Adjuvanticity of β -Glucan for Vaccine Against Trichinella spiralis

Abstract

In the past 30 years, few researches focus on the efficacy of adjuvant against Trichinella spiralis infection. Identifying new, improved vaccine adjuvants for T. spiralis infection are required. β-glucan are effective and safe as adjuvant for infectious diseases. In this paper, we first observed the adjuvanticity of β-glucan as adjuvant for defensing helminth T. spiralis in vivo. We showed that IgG and IgE were elevated in the mice immunized with β-glucan combined with recombinant T. spiralis serine protease inhibitor (rTs-Serpin), which is one of the vaccine candidates. Furthermore, in vitro, the combination of β-glucan and rTs-Serpin enhanced the maturation of bone marrow dendritic cells (BMDCs) compared to rTs-Serpin alone. We showed that β-glucan + rTs-Serpin -treated BMDCs secreted higher production of IL-12 and IL-10. Moreover, β-glucan + rTs-Serpin -treated BMDCs not only promoted the population of CD4⁺ IFN-γ⁺ T cells, but also enhanced the population of CD4⁺ IL-4⁺ T cells. These findings suggested that β-glucan, as an adjuvant, have the capacity to protect against T. spiralis infection via activating both Th1 and Th2 immune response.

Keywords: Th1/Th2 response; Trichinella Spiralis; adjuvant; dendritic cells; β-glucan.

FIGURE 1

Helminth burden in the immunized mice. (A) The number of adults recovered from intestines from immunized mice after challenge with 500 ML of T. spiralis. (B) The reduction rates of adult worms were analyzed based on the mean number of adult worms. (C) The number of muscle larvae (ML) per gram (LPG) in skeletal muscles from immunized mice after challenge with 500 ML of T. spiralis. (D) The reduction rates muscle larvae were analyzed based on the mean number of recovered muscle larvae per gram (LPG) of muscle from vaccinated groups compared with PBS group. Results are expressed as the mean ± SD of 10 mice per group. The data shown are representative of three independent experiments. *P < 0.05 as indicated by the line (Tukey multiple comparison following ANOVA).

FIGURE 2

Analysis of humoral immune responses. (A) The levels of IgG in the serum were measured by ELISA. (B) The levels of IgE in the serum were measured by ELISA. (C) The levels of IgG1 in the serum were measured by ELISA at different time points. (D) The levels of IgG2a in the serum were measured by ELISA at different time points. The values shown for each group are the mean + SD of the antibody levels (n = 10) from three individual experiments *P < 0.05 as indicated by the line (one-way ANOVA with Tukey’s post-test).

FIGURE 3

Analysis of cytokine production from CD4⁺ T cells. (A) The level of IFN-γ was measured by ELISA one week after the final immunization. (B) The level of IL-4 was measured by ELISA one week after the final immunization. The data are the mean ± SD of each group (n = 10) from three independent experiments. *P < 0.05 as indicated by the lines.

FIGURE 4

DC phenotype induced by β-glucan + rTs-Serpin. Immature DCs were treated with rTs-Serpin (10 μg/mL) or combination of rTs-Serpin and β-glucan (50 μg/mL) in vitro for 24 h. (A) (I) gating on viable cells and (II) gating on CD11c⁺ cells. (B) Expression of CD11c⁺ CD86⁺ MHC-II⁺ cells were measured. (C) Cytokines (IL-12p70 and IL-10) levels in the supernatant were quantified by ELISA Data represent mean ± SD deviations (n = 3) of the results from three individual experiments *P < 0.05, **P < 0.01, and ***P < 0.001 vs. the control groups.

FIGURE 5

CD4 + T cells response induced by β-glucan + rTs-Serpin –treated DCs. The purity of CD4⁺ T cells were analysis by FACS after magnetic sorting using anti-CD4 magnetic beads. The purified CD4⁺ T cells had > 90% purity. DCs (1 × 10⁵/well) and CD4⁺ T cells (1 × 10⁶/well) were cocultured for 72 h with OVA (1 mg/mL), then cells were incubated with 10 mg/mL Brefeldin A, 50 ng/mL phorbol 12-myristate 13-acetate (PMA) and 750 ng/mL Ionomycin for 6 h at 37°C. (A) (I) gating on viable cells and (II) gating on CD4⁺cells. (B) Percentage of CD4⁺ IFN-γ⁺ and CD4⁺ IL-4⁺ T cells were determined by flow cytometry. (C) CD4 + T cells were stained with CFSE before co-culture with DCs. The proliferation of CD4 + T cells were determined by flow cytometry. Results are shown as mean ± SD (n = 3) of three different experiments. *P < 0.05, **P < 0.01, ***P < 0.001 as indicated by line (one-way ANOVA with Tukey’s post test).