Vaginal Immunization Research Articles

Slight influence of the estrous cycle stage on the mucosal and systemic specific antibody response induced after vaginal and intraperitoneal immunization with protoxin Cry1Ac from Bacillus thuringiensis in mice

Since the immune response appears to be variable according to the hormonal stage of the mammalian female, the aim of this study was to determine whether estrous cycle stage modifies the mucosal and systemic immune responses induced by intraperitoneal and vaginal immunization of mice with protoxin Cry1Ac. We tested the influence of three immunizations on the specific antibody response elicited at estrus and diestrus, that were the same estrous cycle stages at which the antigen was applied. Both intraperitoneal and vaginal immunization of mice with Cry1Ac either at estrus or diestrus induces specific antibody responses at serum, vagina and large intestine. The stage of the estrous cycle have little or non influence in the magnitude of the response induced, since only at serum the IgM was slightly higher at estrus than at diestrus by both routes. At the large intestine only the IgA response elicited via the intraperitoneal route changed, being higher at diestrus, whereas at the vagina IgA response induced did not change significantly due to the cycle stage. Present results suggest that Cry1Ac may be used as an antigen carrier as it can elicit antibody responses at systemic level and at several mucosal sites including the vagina that are not modified significantly throughout the reproductive cycle.

HIV disease: Working Group Report of the First World Congress of Pediatric Gastroenterology, Hepatology, and Nutrition.

HIV disease: Working Group Report of the First World Congress of Pediatric Gastroenterology, Hepatology, and Nutrition.

Systemic and mucosal immune responses in mice after rectal and vaginal immunization with HIV-DNA vaccine.

We examined the feasibility of inducing local and systemic human immunodeficiency virus (HIV)-specific immune responses by rectal and vaginal application of an HIV-DNA vaccine. Mice were immunized with an HIV-DNA vaccine preparation via a rectal or vaginal route. After several applications, HIV-specific antibodies were detected in sera, fecal extract solutions, and vaginal washes, and these antibodies were potent in inhibiting the syncytium formation of a CD4-positive human T cell line by a cell line capable of inducing HIV-1 infection. Spleen cells from rectally and vaginally immunized mice showed antigen-mediated IFN-gamma-inducing activity. In addition, with rectal immunization, mononuclear cells from both the spleen and the regional lymph nodes of the rectal region were found to be potent at inducing a cytotoxic T lymphocyte response. These humoral and cell-mediated immune responses were enhanced by augmenting the vaccine with granulocyte-macrophage colony-stimulating factor-expressing plasmids or IL-12-expressing plasmid. Our results demonstrated that both rectal and vaginal immunization could induce systemic and mucosal immunity and that these responses were enhanced by the addition of the above cytokine-expressing plasmids.

FEMALE SEX HORMONES AS REGULATORY FACTORS IN THE VAGINAL IMMUNE COMPARTMENT

Sexually transmitted diseases (STD) are now considered to be among the most common human infections. The incidence of STD is on the rise, which is partly due to frequent transmission during the asymptomatic phase of infection. The compounded cost of STD just in the United States is estimated to exceed $10 billion annually. STD are particularly prevalent in teenagers and young adults and the health problems caused by these diseases tend to be more severe and more frequent in woman than in men. Despite considerable efforts, a vaccine that provides protective immunity against sexually transmitted diseases in humans has not been developed. Nonetheless, research in animal models indicates that strong local and regional immune responses can influence the outcome of vaginal challenge with microbial pathogens. Vaginal immunity is an area of basic immunology that has received relatively little attention, but it is already clear that the mucosal and regional immunology of the vagina has unique features. The present review summarizes some of the anatomical, physiological and immunological features of the vagina and uterus that distinguish humans, non-human primates, rats and mice. These interspecies differences need to be taken into account in laboratory efforts to develop effective vaccines for STD in humans.

Human immunodeficiency virus type 1 Gag-specific vaginal immunity and protection after local immunizations with sindbis virus-based replicon particles.

