DKK1单克隆抗体逆转骨髓瘤上清液抑制成骨分化的研究

Study on DKK1 Monoclonal Antibody in Reversing the Inhibition of Osteoblast Differentiation Induced by Myeloma Supernatant

目的探讨dickkopf1(DKK1)单克隆抗体逆转多发性骨髓瘤上清液在体外抑制成骨分化的现象,为以DKK1基因为靶点的骨髓瘤骨病(myeloma bone disease MBD)的靶向治疗研究奠定基础。方法在骨髓瘤上清液抑制成骨分化的体系中加入0.1、1和10ng/ml DKK1单克隆抗体分别培养5天后检测各项成骨分化指标,如碱性磷酸酶(ALP)染色结节计数、碱性磷酸酶(ALP)吸光度测定以及RT-PCR方法检测碱性磷酸酶(ALP)、骨钙素(OC)和核心结合因子1(Cbfa1)的mRNA水平。结果10ng/ml DKK1的高浓度单克隆抗体组与上清液抑制组相比较ALP染色结节计数(P<0.01)和ALP吸光度测定(P<0.01)以及ALP(P<0.05)和cbfa1(P<0.01)的mRNA都有显著意义的升高。结论 DKK1单克隆抗体在体外能够逆转骨髓瘤细胞上清液所致的成骨分化受抑制状态,骨髓瘤细胞能够分泌可溶性DKK1抑制成骨祖细胞分化。

Objective To investigate whether DKK1 mAb has a potential to reverse the inhibition of osteoblast differentiation induced by myeloma cell line supernatant, so as to lay the foundation for the therapeutic research of myeloma bone disease（ MBD）. Methods We added 0.1,1 and 10ng/ml DKK1 mAb into the system of the myeloma supernatant inhibition of osteoblast differentiation. After 5 days, we collected osteoblast to identify ALP staining nodule counting, ALP absorbance and mRNA of ALP, OC and Cbfal with RT - PCR. Results The high concentration of DKK1 mAb for 10ng/ml could significantly elevate ALP staining nodule counting（P 〈 0.01 ） , ALP absorbance and mRNA level of ALP（ P 〈 0.05 ） , and Cbfal （ P 〈 0.01 ） with RT - PCR compared with the supernatant inhibition control group. Conclusion The high concentration of DKK1 mAb could partly reverse the osteoblast differentiation inhibition which was induced by supernatant from mutiple myeloma ceils RPMI8226.