Article Figures & Data
Figures
- Fig. 1.
(A) Production of nitrite by rat peritoneal adherent cells stimulated with different concentrations of E. coliLPS (●), B. abortus S-LPS (▴), and B. melitensis S-LPS (■). Adherent rat peritoneal cells were stimulated with LPS at the concentrations indicated. NO2− production was assayed 24 h after stimulation. Data are the means ± the standard errors of the means of results of four experiments with duplicate samples. (B) Kinetics of nitrite synthesis by adherent peritoneal cells stimulated with 1 μg of E. coli LPS (●), B. abortusS-LPS (▴), or B. melitensis S-LPS (■) per ml. Cells were allowed to adhere to plastic dishes and then were stimulated as indicated. Open circles indicate cells without any stimulus. Data are the means ± the standard errors of the means of results of three experiments performed in duplicate. (C) Production of nitrite by rat peritoneal adherent cells stimulated with different concentrations ofE. coli (●), B. abortus (▴), or B. melitensis (■) lipid A. Nitrite production was assayed after 24 h of incubation. Data are the means ± the standard errors of the means of results of four experiments with duplicate samples. The nitrite levels induced by Brucella S-LPS and lipid A were significantly different from those of E. coli. ∗,P < 0.05; ∗∗, P < 0.01; ∗∗∗, P < 0.001.
- Fig. 2.
Effect of l-NMA on nitrite production induced by E. coli, B. abortus, and B. melitensis LPS and lipid A. Adherent cells were incubated with 1 μg of the indicated LPS (A) or lipid A (B) per ml, in the presence (solid bars) or absence (hatched bars) of 0.5 mM l-NMA. Nitrite production was assayed after 24 h. Open bars show the production by control cells. Data are the means ± the standard errors of the means of results of three experiments performed in duplicate. The nitrite levels induced in the presence ofl-NMA were significantly different from those induced with only the LPS or lipid A. ∗, P < 0.001.
- Fig. 3.
Effect of E. coli, B. abortus, andB. melitensis LPS and lipid A on iNOS mRNA. Adherent cells were treated with 1 μg of LPS or lipid A per ml. (A) Total RNA was isolated after 24 h of stimulation and evaluated by Northern blot analysis with 32P-labeled iNOS and β-actin DNA probes. The relative level of NOS mRNA expression was determined for each lane after normalization to the respective β-actin signal. The iNOS signal obtained in the absence of LPS was assigned a value of 1 to allow calculation of a relative level of mRNA expression for all other treatments. (B) The histogram shows the densitometric analysis of the autoradiograph shown in panel A. This is a representative experiment of four similar ones.
- Fig. 4.
Time course of iNOS mRNA induction by E. coliLPS (A and B), B. abortus S-LPS (C and D), and B. melitensis S-LPS (E and F). Total RNA was isolated at the indicated times and used for Northern blot analysis with32P-labeled iNOS and β-actin DNA probes. The relative level of iNOS mRNA expression was determined for each lane after normalization to the respective β-actin signal. The iNOS signal at 0 h was assigned a value of 1 to allow calculation of a relative level of mRNA expression for all other treatments. Results of scanning densitometric analysis of the autoradiographs in panels A, C, and E are presented in the histograms of panels B, D, and E (hatched bars indicate cells incubated for 24 h in the absence of LPS). These results are representative of those from three similar experiments.
- Fig. 5.
Analysis of iNOS protein in peritoneal macrophages treated with E. coli, B. abortus, and B. melitensis LPS. Adherent cells were incubated with 10 μg of LPS per ml for 24 h. Cell lysates containing 40 μg of protein were analyzed by Western blotting with a rabbit anti-mouse iNOS antibody and peroxidase-conjugated anti-rabbit immunoglobulin G. These results are representative of three experiments.