The majority of human immunodeficiency virus (HIV) infections occur through vaginal and rectal transmission. In seeking a safe, nonreplicating gene-delivery vector that can induce mucosal and systemic immune responses and protection, Sindbis virus-based replicon particles expressing HIV-1 Gag (SIN-Gag) were developed. In mice, after nasal or intramuscular immunization with SIN-Gag and vaginal challenge with vaccinia virus (VV) expressing HIV-1 Gag (VV-Gag), CD8(+) T cell-mediated responses were detected locally, in the vaginal mucosa and in the draining iliac lymph nodes (ILNs), and systemically, in the spleen. However, the mice were not protected against VV-Gag replication in the ovaries. In contrast, after vaginal or rectal immunization with SIN-Gag and vaginal challenge with VV-Gag, despite lower local CD8(+) T cell-mediated responses in the vaginal mucosa and ILNs, the mice were protected against VV-Gag replication in the ovaries. Therefore, local immunization with SIN-Gag induced both local mucosal cell-mediated responses and protection.

Analysis of the CD4 protein on human vaginal T lymphocytes.

Although T lymphocytes at the human vaginal mucosa have been partially characterized, there remains a paucity of information regarding cell-mediated immune mechanisms at this mucosal site. In mice and humans, there are several phenotypic distinctions between vaginal T lymphocytes and those in the peripheral circulation. Recently, we observed as well that the N-terminus of the CD4 protein on murine vaginal T lymphocytes is atypically expressed compared to its systemic counterpart, and that the atypical expression extends to the mRNA level. The purpose of this study was to evaluate the CD4 protein on human vaginal T lymphocytes by flow cytometry and RT-PCR. Results showed that, in contrast to mice, the CD4 protein on human vaginal and peripheral blood T lymphocytes are similar at both the molecular and protein levels. These results indicate that based on several differences between human and mouse vaginal T cells, caution is urged when using mice as a model to study human vaginal immunity.

Polymerase chain reaction for Y chromosome to detect semen in cervicovaginal fluid: a prerequisite to assess HIV-specific vaginal immunity and HIV genital shedding.

Polymerase chain reaction for Y chromosome to detect semen in cervicovaginal fluid: a prerequisite to assess HIV-specific vaginal immunity and HIV genital shedding.

Immunity to vaginal herpes simplex virus-2 infection in B-cell knockout mice.

We investigated the involvement of antibody in protection against vaginal herpes simplex virus type-2 (HSV-2) infection by comparing intact and B-cell knockout (KO) mice. Vaginal immunization of intact mice with attenuated HSV-2 markedly reduced an HSV-2 challenge infection in the vagina. In contrast, immunization of B-cell KO mice produced less immunity against the challenge infection and that immunity occurred in a different pattern. At 20 hr after challenge, immunostaining of virus proteins in the vaginal epithelium and shed virus protein titres in the vaginal secretions were not significantly different between immunized and non-immunized B-cell KO mice and were much greater than in immunized intact mice. At 48 hr after challenge, the vaginal infection in immunized B-cell KO mice was markedly less than at 20 hr but remained approximately sevenfold higher than in intact mice. This pattern of challenge infection in the vagina indicates that B cells, and probably the antibody derived from them, provided significant protection against reinfection in intact mice, especially during the first 20 hr after challenge, while other effector mechanisms became important between 20 and 48 hr after challenge. To determine whether T-cell immunity in immunized B-cell KO mice was equal to that in intact mice, we assessed interferon-gamma (IFN-gamma) secretion by memory T cells in vivo in the vagina at 20 hr after challenge. We found no significant differences in the up-regulation of major histocompatibility complex (MHC) class II antigens in the epithelium, up-regulation of vascular cell adhesion molecule-1 (VCAM-1) in vascular endothelium, or recruitment of T cells to the mucosa, indicating that the memory T-cell response to virus challenge was the same in intact and B-cell KO mice.

Induction of an HPV 6bL1-specific mucosal IgA response by DNA immunization

Human papillomavirus (HPV) plays a crucial role in the development of human anogenital dysplasia. To prevent infection, it is important to induce an HPV-specific mucosal immune response. We investigated whether DNA vaccination would induce an intravaginal mucosal antibody response against HPV 6bL1. New Zealand White rabbits were immunized with an HPV 6bL1 DNA vaccine by one of the three routes: muscular, vaginal, or rectal. We found that vaginal immunization of rabbits with HPV 6bL1 DNA induced 6bL1 virus-like particle-specific lgA antibodies in vaginal secretions. They were detectable until at least 14 weeks after the first immunization. The antibodies also showed neutralizing activity in a hemagglutination inhibition assay. No mucosal immune response was detected in vaginal secretions of rabbits immunized intramuscularly or intrarectally. Our data suggest that vaginal immunization with HPV 6bL1 DNA induces long-lasting IgA responses with neutralizing activity in vaginal secretions of rabbits.

