5-LO deficiency increases the inflammatory response in the lung.

<p>Representative lung sections from WT and 5-LO^−/− mice infected with <i>H. capsulatum</i> (A). Hematoxylin-eosin staining for leukocytes (magnifications ×100) and GMS staining for yeast cells (black arrow) (magnifications ×400). (B) Neutrophils recruitment from lung parenchyma (C) TNF-α production from homogenized lungs. Cells and cytokines were obtained as described in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0031701#s3" target="_blank"><i>Material and Methods</i></a> section from mice after i.t. injection of PBS or i.t. infection with <i>H. capsulatum</i>. Cells were enumerated and identified after Rosenfeld staining, and TNF-α levels were determined by ELISA. Data are expressed as the mean ± SEM from one experiment representative of a total of two experiments (n = 6). *, p<0.05 vs. PBS; #, p<0.05 vs. WT.</p

LTB₄ and CysLTs production in lung tissue.

<p>Enzyme immunoassay quantification of LTB₄ and CysLTs concentrations in lungs from mice that had received either an i.t. PBS injection (uninfected) or an i.t. infection with <i>H. capsulatum</i>. Data are presented as the mean ± SEM and are representative of one of two independent experiments (n = 6). *, p<0.05 vs. PBS.</p

5-LO deficiency affects the ability of macrophages to phagocytose <i>H. capsulatum</i>.

<p>(A) PMs from WT and 5-LO^−/− mice were incubated for 1 h with a yeast:macrophage ratio of 1∶5 in the absence or presence of IgG. PMs were pretreated with LTB₄ (B) and LTC₄ (C) for 5 min before the addition of opsonized <i>H. capsulatum</i>. Phagocytosis was calculated as described in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0031701#s3" target="_blank"><i>Material and Methods</i></a> section and was expressed as a percentage of the control. Data are expressed as the mean ± SEM from one experiment representative of a total of three experiments (n = 6). *, p<0.05 vs. control; #, p<0.05 vs. WT cells; & p<0.05 vs. 5-LO^−/− cells.</p

Effect of 5-LO deficiency on survival, fungal burden and NO₂⁻ production.

<p>(A) 5-LO^−/− and WT mice were infected i.t. with 3×10⁶ yeast <i>H. capsulatum</i> and survival was monitored for 30 days (n = 6). CFU numbers in lungs (B) and spleen (C) were evaluated at 7 and 14 days post <i>H. capsulatum</i> infection. (D) NO₂⁻ levels were quantified in the supernatant of lung homogenates at different time points using a Griess reaction. Data are expressed as the mean ± SEM from one experiment representative of a total of two experiments (n = 6). *, p<0.05 vs. PBS; #, p<0.05 vs. WT.</p

Deficiency of 5-LO impairs T cell recruitment to the lung.

<p>Cells were obtained as described in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0031701#s3" target="_blank"><i>Material and Methods</i></a> section from mice after i.t. injection of PBS or i.t. infection with <i>H. capsulatum</i>. The lymphocyte population was gated for forward/side scatters and analyzed the percentage of T cells expressing a phenotype effector (CD44^high/CD62^low). (A) CD4⁺ T cells, CD8⁺ T cells (B) and T cell proliferation(C). Data are presented as the mean ± the SEM from three experiments. *, WT and 5-LO^−/− vs. PBS; #, WT vs. 5-LO^−/−. p<0.05.</p