direct inhibitory effect of HS on NKCC1 expression in primary astrocytes was also investigated by Western blotting, double immunofluorescence and real time RT-PCR.
  
  
  RESULTS
  BWC and infarct size decreased significantly after 10% HS treatment. TNF-α and IL-1β immunoexpression in microglia was noticeably decreased. Concomitantly, NKCC1 expression in astrocytes was down-regulated. TNF-α and IL-1β released from the primary microglia subjected to hypoxic exposure and treatment with 100 mM HS were decreased. NKCC1 expression in primary astrocytes was concurrently and progressively down-regulated with decreasing concentration of exogenous TNF-α and IL-1β. Additionally, 100 mM HS directly inhibited NKCC1 up-regulation in astrocytes under hypoxic condition.
  
  
  CONCLUSIONS
  The results suggest that 10% HS alleviates cerebral edema through inhibition of the NKCC1 Cotransporter, which is mediated by attenuation of TNF-α and IL-1β stimulation on NKCC1.
authors.forename: Lin-Qiang
authors.initials: LQ
authors.lastname: Huang
authors.forename: Gao-Feng
authors.initials: GF
authors.lastname: Zhu
authors.forename: Yi-Yu
authors.initials: YY
authors.lastname: Deng
authors.forename: Wen-Qiang
authors.initials: WQ
authors.lastname: Jiang
authors.forename: Ming
authors.initials: M
authors.lastname: Fang
authors.forename: Chun-Bo
authors.initials: CB
authors.lastname: Chen
authors.forename: Wei
authors.initials: W
authors.lastname: Cao
authors.forename: Miao-Yun
authors.initials: MY
authors.lastname: Wen
authors.forename: Yong-Li
authors.initials: YL
authors.lastname: Han
authors.affiliation: Department of Emergency & Critical Care Medicine, Guangdong General Hospital, Guangdong Academy of Medical Sciences, Guangzhou 510080, PR China. zenghongke@vip.163.com.
authors.forename: Hong-Ke
authors.initials: HK
authors.lastname: Zeng
mesh_terms: D000818:Animals; D001929:Brain Edema / Q000188:drug therapy* / Q000209:etiology / Q000473:pathology; D002478:Cells, Cultured; D016207:Cytokines / Q000378:metabolism*; D004195:Disease Models, Animal; D007839:Functional Laterality; D005904:Glial Fibrillary Acidic Protein / Q000378:metabolism; D020244:Infarction, Middle Cerebral Artery / Q000150:complications; D053583:Interleukin-1beta / Q000378:metabolism; D008297:Male; D017628:Microglia / Q000187:drug effects*; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D012462:Saline Solution, Hypertonic / Q000627:therapeutic use*; D064506:Solute Carrier Family 12, Member 2 / Q000235:genetics / Q000378:metabolism*; D013997:Time Factors; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism; D015854:Up-Regulation / Q000187:drug effects*
chemical_list: D016207:Cytokines; D005904:Glial Fibrillary Acidic Protein; D053583:Interleukin-1beta; D012333:RNA, Messenger; D012462:Saline Solution, Hypertonic; D064506:Solute Carrier Family 12, Member 2; D014409:Tumor Necrosis Factor-alpha