Fig. 6
From: Galectin-3 impacts Cryptococcus neoformans infection through direct antifungal effects
Gal-3 interacts with the C. neoformans capsule and inhibits the fungal growth. Capsular strain H99 (a and c) and acapsular strain CAP67 (b and d) were incubated for 72 h at 37 °C with Gal-3 (0.001 to 10 μg/ml), and the optical density at 540 nm (OD540) measured every hour (a and b), or the number or colony-forming units (CFU) were counted for determination of cell densities (c and d). e H99 and CAP67 strains were incubated for 72 h at 37 °C with Gal-3 (10 μg/ml), and the frequency of viable cells assessed using propidium iodide staining. Heated fungal cells (90 °C for 10 min) were used as positive control (blue bar). H99 and CAP67 strains were incubated for 40 min at 4 °C with Gal-3 40 μg/ml. After that, both strains were washed with PBS and incubated for 45 min with anti-Gal-3 antibody. Then, washed again with PBS and incubated with anti-rabbit IgG-FITC antibody for 40 min at 4 °C. Labeled cells were acquired on a FACS Guava easyCyte, and the histogram represents the percentage of positive cells recognized by Gal-3 (f). Anti-rabbit IgG-FITC antibody associated or not with Gal-3 was used as control, and use of WGA lectin (30 μg/ml) as control of binding with capsule or cell wall (f). g H99 strain was cultured at 37 °C for 24 h and incubated with Gal-3. C. neoformans were stained for observation of cell wall (CW) with calcofluor white (blue), Gal-3 with anti-Gal-3 antibody (green), and capsule with anti-GXM (red). Body (h), and capsule (i) size of H99 strain cells were assessed by microscopic analysis after cultivation in the presence of Gal-3 10 μg/ml. Data are representative of three experiments. Statistically significant differences are denoted by asterisk (*p < 0.05, unpaired Student’s t-test)
