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Immune response of rats vaccinated orally with various plant-expressed recombinant cysteine proteinase constructs when challenged with Fasciola hepatica metacercariae

View Article: PubMed Central - PubMed

ABSTRACT

Background: Cysteine proteinases of Fasciola hepatica are important candidates for vaccine antigens because of their role in fluke biology and host-parasite relationships. In our previous experiments, we found that a recombinant cysteine proteinase cloned from adult F. hepatica (CPFhW) can protect rats against liver fluke infections when it is administered intramuscularly or intranasally in the form of cDNA. We also observed considerable protection upon challenge following mucosal vaccination with inclusion bodies containing recombinant CPFhW produced in Escherichia coli.

Background: In this study, we explore oral vaccination, which may be the desired method of delivery and is potentially capable of preventing infections at the site of helminth entry. To provide antigen encapsulation and to protect the vaccine antigen from degradation in the intestinal tract, transgenic plant-based systems are used.

Methodology: In the present study, we aimed to evaluate the protective ability of mucosal vaccinations of 12-week-old rats with CPFhW produced in a transgenic-plant-based system. To avoid inducing tolerance and to maximise the immune response induced by oral immunisation, we used the hepatitis B virus (HBV) core protein (HBcAg) as a carrier. Animals were immunised with two doses of the antigen and challenged with 25 or 30 metacercariae of F. hepatica.

Conclusions: We obtained substantial protection after oral administration of the plant-produced hybrids of CPFhW and HBcAg. The highest level of protection (65.4%) was observed in animals immunised with transgenic plants expressing the mature CPFhW enzyme flanked by Gly-rich linkers and inserted into c/e1 epitope of truncated HBcAg. The immunised rats showed clear IgG1 and IgM responses to CPFhW for 4 consecutive weeks after the challenge.

No MeSH data available.


Leukocyte responses in the peritoneal cavity of vaccinated and control rats to challenge infections.(A) eosinophils. (B) monocytes. (C) CD4+ T lymphocytes. (D) CD8+ T lymphocytes. Group G–Lyophilised lettuce expressing the mature CPFhW enzyme flanked by Gly-rich linkers; Group U–Lyophilised lettuce expressing the mature CPFhW protein fused with ubiquitin; Group C–Lyophilised control lettuce; Group N–None. At each timepoint, four rats from each group were euthanised and dissected. *Indicates significantly increased numbers of cells (p<0.05). Error bars indicate standard deviation.
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pntd.0005451.g005: Leukocyte responses in the peritoneal cavity of vaccinated and control rats to challenge infections.(A) eosinophils. (B) monocytes. (C) CD4+ T lymphocytes. (D) CD8+ T lymphocytes. Group G–Lyophilised lettuce expressing the mature CPFhW enzyme flanked by Gly-rich linkers; Group U–Lyophilised lettuce expressing the mature CPFhW protein fused with ubiquitin; Group C–Lyophilised control lettuce; Group N–None. At each timepoint, four rats from each group were euthanised and dissected. *Indicates significantly increased numbers of cells (p<0.05). Error bars indicate standard deviation.

Mentions: The challenge with F. hepatica mc caused remarkable changes in the numbers of eosinophils, monocytes, CD4+ cells and CD8+ cells in both the vaccinated and challenge control rats (Figs 4–6). Significantly (p<0.05) increased numbers of blood eosinophils were observed in the vaccinated rats and challenge controls at 5 weeks after challenge and in groups G and C at 9 weeks after challenge (Fig 4A). Similar cellular response dynamics were observed for the peritoneal fluid (Fig 5A). Blood monocytes were present at similar levels in the vaccinated and control rats at weeks 1 and 5 after challenge, but at the end of the experiment, the rats in group C had the highest numbers of these cells (Fig 4B). In the peritoneal fluid, the numbers of these cells were significantly (p<0.05) higher than that in the challenge controls at the first and fifth weeks after the challenge infection (Fig 5B).


