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Fig. 7 | Journal of Biomedical Science

Fig. 7

From: Identification of human MHC-I HPV18 E6/E7-specific CD8 + T cell epitopes and generation of an HPV18 E6/E7-expressing adenosquamous carcinoma in HLA-A2 transgenic mice

Fig. 7

Activation of HPV18E7- or HPV18E6-specific CD8 + T cells by TC-1/AAD cells transfected with either HPV18 E7 or HPV18E6(delD70) DNA, or by HPV18 positive human cervical cancer cell line, Hela-A2 (AAD). HLA-A2 restricted HPV18 E7 peptide (aa7-15) specific CD8 + T cell line was established using splenocytes derived from HLA-A2(AAD) transgenic mice vaccinated with pcDNA3-CRT/HPV18E7 DNA and boosted with TA-HPV vaccinia. The splenocytes were incubated with irradiated TC-1/AAD cells pulsed with HPV18 E7 peptide (aa7-15) in the presence of murine IL-2. An HLA-A2 restricted HPV18 E6 peptide (aa97-105) specific CD8 + T cell line was established using splenocytes derived from HLA-A2(AAD) transgenic mice vaccinated with pcDNA-3CRT/HPV18E6(delD70) DNA and boosted with TA-HPV vaccinia. The splenocytes were incubated with irradiated TC-1/AAD cells pulsed with HPV18 E6 peptide (aa97-105) in the presence of murine IL-2. For the activation assays using 293-AAD cells, cells were transfected with either pcDNA3-CRT/HPV18E7, or pcDNA3-CRT/HPV18E6(delD70), or pBI-11 using lipofectamine 2000. The transfected cells were then co-cultured with HPV18E7 or HPV18E6 specific CD8 + T cells at the presence of GolgiPlug overnight. For the activation assays using Hela-AAD cells, Hela-AAD, or Hela cells were cocultured with HPV18 E7 or HPV18 E6 specific CD8 + T cells at the presence of GolgiPlug overnight. The cells were harvested, stained with PE-conjugated anti-mouse CD8a. After permeabilization and fixation, the cells were stained with FITC-conjugated anti-mouse IFN-γ. The cells were acquired with FACSCalibur flow cytometer and data were analyzed with CellQuest Pro software. A Representative image (left panel) and summary (right panel) of flow cytometry data from HPV18 E7 peptide (aa7-15)-specific CD8 + T cell activation by transfected 293-AAD cells. B Representative image (left panel) and summary (right panel) of flow cytometry data from HPV18 E7peptide(aa7-15)-specific CD8 + T cell activation by Hela-AAD cells. C Representative image (left panel) and summary (right panel) of flow cytometry data from HPV18 E6 peptide (aa97-105)-specific CD8 + T cell activation by transfected 293-AAD cells. D. Representative image (left panel) and summary (right panel) of flow cytometry data from HPV18 E6 peptide (aa97-105)-specific CD8 + T cell activation by Hela-AAD cells

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