FIGURE 5.

Lentiviral-mediated delivery of anti-TRIM2 shRNAmir reveals a role of TRIM2 in rapid ischemic tolerance-induced neuroprotection. A, neuronal cells were incubated with control or anti-TRIM2 shRNAmir (multiplicity of infection of 10, 48 h). Cell lysates were prepared, and TRIM2 and Bim protein levels were determined by immunoblotting. B, cells were incubated with either nonsilencing control or TRIM2-silencing shRNAmir for 48 h. Cells were then preconditioned with 30-min OGD and recovered for 1 h. Bim protein levels and MAPK phosphorylation were determined by immunoblotting. Representative immunoblots are shown for Bim and phosphorylated p42/p44 MAPK in TRIM2 knockdown cells and control cells subjected to 30-min OGD and 1-h recovery. C, cells were incubated with TRIM2 shRNAmir or nonsilencing control for 48 h. Cells were then subjected to 30-min, 120-min or 30-min followed by 1-h recovery and a subsequent 120-min OGD. Cell death was assessed by propidium iodide assay 24 h later. Data shown are mean ± S.E. (error bars), n = 5. **, p < 0.001 versus nonsilencing treated cells (two-way ANOVA with Bonferroni post hoc test).