Cell adhesion molecule and lymphocyte activation marker expression during experimental vaginal candidiasis

Abstract

Cell-mediated immunity by Th1-type CD4(+) T cells is the predominant host defense mechanism against mucosal candidiasis. However, studies using an estrogen-dependent murine model of vaginal candidiasis have demonstrated little to no change in resident vaginal T cells during infection and no systemic T-cell infiltration despite the presence of Candida-specific systemic Th1-type responses in infected mice. The present study was designed to further investigate these observations by characterizing T-cell activation and cell adhesion molecule expression during primary and secondary C. albicans vaginal infections. While flow cytometry analysis of activation markers showed some evidence for activation of CD3(+) draining lymph node and/or vaginal lymphocytes during both primary and secondary vaginal Candida infection, CD3(+) cells expressing the homing receptors and integrins alpha(4)beta(7), alpha(M290)beta(7), and alpha(4)beta(1) in draining lymph nodes of mice with primary and secondary infections were reduced compared to results for uninfected mice. At the local level, few vaginal lymphocytes expressed integrins, with only minor changes observed during both primary and secondary infections. On the other hand, immunohistochemical analysis of vaginal cell adhesion molecule expression showed increases in mucosal addressin cell adhesion molecule 1 and vascular cell adhesion molecule 1 expression during both primary and secondary infections. Altogether, these data suggest that although the vaginal tissue is permissive to cellular infiltration during a vaginal Candida infection, the reduced numbers of systemic cells expressing the reciprocal cellular adhesion molecules may preempt cellular infiltration, thereby limiting Candida-specific T-cell responses against infection.

FIG. 1

Expression of CD25 following ConA stimulation and collagenase type IV treatment. LNC from four mice were cultured alone (B) or in the presence of ConA (C) for 24 h. Following the culture period, ConA-stimulated cells were incubated with or without collagenase (D) for 45 min. At the conclusion of these treatments, the cells were dually labeled with anti-CD3 and anti-CD25 antibodies or isotype control antibodies (A) and were analyzed by flow cytometry using isotype control antibodies to set fluorescent limits (gates). Numbers within each quadrant represent the percentage of fluorescent positive cells within lymphoid cell limits. Data shown are representative of three separate experiments. FITC, fluorescein isothiocyanate; CYC, cytochrome c.

FIG. 2

Vaginal fungal burden during primary or secondary experimental vaginitis. Mice received a primary vaginal inoculation and were allowed 4 weeks to resolve the infection, followed by a second inoculation in the presence of estrogen. Control mice that were previously naïve received a simultaneous primary inoculation in the presence of estrogen. C. albicans vaginal fungal burden was monitored over a 17-day period in mice with primary infection and a 10-day period in mice with secondary infection. Data are cumulative of four experiments using 10 to 13 mice per group. Separate mice were used for each time point. Asterisks indicate where significant decreases from results found in mice with primary infection were observed. SEM, standard error of the mean.

FIG. 3

Activation marker expression on murine T cells during experimental vaginal candidiasis. Lymphocytes were isolated from the draining lumbar lymph nodes and vaginae of mice with primary and secondary infections as well as of uninfected mice. The LNC (A) and VL (B) were dually labeled with anti-CD3 and anti-CD25 antibodies and analyzed by flow cytometry. Control samples were labeled with isotype control antibodies, and gates were set from these controls. Data shown are cumulative of four experiments using 10 mice per experiment. SEM, standard error of the mean.

FIG. 4

Expression of α₄β₁ on CD3⁺ LNC during experimental vaginal candidiasis. Lymphocytes were isolated from the draining lumbar lymph nodes of uninfected mice (A), mice with primary infection (B), and mice with secondary infection (C) on day 10 postinoculation. The LNC were triply labeled with anti-CD3, anti-α₄, and anti-β₁ antibodies and analyzed by flow cytometry. The numbers within each histogram represent the percentage of gated CD3⁺ cells present within each quadrant. PE, phycoerythrin; CYC, cytochrome c.

FIG. 5

Expression of endothelial cell adhesion molecules at the vaginal mucosa during experimental vaginal candidiasis. Frozen vaginal tissue sections from mice with primary infection and uninfected mice were labeled with anti-MAdCAM-1 (A to C), VCAM-1 (D to F), or ICAM-1 (G to I) antibodies. Frozen sections were also labeled with isotype-matched control antibodies to observe any background staining; however, only labeling with hamster IgG (J) is shown to confirm the positive staining for ICAM-1 in uninfected mice. The figure shows representative staining for several mice from four experiments.