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Journal of Zhejiang University SCIENCE B 2009 Vol.10 No.7 P.512~521


Protective effect of DNA-mediated immunization with liposome-encapsulated GRA4 against infection of Toxoplasma gondii

Author(s):  Rui CHEN, Shao-hong LU, Qun-bo TONG, Di LOU, Dong-yan SHI, Bing-bing JIA, Guo-ping HUANG, Jin-fu WANG

Affiliation(s):  College of Life Sciences, Zhejiang University, Hangzhou 310058, China; more

Corresponding email(s):   wjfu@zju.edu.cn

Key Words:  DNA vaccine, Granule protein 4 (GRA4), Liposome, Toxoplasma gondii

Rui CHEN, Shao-hong LU, Qun-bo TONG, Di LOU, Dong-yan SHI, Bing-bing JIA, Guo-ping HUANG, Jin-fu WANG. Protective effect of DNA-mediated immunization with liposome-encapsulated GRA4 against infection of Toxoplasma gondii[J]. Journal of Zhejiang University Science B, 2009, 10(7): 512~521.

@article{title="Protective effect of DNA-mediated immunization with liposome-encapsulated GRA4 against infection of Toxoplasma gondii",
author="Rui CHEN, Shao-hong LU, Qun-bo TONG, Di LOU, Dong-yan SHI, Bing-bing JIA, Guo-ping HUANG, Jin-fu WANG",
journal="Journal of Zhejiang University Science B",
publisher="Zhejiang University Press & Springer",

%0 Journal Article
%T Protective effect of DNA-mediated immunization with liposome-encapsulated GRA4 against infection of Toxoplasma gondii
%A Shao-hong LU
%A Qun-bo TONG
%A Dong-yan SHI
%A Bing-bing JIA
%A Guo-ping HUANG
%A Jin-fu WANG
%J Journal of Zhejiang University SCIENCE B
%V 10
%N 7
%P 512~521
%@ 1673-1581
%D 2009
%I Zhejiang University Press & Springer
%DOI 10.1631/jzus.B0820300

T1 - Protective effect of DNA-mediated immunization with liposome-encapsulated GRA4 against infection of Toxoplasma gondii
A1 - Rui CHEN
A1 - Shao-hong LU
A1 - Qun-bo TONG
A1 - Di LOU
A1 - Dong-yan SHI
A1 - Bing-bing JIA
A1 - Guo-ping HUANG
A1 - Jin-fu WANG
J0 - Journal of Zhejiang University Science B
VL - 10
IS - 7
SP - 512
EP - 521
%@ 1673-1581
Y1 - 2009
PB - Zhejiang University Press & Springer
ER -
DOI - 10.1631/jzus.B0820300

The dense granule protein 4 (GRA4) is a granular protein from Toxoplasma gondii, and is a candidate for vaccination against this parasite. In this study, the plasmid pcDNA3.1-GRA4 (pGRA4), encoding for the GRA4 antigen, was incorporated by the dehydration-rehydration method into liposomes composed of 16 mmol/L egg phosphatidylcholine (PC), 8 mmol/L dioleoyl phosphatidylethanolamine (DOPE), and 4 mmol/L 1,2-diodeoyl-3-(trimethylammonium) propane (DOTAP). C57BL/6 mice and BALB/c mice were immunized intramuscularly three times with liposome-encapsulated pGRA4 to determine whether DNA immunization could elicit a protective immune response to T. gondii. Enzyme-linked immunosorbent assay (ELISA) of sera from immunized mice showed that liposome-encapsulated pGRA4 generated high levels of IgG antibodies to GRA4. Production of primary interferon (IFN)-γ and interleukin (IL)-2 in GRA4-stimulated splenocytes from vaccinated mice suggested a modulated Th1-type response. 72.7% of C57BL/6 mice immunized with liposome-encapsulated pGRA4 survived the challenge with 80 tissue cysts of ME49 strain, whereas C57BL/6 mice immunized with pGRA4 had only a survival rate of 54.5%. When immunized BALB/c mice were intraperitoneally challenged with 103 tachyzoites of the highly virulent RH strain, the survival time of mice immunized with liposome-encapsulated pGRA4 was markedly longer than that of other groups. Our observations show that liposome-encapsulated pGRA4 enhanced the protective effect against infection of T. gondii.

