Fig. 3

Hes and Nar activate ER-stress and apoptosis. A Venn diagram reporting the differentially expressed genes induced by the two treatments and their comparison. B The plot shows the Gene Ontology (GO) enrichment of the upregulated genes after Hes and Nar treatment. The size of the dot indicates the number of upregulated genes enriched in each pathway, while the color of the dot indicates the p-adjusted value. C q-RT-PCR analysis of ATF4, PERK and sXBP1 mRNAs, in AMO cells, 24 h after treatment with 250 µM of either Hes or Nar. Histogram bars represent the average ± SD of mRNA expression levels after normalization with GAPDH and ΔΔCt calculations. D WB analysis of ATF4, IRE1α, p-EIF2α, PARP and Caspase 3 (right panel) proteins in AMO cells, 48 h after Hes or Nar treatment; normalization was performed using GAPDH or α-tubulin as loading controls. E FACS analysis of Annexin V-positive AMO and AMO-BZB cells, 48 h after Hes or Nar exposure. Data are representative of at least three independent biological replicates (n = 3). Histogram bars reported the percentage of total apoptotic cells. *p < 0.05. F FACS analysis of Annexin V-positive cells, 48 h after treatment with Hes (250 µM) alone or in combination with Z-VAD (20 µM). Data are representative of at least three independent biological replicates (n = 3). Histogram bars report the percentage of apoptotic MM cells. *p < 0.05