Do human myeloma cells directly produce basic FGF?

Basic fibroblast growth factor (bFGF) is a growth factor with proangiogenetic properties. Elevated bone marrow (BM) and peripheral serum bFGF levels have been reported in patients with multiple myeloma (MM) 1-3 ; however, the source of bFGF in patients with MM is not completely elucidated. Recently, Bisping et al, 4 in line with others, 1 have reported that human myeloma cell lines (HMCLs) (RPMI-8226, U266, KMS-11, and KMS-18) and sorted CD38 high /CD138 ϩ cells obtained from 12 of 15 patients with MM produced bFGF, concluding that myeloma cells are the predominant source of bFGF. In contrast, Gupta et al 5 have shown that neither human myeloma cells nor Epstein-Barr virus (EBV)–positive B-cell lines secrete bFGF. In order to better clarify this issue, we wish to present our evidence. Using reverse transcription–polymerase chain reaction (RT-PCR) (bFGF primer pairs: forward, 5Ј-GGCTTCTTCCTGCG-CATCCAT-3; reverse: 5Ј-GGTAACGGTTAGCACACACTCCTTT-3Ј) we found that XG-6, RPMI-8226, OPM-2, as well as EBV-positive cell line ARH-77 did not express bFGF mRNA, whereas U266 was positive and XG-1 expressed bFGF at low intensity (Figure 1A). Similarly, we failed to detect bFGF either in HMCL lysates by Western blot analysis (antipolyclonal bFGF antibody [Ab]; R&D Systems, Minneapolis, MN) or in HMCL (10 6 /mL)– conditioned media by enzyme-linked immunosorbent assay (ELISA; R&D Systems; range of sensitivity, 10 to 640 pg/mL), both in the presence and absence of interleukin-6 (IL-6, 20 ng/mL) with the exception of U266 and XG-1 (Figure 1B-C). Consistently, we previously showed that blocking anti-bFGF Ab failed to inhibit HMCL-induced angiogenesis in an in vitro system, suggesting that any bFGF biologic activity was found in HMCLs. 6 Purified CD138 ϩ MM cells (purity Ͼ 95%) isolated by an immunomagnetic method (magnetic-activated cell sorter [MACS]; Miltenyi Biotec, Bergisch-Gladbach, Germany) were positive for bFGF mRNA expression in 11 of 35 patients with newly diagnosed MM in stages I to III (median age, 64 years [range, 33-88 years]; and median plasmacytosis, 35% [range, 12%-95%]) (Table 1). In contrast, BM stromal cells (BMSCs) obtained from all patients were positive for bFGF mRNA (Figure 1A). bFGF protein has been found in plasma cell lysates in 8 of 30 patients tested. Consistently, bFGF levels were detected by ELISA assay (R&D Systems) in conditioned media of purified MM cells (10 6 /mL) in 7 of 28 patients (Table 1). Furthermore, a nuclear bFGF immunostaining with low cytoplasmic positivity has been found in bone marrow myeloma cells of 3 …