Figure 1.
Morphology and immunophenotype of bone marrow-derived macrophages (BMDM) and dendritic cells (BMDC).
Five-day cultures of BMDM generated with 30% L929-conditioned medium (A) or BMDC generated with 20 ng/mL GM-CSF and IL-4 (B); images were taken at 200x+digital zoom. Over 98% of the BMDM were F4/80+ (C), but less than 4.5% of the macrophage-depleted BMDC were F4/80+ (D).
Figure 2.
CFSE-labeling does not alter phenotype.
A) BMDM and BMDC were fluorescent-labeled with 2 µM CFSE. B) Representative dot plots (1 out of 3 independent experiments) show that control (CTR) and CFSE-labeled macrophages (CFSE) had similar CD11b and CD11c expression profiles.
Figure 3.
CF.SE-labeling does not alter functionality.
The phagocytosis capacity of control and CFSE-labeled BMDM or CFSE-labeled BMDC were similar; 100 cells were counted; the experiment was repeated 3 times; P>0.05.
Figure 4.
Bone marrow-derived macrophages (BMDM) and dendritic cells (BMDC) become lipid-laden after in vitro infection with N. brasiliensis.
Non-infected BMDM and BMDC (A) and BMDM and BMDC infected with N. brasiliensis were stained with Oil Red O as described in Materials and Methods. Only infected macrophages and dendritic cells were Oil Red O positive. (Images were taken at 100×magnification).
Figure 5.
Transferred CFSE-labeled macrophages (Mφ) and dendritic cells (DC) become lipid-laden.
Macrophages (Mφ; in A) and dendritic cells (DC; in B) transferred at day 30 of a N. brasiliensis infection are 7 days later localized in the fibrotic ring (FR) of a microabscess (MA). Non-adherent spleen control cells (NACC; in C) transferred at day 30 of a N. brasiliensis infection are 7 days later localized outside of the FR and MA. Nile Red staining of lipid droplets is observed in transferred Mφ (D) and DC (E). Transferred cells are stained green by CFSE, lipid droplets are stained red by Nile Red, and nuclei are stained blue with DAPI.