Impairment of host defense against disseminated candidiasis in mice overexpressing GATA-3

Abstract

Candida species are the most common source of nosocomial invasive fungal infections. Previous studies have indicated that T-helper immune response is the critical host factor for susceptibility to Candida infection. The transcription factor GATA-3 is known as the master regulator for T-helper type 2 (Th2) differentiation. We therefore investigated the role of GATA-3 in the host defense against systemic Candida infection using GATA-3-overexpressing transgenic mice. The survival of GATA-3-overexpressing mice after Candida infection was significantly lower than that of wild-type mice. Candida outgrowth was significantly increased in the kidneys of GATA-3-overexpressing mice, compared with wild-type mice. The levels of various Th2 cytokines, including interleukin-4 (IL-4), IL-5, and IL-13, were significantly higher while the level of Th1 cytokine gamma interferon was significantly lower in the splenocytes of GATA-3-overexpressing mice after Candida infection. Recruitment of macrophages into the peritoneal cavity in response to Candida infection and their phagocytic activity were significantly lower in GATA-3-overexpressing mice than in wild-type mice. Exogenous administration of gamma interferon to GATA-3-overexpressing mice significantly reduced Candida outgrowth in the kidney and thus increased the survival rate. Administration of gamma interferon also increased the recruitment of macrophages into the peritoneal cavity in response to Candida infection. These results indicate that overexpression of GATA-3 modulates macrophage antifungal activity and thus enhances the susceptibility to systemic Candida infection, possibly by reducing the production of gamma interferon in response to Candida infection.

FIG. 1.

(A) Survival of wild-type (WT; open symbols) mice and GATA-3-overexpressing (GATA-3-tg; closed symbols) mice after intraperitoneal inoculation of 1 × 10⁸ CFU of Candida albicans (circles) or saline (squares). (B) Survival of WT (open symbols) mice and GATA-3-overexpressing (GATA-3-tg; closed symbols) mice after intravenous inoculation of 1 × 10⁶ CFU of Candida albicans (circles) or saline (squares). *, significantly different between wild-type mice and GATA-3-overexpressing mice (P < 0.05; n = 20 in each group).

FIG. 2.

Evaluation of systemic Candida infection in wild-type mice and GATA-3-overexpressing mice. Outgrowth of Candida albicans in the lungs and kidneys of wild-type (WT) mice and GATA-3-overexpressing mice (GATA3-tg) 56 days after intraperitoneal inoculation of Candida albicans (A) and 14 days after intravenous inoculation of Candida albicans (B) is shown. The results are expressed as log₁₀ CFU per organ. The experiments were performed in duplicate with five mice in each group. *, significantly different between wild-type mice and GATA-3-overexpressing mice (P < 0.05). (C) Histopathology of the kidneys of WT mice and GATA-3-overexpressing (GATA3-tg) mice 56 days after intraperitoneal inoculation (i.p.) or 14 days after intravenous inoculation (i.v.) of Candida albicans. Severe hydronephrosis is seen in GATA-3-tg mice but not in WT mice. The boxed area, magnified in the inset, shows Candida albicans colonies with numerous inflammatory cells. PAS staining. Representative photographs of the kidneys from 8 mice in each group are shown (magnification, ×10 [inset, ×100]).

FIG. 3.

The expression of IFN-γ, tumor necrosis factor-α (TNF-α), IL-4, IL-5, and IL-13 in the spleens of wild-type (WT) mice and GATA-3-overexpressing (GATA) mice before and at 3 (Day 3), 7 (Day 7), and 30 (Day 30) days after intraperitoneal inoculation of 1 × 10⁸ CFU of Candida albicans. Experiments were performed in duplicate with five mice in each group. *, significantly different between wild-type mice and GATA-3-overexpressing mice (P < 0.05).

FIG. 4.

