Comparison of HPSE expression between WBM and the corresponding purified MMCs of patients. (A) Expression of HPSE was determined in the WBM of 39 patients with myeloma as well as in the myeloma cells purified from the bone marrow of the same patients, using Affymetrix U1332 Plus 2.0 microarrays. □ indicates that the Affymetrix call is absent; ■ (WBM) and ▩ (MMC) indicate that it is present. (B) HPSE expression was measured by real-time RT-PCR in the whole bone marrow of 7 patients with MM (WBM; ■), in the corresponding MMCs (▩), in the corresponding microenvironment depleted from myeloma cells (< 2% MMCs, ENV; ▨), and in MMCs from 5 patients with plasma cell leukemia (PCL). HPSE expression was normalized to that of GAPDH. For each patient, the WBM sample was used as a reference and was assigned the arbitrary value of 100. For the 5 patients with PCL, the WBM sample of patient 1 was used as a reference. The median percentage of plasma cells in bone marrow aspirates from the 7 patients with intramedullary myeloma was 8.5% (range, 6%-40%). (C) HPSE protein expression was determined in microenvironment cells depleted from MMCs (< 2% MMCs, ENV1-5) of 5 patients and in purified MMCs of 3 other patients (MMC1-3). Cells were lysed, and the lysates were separated on a 12% SDS-PAGE and analyzed by Western blot with a polyclonal anti-HPSE antibody. Both the 65-kDa MW proenzyme and the 50-kDa MW active form were identified. β-Actin was used as a loading control. Results are from 2 separate Western blots, one with 3 purified MMCs, the other one with the 5 microenvironment cells. The HPSE^pos XG-2 HMCL was used as a positive HPSE protein control in the 2 blots (results not shown). kDa indicates molecular weight in thousands.