J Korean Med Sci.  2005 Dec;20(6):1039-1045. 10.3346/jkms.2005.20.6.1039.

The Effects of Intradermal Vaccination with DNA Encoding for the T-cell Receptor on the Induction of Experimental Autoimmune Encephalomyelitis in B10.PL Mice

• Kwon SS
• Kim N
• Yoo TJ

• KMID: 2157759
• DOI: http://doi.org/10.3346/jkms.2005.20.6.1039

Abstract

Intradermal gene administration was found to induce a more profound immune response than direct intramusclular gene injection. We performed intradermal vaccination of B10.PL mice with DNA encoding for the V 8.2 region of the T-cell receptors (TCR). Three weeks later, these mice were immunized with rat myelin basic protein (MBP). Daily mean clinical scores and mortality rate were lower in this group compared with controls. The proliferative responses of lymph node cells to rat MBP were slightly less in the vaccination groups than in the control groups (p<0.05). However, we detected no differences between the two groups with regard to the production of MBP-specific IgG, IgG1, & IgG2a antibodies. The levels of cytokine mRNA expression in the vaccination groups were observed higher than in the control groups without antigen-specific stimulation, but all of cytokine expressions between the vaccination and control groups after antigen-specific stimulation were identical. These results demonstrate that intradermal DNA vaccines encoding for TCR might prove to be useful in the control of autoimmune disease.

Keyword

Vaccines, DNA; Encephalomyelitis, Autoimmune, Experimental; Injections, Intradermal; Receptors, Antigen, T-Cell; Vaccination

MeSH Terms

Animals
Autoantibodies/blood
Base Sequence
Cytokines/genetics
DNA, Complementary/genetics
Encephalomyelitis, Autoimmune, Experimental/etiology/immunology/*prevention and control
Female
Gene Expression
*Genes, T-Cell Receptor beta
In Vitro
Injections, Intradermal
Lymphocyte Activation
Mice
Myelin Basic Proteins/immunology
RNA, Messenger/genetics/metabolism
Rats
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Vaccines, DNA/*administration and dosage/genetics

Figure

• 

  Fig. 1 Preventive effects of DNA vaccination with TCR Vβ8.2 in mice after active immunization with rat MBP on the induction of EAE disease. The vaccination group (n=20) was intradermally vaccinated with DNA encoding for the Vβ8.2 of the T-cell receptor, three times at weekly intervals. The control group (n=20) was injected with pcDNA 3.1 blank vector. After 3 weeks, all mice were immunized with rat MBP. The clinical symptoms of EAE were monitored and scored daily, as described in the Methods section. The control group developed higher daily mean clinical score than the vaccination group between days 20-25 (1.50±0.92, 1.63±0.74, 2.25±1.28, 3.0±1.07, 3.38±1.19, 3.5±1.41 vs. 0.13±0.35, 0.38±0.52, 0.50±0.76, 1.00±1.07, 1.25±1.06, 1.13±1.36, p<0.05). There was a dramatic reduction in mortality in the vaccination group compared with controls; eight of sixteen mice died in the control group, compared with none in the vaccination gorup (p<0.05).

• 

  Fig. 2 The IgG1 (A), IgG2a (B), and IgG (C) antibody responses of the B10.PL mice immunized with rat MBP with DNA coding for Vβ8.2 or pcDNA only. Intradermal vaccination with Vβ8.2 DNA resulted in no different response with regard to MBP-specific IgG1 antibody, as compared with the control group, at 2 and 3 weeks post-immunization with whole MBP (p>0.05). Also, with regard to the response of MBP-specific IgG2a and IgG antibodies, the vaccination group did not significantly increase antibody production, in comparison with the control group (p>0.05). Sera of eigth mice in each group were collected three times on weeks 0, 2, and 3, after immunization. Data are expressed as means±S.D.

