title: Heme activates TLR4-mediated inflammatory injury via MyD88/TRIF signaling pathway in intracerebral hemorrhage.
abstract: BACKGROUND
  Inflammatory injury plays a critical role in intracerebral hemorrhage (ICH)-induced neurological deficits; however, the signaling pathways are not apparent by which the upstream cellular events trigger innate immune and inflammatory responses that contribute to neurological impairments. Toll-like receptor 4 (TLR4) plays a role in inflammatory damage caused by brain disorders.
  
  
  METHODS
  In this study, we investigate the role of TLR4 signaling in ICH-induced inflammation. In the ICH model, a significant upregulation of TLR4 expression in reactive microglia has been demonstrated using real-time RT-PCR. Activation of microglia was detected by immunohistochemistry, cytokines were measured by ELISA, MyD88, TRIF and NF-κB were measured by Western blot and EMSA, animal behavior was evaluated by animal behavioristics.
  
  
  RESULTS
  Compared to WT mice, TLR4(-/-) mice had restrained ICH-induced brain damage showing in reduced cerebral edema and lower neurological deficit scores. Quantification of cytokines including IL-6, TNF-α and IL-1β and assessment of macrophage infiltration in perihematoma tissues from TLR4(-/-), MyD88(-/-) and TRIF(-/-) mice showed attenuated inflammatory damage after ICH. TLR4(-/-) mice also exhibited reduced MyD88 and TRIF expression which was accompanied by decreased NF-κB activity. This suggests that after ICH both MyD88 and TRIF pathways might be involved in TLR4-mediated inflammatory injury possibly via NF-κB activation. Exogenous hemin administration significantly increased TLR4 expression and microglial activation in cultures and also exacerbated brain injury in WT mice but not in TLR4(-/-) mice. Anti-TLR4 antibody administration suppressed hemin-induced microglial activation in cultures and in the mice model of ICH.
  
  
  CONCLUSIONS
  Our findings suggest that heme potentiates microglial activation via TLR4, in turn inducing NF-κB activation via the MyD88/TRIF signaling pathway, and ultimately increasing cytokine expression and inflammatory injury in ICH. Targeting TLR4 signaling may be a promising therapeutic strategy for ICH.
authors.affiliation: Department of Neurology, Daping Hospital, Third Military Medical University, Yuzhong District, Chongqing, China.
authors.forename: Sen
authors.initials: S
authors.lastname: Lin
authors.forename: Qing
authors.initials: Q
authors.lastname: Yin
authors.forename: Qi
authors.initials: Q
authors.lastname: Zhong
authors.forename: Feng-Lin
authors.initials: FL
authors.lastname: Lv
authors.forename: Yu
authors.initials: Y
authors.lastname: Zhou
authors.forename: Jing-Qi
authors.initials: JQ
authors.lastname: Li
authors.forename: Jing-Zhou
authors.initials: JZ
authors.lastname: Wang
authors.forename: Bing-yin
authors.initials: BY
authors.lastname: Su
authors.forename: Qing-Wu
authors.initials: QW
authors.lastname: Yang
mesh_terms: D033942:Adaptor Proteins, Vesicular Transport / Q000172:deficiency / Q000378:metabolism*; D000818:Animals; D000831:Animals, Newborn; D000911:Antibodies, Monoclonal / Q000627:therapeutic use; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology; D002543:Cerebral Hemorrhage / Q000150:complications* / Q000188:drug therapy; D016207:Cytokines / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D004797:Enzyme-Linked Immunosorbent Assay; D006418:Heme / Q000494:pharmacology*; D007249:Inflammation / Q000209:etiology; D008070:Lipopolysaccharides / Q000494:pharmacology; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D017628:Microglia / Q000187:drug effects; D053594:Myeloid Differentiation Factor 88 / Q000172:deficiency / Q000378:metabolism*; D016328:NF-kappa B / Q000378:metabolism; D009460:Neurologic Examination; D012333:RNA, Messenger / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology*; D018709:Statistics, Nonparametric; D013997:Time Factors; D051197:Toll-Like Receptor 4 / Q000172:deficiency / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D015854:Up-Regulation / Q000187:drug effects* / Q000235:genetics / Q000502:physiology
chemical_list: D033942:Adaptor Proteins, Vesicular Transport; D000911:Antibodies, Monoclonal; D016207:Cytokines; D006418:Heme; D008070:Lipopolysaccharides; D053594:Myeloid Differentiation Factor 88; D016328:NF-kappa B; D012333:RNA, Messenger; D051197:Toll-Like Receptor 4; C507780:Myd88 protein, mouse; C476286:Ticam1 protein, mouse; C493487:Tlr4 protein, mouse