Targeted Deletion of HIF-1α Gene in T Cells Prevents their Inhibition in Hypoxic Inflamed Tissues and Improves Septic Mice Survival
Figure 4
Increased NF-κB m-RNA expression and activity of ex vivo activated T cells of mice with T-cell targeted deletion of HIF-1 α.
For all three panels, T-cells from spleens were isolated from age and sex matched lck Cre (−) and lck Cre (+) HIF-1α loxP mice and stimulated as described (WT-S, κΟ−S). Unstimulated cells served as controls (WT, κΟ). A: T cell specific disruption of HIF-1 α gene substantially increases NF-κB binding activity in ex vivo TCR-activated T cells. Nuclear extracts were prepared from harvested cells and EMSA was conducted. The experiment was repeated and representative data of two experiments are shown. All lanes contain hot binding probe for NF-κB. Specificity of EMSA was tested in the presence of 50 fold excess of either unlabeled probe (Con 1) or CRE specific probe (Con 2), respectively. B: T cell specific disruption of HIF-1 α gene increases NF-κB p50 and p65 binding activity in ex vivo TCR-activated T cells. NF-κB-ELISA was conducted with nuclear extracts. *:p<0.05 vs. WT, N = 4. C: T cell specific disruption of HIF-1 α gene increases NF-κB p50 mRNA expression in ex vivo TCR-activated T cells. RNA was prepared and subsided to quantitative RT-PCR. *:p<0.01 vs. WT. N = 4.