大鼠实验性牙移动致牙根吸收龈沟液中DSPP、DSP的表达
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摘要
牙根吸收是正畸牙移动过程中不可避免的一种病理性结果,其致病机制、影响因素尚无统一观点。目前,临床工作中常用x线诊断牙根吸收的发生,但x线检查只能反映牙齿的静态结构,其精确度差,不能对牙根吸收进行早期诊断并及时准确地显示牙根吸收是否处于活动期。寻找一种新的评估方法来早期诊断、监测牙根吸收的发生、发展,对于正畸导致的牙根吸收的基础研究和正畸临床工作都有很大的意义。
本实验以大鼠为研究对象,建立龈沟液提取方法和实验性牙移动致牙根吸收模型。通过SDS-PAGE、Western blot等生物学技术比较对照组、轻力组、重力组动物龈沟液中DSPP、DSP的表达差异,显示重力下牙移动致牙根吸收组动物龈沟液中DSPP、DSP的表达明显高于其他组,提示DSPP、DSP在牙根吸收发生、发展过程中发挥一定作用,为临床利用生物学方法早期诊断、检测牙根吸收提供新的思路和理论依据。
Object
The purpose of this study was to establish the model of gingival crevicular fluid (GCF) collecting and root resorption following experimental tooth movement in rats,to observe the expression of DSPP and DSP in GCF after experimental tooth movement. We are looking for a biological index which relates to root resorption following orthodontic treatment to diagnose root resorption in the early stage.
Methods
Section l:15# absorbent paper points were used to collect saliva and GCF of rats. SDS-PAGE was used to analyze the proteins in two kinds of samples. Section 2:36 Wistar rats were used in this study. The animals were divided into 3 groups on average: A control group、B light force group (40g force) and C heavy force group (100g force). An orthodontic appliance, consisting of a coil spring connected the upper right side first molar and the two incisors by ligature wire was used in experiment. The experimental tooth was drawn mesially by applying a force of 40g in B group and a force of 100g in C group, with both of the maxillary central incisors as anchorage. The gingival crevicular fluid of rats was collected at the 7th day, and the expression of DSPP and DSP was assayed by using the biochemistry techniques of SDS-PAGE and Western blot. The histological slices were made, including the upper jaw, the experimental tooth and periodontal tissue. Sections (5μm thick) were stained with hematoxylin and eosin(H&E), Gomori and Tartrate resistant acid phosphatase (TRAP) to observed the root resorption.
Results
Section l:SDS-PAGE analysis identified proteins at MW 77 kD, 66 kD, 55 kD, 51 kD, 28 kD in GCF samples. There were proteins with MW 66 kD, 60 kD, 48 kD in saliva samples. Independent-samples t test showed there was a significant difference in the number of proteins between the two groups (P<0.05).
Section 2:After 7 days treated by 40g mesial force, the upper right first molar moved obviously in the mesial and palatal direction in B group, and there was no root resorption happening. The experimental tooth didn’t move distinctly in C group. The severe root resorption involving dentin layer could be observed in root apical area and bifurcation region in C group. Western blot revealed there was expression of DSPP and no DSP in A group, and there was the expression of DSPP and DSP in both B and C group. LSD Multiple Comparisons Test showed that there were significant differences in the contents of DSPP and DSP in three groups.
Conclusion
1. It is successful to establish the GCF collecting model by using 15# absorbent paper point to collect GCF in gingival pocket of rat.
2. There are proteins at MW 77 kD, 66 kD, 55 kD, 51 kD, 28 kD in gingival crevicular fluid of healthy rats.
3. After the maxillary first molar of rat is treated by 100g mesial force with the central incisors as anchorage for 7 days, the severe root resorption involving dentin layer can be observed in root apical area and bifurcation region.
4. The expression of DSPP and DSP in the three groups is statistically different, the highest being in heavy force group, followed by the light force group and control group with the least amount of proteins. So DSPP and/or DSP may be suitable biological markers for monitoring root resorption during orthodontic treatment.
本实验以大鼠为研究对象,建立龈沟液提取方法和实验性牙移动致牙根吸收模型。通过SDS-PAGE、Western blot等生物学技术比较对照组、轻力组、重力组动物龈沟液中DSPP、DSP的表达差异,显示重力下牙移动致牙根吸收组动物龈沟液中DSPP、DSP的表达明显高于其他组,提示DSPP、DSP在牙根吸收发生、发展过程中发挥一定作用,为临床利用生物学方法早期诊断、检测牙根吸收提供新的思路和理论依据。
Object
The purpose of this study was to establish the model of gingival crevicular fluid (GCF) collecting and root resorption following experimental tooth movement in rats,to observe the expression of DSPP and DSP in GCF after experimental tooth movement. We are looking for a biological index which relates to root resorption following orthodontic treatment to diagnose root resorption in the early stage.
Methods
Section l:15# absorbent paper points were used to collect saliva and GCF of rats. SDS-PAGE was used to analyze the proteins in two kinds of samples. Section 2:36 Wistar rats were used in this study. The animals were divided into 3 groups on average: A control group、B light force group (40g force) and C heavy force group (100g force). An orthodontic appliance, consisting of a coil spring connected the upper right side first molar and the two incisors by ligature wire was used in experiment. The experimental tooth was drawn mesially by applying a force of 40g in B group and a force of 100g in C group, with both of the maxillary central incisors as anchorage. The gingival crevicular fluid of rats was collected at the 7th day, and the expression of DSPP and DSP was assayed by using the biochemistry techniques of SDS-PAGE and Western blot. The histological slices were made, including the upper jaw, the experimental tooth and periodontal tissue. Sections (5μm thick) were stained with hematoxylin and eosin(H&E), Gomori and Tartrate resistant acid phosphatase (TRAP) to observed the root resorption.
Results
Section l:SDS-PAGE analysis identified proteins at MW 77 kD, 66 kD, 55 kD, 51 kD, 28 kD in GCF samples. There were proteins with MW 66 kD, 60 kD, 48 kD in saliva samples. Independent-samples t test showed there was a significant difference in the number of proteins between the two groups (P<0.05).
Section 2:After 7 days treated by 40g mesial force, the upper right first molar moved obviously in the mesial and palatal direction in B group, and there was no root resorption happening. The experimental tooth didn’t move distinctly in C group. The severe root resorption involving dentin layer could be observed in root apical area and bifurcation region in C group. Western blot revealed there was expression of DSPP and no DSP in A group, and there was the expression of DSPP and DSP in both B and C group. LSD Multiple Comparisons Test showed that there were significant differences in the contents of DSPP and DSP in three groups.
Conclusion
1. It is successful to establish the GCF collecting model by using 15# absorbent paper point to collect GCF in gingival pocket of rat.
2. There are proteins at MW 77 kD, 66 kD, 55 kD, 51 kD, 28 kD in gingival crevicular fluid of healthy rats.
3. After the maxillary first molar of rat is treated by 100g mesial force with the central incisors as anchorage for 7 days, the severe root resorption involving dentin layer can be observed in root apical area and bifurcation region.
4. The expression of DSPP and DSP in the three groups is statistically different, the highest being in heavy force group, followed by the light force group and control group with the least amount of proteins. So DSPP and/or DSP may be suitable biological markers for monitoring root resorption during orthodontic treatment.
引文
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