title: Necrotic Bone Stimulates Proinflammatory Responses in Macrophages through the Activation of Toll-Like Receptor 4.
abstract: In Legg-Calvé-Perthes disease, loss of blood supply results in ischemic osteonecrosis of the femoral head (ONFH). Generally, macrophages play important roles in inflammatory responses to tissue necrosis, but their role in ONFH is not known. The purpose of this study was to determine the macrophage-inflammatory responses after ONFH and the receptor mechanisms involved in sensing the necrotic bone, using a piglet model of Legg-Calvé-Perthes disease. Induction of ONFH resulted in increased numbers of CD14+macrophages in the fibrovascular repair tissue compared with normal, as determined by immunohistochemistry. Quantitative real-time PCR analysis of macrophages isolated by laser capture microdissection showed significantly increased expression of proinflammatory cytokines IL-1β, tumor necrosis factor-α, and IL-6 in ONFH compared with normal. Because Toll-like receptors (TLRs) mediate macrophage-inflammatory responses in other inflammatory conditions, we determined their gene expression in macrophages and found significantly increased levels of TLR4 but not TLR2 and TLR9 in ONFH. Mechanistically, in vitro, bone marrow-derived macrophages treated with necrotic bone showed increased extracellular signal-regulated kinases 1/2 and Iκ kinase-α phosphorylation, increased proliferation, migration, and inflammatory cytokine expression, which were blocked by TLR4 inhibitor, TAK242, and by TLR4 ablation in macrophages using the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein-9 nuclease method. In conclusion, necrotic bone stimulates macrophage-inflammatory responses through TLR4 activation.
authors.affiliation: Center for Excellence in Hip Disorders, Texas Scottish Rite Hospital for Children, Dallas, Texas; Department of Orthopedic Surgery, University of Texas Southwestern Medical Center, Dallas, Texas.
authors.forename: Naga Suresh
authors.initials: NS
authors.lastname: Adapala
authors.affiliation: Center for Excellence in Hip Disorders, Texas Scottish Rite Hospital for Children, Dallas, Texas; Department of Orthopedic Surgery, University of Texas Southwestern Medical Center, Dallas, Texas.
authors.forename: Ryosuke
authors.initials: R
authors.lastname: Yamaguchi
authors.affiliation: Center for Excellence in Hip Disorders, Texas Scottish Rite Hospital for Children, Dallas, Texas.
authors.forename: Matthew
authors.initials: M
authors.lastname: Phipps
authors.affiliation: Center for Excellence in Hip Disorders, Texas Scottish Rite Hospital for Children, Dallas, Texas.
authors.forename: Olumide
authors.initials: O
authors.lastname: Aruwajoye
authors.affiliation: Center for Excellence in Hip Disorders, Texas Scottish Rite Hospital for Children, Dallas, Texas; Department of Orthopedic Surgery, University of Texas Southwestern Medical Center, Dallas, Texas. Electronic address: harry.kim@tsrh.org.
authors.forename: Harry K W
authors.initials: HKW
authors.lastname: Kim
mesh_terms: D000818:Animals; D001853:Bone Marrow / Q000276:immunology / Q000473:pathology; D049109:Cell Proliferation; D016207:Cytokines / Q000378:metabolism; D004195:Disease Models, Animal; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D005271:Femur Head Necrosis / Q000276:immunology* / Q000473:pathology; D007249:Inflammation / Q000276:immunology / Q000473:pathology; D007873:Legg-Calve-Perthes Disease / Q000276:immunology* / Q000473:pathology; D008264:Macrophages / Q000276:immunology*; D013552:Swine; D051195:Toll-Like Receptor 2 / Q000378:metabolism; D051197:Toll-Like Receptor 4 / Q000276:immunology*; D051193:Toll-Like Receptors / Q000378:metabolism
chemical_list: D016207:Cytokines; D051195:Toll-Like Receptor 2; D051197:Toll-Like Receptor 4; D051193:Toll-Like Receptors; D048049:Extracellular Signal-Regulated MAP Kinases