Immune Netw.  2012 Jun;12(3):118-125. 10.4110/in.2012.12.3.118.

Requirement of CD4 Help for Induction of CD8 T Cell Response Specific for Virally Derived H60

• Ryu SJ
• Kang B
• Kim SH
• Kim TW
• Chang J
• Choi EY

• KMID: 2168010
• DOI: http://doi.org/10.4110/in.2012.12.3.118

Abstract

CD40-CD40L-mediated help from CD4 T cells is essential to induce primary CD8 T cell responses specific to the non-inflammatory cell-based antigen H60. In this study, using H60 as a model antigen, we generated recombinant vaccinia viruses (rVVs) expressing the H60 CD8 epitope and investigated whether CD4 help was required to activate the CD8 T cell response specific to the virally expressed H60. The immune response after infection with rVVs expressing H60 was similar to that after immunization with H60 congenic splenocytes, with a peak frequency of H60-specific CD8 T cells detected in the blood on day 10 post-infection. A CD8 T cell response specific for virally derived H60 was not induced in CD4-depleted mice, but was in CD40-deficient mice. These results provide insights into the characterization of the CD8 T cell response specifically for antigens originating from cellular sources compared to viral sources.

Keyword

Recombinant vaccinia virus; Viral epitope; CD4 help; CD8 T cell response

MeSH Terms

Animals
Immunization
Mice
T-Lymphocytes
Vaccinia virus

Figure

• 

  Figure 1 Generation of recombinant vaccinia viruses expressing the H60 CD8 epitope. (A) To generate a recombinant vaccinia virus expressing the H60 CD8 epitope (rVV-H6039), a minigene corresponding to a region encompassing a leader signal peptide to the epitope sequences (39~46 amino acids) of H60 was cloned into a pIRES vector. The DNA fragment encompassing the leader signal to the IRES region containing a multi-cloning site was cloned into a pSC11 vector. rVV-H6039+HY-Dby608 expressed the HY-Dby CD4 epitope peptide (608~622 aa) with an additional Met at the N-terminal, as well as the H60 CD8 epitope. This was achieved by cloning the corresponding minigene into the cloning site located after the IRES region of the rVV-H6039-pSC11 vector. (B) Female B6 mice were immunized with different amounts of rVV-H6039+HY-Dby608 and rVV-H6039 in order to determine the optimal titer of virus needed for the induction of H60-specific CD8 T cell responses, and to compare the ability of each virus to induce the specific immune response. Day 7 post-infection PBLs from immunized mice were stained with H60-Tet-PE, CD11a-FITC, and CD8-APC and analyzed by flow cytometry. Data from the flow cytometry analysis are shown here after gating on CD8+ cells.

• 

  Figure 2 Kinetics of the immune response after infection with rVV-H6039+HY-Dby608. Female B6 mice were immunized with rVV-H6039+HY-Dby608 or male H60 congenic splenocytes and periodic retro-orbital blood samples were obtained. PBLs were stained with H60-Tet-PE, CD11a-FITC, and CD8-APC for flow cytometry. The frequencies of H60-tetramer binding cells in the peripheral blood CD8 T cells at each time point are plotted as kinetics curves. Representative data from flow cytometry performed on day 10 post-immunization are shown after gating on CD8+ cells. These data were obtained from two independent experiments.

• 

  Figure 3 Dependency of the CD8 T cell response to a virally expressed H60 epitope on CD4 help. Female B6 mice were treated with GK1.5 (anti-CD4 mAb) or Rat-IgG (control) three days prior to infection with rVV-H6039+HY-Dby608. PBLs taken periodically from the two different immunized groups were stained with H60-Tet-PE, CD11a-FITC, and CD8-APC. Representative flow cytometry data obtained on day 10 post-infection (peak point) are shown. The peak frequencies observed from four individual mice in each group are plotted. These data were obtained from two independent experiments.

• 

  Figure 4 The CD8 T cell response specific for virally expressed H60 is independent of CD40. This figure shows female CD40-deficient and wild type B6 mice immunized with rVV-H6039+HY-Dby608 (A) and female CD40-deficient and wild type B6 mice immunized with male H60 congenic splenocytes (B). PBLs taken periodically from immunized mice were stained with H60-Tet-PE, CD11a-FITC, and CD8-APC. Representative flow cytometry data obtained on day 10 post-infection (peak point) are shown. The peak frequencies observed from five individual mice in each group are plotted. These data were obtained from two independent experiments.

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