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Problem: Successful immunocontraception using sperm antigens is dependent on achieving sufficient sperm-specific antibody in the reproductive ducts to prevent fertilization. The blood: luminal barrier of the male and female reproductive ducts must be overcome for this to occur. We have, therefore, investigated the relative titers of antigen-specific immunoglobulin G (IgG) in luminal fluids collected from male and female rabbit reproductive ducts following immunization with recombinant rabbit PH-20 (rPH-20). Method of study: Male and female rabbits were immunized subcutaneously with rPH-20 in Freund's adjuvant. Reproductive tract fluids and plasma were collected and assayed for specific IgG by enzyme-linked immunoabsorbent assay (ELISA). Plasma anti-rPH-20 antibodies were tested for their ability to inhibit in vitro fertilization. Results: Plasma rPH-20-specific IgG titers of >21 × 106 were induced in bucks. Antibody levels in the rete testis and cauda epididymidis fluids were only 0.026 and 0.168% of plasma levels, respectively. Plasma IgG titers were > 30 × 106 in does, but antibody levels in free flow vaginal fluid, free flow uterine fluid and free flow oviduct fluid were only 0.016, 0.078 and 0.072% of plasma levels, respectively. Induction of ovulation by administration of human chorionic gonadotrophin (hCG) significantly increased rPH-20-specific IgG only in free flow vaginal fluids. Plasma antibody from immunized rabbits inhibited in vitro fertilization but conception rates following mating of rPH-20 immunized male to untreated female rabbits were not affected. Conclusions: The IgG antibody entry into the reproductive ducts of both male and female rabbits is restricted to less than 0.2% of levels induced in plasma following subcutaneous immunization. This finding raises doubts about the suitability of rPH-20 to induce immunocontraception in rabbits using strategies

Female BALB/c mice were immunized with tetanus toxoid (TT) admixed with cholera toxin by direct application to shaved skin (Transcutaneous immunization, TCI). Tetanus toxoid-specific IgG and IgA in serum, saliva, vaginal lavage and fecal pellets were assayed by ELISA. Tetanus toxoid specific antibody-secreting cell (ASC) numbers were also determined by immunohistochemistry in sections of vagina, uterus, salivary gland and small intestine of immunized mice. TCI elicited significant levels of TT-specific IgG in serum, saliva and vaginal lavage, with the greatest increases over background seen in saliva (80-400 fold) and vaginal lavage (2-87 fold). TCI induced only modest levels of IgA in any of the samples tested (range 2-7 fold increase). In the absence of cholera toxin, application of TT alone did not result in detectable TT-specific antibodies in mucosal secretions. ASCs were found in all tissues following TCI. Cells were most frequent in uterus and vaginal tissues with ASC numbers less frequent in small intestine and salivary gland. This suggests that local production, rather than transudation from serum, is a major contributor of antibody in reproductive tract secretions. Further studies focussed on the role of sex hormones and immune induction following TCI. Animals immunized at the stage of oestrus cycle at which estrogen is abundant (Estrus), showed significantly lower levels of TT-specific IgG in vaginal lavage samples. Collectively, these data confirm the findings of Glenn and colleagues (1998), who showed TCI using cholera toxin can elicit high levels of serum IgG to both the toxin and co-administered antigen and further demonstrates that this route of immunization is particularly effective at eliciting humoral immunity in saliva and in the female reproductive tract. (C) 2000 Elsevier Science Ltd.

A mouse model of Helicobacter pylori infection was used to evaluate the vaccine antigen potential of the citrate synthase homologue protein purified from the H. pylori NCTC 11637 strain. Mice were immunised with the protein by intra-Peyer's patch immunisation. This route gives maximal intestinal immunisation and was used to screen oral vaccine candidate antigens without the added complication of simultaneously testing oral delivery systems. Two weeks post-immunisation mice were infected with Sydney strain H. pylori and 4 weeks after infection the mice were killed and the level of H. pylori infection in the stomach determined. Pre-immunisation with the 50/52-kDa protein led to a 84-91% reduction in H. pylori infection compared to unimmunised controls. Copyright (C) 1999 Federation of European Microbiological Societies.