Darkslateblue:Affiliate; Royal Blue:Author; Turquoise:Article


[1] Aosai, F., Mun, H.S., Norose, K., Chen, M., Hata, H., Kobayashi, M., Kiuchi, M., Stauss, H.J., Yano, A., 1999. Protective immunity induced by vaccination with SAG1 gene-transfected cells against Toxoplasma gondii infection in mice. Microbiology and Immunology, 43(1): 87-91.

[2] Bivas-Benita, M., Laloup, M., Versteyhe, S., Dewitc, J., de Braekeleerc, J., Jongertc, E., Borchard, G., 2003. Generation of Toxoplasma gondii GRA1 protein and DNA vaccine loaded chitosan particles: preparation, characterization, and preliminary in vivo studies. International Journal of Pharmaceutics, 266(1-2):17-27.

[3] Brown, C.R., McLeod, R., 1990. Class I MHC genes and CD8+ T cells determine cyst numbers in Toxoplasma gondii infection. The Journal of Immunology, 145(10): 3438-3441.

[4] Buxton, D., Innes, E.A., 1995. A commercial vaccine for ovine toxoplasmosis. Parasitology, 110(s1):S11-S16.

[5] Buxton, D., Thomson, K., Maley, S., Wright, S., Bos, H.J., 1991. Vaccination of sheep with a live incomplete strain (S48) of Toxoplasma gondii and their immunity to challenge when pregnant. Vet. Rec., 129(5):89-93.

[6] Caplen, N.J., Alton, E.W., Middleton, P.G., Dorin, J.R., Stevenson, B.J., Gao, X., Durham, S.R., Jeffery, P.K., Hodson, M.E., Coutelle, C., 1995. Liposome-mediated CFTR gene transfer to the nasal epithelium of patients with cystic fibrosis. Nature Medicine, 1(1):39-46.

[7] Chen, H.F., Chen, G.J., Zheng, H., Guo, H., 2003. Induction of immune responses in mice by vaccination with liposome-entrapped DNA complexes encoding Toxoplasma gondii SAG1 and ROP1 genes. Chinese Medicine Journal, 116(10):1561-1566.

[8] Chen, R., Lu, S.H., Lou, D., Lin, A.F., Zeng, X.J., Ding, Z.Y., Wen, L.Y., Ohta, N., Wang, J.F., Fu, C., 2008. Evaluation of a rapid ELISA technique for detection of circulating antigens of Toxoplasma gondii. Microbiology and Immunology, 52(3):180-187.

[9] Cong, H., Gu, Q.M., Hong, E.Y., Wang, J.W., Zhao, Q.L., Zhou, H.Y., Li, Y., Zhang, J.Q., 2008. Multi-epitope DNA vaccine linked to the A(2)/B subunit of cholera toxin protect mice against Toxoplasma gondii. Vaccine, 26(31):3913-3921.

[10] Couper, K.N., Nielsen, H.V., Petersen, E., Roberts, F., Roberts, C.W., Alexander, J., 2003. DNA vaccination with the immunodominant tachyzoite surface antigen (SAG-1) protects against adult acquired Toxoplasma gondii infection but does not prevent maternofetal transmission. Vaccine, 21(21-22):2813-2820.

[11] Denkers, E.Y., Gazzinelli, R.T., 1998. Regulation and function of T-cell-mediated immunity during Toxoplasma gondii infection. Clinical Microbiology Reviews, 11(4):569-588.

[12] Desolme, B., Mévélec, M.N., Buzoni-Gatel, D., Bout, D., 2000. Induction of protective immunity against toxoplasmosis in mice by DNA immunization with a plasmid encoding Toxoplasma gondii GRA4 gene. Vaccine, 18(23): 2512-2521.

[13] Fachado, A., Rodriguez, A., Angel, S.O., Pinto, D., Vila, I., Acosta, A., Amendoeira, R.M., Lannes-Vieira, J., 2003. Protective effect of a naked DNA vaccine cocktail against lethal toxoplasmosis in mice. Vaccine, 21(13-14): 1327-1335.

