Conjugation of protein antigen to microparticulate beta-glucan from Saccharomyces cerevisiae: a new adjuvant for intradermal and oral immunizations.
ABSTRACT Immunostimulatory glucose polymers known as beta-glucans have been studied for many years. Our laboratory has prepared and characterized a novel microparticulate beta-glucan (MG) from the budding yeast Saccharomyces cerevisiae. Because MG particles are rapidly phagocytized by murine peritoneal macrophages and induce the expression of B7 costimulatory molecules, we hypothesized that MG could serve as a vaccine adjuvant to enhance specific immune responses. Here, we describe a procedure for conjugating the test vaccine antigen bovine serum albumin (BSA) to MG via water-soluble carbodiimide linkage. Conjugates with up to 0.4 mg of BSA/mg MG were prepared. MG/BSA conjugates were still actively phagocytized by mouse peritoneal macrophages. When used to immunize mice by the intradermal route, these conjugates enhanced the primary IgG antibody response to BSA in a manner comparable to the prototypic complete Freund's adjuvant. Although primary oral immunization with MG/BSA caused no increase in serum anti-BSA antibody titers, booster immunization elicited a significant anti-BSA antibody response. These results suggest that protein antigens can be conjugated to MG via a carbodiimide linkage and that these conjugates provide an adjuvant effect for stimulating the antibody response to the protein antigens.
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ABSTRACT: Trypanosoma cruzi, the causative agent of Chagas' disease, infects humans and animals in tropical, subtropical and some temperature regions of the western hemisphere. At present, there is no effective vaccine for T. cruzi infection. Glucan, a beta-1,3 polyglucose biological response modifier, possesses significant adjuvant activity. The present study investigated the adjuvant activity of particulate glucan when combined with a vaccine of glutaraldehyde-killed T. cruzi culture forms. ICR/HSD mice (20 g) were injected s.c. with glutaraldehyde-killed T. cruzi on days 21, 14 and 7 prior to challenge with 50 T. cruzi blood forms. Particulate glucan (1 mg/mouse) was administered s.c. either alone or in conjunction with T. cruzi vaccine. Isovolumetric dextrose served as control. Dextrose, glucan or T. cruzi vaccine as single treatment regimens showed 100% mortality with 20.5, 21.4 and 21.6 day median survival times, respectively. In contrast, glucan administered with T. cruzi vaccine showed an 85% (P less than 0.01) survival at 275 days post-challenge. In addition, the number of T. cruzi observed in the blood of glucan--T. cruzi immunized mice was lower than the appropriate controls. However, immunized mice which survived at 275 days were positive for the presence of T. cruzi by xenodiagnosis. Histopathologic evaluation of glucan--T. cruzi mice revealed no parasites or cardiac pathology, but a mild splenic hyperplasia and inflammation of skeletal muscle were noted. In subsequent studies, mice were immunized with the same regimen of glucan--T. cruzi and challenged with 500 or 5000 T. cruzi. Glucan significantly (P less than 0.05) increased survival as denoted by 60% and 50% survival in the glucan-T. cruzi group vs 0% in controls.(ABSTRACT TRUNCATED AT 250 WORDS)International Journal of Immunopharmacology 02/1989; 11(4):403-10.
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ABSTRACT: Streptococcus group B (GBS) is usually carried asymptomatically in the vaginal tract of women and can be transferred to the newborn during parturition. Serum antibodies to the capsular polysaccharide (CPS) can prevent invasive diseases, whereas immunity acting at the mucosal surface may be more important to inhibit the mucosal colonization of GBS and thus the risk of infection for the newborn. We prepared different GBS type III CPS-protein conjugate vaccines and evaluated their systemic and mucosal immunogenicity in mice. GBS type III CPS was conjugated to tetanus toxoid (TT) or recombinant cholera toxin B subunit (rCTB) either directly or to rCTB indirectly via TT. The conjugation was performed by different methods: (1) CPS was coupled to TT with 1-ethyl-3 (3-dimethylaminopropyl)-carbodiimide (EDAC), using adipic acid dihydrazide (ADH) as a spacer; (2) CPS was conjugated with rCTB using reductive amination; or, (3) N-succinimidyl 3-(2-pyridyldithio) propionate (SPDP) was used to bind rCTB to the TT of the CPS-TT conjugate. Mice were immunized with these conjugates or purified CPS by subcutaneous (s.c.) and intranasal (i.n.) routes. Antibodies to GBS III in serum, lungs and vagina were measured with ELISA. All of the CPS–protein conjugates were superior to unconjugated CPS in eliciting CPS-specific immune responses in serum and mucosal tissue extracts. The conjugates, when administrated s.c., induced only IgG responses in serum, lung and vagina, while i.n. vaccination also elicited IgA responses in the lungs and vagina. The CPS–TT conjugate administrated i.n. induced a strong serum IgG, but only a weak mucosal IgA response, while the CPS–rCTB conjugate elicited high IgG as well as IgA antibodies in the lungs after i.n. immunization. GBS III CPS–TT conjugated with rCTB produced a strong systemic and local anti-CPSIII response after i.n. administration. Co-administration of CT as adjuvant enhanced the anti-CPS systemic and mucosal immune responses further after i.n. administration with the CPS conjugates. These findings indicate that: (i) i.n. immunization with GBS CPS–protein conjugates was more effective than s.c immunization for stimulating serum as well as mucosal immune responses; (ii) rCTB as a carrier protein for GBS III CPS could markedly improve the mucosal immune response; and (iii) the experimental GBS type III CPS conjugates containing rCTB should be investigated as mucosal vaccine to prevent GBS infection in humans.Vaccine 12/2000; · 3.49 Impact Factor
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ABSTRACT: Beta-1,3-(D)-glucan from a variety of biological sources has been shown to enhance both humoral and cellular immune responses to a variety of antigens, infectious agents, and tumors. Nevertheless, its mode of action has not been fully defined. We sought to determine whether a 1-2 microm diameter microparticulate form of beta-glucan (MG) from the yeast Saccharomyces cerevisiae could regulate expression of B7 family glycoproteins on resident peritoneal macrophages from BALB/c mice. We discovered that MG uregulated B7.2 mRNA expression and enhanced the surface membrane expression of B7.2 glycoprotein. Although B7.1 mRNA was not upregulated above constitutive levels, MG treatment enhanced B7.1 glycoprotein expression on the macrophages, albeit to a lesser extent than B7.2. At the same time, the gene and cell surface expression of B7-H1, a putative negative regulator of T cell activity, was also upregulated by MG. The expression of B7-DC, another B7 family molecule with negative regulatory activity, was not affected by incubation with MG. This study has demonstrated that a microparticulate form of beta-glucan can enhance B7 co-stimulatory molecule expression on macrophages, thereby enabling these antigen-presenting cells to deliver the second signal to T-lymphocytes that express CD28. In addition, because MG also induces the expression of B7-H1, it may enable macrophages to provide a concomitant downregulatory signal to T-lymphocytes expressing PD-1 or related receptors.Immunology Letters 05/2004; 93(1):71-8. · 2.34 Impact Factor