国际口腔医学杂志

国际口腔医学杂志 ›› 2020, Vol. 47 ›› Issue (6): 627-634.doi: 10.7518/gjkq.2020091

• 论著 • 上一篇    下一篇

小鼠正畸牙移动中牙周组织自噬相关基因表达的初步研究

尹圆圆(),马华钰,李昕怡,徐静晨,柳汀,陈嵩,何姝姝()   

  1. 口腔疾病研究国家重点实验室 国家口腔疾病临床医学研究中心 四川大学华西口腔医院正畸科 成都 610041
  • 收稿日期:2020-02-06 修回日期:2020-07-09 出版日期:2020-11-01 发布日期:2020-11-06
  • 通讯作者: 何姝姝
  • 作者简介:尹圆圆,博士,Email： 1042520312@qq.com
  • 基金资助:
    国家自然科学基金(81671021)

Expression of autophagy related genes in mice periodontal tissue during orthodontic tooth movement

Yin Yuanyuan(),Ma Huayu,Li Xinyi,Xu Jingchen,Liu Ting,Chen Song,He Shushu()   

  1. State Key Laboratory of Oral Diseases & National Clinical Research Center for Oral Diseases & Dept. of Orthodontics, West China Hospital of Stomatology, Sichuan University, Chengdu 610041, China
  • Received:2020-02-06 Revised:2020-07-09 Online:2020-11-01 Published:2020-11-06
  • Contact: Shushu He
  • Supported by:
    National Natural Science Foundation of China(81671021)

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摘要：

目的 研究小鼠正畸牙移动模型中移动牙牙周组织自噬相关基因的表达,探讨自噬在正畸牙移动骨塑建中的作用。方法 30只11周龄雄性C57B/6小鼠随机分为3组,每组10只。A组构建小鼠正畸牙移动模型,安放镍钛拉簧并施加矫治力;B组放置拉簧但不加矫治力;C组空白组。加力装置均安放于上颌左侧第一磨牙与左侧切牙之间,加力至第12 d处死小鼠。取左侧上颌骨标本,制作左侧上颌第一磨牙牙周组织标本切片,采用微计算机断层扫描技术测量牙移动距离,使用苏木精-伊红（HE）染色、抗酒石酸酸性磷酸酶（TRAP）染色和实时荧光定量聚合酶联式反应（RT-qPCR）观察牙移动情况,检测牙周组织中骨塑建相关基因[骨保护因子（OPG）和破骨细胞核因子κB受体活化因子配体（RANKL）]和自噬相关基因（包括Atg5、Atg7和Beclin-1）的表达。结果 A组平均牙移动距离0.09 mm,HE染色显示A组牙根张力侧与压力侧的牙周膜间隙不均等,压力侧细胞受到挤压、张力侧细胞受到拉伸。TRAP染色显示A组第一磨牙牙根近中侧见到阳性破骨细胞且A组压力侧破骨细胞数大于张力侧,差异有统计学意义（P<0.05）。RT-qPCR显示A组牙周组织OPG信使RNA（mRNA）表达显著降低,RANKL、Atg5、Atg7及Beclin-1 mRNA表达显著增加,差异有统计学意义（P<0.05）。结论 正畸力作用下牙周组织自噬相关基因表达水平升高,自噬可能通过影响牙周组织中破骨水平参与了正畸牙移动骨塑建过程。

关键词: C57B/6小鼠, 正畸牙移动, 自噬, 信使RNA, 骨塑建

Abstract:

Objective To investigate the level of autophagy related genes in periodontal tissue during orthodontic tooth movement in an experimental C57B/6 mice model and explore the role of autophagy related genes in bone modeling during orthodontic tooth movement. Methods Thirty-11-week-old male C57B/6 mice were randomly divided into 3 groups. Each group contained 10 mice. The experimental model of orthodontic tooth movement was established in group A by applying active force-loading nickel-titanium springs. Springs were placed in group B without force loading. Group C served as blank group. All the springs were placed between the left maxillary first molar and incisor. The mice were sacrificed on day 12, and the left lateral maxillae were collected to prepare sample sections of the periodontal tissue of the left maxillary first molars. Tooth movement was measured via microcomputed tomography. Hematoxylin and eosin (HE) staining, tartrate-resistant acid phosphatase (TRAP) staining, and real-time quantitative polymerase chain reaction (RT-qPCR) were performed, and the expression levels of bone modeling-related genes [osteoprotegerin (OPG) and receptor activator of nuclear factor-κ B ligand (RANKL)] and autophagy related genes (including Atg5, Atg7 and Beclin-1) were detected. Results The mean distance of first molar tooth movement was 0.09 mm in group A. HE staining showed that the amount of periodontal space between the pressure side and tension side of the first molars differed in group A. TRAP staining showed the presence of TRAP-positive cells in the pressure side of the first molar, and the number of osteoclasts in the pressure side was higher than that in the tension side (P<0.05). RT-qPCR results indicated that compared with those in group B, the messenger RNA (mRNA) expression of OPG in the periodontal tissue of group A decreased, whereas the mRNA expression levels of RANKL, Atg5, Atg7 and Beclin-1 increased (P<0.05). Conclusion Autophagy related genes increased in periodontal tissues under orthodontic force. Autophagy may play an important role in bone modeling during orthodontic tooth movement by regulating osteoclastogenesis.

Key words: C57B/6 mice, orthodontic tooth movement, autophagy, messenger RNA, bone modeling

中图分类号: 

  • R783.5

图1

建立小鼠牙移动模型"

表1

小鼠PCR相关引物序列"

引物 序列
Mus GAPDH-F CGAGAATGGGAAGCTTGTCA
Mus GAPDH-R TTGGCTCCACCCTTCAAGT
Mus OPG-F GCACAGTGAGGAGGAAGACA
Mus OPG-R TGTGTTTCGCTCTGGGGTT
Mus RANKL-F ACACCTCACCATCAATGCTG
Mus RANKL-R TCCCGATGTTTCATGATGC
Mus Atg5-F GCCATCAACCGGAAACTC
Mus Atg5-R CAAGTGTGTGCAGCTGTCCA
Mus Atg7-F AAACCCCATGCTCCTCAACA
Mus Atg7-R GATCCAAGCTCACAGGTCCC
Mus Beclin-1-F ACGCTGTTTGGAGATCCTAG
Mus Beclin-1-R TTCGTCATCCAACTCCAGCT

图2

MicroCT检测第一磨牙移动情况 × 400 a、d：A组横截面与纵截面;b、e：B组横截面与纵截面;c、f：C组横截面与纵截面。"

图3

第一磨牙张压力侧牙周膜组织形态观察 HE染色 × 200 a、d：A组张力侧与压力侧;b、e：B组张力侧与压力侧;c、f：C组张力侧与压力侧;AB：牙槽骨;PDL：牙周膜;R：牙根。"

图4

第一磨牙张压力侧破骨细胞观察 TRAP染色 × 200 a、d：A组张力侧与压力侧;b、e：B组张力侧与压力侧;c、f：C组张力侧与压力侧;AB：牙槽骨;R：牙根。"

图5

第一磨牙张压力侧TRAP染色阳性破骨细胞计数 A：加力组;B：无力拉簧组;C：全空白组。**：P<0.01。"

图6

第一磨牙牙周膜OPG、RANKL、Atg5、Atg7和Beclin-1的mRNA相对表达量 A：加力组;B：无力拉簧组;C：全空白组。**：P<0.01。"

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