Aromatase is decreased in the bone marrow of IMiD-treated patients. (A) Immunoblot analysis of aromatase (AROM) in bone marrow samples from the cases listed in Table 2. (B) Serial changes in AROM protein levels in the bone marrow samples from cases 1, 2, and 17 listed in Table 1. Frozen viable patient samples were thawed at 37°C. The cells were lysed in RIPA lysis buffer. Immunoblot analysis was performed using 5 to 10 µg of protein per sample. Because of limited availability of the patient samples, the experiment was performed only once. (C) Cytospin preparations of megakaryocytes isolated from bone marrow samples of a healthy volunteer and case 24 listed in Table 1 (May-Giemsa staining). Images were captured by a BH-2 microscope (Olympus) and ACT-2U imaging software (Nikon) with ×600 magnification (scale bars, 25 µm). (D) Representative immunofluorescence microscopic images of megakaryocytes. Cells were stained with anti-AROM antibody (green). Nuclei were counterstained with 4′,6-diamidino-2-phenylindole (DAPI) (blue). Megakaryocytes were identified as large cells containing a multilobulated nucleus. Images were captured by BZ-X700 (Keyence) with ×40 objective lens (scale bars, 25 µm). Two independent patient-control–matched pair experiments were performed. UPN, unique patient number.