The Effect Of Zoledronic Acid On Osteoblasts Proliferation And RANKL/OPG Expression In Multiple Myeloma

Objective: To investigate (1) [0]The capacity of bone marrow stromal cells (BMSCs) in multiple myeloma (MM) differentiating into osteoblasts (OB) and their expression of RANKL/OPG; (2) The effect of zoledronic acid on MM OB proliferation and their RANKL/OPG expression.Methods: Bone marrow mononuclear cells were isolated from MM patients. BMSCs were formed from adherent cells. After passage, BMSCs were cultured in condition medium containing dexamethasone(10-8 mol/L),VitaminC(50 mg/L), andβ-glycerophosphate(10 mmol/L). Cell morphology was observed under inverted phase contrast microscope and OB were identified by HE staining, Wright's staining, alkaline phosphatase(ALP) staining and VonKossa staining. The OB RANKL/OPG mRNA expression was investigated by means of reverse transcription-polymerase chain reaction (RT-PCR) and compared with controls. In order to evaluate the effect of zoledronic acid, OB were cultured in medium containing different concentration of zoledronic acid. Cells were harvested after 24 h, 48 h and 72 h treatment. Cell couting kit-8 (CCK-8) was used to evaluate cell proliferation, RT-PCR for RANKL/OPG mRNA expression and ELISA for sRANKL/OPG level in culture supernatant.Results: (1) After culture in condition medium for differentiation, BMSCs showed morphology of OB. ALP staining was positive. The calcium nodule was obvious. (2) Expression of RANKL mRNA of OB in MM was higher than control. Relative density to GAPDH were 0.72±0.02 and 0.65±0.01 respectively (P<0.05). [0]Furthermore, OPG mRNA of OB in MM was lower than control. Relative density to GAPDH were 0.43±0.02 and 0.49±0.01 respectively(P<0.05). (3) CCK-8 assay showed the optical density (OD) increase after zoledronic acid treatment.(4) There is no significant change in OB RANKL mRNA expression after 24 h and 48 h zoledronic acid treatment. After 72 h treatment, expression decreased, especially at 10-9 mol/L zoledronic acid concentration. Comapred with no treatment (0 mol/L), the difference was statistically significant (P<0.05). The concentration of sRANKL in culture supernatant changed in the same way. For OB OPG mRNA expression, there is no significant change after 24 h zoledronic acid treatment. After 48 h treatment, expression increased, especially at 10-8 mol/L zoledronic acid concentration.Comapred with no treatment (0 mol/L), the difference was statistically significant (P<0.05). After 72 h treatment, expression increased, especially at 10-7 mol/L zoledronic acid concentration. Comapred with no treatment (0 mol/L), the difference was statistically significant (P<0.05).Conclusion: BMSCs from MM patients are able to differentiate into OB under certain culture condition. In MM patients, OB RANKL mRNA expression is higher than control, while OPG mRNA is lower than control. Zoledronic acid appears to enhance OB proliferation. Zoledronic acid is likely to down-regulate OB RANKL expression and up-regulate OPG expression.