(A) The ratio of CD4-positive cells to CD8-positive cells in the spleens of wild-type (WT) mice and GATA-3-overexpressing (GATA) mice before and 7 days (Day 7) after intraperitoneal inoculation of 1 × 10⁸ CFU of Candida albicans. Experiments were performed in duplicate with five mice in each group. (B) The proportion of gamma-interferon-producing cells among CD4-positive cells (left panel) or among CD8-positive cells (right panel) in the spleens of wild-type (WT) mice and GATA-3-overexpressing (GATA) mice before and 7 days (Day 7) after intraperitoneal inoculation of 1 × 10⁸ CFU of Candida albicans. Experiments were performed in duplicate with five mice in each group. (C) The proportion of CXCR3-positive cells (left panel) and CCR3-positive cells (right panel) among CD4-positive cells in the spleens of WT mice and GATA-3-overexpressing (GATA) mice before and 7 days (Day 7) after intraperitoneal inoculation of 1 × 10⁸ CFU of Candida albicans. Experiments were performed in duplicate with five mice in each group. (D) The proportion of gamma-interferon-producing cells among CXCR3-positive cells in the spleens of WT mice and GATA-3-overexpressing (GATA) mice before and 7 days (Day 7) after intraperitoneal inoculation of 1 × 10⁸ CFU of Candida albicans. Experiments were performed in duplicate with five mice in each group. *, significantly different between wild-type mice and GATA-3-overexpressing mice (P < 0.05).

FIG. 5.

(A) The number of total cells, macrophages, neutrophils, and lymphocytes in the peritoneal lavage fluids of wild-type (WT) mice and GATA-3-overexpressing (GATA) mice before and 3 days (Day 3) and 7 days (Day 7) after intraperitoneal inoculation of 1 × 10⁸ CFU of Candida albicans. Experiments were performed in duplicate with five mice in each group. (B) The expression of monocyte chemotactic protein 1 (MCP-1), keratinocyte-derived chemokine (KC), and macrophage inflammatory protein 2 (MIP-2) in the peritoneal macrophages of WT mice and GATA-3-overexpressing (GATA) mice before and 3 days (Day 3) and 7 days (Day 7) after intraperitoneal inoculation of 1 × 10⁸ CFU of Candida albicans. Experiments were performed in duplicate with five mice in each group. *, significantly different between wild-type mice and GATA-3-overexpressing mice (P < 0.05).

FIG. 6.

(A) Representative photographs of peritoneal lavage cells 3 days after intraperitoneal inoculation of 1 × 10⁸ CFU of Candida albicans. The fungi phagocytosed by macrophages (arrows) and neutrophils (arrowheads) are visible by PAS staining (magnification, ×100). (B and C) The proportions of macrophages (B) and neutrophils (C) phagocytosing Candida albicans in the peritoneal lavage fluids of wild-type (WT) mice and GATA-3-overexpressing (GATA) mice 3 days (Day 3) and 7 days (Day 7) after intraperitoneal inoculation of 1 × 10⁸ CFU of Candida albicans. Experiments were performed in duplicate with five mice in each group. *, significantly different between wild-type mice and GATA-3-overexpressing mice (P < 0.05).

FIG. 7.

Effects of gamma interferon supplementation on systemic Candida infection. (A) Survival of GATA-3-overexpressing mice treated with 8 × 10⁶ U of gamma interferon (GATA-3-tg, Candida, IFN-γ) (triangles) or with vehicle (GATA-3-tg, Candida, vehicle) (circles) after intraperitoneal inoculation of 1 × 10⁸ CFU of Candida albicans. The survival of GATA-3-overexpressing mice after intraperitoneal inoculation of saline was also evaluated with (squares) or without (stars) treatment with gamma interferon. n = 20 in each group. *, significantly different between the GATA-3-tg-Candida-IFN-γ group and GATA-3-tg-Candida-vehicle group (P < 0.05). (B) Outgrowth of Candida albicans in the lungs and kidneys of GATA-3-overexpressing mice treated with 8 × 10⁶ U of IFN-γ (gray bars) or with vehicle (black bars) 56 days after Candida albicans inoculation. The results were expressed as log₁₀ CFU per organ. Experiments were performed in duplicate with five mice in each group. *, significantly different between mice treated with IFN-γ and those treated with vehicle (P < 0.05). (C) The number of total cells, macrophages, neutrophils, and lymphocytes in the peritoneal lavage fluids of GATA-3-overexpressing mice treated with 8 × 10⁶ U of IFN-γ (gray bars) or with vehicle (black bars) 56 days after Candida albicans inoculation. Experiments were performed in duplicate with five mice in each group. *, significantly different between mice treated with IFN-γ and with vehicle (P < 0.05).