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. 2016 Nov 4;7(49):80820–80841. doi: 10.18632/oncotarget.13098

Figure 12. TRAIL induces a time-dependent increase in the caspase-8 to cFLIP ratio at ligand-activated death receptors in TrkAIII SH-SY5Y but not pcDNA SH-SY5Y cells.

Figure 12

(A) Western blots demonstrating reduced cFLIP levels and increased caspase-8 levels in ligand-precipitated DR4 positive death receptor complexes from TrkAIII SH-SY5Y but not pcDNA SH-SY5Y cells treated with biotin-labelled TRAIL (500 ng/ml) for 6 hours compared to 1 hour. Input levels of cFLIP, Caspase 8 (Casp-8) and α-tubulin (α-Tub) in total cell extracts (20 μg) are provided (Input). (B) Histogram displaying differences in the c-FLIP to caspase-8 ratios in the adjacent Western blots, adjusted for differences in DR4 densitometric levels, in ligand-precipitated DR4 positive death receptors purified from pcDNA SH-SY5Y (white) and TrkAIII SH-SY5Y cells (grey), obtained by image J densitometry. (C) Western blots demonstrating similar levels of total and Y380 phosphorylated caspase 8 in NT SH-SY5Y, pcDNA SH SY5Y and TrkAIII SH-SY5Y whole cell extracts (20 μg). (D) Western blots demonstrating no reduction in the levels of Y380 phosphorylated caspase 8 in whole cell extracts (20 μg) from TrkAIII SH-SY5Y treated in the presence or absence of GW441756 (GW, 1 μM for 24 hours) or TRAIL (200 ng/ml for 12 hours), plus TRAIL-induced caspase 8 cleavage to an 18kDa Y380 phosphorylated fragment.