[14] Feigin, R.D., Cherry, J.D., 1998. Toxoplasmosis. In: Bergelson, J. (Ed.), Textbook of Pediatric Infectious Diseases, 4th Ed. WB Saunders, Philadelphia, p.2921.

[15] Gaku, S., Takachika, H., Yoko, N., Kenji, S., Kohich, I., Yoshiki, S., Hidemi, T., Mitsuo, H., Jun, K., Hiroshi, K., 2003. HIV mucosal vaccine: nasal immunization with gp160-encapsulated hemagglutinating virus of Japan-liposome induces antigen-specific CTLs and neutralizing antibody responses. The Journal of Immunology, 170:495-502.

[16] Golkar, M., Shokrgozar, M.A., Rafati, S., Musset, K., Assmar, M., Sadaie, R., Cesbron-Delauw, M.F., Mercier, C., 2007. Evaluation of protective effect of recombinant dense granule antigens GRA2 and GRA6 formulated in monophosphoryl lipid A (MPL) adjuvant against Toxoplasma chronic infection in mice. Vaccine, 25(21):4301-4311.

[17] Gregoriadis, G., Saffie, R., Hart, S.L., 1996a. High yield incorporation of plasmid DNA within liposomes: effect on DNA integrity and transfection. Journal of Drug Targeting, 3(6):469-475.

[18] Gregoriadis, G., Gursel, I., Gursel, M., McCormack, B., 1996b. Liposomes as immunological adjuvants and vaccine carriers. Journal of Controlled Release, 41(1-2):49-56.

[19] Gregoriadis, G., Bacon, A., Caparros-Wanderley, W., McCormack, B., 2002. A role for liposomes in genetic vaccination. Vaccine, 20(S5):B1-B9.

[20] Happe, S., Fischer, A., Heese, C.H., Reichelt, D., Gruneberg, U., Freund, M., 2002. HIV-associated cerebral toxoplasmosis: review and retrospective analysis of 36 patients. Der Nervenarzt, 73(12):1174-1178.

[21] Ismael, A.B., Sekkai, D., Collin, C., Bout, D., Mévélec, M.N., 2003. The MIC3 gene of Toxoplasma gondii is a novel potent vaccine candidate against toxoplasmosis. Infection and Immunity, 71(11):6222-6228.

[22] Khan, I.A., Ely, K.H., Kasper, L.H., 1991. A purified parasite antigen (P30) mediates CD8+ T-cell immunity against fatal Toxoplasma gondii infection in mice. The Journal of Immunology, 147:3501-3506.

[23] Kirby, C., Gregoriadis, G., 1984. Dehydration-rehydration vesicles: a simple method for high yield drug entrapment in liposomes. Biotechnology, 2(11):979-984.

[24] Leyva, R., Herion, P., Saavedra, R., 2001. Genetic immunization with plasmid DNA coding for the ROP2 protein of Toxoplasma gondii. Parasitology Research, 87(1):70-79.

[25] Liu, F., Yang, J., Huang, L., Liu, D., 1996. New cationic lipid formulations for gene transfer. Pharmaceutical Research, 13(12):1856-1860.

[26] Martin, V., Supanitsky, A., Echeverria, P.C., Litwin, S., Tanos, T., de Roodt, A.R. Guarnera, E.A., Angel, S.O., 2004. Recombinant GRA4 or ROP2 protein combined with alum or the gra4 gene provides partial protection in chronic murine models of toxoplasmosis. Clinical and Diagnostic Laboratory Immunology, 11(4):704-710.

[27] Mévélec, M.N., Chardés, T., Mercereau-Puijalon, O., Bourguin, I., Achbarou, A., Dubremetz, J.F., Bout, D., 1992. Molecular cloning of GRA4, a Toxoplasma gondii dense granule protein, recognized by mucosal IgA anti-bodies. Molecular and Biochemical Parasitology, 56(2):227-238.

[28] Mévélec, M.N., Merecerau-Puijalon, O., Buzoni-Gatel, D., Bourguin, I., Chardés, T., Dubremetz, J.F., Bout, D., 1998. Mapping of B epitopes in Gra4, a dense granule antigen of Toxoplasma gondii and protection studies using recombinant proteins administered by the oral route. Parasite Immunology, 20:183-195.

