title: TLR4-HMGB1-, MyD88- and TRIF-dependent signaling in mouse intestinal ischemia/reperfusion injury.
abstract: OBJECTIVE
  To characterize high-mobility group protein 1-toll-like receptor 4 (HMGB1-TLR4) and downstream signaling pathways in intestinal ischemia/reperfusion (I/R) injury.
  
  
  METHODS
  Forty specific-pathogen-free male C57BL/6 mice were randomly divided into five groups (n = 8 per group): sham, control, anti-HMGB1, anti-myeloid differentiation gene 88 (MyD88), and anti-translocating-chain-associating membrane protein (TRIF) antibody groups. Vehicle with the control IgG antibody, anti-HMGB1, anti-MyD88, or anti-TRIF antibodies (all 1 mg/kg, 0.025%) were injected via the caudal vein 30 min prior to ischemia. After anesthetization, the abdominal wall was opened and the superior mesenteric artery was exposed, followed by 60 min mesenteric ischemia and then 60 min reperfusion. For the sham group, the abdominal wall was opened for 120 min without I/R. Levels of serum nuclear factor (NF)-κB p65, interleukin (IL)-6, and tumor necrosis factor (TNF)-α were measured, along with myeloperoxidase activity in the lung and liver. In addition,morphologic changes that occurred in the lung and intestinal tissues were evaluated. Levels of mRNA transcripts encoding HMGB1 and NF-κB were measured by real-time quantitative PCR, and levels of HMGB1 and NF-κB protein were measured by Western blot. Results were analyzed using one-way analysis of variance.
  
  
  RESULTS
  Blocking HMGB1, MyD88, and TRIF expression by injecting anti-HMGB1, anti-MyD88, or anti-TRIF antibodies prior to ischemia reduced the levels of inflammatory cytokines in serum; NF-κB p65: 104.64 ± 11.89, 228.53 ± 24.85, 145.00 ± 33.63, 191.12 ± 13.22, and 183.73 ± 10.81 (P < 0.05); IL-6: 50.02 ± 6.33, 104.91 ± 31.18, 62.28 ± 6.73, 85.90 ± 17.37, and 78.14 ± 7.38 (P < 0.05); TNF-α, 43.79 ± 4.18, 70.81 ± 6.97, 52.76 ± 5.71, 63.19 ± 5.47, and 59.70 ± 4.63 (P < 0.05) for the sham, control, anti-HMGB1, anti-MyD88, and anti-TRIF groups, respectively (all in pg/mL).Antibodies also alleviated tissue injury in the lung and small intestine compared with the control group in the mouse intestinal I/R model. The administration of anti-HMGB1, anti-MyD88, and anti-TRIF antibodies markedly reduced damage caused by I/R, for which anti-HMGB1 antibody had the most obvious effect.
  
  
  CONCLUSIONS
  HMGB1 and its downstream signaling pathway play important roles in the mouse intestinal I/R injury, and the effect of the TRIF-dependent pathway is slightly greater.
authors.affiliation: Jie Wang, Gui-Zhen He, Yu-Kang Wang, Qian-Kun Zhu, Wei Chen, Department of Parenteral and Enteral Nutrition, Peking Union Medical College Hospital, Centre for Translational Medicine, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100730, China.
authors.forename: Jie
authors.initials: J
authors.lastname: Wang
authors.affiliation: Jie Wang, Gui-Zhen He, Yu-Kang Wang, Qian-Kun Zhu, Wei Chen, Department of Parenteral and Enteral Nutrition, Peking Union Medical College Hospital, Centre for Translational Medicine, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100730, China.
authors.forename: Gui-Zhen
authors.initials: GZ
authors.lastname: He
authors.affiliation: Jie Wang, Gui-Zhen He, Yu-Kang Wang, Qian-Kun Zhu, Wei Chen, Department of Parenteral and Enteral Nutrition, Peking Union Medical College Hospital, Centre for Translational Medicine, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100730, China.
authors.forename: Yu-Kang
authors.initials: YK
authors.lastname: Wang
authors.affiliation: Jie Wang, Gui-Zhen He, Yu-Kang Wang, Qian-Kun Zhu, Wei Chen, Department of Parenteral and Enteral Nutrition, Peking Union Medical College Hospital, Centre for Translational Medicine, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100730, China.
authors.forename: Qian-Kun
authors.initials: QK
authors.lastname: Zhu
authors.affiliation: Jie Wang, Gui-Zhen He, Yu-Kang Wang, Qian-Kun Zhu, Wei Chen, Department of Parenteral and Enteral Nutrition, Peking Union Medical College Hospital, Centre for Translational Medicine, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100730, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Chen
authors.affiliation: Jie Wang, Gui-Zhen He, Yu-Kang Wang, Qian-Kun Zhu, Wei Chen, Department of Parenteral and Enteral Nutrition, Peking Union Medical College Hospital, Centre for Translational Medicine, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100730, China.
authors.forename: Tai
authors.initials: T
authors.lastname: Guo
mesh_terms: D033942:Adaptor Proteins, Vesicular Transport / Q000378:metabolism*; D000818:Animals; D004195:Disease Models, Animal; D005786:Gene Expression Regulation; D024243:HMGB1 Protein / Q000235:genetics / Q000378:metabolism*; D007249:Inflammation / Q000209:etiology / Q000235:genetics / Q000378:metabolism*; D018836:Inflammation Mediators / Q000378:metabolism; D015850:Interleukin-6 / Q000378:metabolism; D007421:Intestine, Small / Q000378:metabolism* / Q000473:pathology; D008099:Liver / Q000378:metabolism; D008168:Lung / Q000378:metabolism; D008297:Male; D017538:Mesenteric Artery, Superior / Q000601:surgery; D008641:Mesenteric Vascular Occlusion / Q000209:etiology / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D008810:Mice, Inbred C57BL; D053594:Myeloid Differentiation Factor 88 / Q000378:metabolism*; D009195:Peroxidase / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D015427:Reperfusion Injury / Q000209:etiology / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D015398:Signal Transduction; D051197:Toll-Like Receptor 4 / Q000378:metabolism*; D051996:Transcription Factor RelA / Q000235:genetics / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
chemical_list: D033942:Adaptor Proteins, Vesicular Transport; D024243:HMGB1 Protein; D018836:Inflammation Mediators; D015850:Interleukin-6; D053594:Myeloid Differentiation Factor 88; D009195:Peroxidase; D012333:RNA, Messenger; D051197:Toll-Like Receptor 4; D051996:Transcription Factor RelA; D014409:Tumor Necrosis Factor-alpha; C582808:HMGB1 protein, mouse; C507780:Myd88 protein, mouse; C498144:Rela protein, mouse; C476286:Ticam1 protein, mouse; C493487:Tlr4 protein, mouse; C577686:interleukin-6, mouse
keywords: C57BL/6 mouse; High-mobility group protein 1; Intestinal ischemia-reperfusion injury; Myeloid differentiation gene 88; Nuclear factor-κB translocating-chain-associating membrane protein