Value of Candida polymerase chain reaction and vaginal cytokine analysis for the differential diagnosis of women with recurrent vulvovaginitis

Objectives Recurrent vulvovaginitis remains difficult to diagnose accurately and to treat. The present investigation evaluated the utility of testing vaginal specimens from women with symptomatic recurrent vulvovaginitis for Candida species by polymerase chain reaction (PCR) and for cytokine responses. Methods Sixty-one consecutive symptomatic women with pruritus, erythema, and/or a thick white discharge and a history of recurrent vulvovaginitis and 31 asymptomatic women with no such history were studied. Vaginal swabs were tested for Candida species by PCR, for the antiinflammatory cytokine interleukin (IL)-10, and for the proinflammatory cytokine IL-12. Results C. albicans was detected in 19 (31.1%) of the patients as well as in three (9.7%) controls (P = 0.03). Both IL-10 (31.1% vs. 0%) and IL-12 (42.6% vs. 6.5%) were also more prevalent in the recurrent vulvovaginitis patients (P < 0.001). However, there was no relation between the presence or absence of Candida and either cytokine. Detection of IL-12 in 14 women indicated the stimulation of a vaginal cell-mediated immune response possibly from an infectious agent. The presence of only IL-10 in six patients indicated a suppression of vaginal cell-mediated immunity and was consistent with a possible allergic etiology. The absence of both IL-10 and IL-12 in other patients, similar to that found in healthy controls, suggested a noninfectious, nonallergic etiology of their symptoms. Conclusion Many women with recurrent vulvovaginitis are not infected with Candida. Testing for Candida should be required in this population. Treatment with only anti-Candida medication will clearly be inadequate for the majority of women with this condition. Infect. Dis. Obstet. Gynecol. 8:244–247, 2000. © 2000 Wiley-Liss, Inc.

Value of Candida polymerase chain reaction and vaginal cytokine analysis for the differential diagnosis of women with recurrent vulvovaginitis.

Recurrent vulvovaginitis remains difficult to diagnose accurately and to treat. The present investigation evaluated the utility of testing vaginal specimens from women with symptomatic recurrent vulvovaginitis for Candida species by polymerase chain reaction (PCR) and for cytokine responses. Sixty-one consecutive symptomatic women with pruritus, erythema, and/or a thick white discharge and a history of recurrent vulvovaginitis and 31 asymptomatic women with no such history were studied. Vaginal swabs were tested for Candida species by PCR, for the antiinflammatory cytokine interleukin (IL)-10, and for the proinflammatory cytokine IL-12. C. albicans was detected in 19 (31.1%) of the patients as well as in three (9.7%) controls (P = 0.03). Both IL-10 (31.1% vs. 0%) and IL-12 (42.6% vs. 6.5%) were also more prevalent in the recurrent vulvovaginitis patients (P < 0.001). However, there was no relation between the presence or absence of Candida and either cytokine. Detection of IL-12 in 14 women indicated the stimulation of a vaginal cell-mediated immune response possibly from an infectious agent. The presence of only IL-10 in six patients indicated a suppression of vaginal cell-mediated immunity and was consistent with a possible allergic etiology. The absence of both IL-10 and IL-12 in other patients, similar to that found in healthy controls, suggested a noninfectious, nonallergic etiology of their symptoms. Many women with recurrent vulvovaginitis are not infected with Candida. Testing for Candida should be required in this population. Treatment with only anti-Candida medication will clearly be inadequate for the majority of women with this condition.

Value ofCandida polymerase chain reaction and vaginal cytokine analysis for the differential diagnosis of women with recurrent vulvovaginitis

