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Enhanced Immune Response to DNA Vaccine Encoding Bacillus anthracis PA-D4 Protects Mice against Anthrax Spore Challenge.

Kim NY, Chang DS, Kim Y, Kim CH, Hur GH, Yang JM, Shin S - PLoS ONE (2015)

Bottom Line: The results revealed that PA-D4 protein can be efficiently expressed and secreted at high levels into the culture medium.Furthermore, incorporation of the SV40 enhancer in the plasmid DNA resulted in approximately a 15-fold increase in serum antibody levels in comparison with the plasmid without enhancer.The biodistribution study showed that plasmid DNA was detected in most organs and it rapidly cleared from the injection site.

View Article: PubMed Central - PubMed

Affiliation: Department of Life Science, Sogang University, Seoul, Republic of Korea.

ABSTRACT
Anthrax has long been considered the most probable bioweapon-induced disease. The protective antigen (PA) of Bacillus anthracis plays a crucial role in the pathogenesis of anthrax. In the current study, we evaluated the efficiency of a genetic vaccination with the fourth domain (D4) of PA, which is responsible for initial binding of the anthrax toxin to the cellular receptor. The eukaryotic expression vector was designed with the immunoglobulin M (IgM) signal sequence encoding for PA-D4, which contains codon-optimized genes. The expression and secretion of recombinant protein was confirmed in vitro in 293T cells transfected with plasmid and detected by western blotting, confocal microscopy, and enzyme-linked immunosorbent assay (ELISA). The results revealed that PA-D4 protein can be efficiently expressed and secreted at high levels into the culture medium. When plasmid DNA was given intramuscularly to mice, a significant PA-D4-specific antibody response was induced. Importantly, high titers of antibodies were maintained for nearly 1 year. Furthermore, incorporation of the SV40 enhancer in the plasmid DNA resulted in approximately a 15-fold increase in serum antibody levels in comparison with the plasmid without enhancer. The antibodies produced were predominantly the immunoglobulin G2 (IgG2) type, indicating the predominance of the Th1 response. In addition, splenocytes collected from immunized mice produced PA-D4-specific interferon gamma (IFN-γ). The biodistribution study showed that plasmid DNA was detected in most organs and it rapidly cleared from the injection site. Finally, DNA vaccination with electroporation induced a significant increase in immunogenicity and successfully protected the mice against anthrax spore challenge. Our approach to enhancing the immune response contributes to the development of DNA vaccines against anthrax and other biothreats.

No MeSH data available.


Related in: MedlinePlus

Survival of A/J mice immunized with IgM-D4/SV40 DNA vaccine.Mice were immunized with 50 μg of DNA vaccine intramuscularly via EP followed by two boosters with the same dose at 2-week intervals. A/J mice immunized with DNA vaccines were challenged by either 20 LD50 or 50 LD50 spores 2 weeks after the final immunization.
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pone.0139671.g008: Survival of A/J mice immunized with IgM-D4/SV40 DNA vaccine.Mice were immunized with 50 μg of DNA vaccine intramuscularly via EP followed by two boosters with the same dose at 2-week intervals. A/J mice immunized with DNA vaccines were challenged by either 20 LD50 or 50 LD50 spores 2 weeks after the final immunization.

Mentions: To determine if the immune response generated against PA-D4 was sufficient for providing protection against anthrax, we performed an in vivo protection experiment. A/J mice (n = 5∼6) were immunized IM via EP with 50μg of IgM-D4/SV40. The mice received two boosters with the same dose at a 2-week interval. Two weeks after the last immunization, mice were challenged with live spores. A/J mice received two different doses of B. anthracis Sterne spores subcutaneously, either 3.5 x 103 (20 LD50) or 9 x 103 (50 LD50). As expected, all mice that were immunized with control groups succumbed to challenge by day 3 and all mice that were immunized with IgM-D4/SV40 survived (Fig 8). In addition, survivors demonstrated high geometric titers (1.86 x 104) of PA-D4-specific antibody one week before challenge (data not shown). From these results, we infer that the IgM-D4/SV40 DNA vaccine is able to generate protection against B. anthracis.


