中文摘要：

目的：探讨TNF-α抑制骨髓间充质干细胞（BMMSCs）成骨分化参与SLE骨质疏松的分子机制。方法分离狼疮鼠（MRL/lpr）及对照鼠（C3H/HeJ）股骨制备组织切片，苏木精-伊红（HE）染色观察骨质情况；免疫组织化学法观察骨TNF-α、NF-κB P50、骨形态发生蛋白-2（BMP-2）、磷酸化Smad（PSmad）1/5/8蛋白表达；分离培养狼疮鼠及对照鼠BMMSCs，爬片后免疫组织化学法观察上述4种蛋白表达情况；定向诱导BMMSCs向成骨细胞分化，ALP染色鉴定早期成骨情况，将BMMSCs与羟基磷灰石共孵育后植入裸鼠皮下，行HE染色及Masson染色观察异位成骨情况；将狼疮鼠皮下注射重组人Ⅱ型肿瘤坏死因子受体-抗体融合蛋白（rhTNFR：Fc）及0.9%氯化钠注射液，4周后观察上述蛋白表达及异位成骨情况。图像分析采用Image-pro plus 6.0软件，2组间比较采用t检验。结果 MRL/lpr鼠骨皮质较对照组减少，皮质骨占骨体积百分比较对照鼠减低[（13.96±0.25）%与（23.61±0.71）%，3只，P〈0.01]；狼疮鼠股骨TNF-α、NF-κB P50蛋白的表达较对照组增高（0.643±0.051与0.405±0.022，0.917±0.023与0.650±0.032，3只， P〈0.01），BMP-2表达较对照鼠明显减少（0.52±0.03与0.72±0.03，3只，P〈0.01），PSmad1/5/8蛋白的表达在两者间差异无统计学意义（1.264±0.021与1.301±0.044，3只，P〉0.05）；细胞爬片显示狼疮鼠BMMSCs TNF-α、NF-κB P50表达较对照鼠明显增强（0.184±0.021与0.136±0.013，0.132±0.021与0.097±0.014，3只，P〈0.01），BMP-2、PSmad1/5/8表达则明显减弱（0.128±0.013与0.216±0.221，0.115±0.023与0.196±0.034，3只，P〈0.01）；狼疮鼠BMMSCs成骨诱导7 d ALP活性较对照组减低，HE染色及Masson提示狼疮鼠BMMSCs体内异位成骨能力较对照组减弱。注射rhTNFR：Fc组狼疮鼠骨组织骨小梁较0.9%氯化钠注射液组明显增多（21.8±1.0与14.3±0.6，3只，P〈0.01），股骨TNF-α、NF-κB P50蛋?

英文摘要：

Objective To investigate the mechanism of tumor necrosis factor-α （TNF）-α inhibiting osteo blastdifferentiation of mesenchymal stem cells （BMMSCs） in the pathogenesis of osteoporosis in the mouse model of systemic lupus erythematosus （MRL/lpr）. Methods The femurs of MRL / lpr and C3He/HeJ mice were isolated, the bone structure were examined by hematoxylin-eosin （HE） staining. The proteins of TNF-α, NF-κB P50, bone morphogenetic protein -2 （BMP-2） and PSmad1/5/8 were measured by immunohistochemical stain. Bone marrow mesenchymal stem cells （BMMSCs） were isolated. After BMMSCs grew on the cover slips, the proteins on top of it were evaluated by immunohistochemistry stain. Moreover, the alkaline phosphatase （ALP） staining was employed for the measurement of the early osteogenic differentiation. BMMSCs together with hydroxyapatite were embedded subcutaneously in the nude mice and eight weeks later, the ectopic bone formation was evaluated. The recombinant human tumor necrosis factor receptor type Ⅱantibody fusion protein （etanercept） or normal saline was subcutaneous injected to the mice with lupus. After four weeks, the expression of these proteins was observed and the ectopic bone formation was investigated. Image-Pro plus 6.0 software was employed for imagine analysis, and Studentˊs t-test was used to test the differences between 2 independent groups. Results MRL/lpr mice showed decreased volume of cortex and the percentage of cortex to the volume of bone of MRL/lpr mice was significantly lower compared to control groups and with C3He/HeJ mice （13.96±0.25 vs 23.61±0.71, n=3, P〈0.01）. The protein levels of both TNF-αand NF-κB P50 on the femur of MRL/lprl mice were higher than those of the control group （0.643±0.051 vs 0.405±0.022, 0.917±0.023 vs 0.650±0.032, n=3, P〈0.01）. The expressions of BMP-2 on the femur of MRL/lpr mice were lower than those of the C3He/HeJ mice （0.52 ±0.03 vs 0.72 ±0.03, n=3, P〈0.01）. There was no difference in the expression