Objectives Recurrent vulvovaginitis remains difficult to diagnose accurately and to treat. The present investigation evaluated the utility of testing vaginal specimens from women with symptomatic recurrent vulvovaginitis for Candida species by polymerase chain reaction (PCR) and for cytokine responses. Methods Sixty-one consecutive symptomatic women with pruritus, erythema, and/or a thick white discharge and a history of recurrent vulvovaginitis and 31 asymptomatic women with no such history were studied. Vaginal swabs were tested for Candida species by PCR, for the antiinflammatory cytokine interleukin (IL)-10, and for the proinflammatory cytokine IL-12. Results C. albicans was detected in 19 (31.1%) of the patients as well as in three (9.7%) controls (P = 0.03). Both IL-10 (31.1% vs. 0%) and IL-12 (42.6% vs. 6.5%) were also more prevalent in the recurrent vulvovaginitis patients (P < 0.001). However, there was no relation between the presence or absence of Candida and either cytokine. Detection of IL-12 in 14 women indicated the stimulation of a vaginal cell-mediated immune response possibly from an infectious agent. The presence of only IL-10 in six patients indicated a suppression of vaginal cell-mediated immunity and was consistent with a possible allergic etiology. The absence of both IL-10 and IL-12 in other patients, similar to that found in healthy controls, suggested a noninfectious, nonallergic etiology of their symptoms. Conclusion Many women with recurrent vulvovaginitis are not infected with Candida. Testing for Candida should be required in this population. Treatment with only anti-Candida medication will clearly be inadequate for the majority of women with this condition. Infect. Dis. Obstet. Gynecol. 8:244–247, 2000. © 2000 Wiley-Liss, Inc.

Rectal and vaginal immunization with a macromolecular multicomponent peptide vaccine candidate for HIV-1 infection induces HIV-specific protective immune responses

An effective vaccine for human immunodeficiency virus (HIV) is needed to stimulate the immune response of the genital mucus to prevent mucosal transmission of the virus. We have developed a macromolecular multicomponent peptide vaccine candidate, VC1. Both rectal and vaginal immunization of VC1 mixed with cholera toxin (CT) induced HIV-1-specific IgA antibody in mouse fecal extract solution and vaginal wash. These antibody productions were enhanced by the combination with IL-4 or GM-CSF expressing plasmids. Either fecal extract or vaginal wash solution from immunized mice inhibited production of HIV-1 IIIB p24 protein. The mononuclear cells from spleen, intestinal lymph nodes, or Peyer's patches from VC1-and CT-immunized mice released IFN-γ or IL-4, when these cells were co-cultured with VC1 antigen. In addition, the regional lymphoid cells from rectal and vaginal region of mice immunized with VC1 and CT also elicited a substantial level of HIV-1-specific cytotoxic T cell (CTL) response. This CTL response was enhanced by the addition of IL-12 expressing plasmid. Our results clearly demonstrated that both rectal and vaginal immunization could induce systemic and mucosal immunities specific for HIV-1.

The effect of route of immunization on mucosal immunity and protection.

In macaques, the route of immunization has a profound effect on the immune response. Augmenting rectal or vaginal immunization with oral or nasal immunization enhanced the secretory IgA, serum IgG, and T cell responses. However, targeted iliac lymph node (TILN) immunization with recombinant simian immunodeficiency virus (SIV) gp120 and p27 elicited the most consistent mucosal antibody responses in the rectum, vagina, urine, seminal fluid, and blood. Both mucosal and TILN immunization induced specific CD4+ T cell proliferative responses in the iliac lymph nodes, which drain these mucosal surfaces, and in the splenic and circulating T cells. Rectal mucosal challenge with cell-free SIV induced total protection in 4 of 7 macaques that were immunized by the TILN route, and, compared with unimmunized macaques or those immunized by the mucosal route (P<.001), it induced a >90% decrease in virus load in 3 of them. Protection from mucosal rectal infection with SIV was significantly associated with an increase in the CD8 suppressor factor (which was generated by the iliac lymph node), RANTES, and MIP-1beta (P<.01).

Association of human leucocyte antigen phenotype with vaccine efficacy in patients receiving vaginal mucosal immunization for recurrent urinary tract infection

Immune responses to specific antigens can be influenced by an individual's genetic make-up. We examined whether the efficacy of a vaginal mucosal vaccine for urinary tract infections (UTI) was affected by a patient's human leucocyte antigen (HLA) phenotype. Urinary tract infection data and the HLA phenotypes of 47 women participating in a phase II clinical trial of immunization for recurrent UTI were statistically analysed for associations between HLA-A, -B, -DR, or -DQ phenotype and postimmunization infection course. Women who received the vaccine and had HLA-DR phenotypes other than DR2 had significantly delayed times to re-infection compared with women receiving placebo. Vaccine-treated patients with the HLA-DR2 phenotype had re-infection courses that were not different than women receiving placebo. These results indicate that the efficacy of a vaginal mucosal UTI vaccine may be influenced by an individual's HLA-DR phenotype.

