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. Author manuscript; available in PMC: 2007 Aug 29.
Published in final edited form as: J Bone Miner Res. 2007 Jan;22(1):29–36. doi: 10.1359/JBMR.060914

FIG. 1.

FIG. 1.

Association between an increase of PKR protein expression and cell survival in MG63 osteosarcoma cells. (A) PKR expression. Cytoplasmic extracts prepared from cells treated for 24 h with vehicle (Veh) or 10 µM of 2-ME, E2, 4-OHE, and 16-OHE was analyzed by Western blot hybridization using anti-PKR antibody and anti-actin antibody (Sigma). (B) Cell survival. Cells were treated with Veh or 5 µM of 2-ME, E2, 4-OHE, and 16-OHE for 72 h. The cells were harvested, and the viable cell counts were taken after staining with trypan blue. Values are mean ± SE (N = 3 replicate cultures). *p ≤ 0.05 vs. Veh. (C) Time-course of 2-ME effects on PKR. Cytoplasmic extracts prepared from cells treated for 12, 24, and 48 h with Veh or 10 µM of 2-ME were analyzed by Western blot hybridization using anti-phospho-PKR (pT446; Epitomics), anti-PKR (Santa Cruz Biotechnology), and anti-Actin (Sigma) antibodies. (D) EIF-2α phosphorylation. Cytoplasmic extracts prepared from cells treated with Veh or 10 µM of 2-ME for 24 h were analyzed by Western blot hybridization using anit-phospho-eIF-2α (pT446; Epitomics) and anti-eIF-2α (Cell Signaling Technology).