Frequencies of SOD1-inclusions and nuclear morphology changes after Bim knockdown.

<p>(A) Neuro2a cells were transfected with siRNA targeting Bim (siBim) or with non-specific control RNA (siC). After 24 h transfection of siRNA, cells were transfected with WT or SOD1 A4V-EGFP vectors for a further 48 h, or not further transfected (Untr). Cell lysates were subjected to Western immunoblotting and tested for binding of anti-Bim antibody and anti-GFP antibody. Western blot with β-actin antibody was used for control of equivalent protein loading. (B) Protein content of Bim corresponding to bands in the Western blots of panel A was quantified, first normalized to β-actin band intensity for the relevant lane, and then expressed as percentage of Bim measured in cells expressing control siC. (C) Quantification of prevalence of SOD1-inclusions. In each replicate experiment, 200 cells were scored for each population. (D) Quantified data for cells with apoptotic nuclei. For the populations containing untransfected cells (Untr) or cells expressing dispersed SOD1 (either WT or A4V), 200 cells were scored in each replicate experiment. For inclusion-positive cells (incl), more than 60 cells were scored in each of three replicate experiments. For panels B, C and D, results are expressed as mean ± SD, n = 3. * <i>p</i>&lt;0.05; ** <i>p</i>&lt;0.0001.