Inflammation and clearance of Chlamydia trachomatis in enteric and nonenteric mucosae

Abstract

Immunization(s) fostering the induction of genital mucosa-targeted immune effectors is the goal of vaccines against sexually transmitted diseases. However, it is uncertain whether vaccine administration should be based on the current assumptions about the common mucosal immune system. We investigated the relationship between mucosal sites of infection, infection-induced inflammation, and immune-mediated bacterial clearance in mice using the epitheliotropic pathogen Chlamydia trachomatis. Chlamydial infection of the conjunctival, pulmonary, or genital mucosae stimulated significant changes in tissue architecture with dramatic up-regulation of the vascular addressin, VCAM, a vigorous mixed-cell inflammatory response with an influx of alpha4beta1+ T cells, and clearance of bacteria within 30 days. Conversely, intestinal mucosa infection was physiologically inapparent, with no change in expression of the local MAdCAM addressin, no VCAM induction, no histologically detectable inflammation, and no tissue pathology. Microbial clearance was complete within 60 days in the small intestine but bacterial titers remained at high levels for at least 8 months in the large intestine. These findings are compatible with the notion that VCAM plays a functional role in recruiting cells to inflammatory foci, and its absence from the intestinal mucosa contributes to immunologic homeostasis at that site. Also, expression of type 1 T cell-mediated immunity to intracellular Chlamydia may exhibit tissue-specific variation, with the rate and possibly the mechanism(s) of clearance differing between enteric and nonenteric mucosae. The implications of these data for the common mucosal immune system and the delivery of vaccines against mucosal pathogens are discussed.

Figures

FIG. 1

Integrin profiles of T and B lymphocytes recovered from Chlamydia-infected pulmonary, conjunctival, genital, and intestinal mucosae. Four groups of female C57BL/6 mice were each infected intranasally, conjunctivally, vaginally, or orally with 1.0 to 1.5 × 103 IFU of the MoPn agent. Lymphocytes collected by collagenase disruption of infected tissues were costained with a panel of MAbs recognizing murine CD3, B220, or the indicated integrin chains and were analyzed by flow cytometry as previously described (10, 35). Data summarized from 10 separate experiments represent the mean percent positive T cells (top panel) or B cells (lower panel) for each integrin marker as determined by quadrant analyses of FACS plots. Note the heightened expression of β1 integrins on T but not B lymphocytes from nonintestinal mucosae.

FIG. 2

FACS plots of integrin chains expressed by T and B lymphocytes infiltrating Chlamydia-infected pulmonary mucosae. Graphic representation of the relative fluorescent intensity of staining for the various integrin markers and the proportion of cells falling within each quadrant, for cells gated for CD3 or B220 expression. Markers such as α4, β1, and β7 are expressed on all cells at variable levels. For example, note the differential induction of β1 integrin chains on T versus B lymphocytes.

FIG. 3

Nonintegrin adhesion molecules expressed by T and B cells infiltrating mucosal sites of Chlamydia infection. CD44 is up-regulated on T but not B cells from the pulmonary, genital, and intestinal mucosae, as determined by flow cytometric analyses of cells recovered from collagenase-digested tissues (see legend to Fig. 1). l-Selectin was variably expressed on cells from either subset. l-Selectin was detected using the CD62L-specific MAb clone MEL-14, and CD44 was detected using MAb clone IM7, which recognizes the Pgp-1 isoform (PharMingen).

FIG. 4

Expression of VCAM and MAdCAM at mucosal sites of Chlamydia infection. Tissues collected from mice infected as described in Table 1 were mounted in OCT, frozen in liquid nitrogen, and stored at −80°C. Five-micrometer sections were cut on a cryostat microtome, transferred to slides, air dried, and fixed in 4°C acetone for 30 min. Tissues were stained with primary rat MAbs recognizing murine ICAM-1, VCAM-1, or MAdCAM-1, washed in PBS, and developed using mouse-Ig-absorbed tetramethylrhodamine-conjugated goat anti-rat Ig as previously described (10, 35). Slides were viewed with a Nikon Microphot SA epifluorescence microscope, and photomicrographs were taken with Fujichrome Provia 400 daylight film. Images were digitized using a Polaroid SprintScan 35-mm slide scanner, and figures were assembled in gray scale without further manipulation using Adobe Photoshop, version 4.0. Although the intense autofluorescence of murine conjunctival tissues precluded satisfactory photographic documentation, the staining profile was similar to that of other nonintestinal mucosae in that VCAM-1 but not MAdCAM-1 was detected. Arrows indicate vascular structures in each tissue.

FIG. 5

Inflammatory responses within Chlamydia-infected pulmonary, genital, and intestinal mucosae. Mice were infected as described in Fig. 1, and tissues were collected 10 to 18 days later. Mixed infiltrates comprised of neutrophils and mononuclear cells were present within infected (bottom row) but not in normal (top row) pulmonary, genital, and conjunctival (not shown) mucosae, whereas no evidence of inflammation was detected within infected intestinal mucosa.