title: Recombinant thrombomodulin inhibits lipopolysaccharide-induced inflammatory response by blocking the functions of CD14.
abstract: CD14, a multiligand pattern-recognition receptor, is involved in the activation of many TLRs. Thrombomodulin (TM), a type I transmembrane glycoprotein, originally was identified as an anticoagulant factor that activates protein C. Previously, we showed that the recombinant TM lectin-like domain binds to LPS and inhibits LPS-induced inflammation, but the function of the recombinant epidermal growth factor-like domain plus serine/threonine-rich domain of TM (rTMD23) in LPS-induced inflammation remains unknown. In the current study, we found that rTMD23 markedly suppressed the activation of intracellular signaling pathways and the production of inflammatory cytokines induced by LPS. The anti-inflammatory activity of rTMD23 was independent of activated protein C. We also found that rTMD23 interacted with the soluble and membrane forms of CD14 and inhibited the CD14-mediated inflammatory response. Knockdown of CD14 in macrophages suppressed the production of inflammatory cytokines induced by LPS, and rTMD23 inhibited LPS-induced IL-6 production in CD14-knockdown macrophages. rTMD23 suppressed the binding of LPS to macrophages by blocking the association between monocytic membrane-bound TM and CD14. The administration of rTMD23 in mice, both pretreatment and posttreatment, significantly increased the survival rate and reduced the inflammatory response to LPS. Notably, the serine/threonine-rich domain is essential for the anti-inflammatory activity of rTMD23. To summarize, we show that rTMD23 suppresses the LPS-induced inflammatory response in mice by targeting CD14 and that the serine/threonine-rich domain is crucial for the inhibitory effect of rTMD23 on LPS-induced inflammation.
authors.affiliation: Department of Biochemistry and Molecular Biology, College of Medicine, National Cheng Kung University, Tainan 701, Taiwan; and Cardiovascular Research Center, National Cheng Kung University, Tainan 701, Taiwan.
authors.forename: Chih-Yuan
authors.initials: CY
authors.lastname: Ma
authors.affiliation: Department of Biochemistry and Molecular Biology, College of Medicine, National Cheng Kung University, Tainan 701, Taiwan; and.
authors.forename: Wei-En
authors.initials: WE
authors.lastname: Chang
authors.affiliation: Department of Biochemistry and Molecular Biology, College of Medicine, National Cheng Kung University, Tainan 701, Taiwan; and Cardiovascular Research Center, National Cheng Kung University, Tainan 701, Taiwan halnwu@mail.ncku.edu.tw gyshi@mail.ncku.edu.tw.
authors.forename: Guey-Yueh
authors.initials: GY
authors.lastname: Shi
authors.affiliation: Department of Biochemistry and Molecular Biology, College of Medicine, National Cheng Kung University, Tainan 701, Taiwan; and.
authors.forename: Bi-Ying
authors.initials: BY
authors.lastname: Chang
authors.affiliation: Department of Biochemistry and Molecular Biology, College of Medicine, National Cheng Kung University, Tainan 701, Taiwan; and.
authors.forename: Sheng-En
authors.initials: SE
authors.lastname: Cheng
authors.affiliation: Department of Biochemistry and Molecular Biology, College of Medicine, National Cheng Kung University, Tainan 701, Taiwan; and.
authors.forename: Yun-Tai
authors.initials: YT
authors.lastname: Shih
authors.affiliation: Department of Biochemistry and Molecular Biology, College of Medicine, National Cheng Kung University, Tainan 701, Taiwan; and Cardiovascular Research Center, National Cheng Kung University, Tainan 701, Taiwan halnwu@mail.ncku.edu.tw gyshi@mail.ncku.edu.tw.
authors.forename: Hua-Lin
authors.initials: HL
authors.lastname: Wu
mesh_terms: D000818:Animals; D004195:Disease Models, Animal; D042783:Endothelial Cells / Q000276:immunology; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D006801:Humans; D047468:Immunoprecipitation; D007249:Inflammation / Q000276:immunology*; D018950:Lipopolysaccharide Receptors / Q000276:immunology*; D008070:Lipopolysaccharides / Q000276:immunology; D008264:Macrophages / Q000276:immunology*; D051379:Mice; D008810:Mice, Inbred C57BL; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000276:immunology; D015398:Signal Transduction / Q000276:immunology*; D020349:Surface Plasmon Resonance; D018180:Thrombomodulin / Q000276:immunology*; D014471:Umbilical Veins
chemical_list: D018950:Lipopolysaccharide Receptors; D008070:Lipopolysaccharides; D011994:Recombinant Proteins; D018180:Thrombomodulin