Mammalian DNA is an endogenous danger signal that stimulates local synthesis and release of complement factor B

Mol Med. 2012 Jul 18;18(1):851-60. doi: 10.2119/molmed.2012.00011.

Abstract

Complement factor B plays a critical role in ischemic tissue injury and autoimmunity. Factor B is dynamically synthesized and released by cells outside of the liver, but the molecules that trigger local factor B synthesis and release during endogenous tissue injury have not been identified. We determined that factor B is upregulated early after cold ischemia-reperfusion in mice, using a heterotopic heart transplant model. These data suggested upregulation of factor B by damage-associated molecular patterns (DAMPs), but multiple common DAMPs did not induce factor B in RAW264.7 mouse macrophages. However, exogenous DNA induced factor B mRNA and protein expression in RAW cells in vitro, as well as in peritoneal and alveolar macrophages in vivo. To determine the cellular mechanisms involved in DNA-induced factor B upregulation we then investigated the role of multiple known DNA receptors or binding partners. We stimulated peritoneal macrophages from wild-type (WT), toll-like receptor 9 (TLR9)-deficient, receptor for advanced glycation end products (RAGE)⁻/⁻ and myeloid differentiation factor 88 (MyD88)⁻/⁻ mice, or mouse macrophages deficient in high-mobility group box proteins (HMGBs), DNA-dependent activator of interferon-regulatory factors (DAI) or absent in melanoma 2 (AIM2), with DNA in the presence or absence of lipofection reagent. Reverse transcription-polymerase chain reaction, Western blotting and immunocytochemical analysis were employed for analysis. Synthesis of factor B was independent of TLR9, RAGE, DAI and AIM2, but was dependent on HMGBs, MyD88, p38 and NF-κB. Our data therefore show that mammalian DNA is an endogenous molecule that stimulates factor B synthesis and release from macrophages via HMGBs, MyD88, p38 and NF-κB signaling. This activation of the immune system likely contributes to damage following sterile injury such as hemorrhagic shock and ischemia-reperfusion.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Line
  • Complement Factor B / genetics
  • Complement Factor B / metabolism*
  • DNA / metabolism*
  • DNA-Binding Proteins
  • Glycoproteins / genetics
  • Glycoproteins / metabolism
  • HMGB Proteins / genetics
  • HMGB Proteins / metabolism
  • Macrophages / immunology
  • Macrophages / metabolism
  • Male
  • Mice
  • Mice, Inbred C57BL
  • Mice, Knockout
  • Mitogen-Activated Protein Kinases / metabolism
  • Myeloid Differentiation Factor 88 / genetics
  • Myeloid Differentiation Factor 88 / metabolism
  • Myocardial Reperfusion Injury / genetics
  • Myocardial Reperfusion Injury / metabolism
  • NF-kappa B / metabolism
  • Nuclear Proteins / genetics
  • Nuclear Proteins / metabolism
  • RNA Interference
  • RNA-Binding Proteins
  • Toll-Like Receptor 9 / genetics
  • Toll-Like Receptor 9 / metabolism
  • Up-Regulation / genetics

Substances

  • Aim2 protein, mouse
  • DNA-Binding Proteins
  • Glycoproteins
  • HMGB Proteins
  • Myeloid Differentiation Factor 88
  • NF-kappa B
  • Nuclear Proteins
  • RNA-Binding Proteins
  • Toll-Like Receptor 9
  • Zbp1 protein, mouse
  • DNA
  • Mitogen-Activated Protein Kinases
  • Complement Factor B