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J. Teramachi et al.
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Figure 1. TAK1 is activated to mediate growth and survival of multiple myeloma cells. (A) TAK1 expression and phosphorylation in multiple myeloma (MM) cell lines. TAK1 expression and phosphorylation were examined by western blotting analysis in various MM cell lines and primary MM cells as well as normal peripheral blood mononuclear cells (PBMC). Glyceraldehyde 3-phosphate dehydrogenase (GADPH) was used as a loading control. (B) The indicated MM cell lines were cultured in the absence or presence of LLZ at 1 or 5 mM for 2 days (left). The cells were transduced with scrambled (siCTL) or human TAK1 small interfering RNA (siRNA) (siTAK1) and cultured for 13 hours (right). Cell lysates were then collected, and protein levels of the indicated molecules were analyzed by western blotting analysis. b-actin was used as a loading control. (C) MM cell lines as indicated were incubated in triplicate in the presence of the indicated concentrations of TAK1 inhibitor LLZ1640- 2 (LLZ) for 48 hours (left). The indicated MM cell lines transduced with scrambled (siCTL) or human TAK1 siRNA (siTAK1) were cultured in triplicate for 24 hours (right). Cell viability was measured using a WST-8 assay. Data are expressed as means ± standard deviation. (D) The indicated MM cell lines and CD138-positive cells isolated from MM patients (Pt #1 and Pt #2) were cultured in the presence or absence of LLZ at 3 mM for 24 hours. The induction of apoptosis was analyzed using annexinV and propidium iodide (PI) dual staining. (E) The indicated MM cell lines were cultured alone or in the presence of LLZ at 1 or 5 mM for 2 days, and cell lysates were then collected. Caspase-mediated apoptotic pathways were analyzed using western blotting analysis. b-actin was used as a loading control.
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haematologica | 2021; 106(5)