• 

  Fig. 3 Cytokine gene expression. Four mice from each group were sacrificed 10 days-after MBP immunization. Lymph node cells (1×107 cells per well) were cultured either without antigen, or with rat MBP (100 µg/mL) in vitro, for 18 hr. The cells were washed with PBS buffer and the total RNA was extracted using TRIzol reagent. Using murine leukemia virus reverse transcriptase and random hexanucleotide primer, according to the instructions of the Perkin Elmer Gene Amp RNA PCR kit, first-strand cDNA was generated from 1 µg of total RNA and subjected to RT-PCR analysis. RT-PCR reactions were then conducted using cDNA with different primers, specific for β-actin, IL-2, γ-interferon, IL-4, and IL-5. (1) Lymph node cells from the control mice were cultured without antigen, (2) or in the presence of rat MBP (100 µg/mL). (3) Lymph node cells from the vaccinated mice were cultured without antigen, (4) or in the presence of rat MBP (100 µg/mL). The levels of mRNA expression of IFN-γ, IL-2, IL-4, & IL-5 in the vaccination groups were observed higher than in the control groups without antigen-specific stimulation, but there were no differences all of the cytokine expressions between the two groups after antigen-specific stimulation.

• 

  Fig. 4 Lymph node cell proliferation 10 days after immunization, the lymph nodes of four mice of each group were removed aseptically, and single-cell suspensions were prepared. The cells (2×105 cells per well) were then cultured with serial dilutions of rat MBP (range, 0.01-100 µg/mL). After 4 days of culturing, 1 µCi of [3H] thymidine was added to each well. 18 hr later, the cells were harvested, and measured via liquid scintillation counting. The vaccination group's proliferative responses to MBP of lymph node cells was found to be significantly less pronounced than in the control group (p<0.05 at antigen concentrations 0.01-100 µg/mL, except 10 µg/mL). The values were expressed in counts per minute, as follows: Counts per minute with antigen - Counts per minute without antigen. *p<0.05 vaccination group compared with control group.

Reference

1. Mor F, Kantorowitz M, Cohen IR. The dominant and the cryptic T cell repertoire to myelin basic protein in the Lewis rat. J Neurosci Res. 1996. 45:670–679.

2. Cheng KC, Chiang HJ, Wang K, Krug MS, Yoo TJ, Hood L. TCRV and TCRJ gene usage in MBP responding T cells from (B10.PL×PL/J) F1 mice is biased towards that of B10.PL mice. J Neuroimmunol. 1997. 80:13–22.

3. Matsumoto Y, Tsuchida M, Hanawa J, Abo T. T cell receptor peptide therapy for autoimmune encephalomyelitis: stronger immunization is necessary for effective vaccination. Cell Immunol. 1994. 153:468–478.

4. Bot A, Bot S, Antohi S, Karjalainen K, Bona C. Kinetics of generation and persistence on membrane class II molecules of a viral peptide expressed on foreign and self proteins. J Immunol. 1996. 157:3436–3442.

5. Raz E, Carson DA, Parker SE, Parr TB, Abai AM, Aichinger G, Gromkowski SH, Singh M, Lew D, Yankauckas MA, Baird SM, Rhodes GH. Intradermal gene immunization: The possible role of DNA uptake in the induction of cellular immunity to viruses. Proc Natl Acad Sci USA. 1994. 91:9519–9523.

6. Kwon SS, Kim N, Yoo TJ. The effect of vaccination with DNA encoding murine T-cell epitopes on the Der p 1 and 2 induced immunoglobulin E synthesis. Allergy. 2001. 56:741–748.

7. Waisman A, Ruiz PJ, Hirschberg DL, Gelman A, Oksenberg JR, Brocke S, Mor F, Cohen IR, Steinman L. Suppressive vaccination with DNA encoding a variable region gene of the T-cell receptor prevents autoimmune encephalomyelitis and activates Th2 immunity. Nature Med. 1996. 2:857–859.