Enhanced Immune Response to DNA Vaccine Encoding Bacillus anthracis PA-D4 Protects Mice against Anthrax Spore Challenge.

Kim NY, Chang DS, Kim Y, Kim CH, Hur GH, Yang JM, Shin S - PLoS ONE (2015)

Survival of A/J mice immunized with IgM-D4/SV40 DNA vaccine.Mice were immunized with 50 μg of DNA vaccine intramuscularly via EP followed by two boosters with the same dose at 2-week intervals. A/J mice immunized with DNA vaccines were challenged by either 20 LD50 or 50 LD50 spores 2 weeks after the final immunization.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4591996&req=5

pone.0139671.g008: Survival of A/J mice immunized with IgM-D4/SV40 DNA vaccine.Mice were immunized with 50 μg of DNA vaccine intramuscularly via EP followed by two boosters with the same dose at 2-week intervals. A/J mice immunized with DNA vaccines were challenged by either 20 LD50 or 50 LD50 spores 2 weeks after the final immunization.
Mentions: To determine if the immune response generated against PA-D4 was sufficient for providing protection against anthrax, we performed an in vivo protection experiment. A/J mice (n = 5∼6) were immunized IM via EP with 50μg of IgM-D4/SV40. The mice received two boosters with the same dose at a 2-week interval. Two weeks after the last immunization, mice were challenged with live spores. A/J mice received two different doses of B. anthracis Sterne spores subcutaneously, either 3.5 x 103 (20 LD50) or 9 x 103 (50 LD50). As expected, all mice that were immunized with control groups succumbed to challenge by day 3 and all mice that were immunized with IgM-D4/SV40 survived (Fig 8). In addition, survivors demonstrated high geometric titers (1.86 x 104) of PA-D4-specific antibody one week before challenge (data not shown). From these results, we infer that the IgM-D4/SV40 DNA vaccine is able to generate protection against B. anthracis.

Bottom Line: The results revealed that PA-D4 protein can be efficiently expressed and secreted at high levels into the culture medium.Furthermore, incorporation of the SV40 enhancer in the plasmid DNA resulted in approximately a 15-fold increase in serum antibody levels in comparison with the plasmid without enhancer.The biodistribution study showed that plasmid DNA was detected in most organs and it rapidly cleared from the injection site.

View Article: PubMed Central - PubMed

Affiliation: Department of Life Science, Sogang University, Seoul, Republic of Korea.

ABSTRACT
Anthrax has long been considered the most probable bioweapon-induced disease. The protective antigen (PA) of Bacillus anthracis plays a crucial role in the pathogenesis of anthrax. In the current study, we evaluated the efficiency of a genetic vaccination with the fourth domain (D4) of PA, which is responsible for initial binding of the anthrax toxin to the cellular receptor. The eukaryotic expression vector was designed with the immunoglobulin M (IgM) signal sequence encoding for PA-D4, which contains codon-optimized genes. The expression and secretion of recombinant protein was confirmed in vitro in 293T cells transfected with plasmid and detected by western blotting, confocal microscopy, and enzyme-linked immunosorbent assay (ELISA). The results revealed that PA-D4 protein can be efficiently expressed and secreted at high levels into the culture medium. When plasmid DNA was given intramuscularly to mice, a significant PA-D4-specific antibody response was induced. Importantly, high titers of antibodies were maintained for nearly 1 year. Furthermore, incorporation of the SV40 enhancer in the plasmid DNA resulted in approximately a 15-fold increase in serum antibody levels in comparison with the plasmid without enhancer. The antibodies produced were predominantly the immunoglobulin G2 (IgG2) type, indicating the predominance of the Th1 response. In addition, splenocytes collected from immunized mice produced PA-D4-specific interferon gamma (IFN-γ). The biodistribution study showed that plasmid DNA was detected in most organs and it rapidly cleared from the injection site. Finally, DNA vaccination with electroporation induced a significant increase in immunogenicity and successfully protected the mice against anthrax spore challenge. Our approach to enhancing the immune response contributes to the development of DNA vaccines against anthrax and other biothreats.

No MeSH data available.


Related in: MedlinePlus