[29] Mévélec, M.N., Bout, D., Desolme, B., Marchand, H., Magné, R., Bruneel, O., Buzoni-Gatel, D., 2005. Evaluation of protective effect of DNA vaccination with genes encoding antigens GRA4 and SAG1 associated with GM-CSF plasmid, against acute, chronical and congenital toxoplasmosis in mice. Vaccine, 23(36):4489-4499.

[30] Montgomery, D.L., Ulmer, J.B., Donnelly, J.J., Liu, M.A., 1997. DNA vaccines. Pharmacology & Therapeutics, 74(2):195-207.

[31] Nigro, M., Gutierrez, A., Hoffer, A.M., Clemente, M.H., Kaufer, F., Carral, L., Martin, V., Guarnera, E.A., Angel, S.O., 2003. Evaluation of Toxoplasma gondii recombinant proteins for the diagnosis of recently acquired toxoplasmosis by an immunoglobulin G analysis. Diagnostic Microbiology and Infectious Disease, 47(4): 609-613.

[32] Parker, S.J., Roberts, C.W., Alexander, J., 1991. CD8+ T cells are the major lymphocyte subpopulation involved in the protective immune response to Toxoplasma gondii in mice. Clinical and Experimental Immunology, 84(2): 207-212.

[33] Perrie, Y., Gregoriadis, G., 2000. Liposome-entrapped plasmid DNA: characterisation studies. Biochimica et Biophysica Acta (BBA), 1475(2):125-132.

[34] Perrie, Y., Frederik, P.M., Gregoriadis, G., 2001. Liposome mediated DNA vaccine: the effect of vesicle composition. Vaccine, 19(23-24):3301-3310.

[35] Perrie, Y., McNeil, S., Vangala, A., 2003. Liposome-mediated DNA immunisation via the subcutaneous route. Journal of Drug Targeting, 11(8-10):555-563.

[36] Remington, J.S., Mcleod, R., Thulliez, P., Desmonts, G., 2006. Infection Diseases of the Fetus and Newborn Infant, 6th Ed. Elsevier Saunders, Philadelphia, p.947-1091.

[37] Rolland, A.P., 1998. From genes to gene medicines: recent advances in nonviral gene delivery. Critical Reviews in Therapeutic Drug Carrier Systems, 15(2):143-198.

[38] Schwartzman, J.D., 2001. Toxoplasmosis. Current Infectious Disease Reports, 3(1):85-89.

[39] Stanley, A.C., Buxton, D., Innes, E.A., Huntley, J.F., 2004. Intranasal immunisation with Toxoplasma gondii tachyzoite antigen encapsulated into PLG microspheres induces humoral and cell-mediated immunity in sheep. Vaccine, 22(29-30):3929-3941.

[40] Tenter, A.M., Heckeroth, A.R., Weiss, L.M., 2000. Toxoplasma gondii from animals to humans. International Journal for Parasitology, 30(12-13):1217-1258.

[41] Tighe, H.M., Roman, C.M., Raz, E., 1998. Gene vaccination: plasmid DNA is more than just a blueprint. Immunology Today, 19(2):89-97.

[42] Ulmer, J.B., Donnelly, J.J., Parker, S.E., Rhodes, G.H., Felgner, P.L., Darki, V.J., Gromkowski, S.H., Deck, R.R., DeWitt, C.M., Friedman, A., et al., 1993. Heterologous protection against influenza by injection of DNA encoding a viral protein. Science, 259(5102):1745-1749.

[43] Vercammen, M., Scorza, T., Huygen, K., de Braekeleer, J., Diet, R., Jacobs, D., 2000. DNA vaccination with genes encoding Toxoplasma gondii antigens GRA1, GRA7, and ROP2 induces partially protective immunity against lethal challenge in mice. Infection and Immunity, 68(1): 38-45.

[44] Wheeler, C.J., Felgner, P.L., Tsai, Y.J., Marshall, J., Sukhu, L., Doh, S.G., Hartikka, J., Nietupski, J., Manthorpe, M., Nichols, M., et al., 1996. A novel cationic lipid greatly enhances plasmid DNA delivery and expression in mouse lung. Proceedings of the National Academy of Sciences USA, 93(21):11454-11459.

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