| ObjectiveTo investigate the effect of proteasome inhibitor bortezomib on the cell viability, migratory ability and the cytokines/chemokines mRNA expressions such as monocyte chemotactic protein-1(MCP-1), stromal cell derived factor-1(SDF-1), vascular endothelial growth factor(VEGF), hepatocyte growth factor(HGF), insulin-like growth factor-1 receptor(IGF-1R)and platelet-derived growth factor receptor alpha (PDGFRα) of bone marrow mesenchymal stem cells(BMMSC) derived from multiple myeloma (MM) patients, which can be used to explore the biological differences of BMMSC between MM patients and normal individuals as well as the impacts of bortezomib on the bone marrow microenvironment.MethodMesenchymal stem cells derived from bone marrow sample of newly diagnosed multiple myeloma patients (n=8) and normal individuals (n=11) were isolated using density gradient centrifugation, then cultured and expanded in vitro. The second to fourth passage of mesenchymal stem cells were used to follow-up experiment. Four groups were assigned,which including MM BMMSC group, control BMMSC group, MM BMMSC treated with bortezomib group and control BMMSC treated with bortezomib group. The cell inhibitory rate after incubated with different concentrations of bortezomib was evaluated by MTT assay. Annexin V-FITC/PI double staining was performed to detect apoptosis rate after treated with bortezomib.The mRNA expression levels of MCP-1, SDF-1, VEGF, HGF, IGF-1R and PDGFRαin BMMSC were measured by SYBR Green I dye-based fluorescent quantitative real-time polymerase chain reaction (RQ-PCR). The standard curves of MCP-1, SDF-1, VEGF, HGF, IGF-1R, PDGFRαand glyceraldehyde-3-phosphate dehydrogenase (GAPDH) were generated to calculate the relative expression levels of interest genes. Transwell chamber experiment was employed to evaluate the migratory capacity of mesenchymal stem cells in forementioned four groups.Results1. Mesenchymal stem cells derived from MM patients and normal individuals were similar in morphology. BMMSC from both cohorts were fibroblast-like spindle.2. According to MTT assay and Annexin V-FITC/PI double staining, bortezomib can promote apoptosis and decrease the cell viability of mesenchymal stem cells, which was in correlation to a time- and dose-dependent manner. However, inhibitory effect of bortezomib was significantly less powerful to MM BMMSC group than that to control group(P<0.01).3. The cytokines/chemokines expressions in MM BMMSC were dysregulated.The mRNA expression of SDF-1 in MM BMMSC group was much lower(P<0.05),but VEGF, HGF were much higher(P<0.05) than those of control group. Bortezomib can decrease the mRNA expressions of MCP-1, SDF-1, VEGF, HGF, IGF-1R, PDGFRαboth from MM group(P<0.05)and control group (P<0.05). The inhibitory effect of bortezomib between MM BMMSC group and control group was no significant difference(P>0.05). 4. Migratory capacity between MM BMMSC group and control group measured by transwell assay was no significant difference (P>0.05). The migratory ability was reduced after bortezomib treatment in both groups (P<0.05), but can be enhanced after stimulated with SDF-1 in both groups regardless of incubated in the presence or absence of bortezomib (P<0.05), which is more obviously in MM BMMSC group(P<0.05).ConclusionThere were no significant differences between BMMSC derived from multiple myeloma patients and normal individuals in morphology and migratory capacity. Migratory capacity can be enhanced after stimulated with SDF-1 in both groups, but the cytokines/chemokines expressions such as SDF-1, VEGF and HGF in MM BMMSC group were dysregulated,which may a?ect growth, survival, migration, and drug resistance of multiple myeloma cells. Bortezomib can inhibit the proliferatory and migratory capacity of BMMSC,as well as the mRNA expression levels of MCP-1, SDF-1, VEGF, HGF, IGF-1R and PDGFRα, which suggested bortezomib might execute an anti-tumor effect not only by directly inducing apoptosis in multiple myeloma cells, but also by targeting on bone marrow microenvironment. |
PDF Full Text Request