辛伐他汀对幼鼠骨形成相关因子表达及骨髓基质干细胞成骨分化的影响

发布时间：2018-03-18 19:07

本文选题：辛伐他汀　切入点：骨形成　出处：《华北煤炭医学院》2009年硕士论文　论文类型：学位论文

【摘要】：目的:本实验给予SD大鼠颈部皮下注射辛伐他汀(5mg/kg/d),观察辛伐他汀对幼鼠胫骨上端骨形成相关因子BMP-2、MMP-13、VEGF的表达及骨髓基质干细胞成骨分化过程中BMP-2、RUNX2、Osterix、MSX2、DLX3、DLX5等因子mRNA表达水平的影响。方法:60只1周龄雄性SD大鼠随机分为6组:实验组,给药1周(S1)、给药2周(S2)、给药3周(S3),对照组,对照1周(D1)、对照2周(D2)、对照3周(D3),每组10只。实验组给予辛伐他汀悬液(simvastatin、DMSO、PBS)(5mg/kg/d),对照组给予空白对照液(PBS、DMSO混合液),皮下注射持续1、2、3周。所有实验动物于最后一次注射药物或空白对照液的第2天脱颈处死,立即取胫骨,固定、脱钙、包埋,沿正中矢状面作4μm连续切片,行BMP-2、MMP-13、VEGF免疫组化染色用图像分析仪测量平均光密度值;取双侧股骨BMSCs向成骨方向诱导培养,在细胞培养14天分别进行ALP染色,21天采用Real-time PCR法检测BMP-2、RUNX2、Osterix、MSX2、DLX3、DLX5等成骨过程中相关因子mRNA的表达,28天进行von Kossa染色。结果:(1)胫骨免疫组化检测结果:BMP-2、MMP-13、VEGF等因子的表达各周实验组与对照组无差异(P0.05)。(2)ALP:各组ALP染色阳性细胞(胞浆蓝黑色)比例均在30%左右(D1:0.29±0.13,S1:0.32±0.10,D1:0.28±0.15,S1:0.30±0.13,D1:0.31±0.13,S1:0.29±0.14)无显著差别(P0.05)。(3)BMP-2、RUNX2、Osterix、MSX2、DLX3、DLX5因子mRNA表达水平组间无显著差异(P0.05)。(4)von Kossa染色可见大小不等染为棕黑色的钙化点,其大小和密度组间无显著差别(P0.05)。结论:(1)皮下注射辛伐他汀(5mg/kg/d)不能影响胫骨上端骨形成相关因子BMP-2、MMP-13、VEGF等的表达。(2)皮下注射辛伐他汀(5mg/kg)不能影响BMSCs向成骨细胞诱导分化过程以及成骨分化相关因子BMP-2、RUNX2、Osterix、MSX2、DLX3、DLX5 mRNA表达水平的变化。
[Abstract]:Objective: to observe the expression of simvastatin 5 mg / kg 路kg / d after subcutaneous injection of simvastatin to SD rats and the expression of BMP-2MMP 13 mRNA during osteogenic differentiation of bone marrow stromal stem cells (BMSCs) and the expression level of factors such as BMP-2RUNX2OUNX2OsterixMX2DLX3DLX5 in young rats. Methods Sixty one-week-old male Sprague-Dawley rats were randomly divided into 6 groups: experimental group, The control group was treated with S1 for 1 week, S2 for 2 weeks and S3 for 3 weeks. 10 rats in each group were given simvastatin suspension of simvastatin (DMSON) PBS5 mg / kg / d, while the control group received a mixture of PBSn and DMSO for 12 ~ 3 weeks. All the experimental animals were injected with simvastatin suspension (DMSON) 5 mg / kg / kg / d. All the experimental animals were injected with simvastatin suspension at the last time for 12 ~ 3 weeks, while the control group was given a mixture of PBSn and DMSO, which lasted for 12 ~ 3 weeks. The control group was given simvastatin suspension (DMSON) 5 mg / kg / d ~ (-1). On the second day after the removal of the neck, the subjects or the blank control solution were killed. Tibia was taken immediately, fixed, decalcified, embedded, and 4 渭 m serial sections were made along the median sagittal plane. The mean optical density was measured by using image analyzer with BMP-2MMP 13 BMSCs immunohistochemical staining, and the bilateral femur BMSCs was induced to be cultured in the direction of osteogenesis. After 14 days of culture, the expression of mRNA in osteogenesis was detected by Real-time PCR method. The expression of mRNA in osteogenic process such as MSX2DLX3DLX3DLX5 was detected by von Kossa staining for 28 days. Results Immunohistochemical examination of tibia showed that there was no difference in the expression of VEGF and other factors in the tibia between the experimental group and the control group. There was no significant difference between the experimental group and the control group in the proportion of ALP staining positive cells (cytosol blue black) in the 30% or so groups. The ratio of ALP staining positive cells (cytosol blue black) was about 30%. The ratio of mRNA expression was 0.29 卤0.10D1: 0.28 卤0.15S1: 0.30 卤0.13D1cell 0.31 卤0.13S1cell 0.29 卤0.14). There was no significant difference in the expression of mRNA of RUNX2OsterixMX2DX3DX5. There was no significant difference between the two groups. The calcification spots of brown and black color could be seen in the staining of P0.05P0. 0. 0. 4 von Kossa, and there was no significant difference between the two groups. There was no significant difference in size and density between the two groups (P 0.05). Conclusion injection of simvastatin 5 mg / kg / d subcutaneously does not affect the expression of BMP-2MMP 13, VEGF et al.) injection of simvastatin 5 mg / kg subcutaneously does not affect the differentiation of BMSCs into osteoblasts and the osteogenic differentiation related factor BMP-2RUNX2OsterixMSX2DLX3 DLX5. Changes of mRNA expression level.
【学位授予单位】：华北煤炭医学院
【学位级别】：硕士
【学位授予年份】：2009
【分类号】：R329.2

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