Fig. 2. V1-1 inhibits ER-stress-mediated apoptosis. Rats were subjected to 2 hours MCAO followed by 24 hours of reperfusion (I2/R24). V1-1 and TAT control (0.2 mg/kg) were injected intraperitoneally at the onset of reperfusion. Neuro2a cells were pre-treated with V1-1 or TAT control (1 µM) for 15 minutes followed by Tm (5 µg/ml) or Tg (3 µM) treatment. (A) (Left panel) ER fractions were isolated from brain. (Right panel) After 3 minutes of Tm treatment, ER fractions were isolated from Neuro2a cells. The lysates were subjected to western blot analysis using anti-PKC antibody. Calnexin was used as internal loading control. (B) Induction of JNK phosphorylation was detected in total-cell lysates of rat brains. (C) Phosphorylation of JNK in total lysates of Neuro2a cells was detected by western blot after 24 hours of Tm (left) or 18 hours of Tg (right) treatment. (D) Phosphorylation of Jun, a downstream effector of JNK, was detected by western blotting at the indicated time points after Tm (upper) or Tg (lower) treatment. (E) TUNEL assay was carried out after 30 hours of Tm or Tg treatment. (Left panel) Representative data of three independent experiments. Original magnification was x40. (Right panel) TUNEL-positive cells are expressed as a percentage of the number of total cells, as determined by staining with Hoechst dye. In the animal study, data are expressed as mean ± s.e. from results of six rats per group. ^*P<0.05 vs TAT treatment; ^#P<0.05 vs sham-operated rats. In the cell culture study, the data are presented as mean ± s.e. of three independent experiments. ^#P<0.05 vs control, ^*P<0.05 vs Tm treatment, ^**P<0.01 vs TAT treatment.