国际口腔医学杂志 ›› 2019, Vol. 46 ›› Issue (3): 270-276.doi: 10.7518/gjkq.2019028

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大鼠正畸牙移动过程中转化生长因子-β/Smad信号通路相关蛋白质在Malassez上皮剩余细胞的表达变化

杨亚,陈鹏,戴红卫,张林()   

  1. 重庆医科大学附属口腔医院正畸科 口腔疾病与生物医学重庆市重点实验室重庆市高校市级口腔生物医学工程重点实验室 重庆 401147
  • 收稿日期:2018-09-09 修回日期:2018-12-28 出版日期:2019-05-01 发布日期:2019-06-05
  • 通讯作者: 张林 E-mail:zhanglinfine@126.com
  • 作者简介:杨亚,硕士,Email:1289971655@qq.com
  • 基金资助:
    国家自然科学基金(81400541);重庆高校创新团队建设计划(CXTDG201602006);重庆市高校市级口腔生物医学工程重点实验室资助项目(2014)

Change in expression of transformation growth factor-β/Smad signalling pathway-related proteins in epithelial rests of Malassez during orthodontic tooth movement in rats

Ya Yang,Peng Chen,Hongwei Dai,Lin Zhang()   

  1. Dept. of Orthodontics, Affiliated Stomatological Hospital of Chongqing Medical University, Chongqing Key Laboratory of Oral Diseases and Biomedical Sciences, Chongqing Municipal Key Laboratory of Oral Biomedical Engineering of Higher Education, Chongqing 401147, China
  • Received:2018-09-09 Revised:2018-12-28 Online:2019-05-01 Published:2019-06-05
  • Contact: Lin Zhang E-mail:zhanglinfine@126.com
  • Supported by:
    This study was supported by National Natural Science Foundation of China(81400541);Project Supported by Program for Innovation Team Building at Institutions of Higher Education in Chongqing(CXTDG201602006);Project Supported by Chongqing Municipal Key Laboratory of Oral Biomedical Engineering of Higher Education

摘要:

目的 通过检测大鼠正畸牙移动过程中转化生长因子(TGF)-β/Smad信号通路相关蛋白质在Malassez上皮剩余细胞(ERM)中的表达情况以及ERM的功能变化,探讨TGF-β/Smad信号通路调控ERM参与正畸牙移动的作用机制。方法 30只雄性8周龄健康Sprague-Dawley(SD)大鼠,在上颌右侧第一磨牙与切牙间安装镍钛拉簧,加载50 g力,作为实验组,上颌左侧作为对照组。于加力前以及加力后1、4、7、10、14 d处死大鼠,取下双侧包括上颌第一磨牙及周围牙槽骨在内的组织块,采用免疫组织化学染色方法,通过Image-Pro plus图像分析系统软件对实验组和对照组切片进行分析,在同一光强度下测量染色阳性信号的积分光密度(IOD),观察ERM表达的TGF-β1、Smad2、Smad3、增殖细胞核抗原(PCNA)的IOD值表达变化,并且比较实验组和对照组第一磨牙牙颈部和根分叉区ERM数量以及ERM集群表面积变化。所有数据使用SPSS 17.0软件进行t检验和秩和检验统计分析。结果 加力1 d后,TGF-β1、Smad2、Smad3免疫组织化学染色阳性增强,7 d后达到高峰,随后呈下降趋势;加力1、4、7、10、14 d组,TGF-β1、Smad2、Smad3的IOD值与对照组比较差异有统计学意义。加力4 d后,PCNA免疫组织化学染色阳性增强,加力7 d后明显增强,在第10 d达到高峰,随后呈下降趋势;加力4、7、10、14 d组,PCNA的IOD值与对照组比较差异有统计学意义。加力4、7、10、14 d组,ERM数量和ERM集群表面积与对照组比较差异有统计学意义。结论 在力学刺激的作用下,ERM的数量增多,ERM集群表面积也增大,TGF-β/Smad信号通路相关分子TGF-β1、Smad2、Smad3在ERM内表达,并且随加力时间的增加而呈现趋势性变化,表明ERM在机械力作用下通过TGF-β/Smad信号通路来调控其在正畸牙移动过程中的作用。

关键词: Malassez上皮剩余细胞, 正畸牙移动, 转化生长因子-β/Smad信号通路;

Abstract:

