口腔医学研究

口腔医学研究 ›› 2019, Vol. 35 ›› Issue (6): 531-536.DOI: 10.13701/j.cnki.kqyxyj.2019.06.006

• 牙体牙髓病学研究 • 上一篇    下一篇

细胞自噬在牙髓损伤修复过程中的作用

陈杰1,2,徐华兴1,2,程抒华1,2,张旗1,2*   

  1. 1. 同济大学口腔医学院·同济大学附属口腔医院牙体牙髓科 上海 200072;
    2. 上海牙组织修复与再生工程技术研究中心 上海 200072
  • 收稿日期:2019-03-24 出版日期:2019-06-28 发布日期:2019-06-27
  • 通讯作者: 张旗,E-mail:qizhang@tongji.edu.cn
  • 作者简介:陈杰(1990~ ),女,江苏淮安人,硕士在读,研究方向为牙髓再生与牙齿硬组织再生。
  • 基金资助:
    国家自然科学基金(编号:81870760、81570966)

Effects of Autophagy in Dental Pulp Repair

CHEN Jie1,2, XU Hua-xing1,2, CHENG Shu-hua1,2, ZHANG Qi1,2*   

  1. 1. Department of Endodontics, School and Hospital of Stomatology, Tongji University, Shanghai 200072, China;
    2. Shanghai Engineering Research Center of Tooth Restoration and Regeneration, Shanghai 200072, China.
  • Received:2019-03-24 Online:2019-06-28 Published:2019-06-27

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摘要： 目的:研究大鼠牙髓损伤修复过程中细胞自噬表达的变化,探讨其在这一过程中可能发挥的作用。方法:60只8周龄SD大鼠上颌第一磨牙开髓后直接盖髓,术后第1、3、7、14天取材,苏木素-伊红(hematoxylin-eosin,HE)染色、Masson染色观察牙髓损伤修复情况,免疫组织荧光染色检测炎症因子肿瘤坏死因子-α(tumor necrosis factor-α, TNF-α)、CD68以及自噬蛋白LC3B和Beclin1的表达。分别提取损伤牙髓组织及体外培养牙髓细胞加脂多糖(lipopolysaccharide,LPS),实时荧光定量聚合酶链反应(quantitative Real-time PCR)检测损伤后不同时间点炎症、矿化和自噬相关基因的表达变化。结果:术后第1天大量中性粒细胞浸润,第3天炎症浸润逐渐减轻,第14天炎症明显好转,有新胶原纤维形成(P<0.05)。这一过程可见自噬蛋白LC3B、Beclin1主要表达在牙髓细胞、成牙本质细胞及血管内皮细胞胞质及胞膜上。mRNA水平表明炎症因子白细胞介素-1β(interleukin-1β,IL-1β)、CD68、TNF-α在损伤后3天表达达高峰,随后降低;矿化指标Ⅰ型胶原(collagen type I,COL1)、牙本质涎磷蛋白(dentin sialophosphoprotein,DSPP)及牙本质基质蛋白1 (dentin matrix protein1,DMP1)在损伤修复过程中表达逐渐升高。自噬基因LC3B与Atg5在损伤后12 h表达量至高峰,24 h表达量最低,随后逐渐升高;Atg12与Beclin1在损伤后出现先降低后升高趋势。体外细胞与体内组织mRNA趋势一致,差异有统计学意义(P<0.05)。结论:牙髓损伤修复过程中,自噬参与牙髓炎症的发生发展,并促进牙髓组织修复。

关键词: 细胞自噬, 牙髓炎症, 牙髓损伤修复

Abstract: Objective: To investigate the roles of autophagy in dental pulp injury and repair process by observing the changes of its expression after injury. Methods: Sixty 8-week-old male SD rats were exposed and directly capped on the first maxillary molars. Rats were sacrificed on days 1, 3, 7, and 14, dental pulp repair was detected with HE and Masson's trichrome staining. Inflammation markers TNF-α, CD68, and autophagy proteins LC3B and Beclin1 were detected by immunofluorescence staining. Damaged pulp tissue and dental pulp cells (DPCs) treated with LPS in vitro were extracted to detect the expression of inflammation, mineralization, and autophagy genes by real-time quantitate PCR. Results: Inflammation was increased gradually from 1st to 3rd day after operation, but no obvious inflammation was observed at 14th day. New collagen fibers were formed (P<0.05). Autophagy related proteins were mainly expressed on the cytoplasm and membrane of DPCs, odontoblasts, and vascular endothelial cells. The mRNA level of tissue showed that inflammatory markers IL-1β, CD68, and TNF-α were peaked at 3th day and then gradually decreased; the mineralization markers COL1, DSPP, and DMP1 were increased gradually after injury. LC3B and Atg5 reached the peak after 12 hours, decreased to the bottom after 24 hours, and then gradually increased; Atg12 and Beclin1 showed a trend of decreasing first and then increasing. The trend of mRNA in DPCs was consistent with pulp tissue (P<0.05). Conclusion: During the process of dental pulp injury and repair, autophagy participates in the development of pulp inflammation and promotes the repair of pulp tissue.

Key words: Autophagy, Pulp inflammation, Pulp repair