Figure 5.

Chemoattractant gradients prime human neutrophils to effectively block Aspergillus fumigatus growth. A, In the absence of chemoattractant, low numbers of neutrophils begin accumulating in the chemotaxis chamber at 3 h, soon after conidia start swelling. On average, around 50 neutrophils accumulate per chamber at 6 h (blue line). However, the presence of neutrophils does not hinder hyphal growth (red line). B, In the presence of fungus and fMLP (100 nM), neutrophils begin to accumulate at 1 h. Two-fold more neutrophils migrate to the chemotaxis chamber, compared with fungus alone (blue line; P < .001). Neutrophils phagocytose first the swollen conidia (reflected by the decrease in average size of free conidia in the chemotaxis chamber; red line), and by 3 h all conidia are cleared from the chambers. The growth from conidia to hyphae is blocked for the remaining 14 h of the experiment. C, The average white blood cell (WBC) count (solid bars) and isolated neutrophils (striped bars) per chemotaxis chamber (2 devices with 16 chambers each) is presented at 14 h in the presence of A. fumigatus alone (blue bar) or with fMLP (black bar), LTB₄ (yellow bar), or interleukin 8 (IL-8; green bar). Positive (purple) and negative (red) control bars represent the number of neutrophils migrating toward chemotaxis chambers with and those without fMLP, respectively. D, WBCs (buffy coat) significantly decrease fungal growth alone (P < .001), reducing growth from 80% (red bar) to 40% (blue bar) of conidia into hyphae. In the presence of fMLP (black bar), LTB₄ (yellow bar), or IL-8 (green bar), <1% of conidia convert to hyphal growth. Isolated neutrophils reduce hyphae growth to 40% (blue stripe) and to <5% with fMLP (black stripe). All data are from 18 h. E, Normalized neutrophil fungus growth–blocking capacities in the presence of fMLP (P < .01) and LTB₄ (P = .02, by the nonparametric t test) gradients are significantly larger than in the presence of uniform concentrations. Neutrophil-blocking capacity is calculated as follows: ([percentage of fungi killed with chemoattractant]/[percentage of fungi killed without chemoattractant]) × ([number of neutrophils per well or chamber with chemoattractant]/[number of neutrophils per well or chamber without chemoattractant]).