Necrotic Bone Stimulates Proinflammatory Responses in Macrophages through the Activation of Toll-Like Receptor 4

Naga Suresh Adapala, Ryosuke Yamaguchi, Matthew Phipps, Olumide Aruwajoye, Harry K.W. Kim

Research output: Contribution to journalArticle

2 Citations (Scopus)

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.

Original languageEnglish
Pages (from-to)2987-2999
Number of pages13
JournalAmerican Journal of Pathology
Volume186
Issue number11
DOIs
Publication statusPublished - Nov 1 2016
Externally publishedYes

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Toll-Like Receptor 4
Macrophage Activation
Osteonecrosis
Macrophages
Thigh
Bone and Bones
Clustered Regularly Interspaced Short Palindromic Repeats
Legg-Calve-Perthes Disease
Cytokines
Laser Capture Microdissection
Mitogen-Activated Protein Kinase 3
Toll-Like Receptors
Mitogen-Activated Protein Kinase 1
Interleukin-1
Real-Time Polymerase Chain Reaction
Interleukin-6
Necrosis
Phosphotransferases
Tumor Necrosis Factor-alpha
Immunohistochemistry

All Science Journal Classification (ASJC) codes

  • Pathology and Forensic Medicine

Cite this

Necrotic Bone Stimulates Proinflammatory Responses in Macrophages through the Activation of Toll-Like Receptor 4. / Adapala, Naga Suresh; Yamaguchi, Ryosuke; Phipps, Matthew; Aruwajoye, Olumide; Kim, Harry K.W.

In: American Journal of Pathology, Vol. 186, No. 11, 01.11.2016, p. 2987-2999.

Research output: Contribution to journalArticle

Adapala, Naga Suresh ; Yamaguchi, Ryosuke ; Phipps, Matthew ; Aruwajoye, Olumide ; Kim, Harry K.W. / Necrotic Bone Stimulates Proinflammatory Responses in Macrophages through the Activation of Toll-Like Receptor 4. In: American Journal of Pathology. 2016 ; Vol. 186, No. 11. pp. 2987-2999.
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abstract = "In Legg-Calv{\'e}-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-Calv{\'e}-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.",
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