Immune response of rats vaccinated orally with various plant-expressed recombinant cysteine proteinase constructs when challenged with Fasciola hepatica metacercariae
Leukocyte responses in the peritoneal cavity of vaccinated and control rats to challenge infections.(A) eosinophils. (B) monocytes. (C) CD4+ T lymphocytes. (D) CD8+ T lymphocytes. Group G–Lyophilised lettuce expressing the mature CPFhW enzyme flanked by Gly-rich linkers; Group U–Lyophilised lettuce expressing the mature CPFhW protein fused with ubiquitin; Group C–Lyophilised control lettuce; Group N–None. At each timepoint, four rats from each group were euthanised and dissected. *Indicates significantly increased numbers of cells (p<0.05). Error bars indicate standard deviation.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC5383346&req=5

pntd.0005451.g005: Leukocyte responses in the peritoneal cavity of vaccinated and control rats to challenge infections.(A) eosinophils. (B) monocytes. (C) CD4+ T lymphocytes. (D) CD8+ T lymphocytes. Group G–Lyophilised lettuce expressing the mature CPFhW enzyme flanked by Gly-rich linkers; Group U–Lyophilised lettuce expressing the mature CPFhW protein fused with ubiquitin; Group C–Lyophilised control lettuce; Group N–None. At each timepoint, four rats from each group were euthanised and dissected. *Indicates significantly increased numbers of cells (p<0.05). Error bars indicate standard deviation.
Mentions: The challenge with F. hepatica mc caused remarkable changes in the numbers of eosinophils, monocytes, CD4+ cells and CD8+ cells in both the vaccinated and challenge control rats (Figs 4–6). Significantly (p<0.05) increased numbers of blood eosinophils were observed in the vaccinated rats and challenge controls at 5 weeks after challenge and in groups G and C at 9 weeks after challenge (Fig 4A). Similar cellular response dynamics were observed for the peritoneal fluid (Fig 5A). Blood monocytes were present at similar levels in the vaccinated and control rats at weeks 1 and 5 after challenge, but at the end of the experiment, the rats in group C had the highest numbers of these cells (Fig 4B). In the peritoneal fluid, the numbers of these cells were significantly (p<0.05) higher than that in the challenge controls at the first and fifth weeks after the challenge infection (Fig 5B).

View Article: PubMed Central - PubMed

ABSTRACT

Background: Cysteine proteinases of Fasciola hepatica are important candidates for vaccine antigens because of their role in fluke biology and host-parasite relationships. In our previous experiments, we found that a recombinant cysteine proteinase cloned from adult F. hepatica (CPFhW) can protect rats against liver fluke infections when it is administered intramuscularly or intranasally in the form of cDNA. We also observed considerable protection upon challenge following mucosal vaccination with inclusion bodies containing recombinant CPFhW produced in Escherichia coli.

Background: In this study, we explore oral vaccination, which may be the desired method of delivery and is potentially capable of preventing infections at the site of helminth entry. To provide antigen encapsulation and to protect the vaccine antigen from degradation in the intestinal tract, transgenic plant-based systems are used.

Methodology: In the present study, we aimed to evaluate the protective ability of mucosal vaccinations of 12-week-old rats with CPFhW produced in a transgenic-plant-based system. To avoid inducing tolerance and to maximise the immune response induced by oral immunisation, we used the hepatitis B virus (HBV) core protein (HBcAg) as a carrier. Animals were immunised with two doses of the antigen and challenged with 25 or 30 metacercariae of F. hepatica.

Conclusions: We obtained substantial protection after oral administration of the plant-produced hybrids of CPFhW and HBcAg. The highest level of protection (65.4%) was observed in animals immunised with transgenic plants expressing the mature CPFhW enzyme flanked by Gly-rich linkers and inserted into c/e1 epitope of truncated HBcAg. The immunised rats showed clear IgG1 and IgM responses to CPFhW for 4 consecutive weeks after the challenge.

No MeSH data available.