Objective: Observe the effect of normal and myeloma bone marrow stromal cells (BMSC) to the growth of myeloma cells. Evaluate the function of adhesion molecules in the mutual action of BMSC and myeloma cells.
Methods: Immunomagnetic beads and anti-CD138 antibody were used to separate myeloma cells from patients’ bone marrow of multiple myeloma (MM). U266 and freshly purified myeloma cells respectively co-cultured with normal BMSC and myeloma BMSC. Single U266 or fresh myeloma cells were cultured as control group. Anti-CD11a, anti-CD29, anti-CD44 and anti-CD49d antibodies were respectively added in the co-cultured system and single myeloma cells. After culture for 72 hours, cell cycle of myeloma cells and the ratio of apoptotic cells were tested by flowcytometry. The level of interleukin 6 (IL-6) in the supernate was measured.
Results: Normal BMSC inhibited the growth of myeloma cells while myeloma BMSC improved it. As to U266, S+G2% of myeloma cells in normal BMSC group and myeloma BMSC group were 55.5±4.0% and 65.4±4.5%(p<0.05)respectively. The ratio of apoptotic cells were 3.92±1.83% and 0.86±0.58%(p<0.05)respectively(figure1 from left to right: U266, U266+normal BMSC, U266+myeloma BMSC, red: apoptotic cells). The level of IL-6 in the supernate were 1236±146.2pg/ml and 1661±161.5pg/ml(p<0.05)respectively. After co-cultured with normal BMSC and myeloma BMSC, S+G2% of freshly separated myeloma cells were 5.2±4.1% and 15.9±5.5%(p<0.05)respectively. The ratio of apoptotic cells were 18.7±7.8% and 8±3.9%(p<0.05) respectively (figure2 from left to right: fresh myeloma cells, fresh myeloma cells+normal BMSC, fresh myeloma cells+myeloma BMSC, red: apoptotic cells). The level of IL-6 in the supernate were 1621.2±121.1pg/ml and 2151.1±170.4pg/ml(p<0.05)respectively. The function of four adhesion molecules was different in the co-culture system. Anti-CD11a hardly affected the growth of myeloma cells. But anti-CD29 and anti-CD44 showed their significant inhibition to the growth of myeloma cells, including the decrease of supernatant IL-6 level, percentage of phase S+G2 and the increase of apoptotic cells (fresh myeloma cells). Anti-CD49d indicated partial inhibition, but no significant change.
Conclusion: We firstly proved that normal BMSC inhibited the growth of myeloma cells. Anti-CD29 and anti-CD44 can block the adhesion of myeloma cells and BMSC, resulting in the growth inhibition of myeloma cells.