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Virus-Like Particles Produced in Pichia Pastoris Induce Protective Immune Responses Against Coxsackievirus A16 in Mice

View Article: PubMed Central - PubMed

ABSTRACT

Background: Coxsackievirus A16 (CA16) is one of the main causative agents of hand, foot, and mouth disease (HFMD), and the development of a safe and effective vaccine has been a top priority among CA16 researchers.

Material/methods: In this study, we developed a Pichia pastoris yeast system for secretory expression of the virus-like particles (VLPs) for CA16 by co-expression of the P1 and 3CD proteins of CA16. SDS-PAGE, Western blot, and transmission electron microscopy (TEM) were performed to identify the formation of VLPs. Immunogenicity and vaccine efficacy of the CA16 VLPs were assessed in BABL/c mouse models.

Results: Biochemical and biophysical analysis showed that the yeast-expressed CA16 VLPs were composed of VP0, VP1, and VP3 capsid subunit proteins, and present spherical particles with a diameter of 30 nm, similar to the parental infectious CA16 virus. Furthermore, CA16 VLPs elicited potent humoral and cellular immune responses, and VLPs-immunized sera conferred efficient protection to neonatal mice against lethal CA16 challenge.

Conclusions: Our results demonstrate that VLPs produced in Pichia pastoris represent a safe and effective vaccine strategy for CA16.

No MeSH data available.


The production of cytokines by VLPs-induced splenocytes. The splenocytes derived from different groups of immunized mice were stimulated with purified VLPs, and the cytokine (INF-γ, IL-2, IL-4, IL-6 and IL-10) concentrations in the culture media were measured by ELISA.
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f5-medscimonit-22-3370: The production of cytokines by VLPs-induced splenocytes. The splenocytes derived from different groups of immunized mice were stimulated with purified VLPs, and the cytokine (INF-γ, IL-2, IL-4, IL-6 and IL-10) concentrations in the culture media were measured by ELISA.

Mentions: Cellular immune response elicited by VLPs immunization was evaluated by splenocyte proliferation assay and cytokine production analysis. The splenocytes were isolated from immunized mice at week 2 after the booster immunization and subjected to the above assays. The VLPs-specific splenocyte proliferation assay was performed using CCK-8 methods. Figure 4 shows that the splenocytes collected from VLPs-immunized or VLPs+FA-immunized mice could be significantly induced to proliferate by VLPs compared with splenocytes of negative control groups (p<0.05). Compared to the inactivated CA16 group, the SI values for the VLPs group were statistically higher with ConA or VLPs stimulation (p<0.05). Profiles of cytokines secreted by VLPs-stimulated splenocytes were evaluated by measuring the levels of INF-γ and IL-2 (Th1 cytokines) and IL-4, IL-6, and IL-10 (Th2 cytokines). As shown in Figure 5, splenocytes of VLPs-immunized or VLPs+FA-immunized mice produced significantly higher levels of INF-γ, IL-2, IL-4, IL-6, and IL-10 compared with those from the negative controls (p<0.05). Under the same sample treatment, cytokine profiles of inactivated CA16-immunized mice were similar to those of the VLPs group, except for lower levels of IL-4 (p<0.05). The results suggested that VLPs immunization induced both Th1-type and Th2-type cellular immune response.


Virus-Like Particles Produced in Pichia Pastoris Induce Protective Immune Responses Against Coxsackievirus A16 in Mice
The production of cytokines by VLPs-induced splenocytes. The splenocytes derived from different groups of immunized mice were stimulated with purified VLPs, and the cytokine (INF-γ, IL-2, IL-4, IL-6 and IL-10) concentrations in the culture media were measured by ELISA.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC5036384&req=5

f5-medscimonit-22-3370: The production of cytokines by VLPs-induced splenocytes. The splenocytes derived from different groups of immunized mice were stimulated with purified VLPs, and the cytokine (INF-γ, IL-2, IL-4, IL-6 and IL-10) concentrations in the culture media were measured by ELISA.
Mentions: Cellular immune response elicited by VLPs immunization was evaluated by splenocyte proliferation assay and cytokine production analysis. The splenocytes were isolated from immunized mice at week 2 after the booster immunization and subjected to the above assays. The VLPs-specific splenocyte proliferation assay was performed using CCK-8 methods. Figure 4 shows that the splenocytes collected from VLPs-immunized or VLPs+FA-immunized mice could be significantly induced to proliferate by VLPs compared with splenocytes of negative control groups (p<0.05). Compared to the inactivated CA16 group, the SI values for the VLPs group were statistically higher with ConA or VLPs stimulation (p<0.05). Profiles of cytokines secreted by VLPs-stimulated splenocytes were evaluated by measuring the levels of INF-γ and IL-2 (Th1 cytokines) and IL-4, IL-6, and IL-10 (Th2 cytokines). As shown in Figure 5, splenocytes of VLPs-immunized or VLPs+FA-immunized mice produced significantly higher levels of INF-γ, IL-2, IL-4, IL-6, and IL-10 compared with those from the negative controls (p<0.05). Under the same sample treatment, cytokine profiles of inactivated CA16-immunized mice were similar to those of the VLPs group, except for lower levels of IL-4 (p<0.05). The results suggested that VLPs immunization induced both Th1-type and Th2-type cellular immune response.

View Article: PubMed Central - PubMed

ABSTRACT

Background: Coxsackievirus A16 (CA16) is one of the main causative agents of hand, foot, and mouth disease (HFMD), and the development of a safe and effective vaccine has been a top priority among CA16 researchers.

Material/methods: In this study, we developed a Pichia pastoris yeast system for secretory expression of the virus-like particles (VLPs) for CA16 by co-expression of the P1 and 3CD proteins of CA16. SDS-PAGE, Western blot, and transmission electron microscopy (TEM) were performed to identify the formation of VLPs. Immunogenicity and vaccine efficacy of the CA16 VLPs were assessed in BABL/c mouse models.

Results: Biochemical and biophysical analysis showed that the yeast-expressed CA16 VLPs were composed of VP0, VP1, and VP3 capsid subunit proteins, and present spherical particles with a diameter of 30 nm, similar to the parental infectious CA16 virus. Furthermore, CA16 VLPs elicited potent humoral and cellular immune responses, and VLPs-immunized sera conferred efficient protection to neonatal mice against lethal CA16 challenge.

Conclusions: Our results demonstrate that VLPs produced in Pichia pastoris represent a safe and effective vaccine strategy for CA16.

No MeSH data available.