摘要
目的探讨铁调素对小鼠单核细胞株RAW264.7向成熟破骨细胞分化的影响。方法将不同浓度铁调素加入含有核因子KB受体活化因子配体(RANKL)的RAW264.7细胞培养基,24h后用CCK-8法检测细胞的增生活性;4d后对细胞进行抗酒石酸酸性磷酸酶(TRAP)染色,光镜下观察;用RT—PCR法检测TRAP、组织蛋白酶K(CTK)、金属蛋白酶9(MMM-9)基因表达;用酶联免疫吸附法(ELISA)检测上清液中TRAP-5b含量;24h后用共聚焦显微镜(CLSM)测定细胞内铁离子浓度。结果铁调素在0~800nmol/L浓度围内可增加RAW264.7细胞TRAP染色阳性数目,上调TRAP、CTK和MMP-9基因表达,增加上清液TRAP-Sb含量(P〈0.05),同时增加RAW264.7细胞内铁离子浓度(P〈0.05)。结论铁调素可以促进RAW264.7细胞向破骨细胞分化,其作用机制可能与铁调素增加细胞内铁离子有关。
Objective To investigate the effects of hepcidin on differentiation of mouse RAW264.7 monocytes into osteoclasts. Methods RAW264. 7 cells were treated with different concentrations of bepcidin in the presence of receptor activator of NF-Kb ligand (RANKL). Cell viability, the number of tartrate-resistant acid phosphatase (TRAP) - positive cells, levels of TRAP, cathepsin K (CTK) , and matrix metalloproteinase 9 (MMP-9) mRNA, and levels of TRAP-Sb protein in the supernatant were examined. The intracellular iron ion was measured by a confocal laser scanning microscope. Results Hepcidin at 0-800 nmol/1 could significantly increase the number of TRAP-positive MNCs, and up- regulate gene expression of TRAP, CTK, and MMP-9, and increase the concentration of TRAP-5b in the supernatant, and increase concentrations of intracellular iron of RANKL-induced RAW264. 7 cells (P 〈 0. 05 ). Conclusion Hepcidin can significantly facilitate RANKL-induced differentiation of RAW264.7 into osteoclasts in vitro. The mechanism behind accelerated differentiation involves increased levels of intracellular iron.
出处
《中华骨质疏松和骨矿盐疾病杂志》
2013年第3期225-231,共7页
Chinese Journal Of Osteoporosis And Bone Mineral Research
基金
国家自然科学基金(81273090)
江苏省自然科学基金(BK2012608)
苏州市科技计划项目(510303)
江苏省普通高等学校研究生计划项目(CXLX11_0086)
关键词
铁调素
破骨细胞
细胞分化
骨代谢
hepcidin
osteoclast
differentiation
bone metabolism