Adjuvant Efficacy of the ECMS-Oil on Immune Responses against Bordetella bronchiseptica in Mice through the TLR2/MyD88/NF-κB Pathway

Abstract

Bordetella infection can be efficiently prevented through vaccination. The current study investigated the effects of an extract of Cochinchina momordica seed (ECMS) combined with oil on the immune responses to the inactivated Bordetella vaccine in mice. Serum IgG and IgG1 level was significantly increased in ECMS-oil group compared to any other group ( $P<0.05$ ) 2 weeks after immunization, while groups ECMS200 μg/400 μg-oil had a markedly higher level of serum IgG2b and IgG3 than any other groups ( $P<0.05$ ). Moreover, lipopolysaccharide/ConA-stimulated proliferation of splenocytes was significantly enhanced in ECMS 400 μg-oil immunized mice in comparison with mice in any other group ( $P<0.05$ ). RT-PCR assay revealed that while ECMS800 μg-oil group had significantly higher levels of serum IL-4, IL-10, Toll-like receptor (TLR)2, and IL-1 beta than any other group ( $P<0.05$ ), the levels of serum IL-2, IL-4, and IL-10 were markedly increased in ECMS 400 μg-oil group as compared to any other groups ( $P<0.05$ ). Blood analysis showed that ECMS800 μg-oil and oil groups had a significantly higher number of immunocytes than any other groups ( $P<0.05$ ). There were significant differences in the number of IgG+, IgG2b+, and IgA+ cells in the lung between ECMS800 μg-oil group and any other groups ( $P<0.05$ ). Western blot analysis demonstrated that stimulation with ECMS 25 μg/mL or 50 ng/mL led to a significant increase in the expression of TLR2, MyD88, and NF-κB in Raw264.7 cells ( $P<0.05$ ). Compared with any other group, the expression of MyD88 was markedly increased in the cells stimulated with ECMS 50 ng/mL, as indicated by the RT-PCR analysis ( $P<0.05$ ). Overall, we observed that ECMS-oil efficiently enhanced the humoral or cellular immune responses against Bordetella and suggested that the mechanism of adjuvant activity of ECMS-oil might involve TLR2/MyD88/NF-κB signaling pathway.