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TLR9 and MyD88 are crucial for the maturation and activation of dendritic cells by paromomycin–miltefosine combination therapy in visceral leishmaniasis
Author(s) -
Das Sushmita,
Rani Mukta,
Rabidas Vidyanand,
Pandey Krishna,
Sahoo Ganesh Chandra,
Das Pradeep
Publication year - 2014
Publication title -
british journal of pharmacology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.432
H-Index - 211
eISSN - 1476-5381
pISSN - 0007-1188
DOI - 10.1111/bph.12530
Subject(s) - miltefosine , paromomycin , tlr9 , pharmacology , biology , microbiology and biotechnology , immunology , chemistry , visceral leishmaniasis , leishmaniasis , biochemistry , antibiotics , gene expression , dna methylation , aminoglycoside , gene
Background and Purpose The combination of paromomycin–miltefosine is a successful anti‐leishmanial therapy in visceral leishmaniasis ( VL ). This encouraged us to study its effect on T oll‐like receptor ( TLR )‐mediated immunomodulation of dendritic cells ( DC ), as DC maturation and activation is crucial for anti‐leishmanial activity. Experimental Approach In silico protein–ligand interaction and biophysical characterization of TLR9 –drug interaction was performed. Interaction assays of HEK293 cells with different concentrations of miltefosine and/or paromomycin were performed, and NF ‐ κB promoter activity measured. The role of TLR9 and MyD88 in paromomycin/miltefosine‐induced maturation and activation of DCs was evaluated through RNA interference techniques. The effect of drugs on DCs was measured in terms of counter‐regulatory production of IL‐12 over IL‐10 , and characterized by chromatin immunoprecipitation assay at the molecular level. Key Results Computational and biophysical studies revealed that paromomycin/miltefosine interact with TLR9 . Both drugs, as a monotherapy/combination, induced TLR9 ‐dependent NF ‐ κB promoter activity through MyD88 . Moreover, the drug combination induced TLR9 / MyD88 ‐dependent functional maturation of DCs , evident as an up‐regulation of co‐stimulatory markers, enhanced antigen presentation by increasing MHC II expression, and increased stimulation of naive T ‐cells to produce IFN ‐ γ . Both drugs, by modifying histone H3 at the promoter level, increased the release of IL‐12 , but down‐regulated IL‐10 in a TLR9 ‐dependent manner. Conclusions and Implications These results provide the first evidence that the combination of paromomycin–miltefosine critically modifies the maturation, activation and development of host DCs through a mechanism dependent on TLR9 and MyD88 . This has implications for evaluating the success of other combination anti‐leishmanial therapies that act by targeting host DCs .