重组可溶性核因子κB受体活化因子体外抑制甲状旁腺激素诱导的破骨细胞生成

新近发现的核因子κB受体活化因子配基（receptor activator of nuclear factor-κB ligand,RANKL）,核因子κB受体活化因子（receptor activator ofnuclear factor-κB,RANK）／护骨素（osteoprotegerin,OPG）细胞因子系统提高了对破骨细胞生物学和骨稳态分子水平的认识。RANKL与RANK之间的相互作用决定了破骨细胞的分化。本研究通过体外实验评价可溶性RANK （soluble RANK,sRANK）是否可作为RANKL的拈抗剂下调破骨细胞生成和骨吸收陷窝的形成。构建sRANK的原核表达载体,转化入大肠杆菌表达菌株Origami B（DE3）,成功表达了重组蛋白,亲和层析进行纯化。重组sRANK以剂量依赖方式抑制由甲状旁腺激素（parathyroid hormone,PTH）诱导的破骨细胞生成和骨吸收陷窝形成。RT-PCR实验证实,sRANK可显著抑制PTH刺激的小鼠骨髓细胞碳酸苷酶Ⅱ和抗酒石酸酸性磷酸酶mRNA的表达。结果表明,sRANK具有抗骨吸收功能,可能成为一种治疗以骨吸收加强为特征的骨疾病的新方法。

Recombinant soluble receptor activator of nuclear factor-kappaB inhibits parathyroid hormone-induced osteoclastogenesis in vitro

The recent identification of receptor activator of nuclear factor-kappaB ligand (RANKL)/RANK/osteoprotegerin (OPG) cytokine system has led to a new molecular perspective on osteoclast biology and bone homeostasis. Specifically, the interaction between RANKL and RANK is responsible for osteoclast differentiation. In the present study, we evaluated whether soluble RANK (sRANK) could act as an antagonist of RANKL and down-regulate osteoclastogenesis and bone resorption in vitro. The prokaryotic expression vector coding for sRANK was constructed. Then the construct was introduced into E. coli Origami B (DE3) competent cells and recombinant sRANK was successfully produced and purified through affinity chromatography. sRANK reduced osteoclast-like cell (OLC) formation and resorption pit formation induced by parathyroid hormone (PTH) in a dose-dependent manner. In addition, sRANK significantly inhibited PTH-induced mRNA expression of carbonic anhydrase II and tartrate-resistant acid phosphatase in murine bone marrow cells as confirmed by using semi-quantitative RT-PCR. The down-regulation was highly correlated with the effect of sRANK on OLC formation from marrow cells. These data demonstrate the anti-resorptive effects of sRANK in vitro and highlight the potential of sRANK as a novel therapeutic approach to bone disorders characterized by enhanced bone resorption.