[A multi-gene DNA vaccine encoding non-fusion membrane-anchoring antigen against Schistosoma japonicum].
OBJECTIVE
To construct DNA vaccine (pIRES-Sj97-Sj14-Sj26) and study its immunogenicity and protective immunity against Schistosoma japonicum.


METHODS
The plasmid pIRES-Sj97-Sj14-Sj26 containing fatty binding protein (Sj14), GST (Sj26) and paramyosin (Sj97) was constructed and expressed on the membrane. RT-PCR was used to detect the expression of Sj14 mRNA, Sj26 mRNA and Sj97 mRNA in the Hela cells, and IFA for detecting the expression of trans-membrane Sj14, Sj26 and Sj97. Sixty BALB/c mice were randomly divided into 3 groups. Mice in each group respectively received normal saline, pIRES blank vector and pIRES-Sj97-Sj14-Sj26 by intramuscular injection. Two weeks after the 3rd immunization, 10 mice from each group were sacrificed and total IgG in serum and the level of IFN-y were detected by ELISA and lymphocyte stimulating index (SI) by MTt. FCM was used to analyze the subgroups of splenocytes. The level of NO secreted by peritoneal macrophages was determined by nitrate reductase approaches. The left 10 mice in each group were challenged with (40 +/- 1) cercariae of S. japonicum by abdominal skin penetration. Forty-five days after challenge, mice were sacrificed, and numbers of recovered worms and hepatic eggs were counted.


RESULTS
RT-PCR showed the expression of Sj14 mRNA, Sj26 mRNA and Sj97 mRNA. IFA proved the expression of Sj26, Sj14 and Sj97 protein. Level of total IgG in the vaccination group, saline group and pIRES blank vector group was (5.62 +/- 0.64), (1.22 +/- 0.20) and (1.48 +/- 0.36) mg/ml respectively, showing a statistical significance (P < 0.01, P < 0.05). The NO level in macrophages was (321.19 +/- 18.03), (184.12 +/- 11.05) and (213.51 +/- 15.93) nmol/ml in the 3 groups respectively (P < 0.05), and the lymphocyte stimulating index in the 3 groups was (2.25 +/- 0.29), (1.18 +/- 0.07) and(1.22 +/- 0.09) respectively (P < 0.01). The INF-gamma level was higher in the vaccination group than others (P < 0.01). The percentage of CD4+ and CD8+ T cells increased considerably (P < 0.01). The vaccination group showed a worm reduction rate of 39.9% (P < 0.01) and an egg reduction rate of 43.9% (P < 0.01).


CONCLUSIONS
The vaccine candidate pIRES-Sj97-Sj14-Sj26 induces an immune protection in BALB/c mice against Schistosoma japonicum.
Tang
Cheng-wu
CW
Department of Biochemistry and Molecular Biology, TongJi Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.
Liu
Shuo-jie
SJ
Ma
Yan-Bin
YB
Liang
Liang
L
Guo
Ping
P
Wang
Shu-yu
SY
Gao
Hong
H
Duan
Qiu-hong
QH
Cheng
Ji-zhong
JZ
Dai
Wu-xing
WX
D000818:Animals; D000909:Antibodies, Helminth / Q000097:blood / Q000276:immunology; D000947:Antigens, Helminth / Q000235:genetics / Q000276:immunology*; D006367:HeLa Cells; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood / Q000276:immunology; D007371:Interferon-gamma / Q000276:immunology; D017737:Macrophages, Peritoneal / Q000276:immunology / Q000378:metabolism; D008297:Male; D008565:Membrane Proteins / Q000235:genetics / Q000276:immunology*; D051379:Mice; D008807:Mice, Inbred BALB C; D009569:Nitric Oxide / Q000378:metabolism; D010957:Plasmids; D012549:Schistosoma japonicum / Q000235:genetics / Q000276:immunology*; D014162:Transfection; D019444:Vaccines, DNA / Q000276:immunology*
D000909:Antibodies, Helminth; D000947:Antigens, Helminth; D007074:Immunoglobulin G; D008565:Membrane Proteins; D019444:Vaccines, DNA; D009569:Nitric Oxide; D007371:Interferon-gamma