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. Author manuscript; available in PMC: 2016 Jul 1.
Published in final edited form as: Apoptosis. 2015 Jul;20(7):996–1015. doi: 10.1007/s10495-015-1131-3

Fig. 2. TNFα/TNFR-mediated cell death in NSC and in human melanoma lines WM35, WM793 and WM9 expressing the neural stem cell markers and TNFR1/2.

Fig. 2

(A and B) Confocal analysis of images after immunostaining using rabbit polyclonal Ab to phospho-ERK (green) and monoclonal Ab to Nestin (red) or polyclonal Ab to SOX2 (green) and monoclonal Ab to Nestin (red); merged images in yellow/orange. Bar = 50 μm (C) Surface expression levels of NGFR, CD133 and TNFR1/2 in NSC and melanoma lines; ns - nonstained cells. Percentage of antigen-positive cells is indicated. (D) Western blot analysis of protein expression levels of SOX2 and NANOG, pluripotency markers, as well as signaling proteins and transcription factors in NSC (1) and specified melanoma lines (2-4). (E) Cell cycle-apoptosis analysis of WM35 and WM9 melanoma cells after treatment with cycloheximide (CHX) 1 μM, TNFα (20 ng/ml), zVAD (40 μM) alone or in combination. 48 h after treatment, cells were stained with PI and analyzed by the flow cytometry. (F) Total death levels were determined by Trypan blue exclusion assay. Pooled results of four independent experiments with indicated treatments are shown for NSC, WM35 and WM9 cells. zVAD, 40 μM and Necrostatin (Necro), 40 μM were used. Error bars represent mean ± SD (p < 0.05, Student’s t test). Stars and triangulars indicate significant differences.