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目的研究骨质疏松小鼠模型骨密度(BMD)及骨代谢等指标的变化规律,为揭示骨质疏松症的机理提供新的实验方法及思路。方法 C57BL/6J雌性小鼠随机分为模型组(Ovx组,n=25)和假手术组(Sham组,n=5),建立骨质疏松模型。以微计算机断层扫描技术(Micro-CT)测量其BMD,根据测量结果选取模型组中BMD最低和最高的5只小鼠分别作为低骨密度组(L组)和高骨密度组(H组)。应用酶联免疫吸附法(ELISA)检测其血清雌二醇(E~2)水平和碱性磷酸酶(ALP)活性;提取骨质疏松小鼠的骨髓间充质干细胞(BMSCs)并鉴定其表面抗原;通过使用茜素红染色观察钙化结节形成情况,对比不同组小鼠BMSCs的骨矿化能力。结果 L组、H组和Sham组的BMD分别为(297.82±10.22)、(365.26±3.25)、(442.76±21.96)mg/cm~2,各组间比较差异有统计学意义(P<0.05);Ovx组、L组、H组的E~2水平分别为(142.75±25.26)、(142.49±17.01)、(133.54±24.58)pg/ml,均低于Sham组的(240.38±36.54)pg/ml,差异有统计学意义(P<0.05);L组、H组、Sham组的ALP活性分别为(221.82±33.58)、(155.65±14.99)、(117.30±20.96)U/L,各组间小鼠血清ALP活性差异有统计学意义(P<0.05);流式细胞仪分析显示小鼠BMSCs不表达的表面标记包括:内皮细胞表面标记CD31(阳性表达率:1.4%),巨噬细胞及单核细胞表面标记CD11b(阳性表达率:0.8%),白细胞及淋巴细胞表面标记CD45(阳性表达率:0.3%),而表达黏附分子和间充质干细胞表面标记Ly-6A/E(阳性表达率:85.0%)。茜素红染色实验反应强弱的顺序为Sham组>H组>L组>Blank组。结论受个体遗传因素的影响,骨质疏松小鼠可表现出具有明显差异的BMD和骨代谢水平,其机制可能与BMSCs成骨分化基因的差异表达有关。
Objective To study the changes of bone mineral density (BMD) and bone metabolism in osteoporotic mouse model and to provide new experimental methods and ideas for revealing the mechanism of osteoporosis. Methods C57BL / 6J female mice were randomly divided into model group (Ovx group, n = 25) and sham operation group (n = 5), and osteoporosis model was established. The BMD was measured by Micro-CT, and the lowest and highest BMD among the model groups were selected as low BMD group (L group) and high BMD group (H group) according to the measurement results. . Serum estradiol level and alkaline phosphatase (ALP) activity were measured by enzyme-linked immunosorbent assay (ELISA). Bone marrow mesenchymal stem cells (BMSCs) from osteoporotic mice were extracted and their surface Antigen was measured by alizarin red staining. The bone mineralization of BMSCs in different groups was compared. Results The BMD of L group, H group and Sham group were (297.82 ± 10.22), (365.26 ± 3.25) and (442.76 ± 21.96) mg / cm ~ 2 respectively. There was significant difference between the two groups (P <0.05) (142.75 ± 25.26), (142.49 ± 17.01) and (133.54 ± 24.58) pg / ml respectively in Ovx group, L group and H group were lower than those in Sham group (240.38 ± 36.54) pg / ml, the difference was statistically significant (P <0.05). The ALP activities of L group, H group and Sham group were (221.82 ± 33.58), (155.65 ± 14.99) and (117.30 ± 20.96) U / L, The serum ALP activity of mice was significantly different (P <0.05). Flow cytometry analysis showed that the surface markers not expressed in BMSCs of mice include CD31 (positive expression rate of endothelial cells), macrophages Monocyte surface marker CD11b (positive rate: 0.8%), leukocyte and lymphocyte surface marker CD45 (positive rate: 0.3%), while the expression of adhesion molecules and mesenchymal stem cell surface marker Ly-6A / E Rate: 85.0%). Alizarin red staining reaction intensity of the order of Sham group> H group> L group> Blank group. CONCLUSIONS: Osteoporotic mice exhibit markedly different levels of BMD and bone metabolism, which may be related to the differential expression of osteogenic differentiation genes in BMSCs.