The reduced bactericidal function of complement C5-deficient murine macrophages is associated with defects in the synthesis and delivery of reactive oxygen radicals to mycobacterial phagosomes

D. Sundarsingh Daniel, Guixiang Dai, Christopher R. Singh, Devin R. Lindsey, Amanda K. Smith, Subramanian Dhandayuthapani, Robert L. Hunter, Chinnaswamy Jagannath

Research output: Contribution to journalArticle

28 Scopus citations

Abstract

Complement C5-deficient (C5-/-) macrophages derived from B.10 congenic mice ware found to be defective to killing intracellular Mycobacterium tuberculosis (MTB). They were bacteriostatic after activation with IFN-γ alone but bactericidal in the combined presence of IFN-γ and C5-derived C5a anaphylatoxin that was deficient among these macrophages. Reduced killing correlated with a decreased production of reactive oxygen species (ROS) in the C5-/- macrophages measured using fluorescent probes. Furthermore, a lack of colocalizatlon of p47phox protein of the NADPH oxidase (phox) complex with GFP-espressing MTB (gfpMTB) indicated a defective assembly of the phox complex on phagosomes. Reconstitution with C5a, a known ROS activator, enhanced the assembly of phox complex on the phagosomes as well as the production of ROS that inhibited the growth of MTB. Protein kinase C (PKC) isoforms are involved in the phosphorylation and translocation of p47 phox onto bacterial phagosomes. Western blot analysis demonstrated a defective phosphorylation of PKC (α, β, δ) and PKC-ζ in the cytosol of C5-/- macrophages compared with C5 intact (C5 +/+) macrophages. Furthermore, in site fluorescent labeling of phagosomes indicated that PKC-β and PKC-ζ were the isoforms that are not phosphorylated in C5-/- macrophages. Because Fc receptor-mediated phox assembly was normal in both C5-/- and C5+/+ macrophages, the defect in phox assembly around MTB phagosomes was specific to C5 deficiency. Reduced bactericidal function of C5-/- macrophages thus appears to be due to a defective assembly and production of ROS that prevents effective killing of intracellular MTB.

Original languageEnglish (US)
Pages (from-to)4688-4698
Number of pages11
JournalJournal of Immunology
Volume177
Issue number7
DOIs
StatePublished - Oct 1 2006

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology

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