Effects of macrophage colony-stimulating factor (M-CSF) on anti-fungal activity of mononuclear phagocytes against Trichosporon asahii

Abstract

Trichosporon asahii is an emerging opportunistic pathogen in immunocompromised patients. Little is known about the mechanisms of host defence against T. asahii. We investigated the fungicidal activity of human peripheral blood monocytes and murine peritoneal macrophages against T. asahii isolates, and the effects of M-CSF on the anti-fungal activity of mononuclear phagocytes. We also established a neutropenic mouse model of disseminated trichosporonosis with T. asahii. M-CSF enhanced the phagocytic fungicidal activity of mononuclear cells, and infected mice treated with human M-CSF at 10 x 106 U/kg showed a significant improvement in survival rate, with fewer fungal colony counts in the lung compared with control mice. Mice treated with human M-CSF showed higher concentrations of tumour necrosis factor-alpha (TNF-alpha) in the lung and plasma compared with control mice. The survival rate was significantly reduced in mice treated with anti-mouse TNF-alpha. Our results showed that M-CSF enhanced the fungicidal activity of mononuclear phagocytes partly by production of TNF-alpha, and suggest that the administration of M-CSF to patients with disseminated trichosporonosis may be a useful adjunct to conventional anti-microbial therapy and prophylaxis.

Fig. 1

M-CSF enhanced the fungicidal activity of human peripheral monocytes against Trichosporon asahii after 60 min incubation. Effector cell-to-target ratio used was 10:1. Data are mean ± s.e.m. of five experiments.

Fig. 2

The percentage killing of Trichosporon asahii by murine peritoneal macrophages harvested from mice treated with human M-CSF (10 × 10⁶ U/kg per day) after incubation for 60, 120 and 180 min. Effector cell-to-target ratio used was 10:1. Data are mean ± s.e.m. of five experiments. ▪, M-CSF; □, control.

Fig. 3

Organ clearance of trichosporonosis murine survival model (a) and lethal model (b). The survival model was inoculated intravenously with Trichosporon asahii at 2 × 10⁵ colony-forming units (CFU)/mouse. Four mice were killed at 12, 24, 72, 96 h and 7 and 14 days after infection. The lethal model was inoculated intravenously with T. asahii at 2 × 10⁶ CFU/mouse, and four mice were killed at 12, 24, 72, 96 h after infection. The lung (•), kidney (○), liver (Δ), heart (▴) and spleen (□) of each animal were removed aseptically and homogenized in sterile saline. The homogenate was diluted with sterile saline and incubated on Sabouraud dextrose agar (SDA) at 37°C. Data are mean ± s.e.m. of four experiments.

Fig. 4

Organ clearance of human M-CSF-treated trichosporonosis murine model. Five mice were killed at 12, 24, 48, 72 h after infection. In the human M-CSF-treated model (•) and control model (○), data are mean ± s.e.m. of five experiments. *P < 0.05 versus human M-CSF at 1 × 10⁶ U/kg per day group and control group.

Fig. 5

Effect of M-CSF on survival rates of mice with disseminated trichosporonosis. Mice immunosuppressed with two injections of 100 mg of cyclophosphamide were treated with M-CSF at 10 × 10⁶ U/kg per day (○), 1 × 10⁶ U/kg per day (•) or sterile saline (□) for 5 consecutive days starting 2 days before infection with Trichosporon asahii (2 × 10⁶ colony-forming units (CFU)/mouse). Treatment with M-CSF at 10 × 10⁶ U/kg per day significantly increased the survival rate. *P < 0.05 versus M-CSF at 1 × 10⁶ U/kg per day group and control group.

Fig. 6

Effect of anti-mouse tumour necrosis factor-alpha (TNF-α) on survival rates of trichosporonosis murine model. Every 10 mice immunosuppressed with two injections of 100 mg/kg of cyclophosphamide were treated with human M-CSF at 10 × 10⁶ U/kg per day (○), anti-mouse TNF-α at 200 μg/kg on 0 and 2 days after infection and human M-CSF at 10 × 10⁶ U/kg per day (•) or saline (□). *P < 0.01 versus anti-TNF-α with M-CSF group and M-CSF alone group.