Expression and phosphorylation of PDGFRβ and c-Src TKs in MM patient-derived plasma cells and ECs. (A) FACS analysis of BMMCs from a representative patient with MGUS or active MM at diagnosis and relapse. Numbers on plots are percentages (± SD) of total cells gated. RT-PCR (B) and Western blotting analysis (C) of PDGF-BB, PDGFRβ, and pp60c-Src expression in control BMMCs (CTR), plasma cells isolated from MGUS (MGUS-PC) MM at diagnosis (MM-PC, D), and relapse (MM-PC, R). HUVEC (D) and ECs isolated from a representative patient with MGUS (MGEC), MM at diagnosis (MMEC, D), and MM in relapse (MMEC, R) were also included. All cells were serum-starved for 16 hours before immunoblotting analysis of constitutive phosphorylation of PDGFRβ (phosphoPDGFRβ) and pp60c-Src (phosphoSrc). Vertical lines have been inserted to indicate repositioned gel lanes. (D) Total cell lysates from control (1) and serum-starved MMECs at different time points (1-4) were probed with the indicated antibodies. (E) MMECs cultured in the presence of serum (1) were starved for 2 hours (2) and incubated with the conditioned medium (CM) harvested from MM plasma cells (CM-PC; 3) for 15 minutes. Whole cell lysates were then prepared and analyzed by SDS-PAGE analysis as shown.