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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

PA-D4-specific antibody responses in guinea pigs immunized with IgM-D4/SV40 DNA vaccine by EP.Guinea pigs were immunized 3 times (week 0, 3, and 6) with different doses of DNA vaccine. (A) Anti-PA-D4 IgG levels are expressed as OD450 values after the serum samples were diluted 100-fold. (B) Anti-PA-D4 IgG levels are expressed as endpoint titers obtained from the ELISA assay. Each symbol represents data from an individual guinea pig. The bars represent the mean titer for each group.
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pone.0139671.g007: PA-D4-specific antibody responses in guinea pigs immunized with IgM-D4/SV40 DNA vaccine by EP.Guinea pigs were immunized 3 times (week 0, 3, and 6) with different doses of DNA vaccine. (A) Anti-PA-D4 IgG levels are expressed as OD450 values after the serum samples were diluted 100-fold. (B) Anti-PA-D4 IgG levels are expressed as endpoint titers obtained from the ELISA assay. Each symbol represents data from an individual guinea pig. The bars represent the mean titer for each group.

Mentions: Next, to assess the effect of IgM-D4/SV40 DNA vaccine in a species of large body mass, guinea pigs were immunized with vaccine doses of 100, 300, and 500μg. Anti-PA-D4 IgG was measured for 44 weeks following three immunizations with EP. As shown in Fig 7A, administration by EP resulted in the rapid and consistent development of high titers of antibodies to PA-D4 for approximately 1 year. However, we noted no significance difference (P>0.05) in the anti-PA-D4 ELISA titers among the groups vaccinated with the 100, 300 and 500μg dose (Fig 7B).


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)

PA-D4-specific antibody responses in guinea pigs immunized with IgM-D4/SV40 DNA vaccine by EP.Guinea pigs were immunized 3 times (week 0, 3, and 6) with different doses of DNA vaccine. (A) Anti-PA-D4 IgG levels are expressed as OD450 values after the serum samples were diluted 100-fold. (B) Anti-PA-D4 IgG levels are expressed as endpoint titers obtained from the ELISA assay. Each symbol represents data from an individual guinea pig. The bars represent the mean titer for each group.
© Copyright Policy
Related In: Results  -  Collection

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

pone.0139671.g007: PA-D4-specific antibody responses in guinea pigs immunized with IgM-D4/SV40 DNA vaccine by EP.Guinea pigs were immunized 3 times (week 0, 3, and 6) with different doses of DNA vaccine. (A) Anti-PA-D4 IgG levels are expressed as OD450 values after the serum samples were diluted 100-fold. (B) Anti-PA-D4 IgG levels are expressed as endpoint titers obtained from the ELISA assay. Each symbol represents data from an individual guinea pig. The bars represent the mean titer for each group.
Mentions: Next, to assess the effect of IgM-D4/SV40 DNA vaccine in a species of large body mass, guinea pigs were immunized with vaccine doses of 100, 300, and 500μg. Anti-PA-D4 IgG was measured for 44 weeks following three immunizations with EP. As shown in Fig 7A, administration by EP resulted in the rapid and consistent development of high titers of antibodies to PA-D4 for approximately 1 year. However, we noted no significance difference (P>0.05) in the anti-PA-D4 ELISA titers among the groups vaccinated with the 100, 300 and 500μg dose (Fig 7B).

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