The role of the host immune system in combating candidal infections in the vagina is poorly understood. A murine model of Candida vaginitis was used to elucidate the role of T cells in a candidal infection. Athymic BALB/c nu/nu mice or normal BALB/c mice were induced into estrus and then infected with 1 x 106 Candida albicans intravaginally. The infection was monitored over 1 week. Samples from blood, small intestine, tongue, kidney, spleen, liver, uterus and vagina were tested for recoverable C. albicans. Histology of the vagina was assessed for both inflammation and extent of infection. Results indicated that the BALB/c nu/nu mice had similar levels of vaginal yeast load to the normal BALB/c mice. In 25-30% of nude mice Candida was also recovered from extra vaginal sites (kidney, liver, small intestine), however, extra vaginal dissemination was not observed in any normal BALB/c animals. Histologically, both the nu/nu and control BALB/c had similar levels of vaginal inflammation; however, the nu/nu mice had more florid fungal growth in the vaginal epithelium. Adoptive transfer of either immune or non-immune BALB/c T cells into nude mice had no affect on either infection or vaginal inflammation. Immunohistochemical staining of vaginal tissues from normal BALB/c mice or nude mice adoptively transferred with either immune or non- immune T cells with anti-CD3 monoclonal antibody revealed no significant difference between groups in the numbers of CD3+ vaginal T cells. However, in mice receiving either immune or non-immune T cells no yeast was recovered from any tissues except the vagina. These data show that T cells have a limited role in protecting the vagina from C. albicans infection.

Peritoneal cavity cells were isolated from IL6-gene knockout (IL6(-/-)) and wild-type mice and stained for expression of IgM, CD5, and CD23. B1 cell (IgM+/CD23-, CD5+/IgM+) numbers were increased twofold in IL6(-/-) mice compared to normals while IgM+/CD23+ (B2) cell numbers were reduced threefold. Intestinal antibody levels were also determined for both total immunoglobulin and phosphorylcholine (PC)-specific and LPS-specific antibody following oral challenge with attenuated Salmonella typhimurium. Total immunoglobulin levels (IgM, IgG, and IgA) were reduced 60-80% in intestinal secretions of IL6(-/-) mice compared to wild-type controls; however, PC- specific antibody was significantly higher in IL6(-/-) mice. Anti-LPS antibodies were also three- to sevenfold higher in IL6(-/-) mice compared to controls following Salmonella challenge. These data suggest that in IL6(-/-) mice the development of mucosal B2 cells is impaired but that intestinal B1 cells responding to microbial antigens such as PC and LPS develop normally and are fully functional. (C) 1999 Academic Press.

We have used a mouse model of vaginal candidiasis to determine the effect of neutrophil depletion on (a) the clearance of Candida albicans and (b) the degree of inflammation associated with infection. No differences in recoverable yeast number or rate of clearance were observed between normal and neutrophil-depleted mice; however, vaginal inflammation was significantly decreased in neutrophil-depleted animals.

An increasing body of evidence suggests that certain T cell subsets mature extrathymically in the epithelium of the intestine. In the studies reported here, the authors have analysed cytokine/growth factor gene expression, recombinase-activating gene RAG-1 and RAG-2 gene expression and terminal deoxynucleotidyl transferase (TdT) gene expression in mouse fetal intestine and fetal thymus and liver, two known haematopoietic tissues. Stem cell factor (SCF) and interleukin 7 (IL-7) message was abundant in all three tissues during fetal development. IL-2 and IL-4 were not expressed in fetal gut but IL-4 was weakly detected in fetal liver and thymus. IL-9 and IL-13 mRNA was detected in all fetal tissues and IL-15 mRNA was abundant in fetal intestine but only weakly expressed in fetal liver and thymus. mRNA for SCF, IL-7, IL-13 and IL-15 was also detected in fibroblast-like cell lines derived from fetal intestine. RAG-1 and RAG-2 mRNA was detected in all three fetal tissues. TdT mRNA was not detected in fetal gut or liver but was weakly expressed in (fetal day) fd19-20 fetal thymus. Long-term (> 6 weeks) in vitro growth of IEL was achieved by coculturing intraepithelial lymphocytes (IEL) with IL-7-secreting fibroblasts in the presence of SCF and IL-2. The data show that the fetal mouse gut provides a suitable environment for lymphocyte development and receptor rearrangement, similar to fetal thymus and liver, even though expansion of intestinal IEL is delayed until 2-3 weeks after birth.