Mycobacterium indicus pranii (Mw)-mediated protection against visceral leishmaniasis: involvement of TLR4 signalling

J Antimicrob Chemother. 2012 Dec;67(12):2892-902. doi: 10.1093/jac/dks315. Epub 2012 Aug 9.


Objectives: The aim of this study was to characterize the antileishmanial activity of heat-killed Mycobacterium indicus pranii (Mw) alone or in combination with a subtoxic dose of amphotericin B [AMB(st)].

Methods: Mw- and Mw + AMB(st)-mediated antileishmanial activity was evaluated by microscopic counting of intracellular amastigotes in Giemsa-stained macrophages and real-time PCR analysis of inducible nitric oxide synthase (iNOS) expression and measurement of nitric oxide generation by Griess reagent. The relationship between Mw and Toll-like receptor 4 (TLR4) signalling was studied by fluorescence-activated cell sorting, western blot and confocal microscopy. The effect of Mw alone or in combination with AMB(st) on the expression and production of interleukin (IL)-12, tumour necrosis factor-α, IL-10 and transforming growth factor-β was analysed by real-time PCR and ELISA, respectively.

Results: Mw treatment alone or with AMB(st) caused a significant increase in TLR4 expression of L. donovani-infected macrophages along with the activation of TLR4 downstream signalling, facilitating active nuclear translocation of nuclear factor κB (NF-κB). These events culminated in the up-regulation of the proinflammatory response, which was abrogated by treatment with TLR4-specific small-interfering RNA. In addition, this study demonstrates that this chemoimmunotherapeutic strategy confers protection against leishmanial pathogenesis via TLR4-dependent counter-regulation of inducible nitric oxide synthase (iNOS) and arginase1 activity.

Conclusions: These results provide a mechanistic understanding of Mw- or Mw + AMB(st)-mediated protection against leishmanial parasites within host macrophages.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amphotericin B / administration & dosage
  • Animals
  • Antiprotozoal Agents / administration & dosage
  • Blotting, Western
  • Cytokines / metabolism
  • Enzyme-Linked Immunosorbent Assay
  • Flow Cytometry
  • Gene Expression Profiling
  • Humans
  • Leishmaniasis, Visceral / immunology*
  • Leishmaniasis, Visceral / prevention & control*
  • Macrophages / parasitology
  • Mice
  • Mice, Inbred BALB C
  • Microscopy
  • Microscopy, Confocal
  • Mycobacterium / immunology*
  • Nitric Oxide / analysis
  • Nitric Oxide Synthase Type II / biosynthesis
  • Parasite Load
  • Real-Time Polymerase Chain Reaction
  • Signal Transduction*
  • Toll-Like Receptor 4 / immunology*


  • Antiprotozoal Agents
  • Cytokines
  • Tlr4 protein, mouse
  • Toll-Like Receptor 4
  • Nitric Oxide
  • Amphotericin B
  • Nitric Oxide Synthase Type II
  • Nos2 protein, mouse