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Caspase-1 Dependent IL-1β Secretion and Antigen-Specific T-Cell Activation by the Novel Adjuvant, PCEP.

Awate S, Eng NF, Gerdts V, Babiuk LA, Mutwiri G - Vaccines (Basel) (2014)

Bottom Line: Caspase inhibitor YVAD-fmk-treated splenic DCs showed significantly reduced IL-1β and IL-18 secretion in response to PCEP stimulation.Further, PCEP had no effect on the expression of MHC class II or co-stimulatory molecules, CD86 and CD40, suggesting that PCEP does not induce DC maturation.We conclude that PCEP activates innate immunity, leading to increased antigen-specific T-cell responses.

View Article: PubMed Central - PubMed

Affiliation: Vaccinology and Immunotherapeutics program, School of Public Health, 107 Wiggins Road, University of Saskatchewan, Saskatoon, SK S7N 5E5, Canada. sunita.awate@usask.ca.

ABSTRACT
The potent adjuvant activity of the novel adjuvant, poly[di(sodiumcarboxylatoethylphenoxy)phosphazene] (PCEP), with various antigens has been reported previously. However, very little is known about its mechanisms of action. We have recently reported that intramuscular injection of PCEP induces NLRP3, an inflammasome receptor gene, and inflammatory cytokines, including IL-1β and IL-18, in mouse muscle tissue. Caspase-1 is required for the processing of pro-forms of IL-1β and IL-18 into mature forms and is a critical constituent of the NLRP3 inflammasome. Hence, in the present study, we investigated the role of caspase-1 in the secretion of IL-1β and IL-18 in PCEP-stimulated splenic dendritic cells (DCs). Caspase inhibitor YVAD-fmk-treated splenic DCs showed significantly reduced IL-1β and IL-18 secretion in response to PCEP stimulation. Further, PCEP had no effect on the expression of MHC class II or co-stimulatory molecules, CD86 and CD40, suggesting that PCEP does not induce DC maturation. However, PCEP directly activated B-cells to induce significant production of IgM. In addition, PCEP+ovalbumin (OVA) immunized mice showed significantly increased production of antigen-specific IFN-γ by CD4⁺ and CD8⁺ T-cells. We conclude that PCEP activates innate immunity, leading to increased antigen-specific T-cell responses.

No MeSH data available.


The role of caspase-1 in PCEP-stimulated IL-1β and IL-18 secretion. Enriched splenic dendritic cells (DCs) were treated with or without the caspase-1 inhibitor (CI) YVAD-fmk (40 μM) and then incubated with media, PCEP (50 μg/mL), alum (40 mg/mL), lipopolysaccharide (LPS) (0.1 μg/mL), PCEP+LPS or alum+LPS. Supernatants were collected after 12 h of stimulation and were analyzed for IL-1β and IL-18 by ELISA. Data were analyzed by one-way ANOVA, and the comparisons between the treatments were done by Tukey’s multiple comparison test; ***p < 0.0001, **p < 0.001, *p < 0.05.
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vaccines-02-00500-f002: The role of caspase-1 in PCEP-stimulated IL-1β and IL-18 secretion. Enriched splenic dendritic cells (DCs) were treated with or without the caspase-1 inhibitor (CI) YVAD-fmk (40 μM) and then incubated with media, PCEP (50 μg/mL), alum (40 mg/mL), lipopolysaccharide (LPS) (0.1 μg/mL), PCEP+LPS or alum+LPS. Supernatants were collected after 12 h of stimulation and were analyzed for IL-1β and IL-18 by ELISA. Data were analyzed by one-way ANOVA, and the comparisons between the treatments were done by Tukey’s multiple comparison test; ***p < 0.0001, **p < 0.001, *p < 0.05.

Mentions: Caspase-1 is a critical component of the NLRP3 inflammasome, which cleaves the pro-form of IL-1β and IL-18 into mature forms. Hence, we examined the role of caspase-1 in the secretion of IL-1β and IL-18 by splenic DCs. Enriched splenic DCs were treated with or without the caspase-1 inhibitor (CI) YVAD-fmk and then stimulated with media, PCEP, alum, LPS, PCEP+LPS or alum+LPS for 12 h. The secretion of IL-1β and IL-18 was analyzed in culture supernatants. Pre-treatment with YVAD-fmk significantly inhibited IL-1β and IL-18 secretion (Figure 2). The most significant reduction in IL-1β and IL-18 secretion was observed in YVAD-fmk-treated enriched DCs that were stimulated with PCEP+LPS (Figure 2). The same was observed with alum+LPS-treated enriched splenic DCs. These results suggest that PCEP- and alum-mediated secretion of IL-1β and IL-18 in splenic DCs was caspase-1 dependent.


