Mycobacterium lepraemuriumuses TLR-6 and MR, but not lipid rafts or DC-sign, to gain access into mouse macrophages.
Mycobacterium lepraemurium (MLM), the etiologic agent of murine leprosy, is an intracellular parasite of macrophages; the mechanism used by this bacterium to enter macrophages is not known. The fate of the MLM phagosome inside macrophages is also unknown. This study was conducted to investigate how MLM enters macrophages and to define the maturation process of MLM phagosome inside macrophages.



Peritoneal macrophages were incubated in the presence of mannan-bovine serum albumin (BSA), and antibodies to known macrophage receptors, including, anti-FcγRIII/RII (anti-CD16/32), anti-CD35 (anti-CR1), anti-TLR2, anti-TLR4, anti-TLR6, anti-CD14, and anti-dendritic cell-specific intercellular adhesion molecule-3-grabbing nonintegrin (DC-SIGN). Then, macrophages were challenged with Iris Fuchsia-stained MLM, at a multiplicity of infection of 50:1. The blocking effect of the antibodies (and mannan-BSA) used was analyzed using direct microscopy and flow cytometry. The maturation process of MLM phagosomes was visualized by their interaction with antibodies to Rab5, Rab7, proton ATPase, and cathepsin D, by confocal microscopy.



Only mannan-BSA and anti-TLR6 antibody significantly blocked the entry of MLM into macrophages. None of the other antibodies, including that for DC-SIGN, meaningfully inhibited the endocytic process. We also found that MLM is a fusiogenic mycobacterium. This was deduced from the orderly association of MLM phagosomes with Rab5, Rab7, Proton ATPase, and lysosomes (cathepsin D).



Fusion of MLM phagosomes with lysosomes seems to be a necessary event for the intracellular multiplication of MLM; similar to Mycobacterium leprae, this microorganism hardly grows on artificial, synthetic, bacteriologic media.
Department of Immunology, Biomedical Research Institute, National Autonomous University of Mexico, Ciudad de, México.
Mayra
M
Silva-Miranda
Departamento de Inmunología, Escuela Nacional de Ciencias Biológicas, Instituto Politécnico Nacional, Ciudad de, México.
Patricia
P
Arce-Paredes
Departamento de Inmunología, Escuela Nacional de Ciencias Biológicas, Instituto Politécnico Nacional, Ciudad de, México.
Oscar
O
Rojas-Espinosa
D000818:Animals; D015815:Cell Adhesion Molecules / Q000276:immunology / Q000378:metabolism*; D037181:Lectins, C-Type / Q000276:immunology / Q000378:metabolism*; D008247:Lysosomes / Q000382:microbiology; D017737:Macrophages, Peritoneal / Q000187:drug effects / Q000382:microbiology*; D000090323:Mannose Receptor; D037241:Mannose-Binding Lectins / Q000276:immunology / Q000378:metabolism*; D021962:Membrane Microdomains / Q000502:physiology; D051379:Mice; D009167:Mycobacterium lepraemurium / Q000187:drug effects / Q000276:immunology / Q000502:physiology*; D010588:Phagosomes / Q000276:immunology / Q000382:microbiology; D011956:Receptors, Cell Surface / Q000276:immunology / Q000378:metabolism*; D017452:Receptors, IgG / Q000276:immunology; D051216:Toll-Like Receptor 6 / Q000276:immunology / Q000378:metabolism*; D000099215:DC-Specific ICAM-3 Grabbing Nonintegrin
D015815:Cell Adhesion Molecules; D037181:Lectins, C-Type; D000090323:Mannose Receptor; D037241:Mannose-Binding Lectins; D011956:Receptors, Cell Surface; D017452:Receptors, IgG; D051216:Toll-Like Receptor 6; D000099215:DC-Specific ICAM-3 Grabbing Nonintegrin; C495369:Tlr6 protein, mouse