Gelatin-Coated Insert-Well Culture Markedly Improved In Vitro Culture of Primary Myeloma Cells.

In order to clarify the pathogenesis of multiple myeloma (MM) and identify the molecular target for MM treatment, it is important to understand the biology and molecular mechanism of survival and proliferation of myeloma cells in vitro and in vivo. Still, it is hard to keep primary myeloma cells viable at least for 2 weeks in vitro, and these cells rapidly enter apoptosis even in the presence of IL-6. Co-culture with BM stromal cells is considered to be one of the most important factors as well as addition of cytokines for improvement of in vitro culture of primary myeloma cells. Based on our previous data, we devised a new method where bone marrow mononuclear cells (BMMNC) from BM aspirates were put inside insert-wells (8.0 um pore size, #3182, BD) coated with gelatin in the presence of the mixture of cytokines (galectin-1, SDF-1, IL-6 and IGF-1) in the serum-free synthetic medium; addition of galectin-1 and SDF-1 was essential especially at the beginning of this culture. By this method, we observed that BM stromal cells attached to gelatin and survived well with rather low proliferation; myeloma cells interacted well with these stromal cells. Thus, we could maintain viability of primary myeloma cells at least for 4 weeks and the recovery of viable myeloma cells (CD38++ cell) appeared to be about 80% in 30 cases of MM we examined. Phenotypic data also showed that ratio of immature (MPC-1−) and mature (MPC-1+) myeloma cells in the BMMNC before culture was approximately maintained after 2 or 4 weeks in this method. Therefore, these results suggest that gelatin-coated insert-well culture can control the viability of stromal cells and thus maintain the culture of primary myeloma cells in vitro. This new method would contribute to the further understanding of biology and drug sensitivity of primary myeloma cells.