Objective This study aimed to investigate the expression of transformation growth factor (TGF)-β/Smad signal pathway-related proteins in the epithelial rests of Malassez (ERM) and the functional change in ERM during orthodontic tooth movement in rats to explore the mechanism by which the TGF-β/Smad signal pathway regulates ERM that is involved in tooth movement. Methods Tooth movement was achieved in 30 healthy male Sprague-Dawley rats (weighing 200-250 g each) by placing NiTi coil springs between the maxillary right first molar and incisor with the force of 50 g. The left side served as controls. Rats were killed at 0, 1, 4, 7, 10 and 14 days. Bilateral masses, including the maxillary first molars and the surrounding alveolar bone, were removed. Immunohistochemical staining was used to analyse the sections with the Image-Pro Plus Image Analysis System. The integrated optical density (IOD) of the stained positive signal was measured under the same light intensity. The IOD values of TGF-β1, Smad2, Smad3 and proliferative cell nuclear antigen (PCNA) were measured for all groups. The number of ERM was counted, and the surface areas of the ERM cluster were measured in the cervical and furcational regions of the first molar for all groups. The results were evaluated using the t-test and Kruskal-Wallis test with SPSS 17.0 software. Results The TGF-β1, Smad2, Smad3 and PCNA expression was weak in control rats. The immunofluorescence staining of TGF-β1, Smad2 and Smad3 increased after the 1st day, peaked on the 7th day and then decreased. On the 1st, 4th, 7th, 10th and 14th day, the IOD values of TGF-β1, Smad2 and Smad3 were statistically significant compared with those of the control group. After 4 days, PCNA immunohistochemistry positive staining increased. After 7 days, the PCNA expression level significantly increased, peaked on the 10th day and then decreased afterwards. On the 4th, 7th, 10th and 14th day, the IOD values of PCNA were statistically significant compared with those of the control group. The number of ERM and the surface area of the ERM cluster were statistically significant compared with those of the control group on the 4th, 7th, 10th and 14th day. Conclusion The number of ERM and the surface area of the ERM cluster both increased under a mechanical force. The TGF-β/Smad signalling pathway-related proteins (namely, TGF-β1, Smad2 and Smad3) were expressed in ERM and showed a trend change with the increase in force time. This result indicated that ERM can regulate its function in tooth movement through the TGF-β/Smad signalling pathway under a mechanical force.

Key words: epithelial cell rests of Malassez, orthodontic tooth movement, transformation growth factor-β/Smad signalling pathway;

中图分类号: 

  • R783.5

图 1

对照组ERM的TGF-β1、Smad2、Smad3表达情况 免疫组织化学染色 × 40 A:TGF-β1;B:Smad2;C:Smad3。PDL:牙周膜;R:牙根;AL:牙槽骨。箭头示ERM。"

图 2

加力1 d实验组ERM的TGF-β1、Smad2、Smad3表达情况 免疫组织化学染色 × 40 A:TGF-β1;B:Smad2;C:Smad3。PDL:牙周膜;R:牙根;AL:牙槽骨。箭头示ERM。"

图 3

加力7 d实验组ERM的TGF-β1、Smad2、Smad3表达情况 免疫组织化学染色 × 40 A:TGF-β1;B:Smad2;C:Smad3。PDL:牙周膜;R:牙根;AL:牙槽骨。箭头示ERM。"

图 4

加力14 d实验组ERM的TGF-β1、Smad2、Smad3表达情况 免疫组织化学染色 × 40 A:TGF-β1;B:Smad2;C:Smad3。PDL:牙周膜;R:牙根;AL:牙槽骨。箭头示ERM。"

表 1

正畸牙移动大鼠ERM的TGF-β1、Smad2、Smad3、PCNA表达"

时间/d 分组 TGF-β1 Smad2 Smad3 PCNA
0 实验组 47.7±7.0 38.0±10.0 38.2±9.7 23.0±2.6
对照组 43.8±3.2 34.4±1.5 37.2±1.5 21.2±1.5
1 实验组 72.8±4.8 68.6±4.2 69.3±4.9 22.8±2.4
对照组 48.0±2.4 42.2±2.4 39.0±2.1 21.5±1.4
4 实验组 136.2±6.4 142.9±4.9 143.5±5.2 41.2±7.7
对照组 53.4±0.7 45.1±3.1 43.1±1.4 25.1±1.2
7 实验组 276.0±5.9 253.6±11.3 248.2±8.6 76.2±7.1
对照组 52.9±1.6 43.7±4.0 52.8±2.0 32.5±1.5
10 实验组 201.7±5.9 178.9±1.5 179.8±2.0 155.0±5.3
对照组 52.9±1.6 51.1±2.6 38.6±1.0 30.4±2.1
14 实验组 135.9±5.0 123.8±6.7 122.2±5.2 127.6±3.0
对照组 55.6±1.9 41.2±1.6 44.1±1.3 35.8±2.1

图 5

ERM的PCNA表达情况 免疫组织化学染色 × 40 A:对照组;B:加力4 d实验组;C:加力10 d实验组。PDL:牙周膜;R:牙根;AL:牙槽骨。箭头示ERM。"

表 2

正畸牙移动大鼠ERM数量和ERM集群表面积"

时间/d 分组 ERM细胞数量 ERM细胞集群表面积
0 实验组 5.3±0.6 525.4±6.2
对照组 5.0±0.0 518.4±3.9
1 实验组 5.0±1.0 566.3±5.2
对照组 4.7±1.2 536.4±5.2
4 实验组 7.3±0.6 803.2±11.3
对照组 5.3±0.6 542.5±2.7
7 实验组 10.0±1.0 1 071.3±41.9
对照组 6.0±1.0 565.4±7.9
10 实验组 9.3±1.5 1 141.5±55.8
对照组 5.7±0.6 559.4±6.2
14 实验组 8.0±1.0 947.4±11.3
对照组 5.3±0.6 569.0±4.2
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