破骨细胞分化增强参与慢性砷暴露导致的小鼠骨密度下降  

Augmentation of osteoclast differentiation is involved in chronic arsenic exposure-induced decrease of bone mineral density

作　　者：刘志远 左卓 高天畅 杨杨[2] 侯永永 王惠惠 孙永新[2] 皮静波 

机构地区：[1]中国医科大学公共卫生学院环境毒理学研究室,沈阳110122 [2]中国医科大学附属第一医院康复医学科,沈阳110001

出　　处：《中华地方病学杂志》2017年第11期792-797,共6页Chinese Journal of Endemiology

基　　金：国家自然科学基金（81573106、81572221）

摘　　要：目的研究慢性饮水型砷暴露对小鼠长骨骨密度的影响及其相关机制。方法将5月龄雌性C57BL／6小鼠按体质量采用随机数字表法分为假手术组和摘除卵巢（去势手术）组，每组19只。每组分为3个亚组：①对照：饮用蒸馏水（rt=6）；②5mg／L砷处理：饮用蒸馏水中含三价无机砷（iAsⅢ）和五价无机砷（iAsⅢ）各2．5mg／L（n=7）；③20mg／L砷处理：饮用蒸馏水中含iAs“和iAsⅢ各10mg／L（n=6）。自由进食和饮水饲养3个月后，采用双能X射线检测小鼠股骨骨密度。体外实验处理条件：①0．00、0．25、0．50、0．75、1．00、1．50μmol／LiAsⅢ处理单核巨噬白血病（RAW264．7）细胞系或0．0、0．2、0．4、0．6、0．8、1．0μmol／LiAsⅢ处理原代培养小鼠骨髓造血干细胞6d，同时给予50μg／L核因子KB受体活化因子配体（RANKL）和30μg／L巨噬细胞集落刺激因子（M．CSF）进行诱导分化。②根据iAsⅢ处理RAW264．7细胞系诱导分化破骨细胞数量检测结果，选择0．6Ixmol／LiAsⅢ处理组，同时给予0（对照）、5、10mmol／L氧化抑制剂N-乙酰半胱氨酸（NAC）。采用抗酒石酸酸性磷酸酶（TRAP）染色观察iAsUI和NAC对破骨细胞分化的影响。结果①与假手术对照组[（84．44±4．40）mg／cm2]相比，假手术20mg／L砷处理组小鼠股骨骨密度[（80．04±4．06）mg／cm2]呈下降趋势，而去势手术对照组[（76．36±3．36）mg／cm2]明显下降（P〈0．05）；与去势手术对照组比较，去势手术5、20mg／L砷处理组未见明显变化[（77．74±4．91）、（75．56±3．71）mg／cm2,P均〉0．05]。②体外实验显示，在原代培养骨髓造血干细胞向破骨细胞诱导分化过程中，各iAsⅢ处理组破骨细胞数量比较，差异有统计学意义（F=1522，P〈0．05），0．50μmol／LiAsⅢ处理组破骨细胞数量达峰值；在RAW264．7细胞向破骨细胞诱导分化过程中，各iAsⅢObjective To study the effects of chronic exposure to inorganic arsenic （iAs） in drinking water on bone mineral density （BMD） in mice and its underlying mechanisms. Methods Five-month-old female C57BL/6 mice were randomly divided into sham groups and ovarectomy （OVX） groups （n = 19 mice each group）, which were further randomly assigned into control group （distilled water） and iAs exposure groups [5 mg/L and 20 mg/L, inorganic arsenite （iAsⅢ）： inorganic arsenate （iAsv） = 1 ： 1]. Following 3 months of exposure to iAs, BMD of the mice were determined by the dual energy X-ray detector. RAW 264.7 cell line and bone marrow hematopoietic stem cells（BMHSC） primarily isolated from C57BL/6 mice were used to study the in vitro effects of iAs on osteoclast differentiation and underlying mechanisms. During differentiation induced by receptor activator of nuclear factor-K B ligand （RANKL, 50 μg/L） and macrophage colony-stimulating factor （M-CSF, 30 μg/L）, RAW 264.7 cell line were treated with 0.00, 0.25, 0.50, 0.75, 1.00, 1.50 μmol/L iAsⅢ, while BMHSC with 0.0, 0.2, 0.4, 0.6, 0.8, 1.0 μmol/L iAsⅢ for 6 days. Based on the effect of iAsnt on the differentiation of RAW cells, RAW 264.7 cell line were treated by 0.6 μmol/L iAsm combined with 0, 5, 10 mmol/L of N-acetyl-cysteine （NAC）. Tartrate resistant acid phosphatase （TRAP）-positive red-colored cells with 3 or more nuclei were considered mature osteoclast. Results The femoral BMD of the mice [（80.04 ± 4.06） mg/cm2] that had been exposed to 20 mg/L of iAs for 3 months was substantially decreased compared to that of sham control mice [（84.44 ± 4.40） mg/cm2]. As expected, the BMD of the OVX group [（76.36 ± 3.36） mg/cm2] was significant decreased compared to that of the sham control group （P 〈 0.05）. However, the BMD among the OVX groups showed no significant difference [5 mg/L： （77.74 ± 4.91） mg/cm2; 20 mg/L： （75.56 ± 3.71） mg/cm2, P 〉 0.05]. In vitro studies, the iAsm evidently aff

关 键 词：砷 破骨细胞 分化 骨密度 

分 类 号：R599.1[医药卫生—内科学]