Co-delivery of plasmid-encoded cytokines modulates the immune response to a DNA vaccine delivered by in vivo electroporation

doi:10.1016/j.vaccine.2006.12.025

Vaccine

Volume 25, Issue 14, 30 March 2007, Pages 2575-2582

https://doi.org/10.1016/j.vaccine.2006.12.025 Get rights and content

Abstract

In this study, in vivo electroporation of a DNA vaccine adjuvanted with plasmids encoding different cytokines was investigated in large animals. Sheep were injected intramuscularly with a DNA vaccine encoding an antigen of Haemonchus contortus (pNPA) and plasmids encoding different cytokines followed by in vivo electroporation. Plasmids (pCI) carrying the genes of different cytokines including ovine IL-4(pCI-IL4), IL-10(pCI-IL10), GM-CSF(pCI-GMCSF), and MCP-1α(pCI-MCP1α), and pCI-IL4 + pCI-GMCSF were co-delivered with pNPA. The results showed that co-delivery of pCI-GMCSF or pCI-IL4 + pCI-GMCSF significantly enhanced both antibody responses and T cell proliferation responses to the antigen after two DNA immunisations compared to co-delivery of pCI. In contrast, antibody responses of the sheep that received pCI-IL10 were decreased significantly. Other cytokine expressing plasmids did not significantly alter the measured immune responses. Furthermore, co-delivery of pCI-GMCSF increased IgG2 response more than IgG1 responses, suggesting a Th1 bias. However, the increase in IgG2 over IgG1 was less apparent when co-delivery of pCI-IL4 with pCI-GMCSF. Interestingly, the co-delivery of pCI-IL4 alone did not increase the IgG1 titre, suggesting that both pCI-GMCSF and pCI-IL4 are required for optimal IgG1 production. Thus, co-delivery of plasmid-encoded cytokine genes with in vivo electroporation has the ability to effectively modulate immune responses to a DNA vaccine in a large animal.

Introduction

DNA vaccination has been shown to be an effective strategy to induce protection against many diseases in mice [1], [2]. However, results of DNA vaccination in mice have not generally been as successful in large animals, including domestic animals and humans. This suggests that further optimisation of DNA vaccination protocols is necessary to achieve successful commercial vaccinations in large bodied species.

Numerous reports have illustrated that cytokine adjuvants have significant effects on modulating the immune responses to DNA vaccination (reviewed in Refs. [3], [4], [5], [6]). In large animals, the effect of cytokine-encoded plasmid co-delivery is not as marked as it is in mice. Indeed, in our previous study in sheep [7], we found that co-delivery of GM-CSF encoding plasmid to an intra-muscularly delivered DNA vaccine did increase the humoral immune memory response following a protein boost, but no quantitative or qualitative effects were observed with plasmids encoding IL-4, IL-5, IL-15 or IFN-γ. We proposed that GM-CSF increased the immune memory response by attracting dendritic cells (DC) to the site of immunisation hence facilitating the uptake and transport of the minute amounts of antigen produced by transfected muscle cells. In contrast, the other cytokines are more likely to affect immune responses at distal sites such as the draining lymph nodes. Consequently, particularly in larger animals, greater amounts of cytokines are required to achieve a physiologically relevant concentration and such amounts may not be available from DNA injected muscle cells. From this perspective, improving the amount of cytokine produced following the delivery of cytokine encoding plasmid, for example by in vivo electroporation [8], [9], [10], [11] might improve the adjuvant effect of the co-delivered cytokine genes. We therefore set out to investigate whether plasmid-encoded cytokines could modulate immune responses of a DNA vaccine delivered by in vivo electroporation and hence further improve immune responses induced in a large animal model by this mode of DNA vaccine delivery [11], [12].

Section snippets

Experimental animals and reagents

Merino ewes were housed in pens within the School of Veterinary Science animal facility, The University of Melbourne, Parkville, fed Lucerne chaff mixed with commercial pellets and allowed access to water ad libitum. All experimental procedures were approved by the University of Melbourne Animal Experimentation Ethics Committee.

A plasmid with a CMV promoter carrying the gene encoding for the NPA protein of Haemonchus contortus (pNPA, provided by Department of Primary Industry (DPI)) [11], was

Antibody responses

Groups of sheep were immunised with pNPA combined with plasmids containing the genes of different cytokines followed by in vivo electroporation at weeks 0 and 4. A protein boost was given at 12 weeks post-primary DNA immunisation. Antibody responses of each sheep at week 0, 6, 12, 13 and 15 are depicted in Fig. 2. In general, median antibody responses were increased in each group at 1-week post-protein boost. The anti-NPA antibody response induced at week 6 after the secondary immunisation of

Discussion

A key finding of this study was that including in vivo electroporated pCI-GMCSF in the presence or absence of pCI-IL4 in DNA vaccines significantly increased not only the humoral immune response but also the T cell proliferative responses in the blood of sheep. There is only one other study [19] showing that T cell proliferative responses can be induced by the DNA vaccines in sheep. In this study, Kennedy et al., targeted the antigen using ovine CTLA-4 and adjuvanted their DNA vaccine with

Acknowledgments

This work was supported by the Australian Research Council and Novartis Animal Health. The authors thank Mr. Bob Geyer for his care of experimental animals and DPI for assistance in producing NPA. The authors also thank Inovio Biomedical Corporation for the in vivo electroporator.

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