Nitric oxide and TNF-alpha production by murine peritoneal macrophages activated with a novel 20-kDa protein isolated from Bacillus thuringiensis var. thuringiensis parasporal bodies.
The effects of a novel 20-kDa protein isolated from Bacillus thuringiensis var. thuringiensis (BTp20) parasporal bodies on macrophage activation were investigated and compared with the activity of LPS. Murine resident or IFN-gamma-primed peritoneal macrophages (50 U/ml of IFN-gamma for 16 h) were treated with 50 microg/ml of BTp20, alone or in combination with LPS (1 to 100 ng/ml). BTp20 was not toxic for macrophages as determined by the MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) reduction assay, however, 16% reduction of viability was observed when macrophages were treated with a combination of IFN-gamma, BTp20, and LPS at 100 ng/ml. BTp20 was able to stimulate significant cellular adherence and spreading, and significant production of nitrite and TNF-alpha by resident macrophages after 1.5 h of treatment; nitrite release, however, was induced with only 10 min of macrophage exposure to BTp20. BTp20 activities were significantly potentiated by IFN-gamma pretreatment. It was also observed that nitrite release by IFN-gamma-primed macrophages activated with LPS (1-100 ng/ml) was 20 times lower than that induced by IFN-gamma + BTp20. BTp20 and LPS independently stimulated significant TNF-alpha production by IFN-gamma-primed macrophages, the effect of BTp20 + LPS (1 and 10 ng/ml) combination was additive. In summary, this study demonstrated that BTp20 activates macrophage functions in the absence of IFN-gamma or LPS, and that IFN-gamma enhances BTp20 activities.
Gomez-Flores
R
R
Department of Bioimmunotherapy, The University of Texas M.D. Anderson Cancer Center, Houston 77030, USA.
Rodriguez-Padilla
C
C
Mehta
R T
RT
Galan-Wong
L
L
Mendoza-Gamboa
E
E
Tamez-Guerra
R
R
D000818:Animals; D001413:Bacillus thuringiensis / Q000378:metabolism*; D001426:Bacterial Proteins / Q000302:isolation & purification / Q000494:pharmacology*; D002478:Cells, Cultured; D008262:Macrophage Activation / Q000187:drug effects*; D017737:Macrophages, Peritoneal / Q000378:metabolism*; D051379:Mice; D009569:Nitric Oxide / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*
D001426:Bacterial Proteins; D014409:Tumor Necrosis Factor-alpha; D009569:Nitric Oxide