Expression and functional role of c-kit ligand (SCF) in human multiple myeloma cells.
In this study we investigated the proliferation of three well-documented MM lines and 10 bone marrow samples from myeloma patients in response to rh-SCF alone and combined with Interleukin-6 (IL-6), IL-3 and IL-3/GM-CSF fusion protein PIXY 321. Neoplastic plasma cells were highly purified (> 90%) by immunomagnetic depletion of T, myeloid, monocytoid and NK cells. The number of S-phase cells was evaluated after 3 and 7 d of liquid culture by the bromodeoxyuridine (BRDU) incorporation assay. The proliferation of RPMI 8226 and U266 cell lines was also assessed by a clonogenic assay. All the experiments were performed in serum-free conditions. RPMI 8226 cell line was not stimulated by SCF which also did not augment the proliferative activity of IL-6, IL-3 and PIXY-321. Conversely, SCF addition resulted in 2.4-fold increase of the number of U266 colonies and in a higher number of U266 and MT3 cells in S-phase (24.5 +/- 2% SEM v 14.5 +/- 1% SEM and 32 +/- 3% SEM v 21 +/- 4% SEM, respectively; P < 0.05). The c-kit ligand also enhanced the proliferation of MT3 and U266 cells mediated by the other cytokines. Anti-SCF polyclonal antibodies completely abrogated the proliferative response of MT3 cells to exogenous SCF and markedly reduced the spontaneous growth of the same cell line. Reverse transcriptase-polymerase chain reaction amplification (RT-PCR) did detect SCF mRNA in MT3 and RPMI 8226 cells. Moreover, secreted SCF was found, in a biologically active form, in the supernatant of the two cell lines by the MO7e proliferation assay. When tested on fresh myeloma samples, SCF increased the number of S-phase plasma cells (4.7 +/- 1.6% v 3.4 +/- 1.3% in control cultures: P = 0.02). Significant proliferation was also induced by IL-6 (7 +/- 2.3% of BRDU+ cells; P = 0.006), IL-3 (5.3 +/- 1.3%; P = 0.01) and PIXY-321 (5.4 +/- 1.6%; P = 0.02). The addition of SCF significantly enhanced the proliferation of myeloma cells responsive to IL-6. In summary, our results indicate that SCF is expressed in MM cells and stimulates the proliferation of neoplastic plasma cells.
Lemoli
R M
RM
Institute of Haematology, University of Bologna, Italy.
Fortuna
A
A
Grande
A
A
Gamberi
B
B
Bonsi
L
L
Fogli
M
M
Amabile
M
M
Cavo
M
M
Ferrari
S
S
Tura
S
S
D000368:Aged; D001483:Base Sequence; D015139:Blotting, Southern; D002455:Cell Division; D016207:Cytokines / Q000494:pharmacology; D005260:Female; D015870:Gene Expression; D016298:Hematopoietic Cell Growth Factors / Q000096:biosynthesis* / Q000494:pharmacology; D006801:Humans; D008297:Male; D008875:Middle Aged; D008969:Molecular Sequence Data; D009101:Multiple Myeloma / Q000378:metabolism* / Q000473:pathology; D015345:Oligonucleotide Probes; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000235:genetics; D012334:RNA, Neoplasm / Q000235:genetics; D011994:Recombinant Proteins / Q000494:pharmacology; D019089:Stem Cell Factor; D014407:Tumor Cells, Cultured
D016207:Cytokines; D016298:Hematopoietic Cell Growth Factors; D015345:Oligonucleotide Probes; D012333:RNA, Messenger; D012334:RNA, Neoplasm; D011994:Recombinant Proteins; D019089:Stem Cell Factor