柚皮苷对成骨细胞系MC3T3-E1和小鼠颅顶骨骨形成的作用研究

发布时间：2018-04-14 17:07

本文选题：柚皮苷 + MC3T3-E1　；参考：《重庆医科大学》2017年硕士论文

【摘要】：骨组织工程中运用各种生物因子及支架研究骨代谢过程中的骨形成及骨吸收成为当下科学研究及临床关注的重点。鉴于现有临床药物在骨改建过程中促骨形成时的副作用,植物源性促骨形成药物成为当下科研焦点,然而研究药物对单一离体细胞分化成骨作用并不能为药物的临床使用提供有力的证据。本实验拟研究骨碎补主要活性物质-柚皮苷对成骨细胞系MC3T3-E1的促骨形成作用,同时选择药物与颅顶骨器官建立共培养模型,运用Micro-CT扫描技术分析药物对系列骨细胞(骨细胞、成骨细胞及破骨细胞)、骨基质等共存下的颅顶骨器官骨形成的作用。目的:1.在分析区域包含整个颅顶骨的前提下,改变Micro-CT扫描后二维图像的重建层数或者位置,比较进行重建后的骨量参数(骨表面积、骨体积)是否出现改变。2.分析柚皮苷对成骨细胞系MC3T3-E1和颅顶骨骨形成的作用。方法:1.Micro-CT扫描4日龄CD-1小鼠颅顶骨,三维重建时改变空白分析区域层数或位置来分别分析:(1)996~22层组与800~50层组,分析层数不同;(2)996~22层组与随机选择层组,分析层数不同;(3)近边界上缘组与近边界下缘组,分析层数相同的骨量参数变化。2.不同浓度柚皮苷作用于MC3T3-E1后通过CCK-8法检测细胞生长、定量分析细胞分泌蛋白ALP、RT-PCR法检测MC3T3-E1细胞RUNX2、OCN和OPN基因m RNA表达量、茜素红染色柚皮苷作用下细胞矿化结节形成。3.不同浓度柚皮苷作用于颅顶骨后通过Micro-CT扫描分析颅顶骨骨量参数(骨表面积、骨体积)的变化。结果:1.在分析区域包含整个颅顶骨的前提下:(1)增加或者减少空白分析层数会导致颅顶骨骨表面积变化,而骨体积无变化。(2)随机选择层数组与996~22层数组分析比较时颅顶骨骨表面积、骨体积均有变化。(3)重建层数相同的前提下,改变分析区域位置时颅顶骨骨表面积、骨体积无变化。2.不同浓度柚皮苷(0mg/L、1mg/L、10mg/L)均可促进MC3T3-E1的RUNX2、OCN、OPN表达上调及细胞矿化结节形成,但1mg/L浓度柚皮苷促成骨作用较10 mg/L组显著。3.不同浓度柚皮苷作用的离体培养颅顶骨骨表面积和骨体积无明显变化。结论:1.柚皮苷可促体外培养MC3T3-E1细胞成骨向分化,而对体外培养颅顶骨骨形成无明显作用。2.Micro-CT三维重建时,改变重建层数可影响骨量参数,而保证分析层数相同仅改变空白分析区域的位置不会影响骨量参数。
[Abstract]:The use of various biological factors and scaffolds in bone tissue engineering to study bone formation and bone resorption during bone metabolism has become the focus of current scientific and clinical research.In view of the side effects of existing clinical drugs in promoting bone formation in the process of bone remodeling, plant-derived osteogenic drugs have become the focus of current scientific research.However, the study of the effect of drugs on the differentiation and osteogenesis of in vitro cells does not provide strong evidence for the clinical use of drugs.The aim of this study was to study the effect of naringin, the main active substance of osteoclastic, on osteogenesis of osteoblast MC3T3-E1. At the same time, the drug was selected to establish coculture model with cranio-parietal organs. Micro-CT scanning technique was used to analyze the effects of naringin on osteoblasts (bone cells).The role of osteoblasts and osteoclasts and bone matrix in the formation of bone in parietal organs.Purpose 1.Under the condition that the region contains the whole cranio-parietal bone, the number or location of the reconstructed layers in the two-dimensional image after Micro-CT scanning is changed, and the bone mass parameters (bone surface area, bone volume) after reconstruction are compared whether the bone mass parameters (bone surface area, bone volume) change. 2.To investigate the effects of naringin on osteoblast MC3T3-E1 and parietal bone formation.Methods: 1. Micro-CT scanning of the parietal bone of 4-day-old CD-1 mice was performed. The number or position of the blank analysis area was changed during 3D reconstruction to analyze the two groups, the two groups were divided into two groups, one was divided into two groups, one was divided into two groups, the other was divided into two groups, one was divided into two groups, the other was divided into two groups, one was divided into two groups, the other was divided into two groups.The changes of bone mass parameters of the upper and lower boundary groups with the same layer number were analyzed.After different concentrations of naringin was treated with MC3T3-E1, cell growth was detected by CCK-8 assay, and the expression of RUNX2OCN and OPN gene m RNA in MC3T3-E1 cells was detected by RT-PCR. The mineralized nodule was formed by alizarin red staining with naringin.The changes of bone mass parameters (bone surface area and bone volume) of parietal bone were analyzed by Micro-CT scanning after different concentrations of naringin were applied to parietal bone.The result is 1: 1.(1) increasing or decreasing the number of blank analytical layers will result in a change in the surface area of the parietal bone, while the bone volume does not change. 2) the surface area of the parietal bone is compared with that of the random array of selected layers compared with the array of 996 ~ 22 layers.On the premise of the same number of reconstructed layers, the surface area of cranio-parietal bone was changed and the volume of bone did not change. 2.Different concentrations of naringin (0 mg / L ~ (-1) mg / L ~ (10) mg / L) could promote the up-regulation of RUNX2 ~ (2) MC3T3-E1 and the formation of mineralized nodule in MC3T3-E1. However, the effect of naringin at 1mg/L concentration on bone formation was significantly higher than that of 10 mg/L group.There was no significant change in surface area and bone volume of in vitro cultured parietal bone with different concentrations of naringin.Conclusion 1.Naringin could promote the osteogenic differentiation of cultured MC3T3-E1 cells, but had no obvious effect on the formation of cranio-parietal bone in vitro .2.When the three-dimensional reconstruction of Micro-CT, changing the number of reconstructed layers could influence the bone mass parameters.But ensuring that the same number of analysis layers only changes the position of blank analysis area will not affect the bone mass parameters.
【学位授予单位】：重庆医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R782

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