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IL-33 released by alum is responsible for early cytokine production and has adjuvant properties.

Rose WA, Okragly AJ, Patel CN, Benschop RJ - Sci Rep (2015)

Bottom Line: Alum can induce the release of endogenous danger signals via cellular necrosis which elicits inflammation-associated cytokines resulting in humoral immunity.Furthermore, IL-33 itself functions as an adjuvant that, while only inducing a marginal primary response, facilitates a robust secondary response comparable to that observed with alum.Our results provide novel insights into the mechanism of action behind alum-induced cytokine responses and show that IL-33 is sufficient to provide a robust secondary antibody response independently of alum.

View Article: PubMed Central - PubMed

Affiliation: Biotechnology Discovery Research, Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, IN, 46285.

ABSTRACT
Human vaccines have used aluminium-based adjuvants (alum) for >80 years despite incomplete understanding of how alum enhances the immune response. Alum can induce the release of endogenous danger signals via cellular necrosis which elicits inflammation-associated cytokines resulting in humoral immunity. IL-33 is proposed to be one such danger signal that is released from necrotic cells. Therefore, we investigated whether there is a role for IL-33 in the adjuvant activity of alum. We show that alum-induced cellular necrosis results in elevated levels of IL-33 following injection in vivo. Alum and IL-33 induce similar increases in IL-5, KC, MCP-1, MIP-1α and MIP-1β; many of which are dependent on IL-33 as shown in IL-33 knockout mice or by using an IL-33-neutralizing recombinant ST2 receptor. Furthermore, IL-33 itself functions as an adjuvant that, while only inducing a marginal primary response, facilitates a robust secondary response comparable to that observed with alum. However, IL-33 is not absolutely required for alum-induced antibody responses since alum mediates similar humoral responses in IL-33 knockout and wild-type mice. Our results provide novel insights into the mechanism of action behind alum-induced cytokine responses and show that IL-33 is sufficient to provide a robust secondary antibody response independently of alum.

No MeSH data available.


Related in: MedlinePlus

IL-33 injection produces antigen specific antibody responses.(a) C57BL/6 mice (n = 5 mice/group) were pre-treated with mouse IgG1 isotype control antibody (Ctrl) or ST2-Fc injected i.p. one hour prior to i.p. injection of PBS/NP-CGG or IL-33/NP-CGG then boosted i.p. on day 14 with PBS/NP-CGG. (b) BALB/c mice (n = 5 mice/group) were injected i.p. with PBS/NP-CGG or IL-33/NP-CGG then boosted i.p. on day 14 with PBS/NP-CGG. (c) Mice were treated as in (b) except PE was used as the antigen. (d) Mice were treated as in (b) except OVA was used as the antigen. Antigen specific IgM, IgG1 and IgE serum titers were quantified via ELISA. Data in (a) are representative of four independent experiments and data in (b–d) are from one experiment. ***p < 0.001 PBS compared to Ctrl/IL-33 groups (Two-way ANOVA with Bonferroni multiple comparison test). ###p < 0.001 Ctrl/IL-33 compared to ST2-Fc/IL-33 groups (Two-way ANOVA with Bonferroni multiple comparison test).
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f4: IL-33 injection produces antigen specific antibody responses.(a) C57BL/6 mice (n = 5 mice/group) were pre-treated with mouse IgG1 isotype control antibody (Ctrl) or ST2-Fc injected i.p. one hour prior to i.p. injection of PBS/NP-CGG or IL-33/NP-CGG then boosted i.p. on day 14 with PBS/NP-CGG. (b) BALB/c mice (n = 5 mice/group) were injected i.p. with PBS/NP-CGG or IL-33/NP-CGG then boosted i.p. on day 14 with PBS/NP-CGG. (c) Mice were treated as in (b) except PE was used as the antigen. (d) Mice were treated as in (b) except OVA was used as the antigen. Antigen specific IgM, IgG1 and IgE serum titers were quantified via ELISA. Data in (a) are representative of four independent experiments and data in (b–d) are from one experiment. ***p < 0.001 PBS compared to Ctrl/IL-33 groups (Two-way ANOVA with Bonferroni multiple comparison test). ###p < 0.001 Ctrl/IL-33 compared to ST2-Fc/IL-33 groups (Two-way ANOVA with Bonferroni multiple comparison test).

