Resveratrol inhibits the proliferation of CD138⁺ cells from patients with MM and down-regulates NF-κB and STAT3 activation. (A) Enriched CD138⁺ cells (2 × 10⁵/0.1 mL) from bone marrow aspirates of patients with multiple myeloma were cultured in the absence or presence of indicated concentrations of resveratrol for 24 hours, and cell proliferation was measured by MTT assay as described in “Materials and methods.” Values represent the mean ± SD of triplicate cultures. *P < .05. (B) Enriched CD138⁺ cells (2 × 10⁶ cells) from bone marrow aspirates of patients with MM as indicated were cultured in the absence or presence of resveratrol (50 μM) for 12 hours and then tested for NF-κB activity in the nuclei by electrophoretic mobility shift assay as described in “Materials and methods.” (C) STAT3 activation status was determined by fixing the untreated and 12-hour resveratrol-treated (50 μM, 100 μM) enriched CD138⁺ cells (2 × 10⁶ cells) on slides by cytospin followed by immunocytochemistry for STAT3 as described in “Materials and methods.” Patient's numbers are indicated beside each panel. Original magnification, × 200. One hundred cells were counted for each patient. Grading was as follows: + indicates less than 10% cells with nuclear positivity; ++, 10% to 50% cells with nuclear positivity; +++, greater than 51% cells with nuclear positivity. The ++ and + indicate significantly different nuclear (P < .05) positivity than untreated cells. (D) CD138⁺ cells (2 × 10⁴ cells) from patients no. 8 and no. 9 were treated with 25 μM resveratrol, 20 nM bortezomib, 10 μg/mL thalidomide either alone or in combination for 24 hours at 37°C. Cells were incubated with anti–annexin V antibody conjugated with FITC and then analyzed by a flow cytometer for early apoptotic effects.