8. Steinman RM. The dendritic cell system and its role in immunogenicity. Annu Rev Immunol. 1991. 9:271–296.

9. Smith ME. An in vitro system for the study of myelin synthesis. J Neurochem. 1969. 16:83–92.

10. Watanabe T, Cheng KC, Krug MS, Yoo TJ. Brain stem auditory-evoked potentials of mice with experimental allergic encephalomyelitis. Ann Otol Rhinol Laryngol. 1996. 105:905–915.

11. Chen KC, Lee KM, Krug MS, Watanabe T, Suzuki M, Choe IS, Yoo TJ. Houst dust mite-induced sensitivity in mice. J Allergy Clin Immunol. 1998. 101:51–59.

12. Gorelik L, Flavell RA. Abrogation of TGF-beta signaling in T cells leads to spontaneous T cell differentiation and autoimmune disease. Immunity. 2000. 12:171–181.

13. Wolff JA, Malone RW, Williams P, Chong W, Acsadi G, Jani A, Felgner PL. Direct gene transfer into mouse muscle in vivo. Science. 1990. 247:1465–1468.

14. Martin R, McFarland HF. Immunological aspects of experimental allergic encephalomyelitis and multiple sclerosis. Crit Rev Clin Lab Sci. 1995. 32:121–182.

15. Lobell A, Weissert R, Storch MK, Svanholm C, de Graaf KL, Lassmann H, Andersson R, Olsson T, Wigzell H. Vaccination with DNA encoding an immunodominant myelin basic protein peptide targeted to Fc receptor of immunoglobulin G suppresses experimental autoimmune encephalomyelitis. J Exp Med. 1998. 187:1543–1548.

16. Genain CP, Abel K, Belmar N, Villinger F, Rosenberg DP, Linington C, Raine CS, Hauser SL. Late complications of immune deviation therapy in a nonhuman primate. Science. 1996. 274:2054–2057.

17. Marusic S, Tonegawa S. Tolerance induction and autoimmune encephalomyelitis amelioration after administration of myelin basic protein-derived peptide. J Exp Med. 1997. 186:507–515.

18. Singer GG, Abbas AK. The Fas antigen is involved in peripheral but not thymic deletion of T lymphocytes in T cell receptor transgenic mice. Immunity. 1994. 1:365–371.

19. Tighe H, Corr M, Roman M, Raz E. Gene vaccination: plasmid DNA is more than just a blueprint. Immunol Today. 1998. 19:89–97.

20. Kumar V, Sercarz E. Self-determinant selection and selective regulation. Chem Immunol. 1995. 60:1–19.

21. Furtado GC, Olivares-Villagomez D, Curotto de Lafaille MA, Wensky AK, Latkowski J, Lafaille JJ. Regulatory T cells in spontaneous autoimmune encephalomyelitis. Immunol Rev. 2001. 182:122–134.

22. Mathisen PM, Tuohy VK. Gene therapy in experimental autoimmune encephalomyelitis. J Clin Immunol. 2000. 20:327–333.

23. Jiang H., Curran S, Ruiz-Vazquez E, Liang B, Winchester R, Chess L. Regulatory CD8+ T cells fine-tune the myelin basic protein-reactive T cell receptor Vβrepertoire during experimental autoimmune encephalomyelitis. Proc Natl Acad Sci USA. 2003. 100:8378–8383.

24. Braciak TA, Pedersen B, Chin J, Hsiao C, Ward ES, Maricic I, Jahng A, Graham FL, Gauldie J, Sercarz EE, Kumar V. Protection against experimental autoimmune encephalomyelitis generated by a recombinant adenovirus vector expressing the Vβ 8.2 TCR is disrupted by coadministration with vectors expressing either IL-4 or IL-10. J Immunol. 2003. 170:765–774.

25. Wefer J, Harrisa RA, Lobell A. Protective DNA vaccination against experimental autoimmune encephalomyelitis is associated with induction of IFN-β. J Neuroimmunol. 2004. 149:66–76.