Figure 4
Induction and role of interleukin (IL)-10 and type 1 regulatory T (Tr1) cells in mice during vaginal gonococcal infection. (a) IL-10 production by iliac lymph node (ILN) cells isolated from Neisseria gonorrhoeae (Ngo)-infected or sham-infected mice 5 days after inoculation and analyzed by flow cytometry; n=8 mice per group. (b) Intracellular staining for IL-4 in CD4+/IL-10+ T cells isolated from draining lymph nodes of N. gonorrhoeae–infected mice at day 5. The horizontal line shows the gate of IL-4 positivity set by reference to the isotype control. (c) Intracellular staining for forkhead box p3 (Foxp3) in CD4+/IL-4–/IL-10+ or CD4+/IL-4–/IL-10– T cells isolated from ILN of N. gonorrhoeae–infected mice at day 5. (d) Time course of N. gonorrhoeae infection in anti-IL-10 antibody-treated, anti-IL-10 plus anti-TGF-β (anti-transforming growth factor-β) antibody-treated, and control immunoglobulin G (IgG)-treated mice; n=8 mice per group. (e) Cytokine expression in isolated ILN cells from sham-infected or infected mice treated with anti-IL-10 antibody, anti-IL-10 plus anti-TGF-β antibody, or control IgG; n=8 mice per group. Expression of interferon (IFN)-γ, IL-4, and IL-17 in CD4+ T cells at day 5 was analyzed by flow cytometry. (f) Reverse transcriptase–PCR (RT-PCR) analysis of IFN-γ, IL-4, and IL-17 mRNA levels in vaginal tissue harvested at day 3 from sham-infected or from infected mice treated with anti-IL-10 antibody, anti-IL-10 plus anti-TGF-β antibody, or control IgG; n=7 mice per group. Cytokine gene expression levels detected by RT-PCR were normalized relative to expression of β-actin and set at 1.0 for unstimulated cells. (g) Phenotypic profile of vaginal cells isolated on day 5 from sham-infected or infected mice treated with anti-IL-10 antibody or control IgG; n=7 mice per group. (h) Time course of N. gonorrhoeae infection in wild-type vs. IL-10-knockout (IL-10KO) C57BL/6 mice; n=8 mice per group. (i) Time course of N. gonorrhoeae infection mice receiving adoptively transferred IL-10-induced Tr1 cells, gonococcus-induced Tr1 cells, or control CD4+ T cells; n=8 mice per group. Results from one representative out of three independent experiments are shown. #P<0.05; *P<0.01 N. gonorrhoeae- vs. sham-infected groups, anti-IL-10 vs. control IgG treatments, or anti-IL-10 plus anti-TGF-β vs. anti-IL-10 treatments.
