Two different homing pathways involving integrin β7 and E-selectin significantly influence trafficking of CD4 cells to the genital tract following Chlamydia muridarum infection

Abstract

Problem: Chlamydia trachomatis causes STI and reproductive dysfunction worldwide which is not preventable with antibiotics. Identifying a population of endocervical T cells to target in vaccine development would enhance efficacy.

Method of study: Trafficking of murine CD4+ lymphocytes to Chlamydia muridarum infected genital tract (GT) tissue in vivo was measured using adoptive transfer studies of fluorescent CD4+ T cells from integrin β7-/- mice or mice which lack E-selectin on endothelial cells.

Results: Murine in vivo migration studies showed that lack of α4β7 or E-selectin significantly reduced trafficking of CD4 T cells to the GT of mice infected with C. muridarum.

Conclusion: CD4+ T cells use at least two different adhesive mechanisms involving an integrin of the mucosal homing pathway and selectin pathway to accumulate in the GT during C. muridarum infection.

Figure 1

Beta 7 integrin, contributes to lymphocyte migration to the murine genital tract. A) CD4 cells purified from beta 7 integrin KO and WT C57BL/6 mice 7 days after infection with C. muridarum. Cells were label with the red fluorescent dye, PKH-26. To control for individual variability in the migration assay, CD4 cells were also purified from WT mice and labeled with the green fluorescent dye, BODIPY-green. The labeled red (beta 7+ or beta 7−/−) and green cells were mixed in equal proportions and adoptively transferred into 7 day post-C. muridarum infected WT C57BL/6 mice. Cells were harvested 18 hours later. n = 3 mice per group, *p<0.001 by t-test. B) Shedding of C. muridarum was determined every three days after vaginal inoculation with 1×10⁵ IFU. Data points denote means from 5 mice. There were no significant differences between groups. Representative of two experiments with similar results.

Figure 2

The absence of E-selectin on endothelial cells influences the trafficking of CD4 cells to the GT. A) CD4 Th1 clone specific for C. muridarum was labeled with PKH-26 and adoptively transferred into BALB/c mice infected 7 days previously with C. muridarum. The cells were harvested 18 hours later. n = 4 mice per group, *p=0.014 by ANOVA. B) Shedding of C. muridarum was determined every three days after vaginal inoculation with 1×10⁷ IFU. Data points represent the mean from 5 mice. There were no significant differences between groups. Representative of two experiments with similar results.

Figure 3

E-selectin is present in the upper GT. GT tissues were harvested 7 days following vaginal infection as described and homogenates were prepared of the upper GT (oviducts, UGT), middle GT (uterine horns, MGT) and lower GT (cervico-vaginal region, LGT) from two representative mice (1 & 2). Homogenates of the spleen from a TNFα injected mouse served as positive control. PCR analysis was also performed on GT tissue using primers against the housekeeping gene, cyclophilin.