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Figure 1. Time-dependent effects of SOD1 down-regulation. A) SH-SY5Y cells were transfected with siSOD or with siScr. After transfection, total protein extracts were used for Western blot analysis: SOD1 (3 µg) and Bcl-2 (20 µg). 
-Tubulin was used as loading control. Lane 1: siScr at 12 h; lane 2: siSOD at 12 h, lane 3: siSOD at 24 h; lane 4: siSOD at 36 h; lane 5: siSOD at 48 h; lane 6: siScr at 48 h. Immunoblot shown is representative of 6 which gave similar results. B) After transfection of siRNA against SOD1, concentration of intracellular superoxide was determined by cytofluorimetric analyses after incubation with DHE. Histograms represent 6 with similar results. C) After transfection of siRNA against SOD1, time-dependent changes of SOD1, Bcl-2, 
, GSH, and superoxide were evaluated and expressed as percentage of controls. SOD1 and Bcl-2 protein content was determined by Western blot and quantified by densitometric analysis using -tubulin as loading control; GSH was measured by HPLC analysis; superoxide concentration and were determined cytofluorimetrically after staining with DHE and TMRE, respectively. All data reported were statistically significant, except values of at 12 and 24 h (n=6).
