香豆雌酚对体外破骨细胞分化和骨吸收活性的影响  

Effect of coumestrol on in vitro differentiation of osteoclast and activity of bone resorption

作　　者：陈晓钢[1] 吴小涛[1] 许勇[1] 

机构地区：[1]东南大学附属中大医院骨科,江苏省南京市210009

出　　处：《中国临床康复》2006年第39期86-88,共3页Chinese Journal of Clinical Rehabilitation

摘　　要：目的:观察植物雌激素香豆雌酚对破骨细胞的影响,分析其抗骨质疏松的作用方式。方法:实验于2005-07/2006-03在东南大学医学院临床实验中心和分子影像实验室、南京师范大学物理实验中心完成。以1,25(OH)2D3诱导大鼠骨髓细胞形成破骨样细胞,通过相差显微镜下的形态观察,抗酒石酸酸性磷酸酶染色和骨片上骨吸收陷窝的形成来鉴定破骨细胞。分别加入0,10-9,10-8,10-7,10-6,10-5mol/L香豆雌酚,以10-9mol/L17β-雌二醇为阳性对照,于培养3,6和9d分别计数抗酒石酸酸性磷酸酶阳性细胞个数,磷酸苯二钠法测定破骨细胞抗酒石酸酸性磷酸酶的活性,于培养12d利用图像分析系统对骨吸收陷窝的面积进行测量分析,原子吸收光谱法测定破骨细胞和骨片联合培养的上清中钙的浓度。结果:①利用1,25(OH)2D3成功诱导出破骨样细胞。②香豆雌酚抑制破骨细胞的形成和分化:随香豆雌酚浓度增加,每孔抗酒石酸酸性磷酸酶染色阳性的破骨细胞数量呈剂量依赖性减少(香豆雌酚10-9,10-8,10-7,10-6,10-5mol/L组分别为357±8,347±11,303±17,275±18,268±6,空白对照组为360±15,17β-雌二醇组为200±12,P<0.05)。③破骨细胞噬骨活性:骨吸收陷窝面积香豆雌酚各浓度组与空白对照组相比均有显著下降,香豆雌酚10-9与10-8,10-7,10-6,10-5mol/L组之间,香豆雌酚10-8,10-7mol/L组与10-6,10-5mol/L组之间差异均有显著性。④破骨细胞和骨片联合培养的上清中钙的浓度:应用香豆雌酚10-8～10-5mol/L处理后共培养体系中的钙浓度与空白对照组相比分别下降了24.7%,36.2%,36.9%,39.1%。结论:香豆雌酚可通过抑制破骨细胞的形成、抗酒石酸酸性磷酸酶的活性和骨吸收作用,抑制破骨细胞分化和骨吸收活性,从而具有抗骨质疏松作用。AIM： To investigate the effect of coumestrol, a kind of phytoestrogen, on osteoclast, and analyze its mechanism of anti-osteoporosis. METHODS： The experiment was conducted in the Clinical Experimental Center and Laboratory of Molecular Imaging of Medical College of Southeast University as well as the Experimental Center of Physics of Nanjing Normal University from June 2005 to March 2006. Osteoclast-like cells were isolated from rat marrow cells and induced by the 1, 25 （OH）2D3, and the morphological changes were studied under contrast phase microscope. The osteoclasts were identified by tartrate-resistant acid phos-phatase （TRAP） staining and the formation of resorptionlacuna in bone slices. OLCs were exposed to coumestrol at the concentrations of 0 mol/L,10^-9 mol/L,10^-8 mol/L, 10^-7 mol/L, 10^-6 mol/L and 10^-5 mol/L respectively, and 17 β-oestradiol was taken as thepositive control. The number of TRAP positive cells were counted respectively on the 3^rd, 6^th and 9^th day and the activity of TRAP was measured by P-nitmphenyl sodium hosphate assay. The resorption pit area on the bone slices formed by osteoclasts was measured by computer image processing. The calcium concentration in co-cultured supernatant of bone slices and osteoclasts was determined by atomic absorption spectra. RESULTS： ①The osteoclast-like cells were induced by 1, （25 （OH）2D3 successfully.②Coumestrol inhibited the formation and differentiation of osteoclast： The number of TRAP-positive muhinucleated cells in each pore was decreased with the increased concentration of coumestrol and was in a dose-dependent manner （Those in the 10^-9, 10^-8, 10^-7, 10^-6, 10^-5 mol/L coumestrol groups were respectively 357 ±8,347 ± 11,303 ± 17,275 ± 18, 268±6, and it was 360±15 in the blank control group, and it was 200±12 in the 17 β-oestradiol group, P 〈 0.05）.③The phagocytotic activity of osteoclast on bone： The area of bone resorption pits in all coumestrol groups were obviously decreased than that in the

关 键 词：骨质疏松 香豆雌酚 破骨细胞 骨质吸收 

分 类 号：R681.4[医药卫生—骨科学]