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. 2003 Sep;110(1):10–17. doi: 10.1046/j.1365-2567.2003.01710.x

Figure 5.

Figure 5

Retroviral transduction of antisense BNIP3 RNA diminishes effector CTL apoptosis. P14 splenocytes were stimulated with gp33–41 peptide and 2 days later, were infected with retroviruses expressing GFP alone (GFP-RV) or expressing GFP along with sense or antisense BNIP3 RNA. The cells were cultured for 4 days in the presence of 10 ng/ml IL-2, stimulated with 5 μg/ml plate-bound αCD3 for 6 hr and stained with CD8-Cy5 and AnnexinV-PE and examined by flow cytometry. In (a), gates set to analyse GFP and AnnexinV positivity are shown using GFP-RV transfected (filled) or untransfected (open) P14 cells and annexin-stained (filled) and isotype control antibody-stained (open) P14 effectors stimulated with αCD3. (b) shows AnnexinV staining on retrovirally nontransduced (GFP−, top panel) and transduced (GFP+, bottom panel) CD8-gated cells. The percentage of AnnexinV+ cells is indicated. (c) shows RT-PCR analysis for BNIP3 gene expression using sorted GFP-RV and GFP-RV-BNIP3 antisense retrovirally transduced P14 cells. Densitometric analysis after normalization to β-actin expression revealed a 2·3-fold decrease in BNIP3 gene expression in GFP-RV BNIP3 antisense transduced cells compared to GFP-RV vector transduced cells.