Fig. 2

TRPV1 inhibition using AMG9810 induces increase in mitochondrial calcium levels and mitochondrial ROS levels in MM cells. MM cells were treated with AMG9810 (10 µM), with or without BAPTA-AM (5 µM) for 1 h, and mitochondrial calcium and mitochondrial ROS levels were evaluated. a RPMI8226 cells were labeled with fluorescent calcium indicator Rhod-2 (red fluorescence). To ensure the localization of mitochondria, cells were stained for mitochondrial marker COX IV (green fluorescence). The nucleus was stained with DAPI (blue fluorescence). Representative microscopy images show mitochondrial calcium, × 40 magnification. b, c RPMI8226, CAG and OPM-1 cells were treated with AMG9810 (10 µM), with or without BAPTA-AM (5 µM) for 1 h, stained with Rhod-2 and analyzed by flow cytometry. b Representative plots showing mitochondrial calcium change in response to various treatments, using the mean fluorescent intensity (MFI) of RhoD2 staining in RPMI8226 cells. c Mitochondrial calcium level in RPMI8226, CAG and OPM1 cells is presented as MFI of Rhod-2-stained cells. Data are presented as mean of triplicates ± SD (**p < 0.01). d, e Mitochondrial ROS levels and apoptosis were measured using MitoSOX and Annexin V staining. d Representative images showing MitoSOX/Annexin V staining in RPMI8226 cells (e)