Local vaginal immunity in patients with uterine myoma before and after hysterectomy

Vaginal biopsy specimens from premenopausal women with uterine myoma were studied by immunohistochemical methods using monoclonal antibodies to the differentiation antigens before and after hysterectomy. Immunocompetent cells (mainly CD8+ antigens) were detected in vaginal mucosa of controls and were virtually absent from specimens from the patients with uterine myoma. The content of immunocompetent cells in the vaginal mucosa did not change for at least 12 months after hysterectomy irrespective of the scope of intervention.

Immunoglobulin G, plasma cells, and lymphocytes in the murine vagina after vaginal or parenteral immunization with attenuated herpes simplex virus type 2.

This investigation evaluated immunity to vaginal herpes simplex virus type 2 (HSV-2) infection after local or parenteral immunization with attenuated HSV-2. Vaginal immunization induced sterilizing immunity against challenge with a high dose of wild-type virus, whereas parenteral immunizations protected against neurologic disease but did not entirely prevent infection of the vagina. Vaginal immunization caused 86- and 31-fold increases in the numbers of immunoglobulin G (IgG) plasma cells in the vagina at 6 weeks and 10 months after immunization, whereas parenteral immunizations did not increase plasma cell numbers in the vagina. Vaginal secretion/serum titer ratios and specific antibody activities in vaginal secretions and serum indicated that IgG viral antibody was produced in the vagina and released into vaginal secretions at 6 weeks and 10 months after vaginal immunization but not after parenteral immunizations. In contrast to the case for plasma cells, the numbers of T and B lymphocytes in the vagina were similar in vaginally and parenterally immunized mice. Also, lymphocyte numbers in the vagina were markedly but similarly increased by vaginal challenge with HSV-2 in both vaginally and parenterally immunized mice. Lymphocyte recruitment to the vagina after virus challenge appeared to involve memory lymphocytes, because it was not observed in nonimmunized mice. Thus, local vaginal immunization with attenuated HSV-2 increased the number of IgG plasma cells in the vagina and increased vaginal secretion/serum titer ratios to 3.0- to 4.7-fold higher than in parenterally immunized groups but caused little if any selective homing of T and B lymphocytes to the vagina.

Vaginal immunization with recombinant gram-positive bacteria.

Many viral and bacterial pathogens enter the body through the genital mucosa. Therefore, one of the major goals of a vaccine against sexually transmitted diseases (STDs) should be to induce an immune response in the genital mucosa capable of controlling the entry of the pathogen. Our approach for the development of vaccines against STDs is based on the use of nonpathogenic Gram-positive bacteria as live vaccine vectors. Recombinant Gram-positive bacteria expressing vaccine antigens were constructed using genetic systems developed in our laboratory. Balb/c mice and Cynomolgus monkeys were inoculated by the vaginal route and vaginal samples were collected using absorbent wicks. Colonization was evaluated by the presence of recombinant bacteria in the vaginal samples. Local and systemic immune responses were studied. We have developed genetic systems for the expression of heterologous antigens on the surface of the human commensals Streptococcus gordonii and Lactobacillus spp. Both S. gordonii and L. casei stably colonized the murine vagina after a single inoculum. Vaginal colonization of mice with recombinant strains of S. gordonii, expressing human papillomavirus (HPV) and human immunodeficiency virus (HIV) antigens, induced antigen-specific vaginal immunoglobulin A (IgA) and serum IgG. Local and systemic immune responses also were detected in monkeys immunized intravaginally with recombinant S. gordonii. The results obtained indicated that the approach of using colonizing Gram-positive bacteria as live vectors has a great potential for the development of vaccines against STDs.

Vaginal immunization of Cynomolgus monkeys with Streptococcus gordonii expressing HIV-1 and HPV 16 antigens

Cynomolgus monkeys ( Macaca fascicularis) were immunized by intravaginal administration of live recombinant Streptococcus gordonii. The vaccine strains of S. gordonii expressed the V3 domain of the gp120 of human immunodeficiency virus type 1 (HIV-1), and the E7 protein of human papillomavirus type 16 (HPV 16). Multiple inocula of recombinant bacteria were used, since S. gordonii could persist for no longer than 3 days in the monkey vagina. Vaginal immunization was found to induce a local and systemic immune response specific for the heterologous antigen expressed by the bacteria. This antigen-specific immune response consisted of vaginal IgA, serum IgG, and a T-cell proliferative response measured on PBMCs. Vaginal IgG and serum IgA were not detected.

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

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.