Primary role for CD4(+) T lymphocytes in recovery from oropharyngeal candidiasis

Abstract

Oropharyngeal candidiasis is associated with defects in cell-mediated immunity and is commonly seen in human immunodeficiency virus positive individuals and AIDS patients. A model for oral candidiasis in T-cell-deficient BALB/c and CBA/CaH nu/nu mice was established. After inoculation with 10(8) Candida albicans yeasts, these mice displayed increased levels of oral colonization compared to euthymic control mice and developed a chronic oropharyngeal infection. Histopathological examination of nu/nu oral tissues revealed extensive hyphae penetrating the epithelium, with polymorphonuclear leukocyte microabscess formation. Adoptive transfer of either naive or immune lymphocytes into immunodeficient mice resulted in the recovery of these animals from the oral infection. Reconstitution of immunodeficient mice with naive CD4(+) but not CD8(+) T cells significantly decreased oral colonization compared to controls. Interleukin-12 and gamma interferon were detected in the draining lymph nodes of immunodeficient mice following reconstitution with naive lymphocytes. This study demonstrates the direct requirement for T lymphocytes in recovery from oral candidiasis and suggests that this is associated with the production of cytokines by CD4(+) T helper cells.

FIG. 1.

Time course of C. albicans clearance in BALB/c nu/nu (A) and CBA/CaH nu/nu (B) immunodeficient mice compared to wild-type controls following oral infection with 10⁸ C. albicans yeasts. Bars represent scores (mean ± SEM) for a minimum of 30 mice (10 mice/group in three separate experiments). SEM is 0 if error bars are not shown. All groups were significantly different (P < 0.001) except CBA/CaH nu/nu and CBA/CaH on day 1.

FIG. 2.

Time course of C. albicans clearance in heterozygous BALB/c nu/+ and wild-type +/+ counterparts following oral infection with 10⁸ C. albicans yeasts. Bars represent scores (mean ± SEM) for a minimum of 10 mice/group. Each experiment was repeated twice. ∗, significantly different at P < 0.05.

FIG. 3.

Histopathological sections of tongue from immunodeficient BALB/c nu/nu (A) and CBA/CaH (B) nu/nu mice on day 6 after oral C. albicans infection with 10⁸ yeasts. There is heavy infiltration of Candida hyphae penetrating the superficial hyperkeratotic epithelium, and large numbers of PMNLs forming microabscesses (C). Sections were stained with periodic acid-Schiff. Bars, 50 μm (A) and 100 μm (B). Total magnification, ×600 (C).

FIG. 4.

Time course of C. albicans clearance in BALB/c nu/nu mice after thymus transplantation. Bars represent scores (mean ± SEM) for a minimum of 10 mice/group. Each experiment was repeated twice. Comparison was made between animals that received a thymus transplant (Thymus +) and controls (Thymus −) that received no thymus. ∗, significantly different at P < 0.001. SEM is 0 if error bars are not shown.

FIG. 5.

Time course of C. albicans clearance in BALB/c nu/nu (A) and CBA/CaH nu/nu (B) mice reconstituted with 3 × 10⁷ immune (orally primed) or naive lymphocytes. Bars represent scores (mean ± SEM) for a minimum of 10 mice/group. Each experiment was repeated twice. Comparison was made between animals that received lymphocytes and controls that received no cells. ∗, significantly different at P < 0.001. SEM is 0 if error bars are not shown.

FIG. 6.

Time course of C. albicans clearance in BALB/c nu/nu mice reconstituted with 2.5 × 10⁶ naive CD4⁺ or CD8⁺ T cells. The positive control group received 3 × 10⁷ naive lymphocytes, while the negative control group received no cells. Bars represent scores (mean ± SEM) for a minimum of 10 mice/group. Each experiment was repeated twice. Comparison was made between animals that received either CD4⁺ or CD8⁺ T cells and negative controls. ∗, significantly different at P < 0.001 for CD4 group; •, significantly different at P < 0.05 for CD8 group. Positive control group was significantly different from negative control group at P < 0.001 from days 21 to 70.

FIG. 7.

Histopathological section of tongue from immunodeficient BALB/c nu/nu mouse following lymphocyte reconstitution (A) (day 35 posttransfer), and infected control (B) that received no cells. Mice were infected with 10⁸ C. albicans yeasts and reconstituted with 3 × 10⁷ naive lymphocytes on day 6 postinfection. Sections show fewer hyphae penetrating the epithelium in the reconstituted mouse, with some intraepithelial PMNLs (A), compared to the heavy hyphal penetration and PMNL microabscess formation in the untreated control (B). Sections were stained with periodic acid-Schiff. Bars, 45 μm.

FIG. 8.

Representative agarose gel stained with ethidium bromide showing CD4 and CD8 gene products in BALB/c nu/nu mice following lymphocyte reconstitution. RT-PCR was performed on BALB/c tongue and oral mucosa. Lane 1 shows size markers, while lane 2 shows the cDNA negative control for each gel. CD4 is not present in control nude mice (lane 3), whereas CD8 is present at all times (lanes 4, 6, and 8). CD4 is present in oral mucosa on day 4 (lane 5) and day 8 (lane 7) following lymphocyte transfer, but is not seen in the tongue until day 21 (data not shown).