Fig. 5From: Blocking of Transient Receptor Potential Vanilloid 1 (TRPV1) promotes terminal mitophagy in multiple myeloma, disturbing calcium homeostasis and targeting ubiquitin pathway and bortezomib-induced unfolded protein responseAMG9810 in combination with bortezomib elevates mitochondrial ROS and calcium levels, induces mitochondrial and lysosomal destabilization and promotes ER stress. RPMI8226 and OPM1 cells were exposed to low-dose bortezomib (BTZ) (4 nM) in the absence or presence of AMG9810 (10 µM) for 24 h. a Mitochondrial ROS levels and apoptosis were measured using MitoSOX and Annexin V staining. b Mitochondrial calcium levels were assessed using Rhod-2 staining. c Mitochondrial depolarization (ΔΨm loss) was detected by DiOC6 staining and flow cytometry analysis. d Lysosomal membrane destabilization was assessed using LysoTracker staining. Representative images with RPMI8226 cells are shown in upper panels. Bars showing mean of triplicates ± SD (**p < 0.01) in RPMI8226 and OPM-1 cells are presented in lower panels. e RPMI8226, CAG and OPM-1 cells were treated with bortezomib (5 nM), AMG9810 (10 µM) or their combination for 24 h. RNA was extracted and was subjected to qRT-PCR analysis. CHOP, GADD34 and HO-1 mRNA levels were evaluated, using β2-microglobulin as endogenous control gene. f Western blot analysis of ER stress response chaperons HSP70 and HSP40 in RPMI826, CAG and OPM-1 cells before and after 24-h treatment with bortezomib (5 nM), AMG9810 (10 µM) or their combination. β-Actin was used as internal control. Representative data from at least two independent experiments are shownBack to article page