1. In DNA vaccine, After plasmid construction and preparation adjuvants ,The mice were divided into 7 groups [pcDNA3/TSA, pcDNA3/TSA +dendrimer and pcDNA3/TSA +PMMA and pcDNA3, dendrimer, PMMA and PBS].The Immunization schedule was performed with 3 inoculations (equal doses of 100 µg) with intramuscular injections at 3 week interval . The booster peptide (20 µg) and incomplete Freund’s adjuvant were subcutaneously injected two weeks after the last nano-vaccines injection. The mice were challenged intradermally with parasites (1*106 promastigotes of L.major ) , 21 days after the final immunization . The animals were then sacrificed the spleens and serum samples were harvested for immunological analysis 3 weeks after the last immunization and 3 weeks after the challenge infection. The lymphocyte proliferation was assayed with Brdu method. Cytokines( IFN-γ, IL-4) and antibodies ( Total IgG, IgG1, IgG2a) responses were assayed with ELISA method. The parasite burden was determined by draining their spleens using the limiting dilution method. Statistical comparisons between the experimental groups were carried out using analysis of variance (ANOVA) and post hoc Turkey tests. Differences were statistically considered significant when p values were less than 0.05.
Results: This study showed that the new formulated vaccines were strong and effective in inducing the specific antibody and cellular responses and reducting the parasite burden in spleen compared to control groups .
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Fig.1 Cytokine yield [IFN- γ and IL-4] by spleen cells of BALB/c mice after immunization periods and later than challenge A B
Comparison of immune responses elicited by thiol-specific antioxidant-based protein vaccines and DNA vaccines with adjuvants in BALB /c mice infected with Leishmania major
Fatemeh Tabatabaie1* , Narges Khabazzade Tehrani 2, Somayeh Zarrati 2 , Mehdi Mahdavi 3, Abbas ali Imani fooladi4
1- Department of Parasitology and Mycology, Faculty of Medicine, Iran University of Medical Sciences, Tehran, Iran
2-Microbiology Department, Science & Research Branch, Islamic Azad University, Tehran, Iran
3- Immunology Department, Pasteur Institute of Iran, Tehran, Iran
4-Applied Microbiology, Research Center, Baqiyatallah University of Medical Science, Teheran, Iran.
*Corresponding author: Dr. Fatemeh Tabatabaie, Department of Parasitology and Mycology, Faculty of Medicine, Iran University of Medical Sciences, Tehran, Iran.
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Background and purpose: Leishmaniasis is caused by parasitic protozoa of the genus Leishmania. No effective vaccine has been developed yet against the disease. The purpose of this study was to study efficacy of DNA and protein vaccines with various adjuvants in BALB /c mice infected with Leishmania major .
Material/methods: Protein vaccines were included with the recombinant TSA protein (TSA , thiol-specific antioxidant, a main antigen of Leishmania major ) and montanide ISA 70 oil adjuvant and chitosan nanoparticle . DNA vaccines were designed with TSA recombinant plasmid and dendrimer and PMMA nanoparticles (as adjuvant) . After expression and purification of the recombinant TSA protein and preparation adjuvants , BALB/c mice were divided into six groups [TSA/ high chitosan , TSA/ low chitosan , TSA/ montanide and high chitosan, low chitosan, PBS groups] and subcutaneously immunized with 20 μg of recombinant protein TSA vaccine and 40 μg of adjuvants at three times intervals on days 0, 14 and 28.
Conclusions: Based on this results, we may suggest that the formulated vaccines are suitable candidates to undergo further studies in the context of leishmaniasis control. Keywords: Leishmaniasis , BALB/c mice , Adjuvants, DNA vaccines , Protein vaccines
Fig.2 Cytokine production [IFN- γ(A) and IL-4(B)] by the splenocytes in vaccinated and control groups
References:
Badiee A, Heravi Shargh V, Khamesipour A, Jaafari MR , Micro/nanoparticle adjuvants for antileishmanial vaccines: present and future trends. Vaccine ,2013 ,31:735–749
Ghaffarifar F, Tabatabaie F, Sharifi Z, Dalimiasl A, Zahir Hassan M, Mahdavi M, Cloning of a Recombinant Plasmid Encoding Thiol-Specific Antioxidant Antigen [TSA] Gene of Leishmania major and Expression in the Chinese Hamster Ovary Cell Line, Malays J Med Sci. 2012; 19[1]: 15-19
Zarrati S, Maleki F, Mahdavi M, Khabaz zadeh Tehrani N, Abrehdari Tafreshi Z , Asadi A.h. Tabatabaie F ,Humoral immune responses in DNA vaccine formulated with poly [methyl methacrylate] against Leishmania major ,India Journal of Entomology and Zoology Studies ,2014; 2 [5]:
Zarrati S, Mahdavi M, Tabatabaie F,Immune responses in DNA vaccine formulated with PMMA following immunization and after challenge with Leishmania major, J Parasit Dis ,2014, pp 1-9, DOI /s