Role ofmiR33b asatumor suppressor in MM.1S MM cells. (A-C) MM.1S cells were transiently transfected with either pre-miR33b or scrambled probe using the Cell Line Nucleofector Kit V. The transfected cells were examined for miR33b expression by qRT-PCR (A), for cell viability using the CellTiter-Glo assay (B); and for apoptosis by annexin V–FITC/propidium iodide double staining (C). (D) RNA was isolated from GFP vector controls or miR33b-overexpressing MM.1S cells, and miR33b expression was analyzed by qRT-PCR. (E-F) 2 × 104/mL of MM.1S cells stably expressing either vector or miR33b (5mL) were used in forming the cell layer; colonies were stained with MTT (E), and counted 4 weeks later (F). (G) MM.1S cells stably expressing either vector or miR33b were analyzed for their effect on serum-induced migration ability. The migration assay was performed using a 24-well Transwell plate. The migration cells were quantified by measuring the intensity of fluorescence. (H) MM.1S cells stably expressing either vector or miR33b were treated with MLN2238 (12nM) for 24 hours, and cell viability was examined with the CellTiter-Glo assay. Results shown are representative or means ± SD (n = 3). The experiments were repeated at least 3 times. ***P ≤ .001.
