Antibodies and antibody-secreting cells in the female genital tract after vaginal or intranasal immunization with cholera toxin B subunit or conjugates

Abstract

We studied the antibody response including antibody-secreting cells (ASC) in the female genital tract of mice after mucosal immunizations with the recombinant B subunit of cholera toxin (rCTB) perorally, intraperitoneally, vaginally, and intranasally (i.n.). The strongest genital antibody responses as measured with a novel perfusion-extraction method were induced after vaginal and i.n. immunizations, and these routes also gave rise to specific immunoglobulin A (IgA) and IgG ASC in the genital mucosa. Specific ASC in the iliac lymph nodes, which drain the female genital tract, were seen only after vaginal immunization. Progesterone treatment increased the ASC response in the genital tissue after all mucosal immunizations but most markedly after vaginal immunization. We also tested rCTB as a carrier for human gamma globulin (HGG) and the effect of adding cholera toxin (CT) as an adjuvant for the induction of systemic and genital antibody responses to HGG after vaginal and i.n. immunizations. Vaginal immunizations with HGG conjugated to rCTB resulted in high levels of genital anti-HGG antibodies whether or not CT was added, while after i.n. immunization the strongest antibody response was seen with the conjugate together with CT. In summary, vaginal and i.n. immunization give rise to a specific mucosal immune response including ASC in the genital tissue, and vaginal immunization also elicits ASC in the iliac lymph nodes. We have also shown that rCTB can act as an efficient carrier for a conjugated antigen for induction of a specific antibody response in the genital tract of mice after vaginal or i.n. immunization.

FIG. 1

CTB-specific IgA and IgG titers in the genital mucosa after three p.o., i.p., i.n., or vaginal (VAG) immunizations with rCTB admixed with a small amount of CT. Antibody titers are given as log₁₀ of the GM of titers ± SEM. Each group contain 8 to 12 mice. White bars, titers in the fallopian tubes; striped bars, titers in the uterus; black bars, titers in the vagina. The CTB and CT doses used were 135 μg of CTB plus 5 μg of CT (p.o.), 20 μg of CTB plus 1.25 μg of CT (i.p.), and 85 μg of CTB plus 5 or 2.5 μg of CT (vaginal or i.n., respectively). Student’s t test corrected for multiple comparisons shows that vaginal IgA titers after i.n. or vaginal immunizations were significantly (P < 0.01) higher than after p.o. or i.p. immunizations.

FIG. 2

CTB-specific IgA and IgG ASC in the vagina and uterus and ILN after p.o. or vaginal immunization with 20 μg of CTB plus 5 μg of CT or i.n. immunization with 20 μg of CTB plus 2.5 μg of CT. Each group consisted of a pool of three mice, and values are given as the GM of specific ASC/10⁶ MNC from two independent experiments.

FIG. 3

HGG-specific IgA and IgG titers in serum after i.n. or vaginal immunizations with 12 μg of HGG ± 3 μg of CTB ± 2 μg of CT. Antibody titers are given as log₁₀ of the GM of titers ± SEM. Each group contained four mice. Black bars, i.n. immunization; white bars, vaginal immunization.

FIG. 4

HGG-specific IgA and IgG titers in vagina and uterus after i.n. or vaginal immunizations with 12 μg of HGG ± 3 μg of CTB ± 2 μg of CT. Antibody titers are given as log₁₀ of the GM of titers ± SEM. Each group contained four mice. Black bars, i.n. immunization; white bars, vaginal immunization.