重组人骨保护素对体外培养兔破骨细胞的影响

Effect of recombinant human osteoprotegerin on rabbit osteoclasts in vitro

目的观察重组人骨保护素(recombinanthumanosteoprotegerin,rhOPG)对体外培养兔破骨细胞(osteoclast,OC)生存和功能的影响。方法将rhOPG用于干预从新生兔分离出的OC,于干预后1、3、7d取细胞玻片、皮质骨片进行HE、Giemsa、甲苯胺蓝染色,并观察抗酒石酸酸性磷酸酶(TRAP)染色阳性细胞和骨片吸收陷窝,扫描电镜进一步观察吸收陷窝形态。结果分离培养的兔OC多核形态明显;rhOPG对TRAP阳性OC生存的影响,1、3d结果差别不明显,7d的结果差异有显著性(P<0.05);rhOPG能够明显地抑制OC在骨片上形成吸收陷窝,三个时点计数结果差异均有显著性(P<0.01)。结论rhOPG可明显地抑制体外培养兔OC的骨吸收功能。

Objective Osteoprotegerin secreted by osteoblast was an important modulator for os-teoclast.The study is to explore the effectiveness of recombinant human osteoprotegein(rhOPG)on cul -tured rabbit osteoclasts in vitro.Methods New born New Zealand rabbits were sacrificed and long bones such as femur,tibia and humerus were cleaned off soft tissue and cartilage.Bone marrow and endosteum were harvested to obtain osteoclast.The glass cover slips(10mm×10mm)and bone slices(8mm×8mm×0.1mm)were embedded into the cultured plate with24holes before the osteoclasts were added.The con-ditioned medium in the presence or absence of rhOPG100ng /ml was mixed into the osteoclasts.The glass cover slips and bone slices were obtained for HE,Giemsa and toluidine blue staining at1,2and7days after interruption,and tartrate-resistant acid phosphatase(TRAP)positive multineucleated cells and bone resorp-tion pits were counted,scanning electron microscopy was used to observe the pits of bone resorption.Re-sults The ap pearance of isolated rabbit osteoclasts were typical,rhOPG had no effect on the survival rate of osteo clast at day1and day3,but decreased the number of TRAP-positive multinucleated cells significantly at day7(P<0.05).At the same time,rhOPG inhibited significantly the formation of bone resorption pits on bone slices at all time points(P<0.01).Conclusion rhOPG could inhibit the bone resorption by rabbit os-teo clasts.