Mentions: Given the similarities between cytokine patterns induced by alum and IL-33 (Fig. 2), we hypothesized that IL-33 itself could serve as an adjuvant. Mice were immunized with a single injection of IL-33/4-Hydroxy-3-nitrophenylacetyl hapten conjugated to chicken gamma globulin at a 30-39 ratio (NP-CGG) or PBS/NP-CGG, received a PBS/NP-CGG boost on day 14 and serum antibody titers were quantified on days 5, 10, 17 and 21 post-immunization. Mice injected with NP-CGG in the presence of IL-33 showed significantly increased NP-specific IgM titers on day 10 which were absent in control mice (Fig. 4a). Following antigen boost, the IgM titers continued to increase on days 17 and 21. Interestingly, little to no NP-specific IgG1 antibodies (the predominant antigen-specific IgG isotype produced with NP-CGG (data not shown)), were observed during the primary response on days 5 or 10 post-immunization. However, IgG1 titers significantly increased on days 17 and 21 following an antigen boost. IL-13 is a critical mediator of IgE production30 and IL-33 injection induced release of IL-13 in mice (Fig. 2), therefore we also quantified IgE titers. NP-specific IgE antibody response kinetics showed no significant increases on day 10 but boosting resulted in a significant increase that plateaued on days 17 and 21. To demonstrate that the observed antigen-specific antibody responses were due to IL-33 activity, mice were treated with ST2-Fc prior to immunization with IL-33/NP-CGG. This resulted in a complete absence of NP-specific antibody titers (Fig. 4a), indicating that IL-33 was solely responsible for the observed humoral responses.


IL-33 released by alum is responsible for early cytokine production and has adjuvant properties.

Rose WA, Okragly AJ, Patel CN, Benschop RJ - Sci Rep (2015)

