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. 2016 May 10;7(24):36461–36473. doi: 10.18632/oncotarget.9263

Figure 3. Autophagy activation, energy and ER stress induction in cells treated with FF or 2-DG alone or in combination.

Figure 3

FF (40 uM), 2-DG (2 mM), Rapamycin (Rap) (0.1 ug/ml) alone or combination were used to treat NM2C5 cells for 24 h and western blot analysis was performed to detect protein levels of: (A) LC3B II (as an indicator of autophagy activation); (B) Grp78 and CHOP (indicating activation of UPR) and p-eIF2α (indicating ER stress as well as other possibilities, please see text), p-AMPK (energy stress) and cleaved caspase 3 (apoptosis); (C) p-P70s6k and p-4EBP1 (indicative of mTOR activity); (D) Grp78 and CHOP. β-Actin was used as a loading control in each; (E) Human osteosarcoma 143b cell line treated with either 2-DG, FF, or both, for 24 h and western blot analysis was performed to detect protein levels of Grp78, CHOP, p-AMPK and cleaved caspase 3. Representative blots of one of at least two independent experiments are shown.