Caspase-1 Dependent IL-1β Secretion and Antigen-Specific T-Cell Activation by the Novel Adjuvant, PCEP.

Awate S, Eng NF, Gerdts V, Babiuk LA, Mutwiri G - Vaccines (Basel) (2014)

The role of caspase-1 in PCEP-stimulated IL-1β and IL-18 secretion. Enriched splenic dendritic cells (DCs) were treated with or without the caspase-1 inhibitor (CI) YVAD-fmk (40 μM) and then incubated with media, PCEP (50 μg/mL), alum (40 mg/mL), lipopolysaccharide (LPS) (0.1 μg/mL), PCEP+LPS or alum+LPS. Supernatants were collected after 12 h of stimulation and were analyzed for IL-1β and IL-18 by ELISA. Data were analyzed by one-way ANOVA, and the comparisons between the treatments were done by Tukey’s multiple comparison test; ***p < 0.0001, **p < 0.001, *p < 0.05.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4494223&req=5

vaccines-02-00500-f002: The role of caspase-1 in PCEP-stimulated IL-1β and IL-18 secretion. Enriched splenic dendritic cells (DCs) were treated with or without the caspase-1 inhibitor (CI) YVAD-fmk (40 μM) and then incubated with media, PCEP (50 μg/mL), alum (40 mg/mL), lipopolysaccharide (LPS) (0.1 μg/mL), PCEP+LPS or alum+LPS. Supernatants were collected after 12 h of stimulation and were analyzed for IL-1β and IL-18 by ELISA. Data were analyzed by one-way ANOVA, and the comparisons between the treatments were done by Tukey’s multiple comparison test; ***p < 0.0001, **p < 0.001, *p < 0.05.
Mentions: Caspase-1 is a critical component of the NLRP3 inflammasome, which cleaves the pro-form of IL-1β and IL-18 into mature forms. Hence, we examined the role of caspase-1 in the secretion of IL-1β and IL-18 by splenic DCs. Enriched splenic DCs were treated with or without the caspase-1 inhibitor (CI) YVAD-fmk and then stimulated with media, PCEP, alum, LPS, PCEP+LPS or alum+LPS for 12 h. The secretion of IL-1β and IL-18 was analyzed in culture supernatants. Pre-treatment with YVAD-fmk significantly inhibited IL-1β and IL-18 secretion (Figure 2). The most significant reduction in IL-1β and IL-18 secretion was observed in YVAD-fmk-treated enriched DCs that were stimulated with PCEP+LPS (Figure 2). The same was observed with alum+LPS-treated enriched splenic DCs. These results suggest that PCEP- and alum-mediated secretion of IL-1β and IL-18 in splenic DCs was caspase-1 dependent.

Bottom Line: Caspase inhibitor YVAD-fmk-treated splenic DCs showed significantly reduced IL-1β and IL-18 secretion in response to PCEP stimulation.Further, PCEP had no effect on the expression of MHC class II or co-stimulatory molecules, CD86 and CD40, suggesting that PCEP does not induce DC maturation.We conclude that PCEP activates innate immunity, leading to increased antigen-specific T-cell responses.

View Article: PubMed Central - PubMed

Affiliation: Vaccinology and Immunotherapeutics program, School of Public Health, 107 Wiggins Road, University of Saskatchewan, Saskatoon, SK S7N 5E5, Canada. sunita.awate@usask.ca.

ABSTRACT
The potent adjuvant activity of the novel adjuvant, poly[di(sodiumcarboxylatoethylphenoxy)phosphazene] (PCEP), with various antigens has been reported previously. However, very little is known about its mechanisms of action. We have recently reported that intramuscular injection of PCEP induces NLRP3, an inflammasome receptor gene, and inflammatory cytokines, including IL-1β and IL-18, in mouse muscle tissue. Caspase-1 is required for the processing of pro-forms of IL-1β and IL-18 into mature forms and is a critical constituent of the NLRP3 inflammasome. Hence, in the present study, we investigated the role of caspase-1 in the secretion of IL-1β and IL-18 in PCEP-stimulated splenic dendritic cells (DCs). Caspase inhibitor YVAD-fmk-treated splenic DCs showed significantly reduced IL-1β and IL-18 secretion in response to PCEP stimulation. Further, PCEP had no effect on the expression of MHC class II or co-stimulatory molecules, CD86 and CD40, suggesting that PCEP does not induce DC maturation. However, PCEP directly activated B-cells to induce significant production of IgM. In addition, PCEP+ovalbumin (OVA) immunized mice showed significantly increased production of antigen-specific IFN-γ by CD4⁺ and CD8⁺ T-cells. We conclude that PCEP activates innate immunity, leading to increased antigen-specific T-cell responses.

No MeSH data available.