IL-33 injection produces antigen specific antibody responses.(a) C57BL/6 mice (n = 5 mice/group) were pre-treated with mouse IgG1 isotype control antibody (Ctrl) or ST2-Fc injected i.p. one hour prior to i.p. injection of PBS/NP-CGG or IL-33/NP-CGG then boosted i.p. on day 14 with PBS/NP-CGG. (b) BALB/c mice (n = 5 mice/group) were injected i.p. with PBS/NP-CGG or IL-33/NP-CGG then boosted i.p. on day 14 with PBS/NP-CGG. (c) Mice were treated as in (b) except PE was used as the antigen. (d) Mice were treated as in (b) except OVA was used as the antigen. Antigen specific IgM, IgG1 and IgE serum titers were quantified via ELISA. Data in (a) are representative of four independent experiments and data in (b–d) are from one experiment. ***p < 0.001 PBS compared to Ctrl/IL-33 groups (Two-way ANOVA with Bonferroni multiple comparison test). ###p < 0.001 Ctrl/IL-33 compared to ST2-Fc/IL-33 groups (Two-way ANOVA with Bonferroni multiple comparison test).
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f4: IL-33 injection produces antigen specific antibody responses.(a) C57BL/6 mice (n = 5 mice/group) were pre-treated with mouse IgG1 isotype control antibody (Ctrl) or ST2-Fc injected i.p. one hour prior to i.p. injection of PBS/NP-CGG or IL-33/NP-CGG then boosted i.p. on day 14 with PBS/NP-CGG. (b) BALB/c mice (n = 5 mice/group) were injected i.p. with PBS/NP-CGG or IL-33/NP-CGG then boosted i.p. on day 14 with PBS/NP-CGG. (c) Mice were treated as in (b) except PE was used as the antigen. (d) Mice were treated as in (b) except OVA was used as the antigen. Antigen specific IgM, IgG1 and IgE serum titers were quantified via ELISA. Data in (a) are representative of four independent experiments and data in (b–d) are from one experiment. ***p < 0.001 PBS compared to Ctrl/IL-33 groups (Two-way ANOVA with Bonferroni multiple comparison test). ###p < 0.001 Ctrl/IL-33 compared to ST2-Fc/IL-33 groups (Two-way ANOVA with Bonferroni multiple comparison test).
Mentions: Given the similarities between cytokine patterns induced by alum and IL-33 (Fig. 2), we hypothesized that IL-33 itself could serve as an adjuvant. Mice were immunized with a single injection of IL-33/4-Hydroxy-3-nitrophenylacetyl hapten conjugated to chicken gamma globulin at a 30-39 ratio (NP-CGG) or PBS/NP-CGG, received a PBS/NP-CGG boost on day 14 and serum antibody titers were quantified on days 5, 10, 17 and 21 post-immunization. Mice injected with NP-CGG in the presence of IL-33 showed significantly increased NP-specific IgM titers on day 10 which were absent in control mice (Fig. 4a). Following antigen boost, the IgM titers continued to increase on days 17 and 21. Interestingly, little to no NP-specific IgG1 antibodies (the predominant antigen-specific IgG isotype produced with NP-CGG (data not shown)), were observed during the primary response on days 5 or 10 post-immunization. However, IgG1 titers significantly increased on days 17 and 21 following an antigen boost. IL-13 is a critical mediator of IgE production30 and IL-33 injection induced release of IL-13 in mice (Fig. 2), therefore we also quantified IgE titers. NP-specific IgE antibody response kinetics showed no significant increases on day 10 but boosting resulted in a significant increase that plateaued on days 17 and 21. To demonstrate that the observed antigen-specific antibody responses were due to IL-33 activity, mice were treated with ST2-Fc prior to immunization with IL-33/NP-CGG. This resulted in a complete absence of NP-specific antibody titers (Fig. 4a), indicating that IL-33 was solely responsible for the observed humoral responses.

Bottom Line: Alum can induce the release of endogenous danger signals via cellular necrosis which elicits inflammation-associated cytokines resulting in humoral immunity.Furthermore, IL-33 itself functions as an adjuvant that, while only inducing a marginal primary response, facilitates a robust secondary response comparable to that observed with alum.Our results provide novel insights into the mechanism of action behind alum-induced cytokine responses and show that IL-33 is sufficient to provide a robust secondary antibody response independently of alum.

View Article: PubMed Central - PubMed

Affiliation: Biotechnology Discovery Research, Lilly Research Laboratories, Eli Lilly and Company, Indianapolis, IN, 46285.

ABSTRACT
Human vaccines have used aluminium-based adjuvants (alum) for >80 years despite incomplete understanding of how alum enhances the immune response. Alum can induce the release of endogenous danger signals via cellular necrosis which elicits inflammation-associated cytokines resulting in humoral immunity. IL-33 is proposed to be one such danger signal that is released from necrotic cells. Therefore, we investigated whether there is a role for IL-33 in the adjuvant activity of alum. We show that alum-induced cellular necrosis results in elevated levels of IL-33 following injection in vivo. Alum and IL-33 induce similar increases in IL-5, KC, MCP-1, MIP-1α and MIP-1β; many of which are dependent on IL-33 as shown in IL-33 knockout mice or by using an IL-33-neutralizing recombinant ST2 receptor. Furthermore, IL-33 itself functions as an adjuvant that, while only inducing a marginal primary response, facilitates a robust secondary response comparable to that observed with alum. However, IL-33 is not absolutely required for alum-induced antibody responses since alum mediates similar humoral responses in IL-33 knockout and wild-type mice. Our results provide novel insights into the mechanism of action behind alum-induced cytokine responses and show that IL-33 is sufficient to provide a robust secondary antibody response independently of alum.

No MeSH data available.


Related in: MedlinePlus