中国组织工程研究

中国组织工程研究 ›› 2023, Vol. 27 ›› Issue (10): 1521-1527.doi: 10.12307/2023.284

• 牙髓及牙周膜干细胞 Dental pulp and periodontal ligament stem cells • 上一篇    下一篇

巨噬细胞炎性蛋白1α对人牙周膜干细胞生物学行为的影响

王钊鑫，尼加提·吐尔逊，代慧娟，王俊祥，霞黒达·依拉尔江，李淑慧   

  1. 新疆医科大学第二附属医院口腔科，新疆维吾尔自治区乌鲁木齐市   830063
  • 收稿日期:2022-03-04 接受日期:2022-05-20 出版日期:2023-04-08 发布日期:2022-09-08
  • 通讯作者: 李淑慧，博士，副主任医师，新疆医科大学第二附属医院口腔科，新疆维吾尔自治区乌鲁木齐市 830063
  • 作者简介:王钊鑫，女，1997年生，湖北省麻城市人，汉族，新疆医科大学在读硕士，主要从事口腔种植与修复研究。
  • 基金资助:
    新疆维吾尔自治区科学基金(2019D01C231)，项目负责人：李淑慧

Effect of macrophage inflammatory protein-1 alpha on the biological behavior of human periodontal ligament stem cells

Wang Zhaoxin, Nijati•Tursun, Dai Huijuan, Wang Junxiang, Xiaheida•Yilaljiang, Li Shuhui   

  1. Department of Stomatology, Second Affiliated Hospital of Xinjiang Medical University, Urumqi 830063, Xinjiang Uygur Autonomous Region, China
  • Received:2022-03-04 Accepted:2022-05-20 Online:2023-04-08 Published:2022-09-08
  • Contact: Li Shuhui, MD, Associate chief physician, Department of Stomatology, Second Affiliated Hospital of Xinjiang Medical University, Urumqi 830063, Xinjiang Uygur Autonomous Region, China
  • About author:Wang Zhaoxin, Master candidate, Department of Stomatology, Second Affiliated Hospital of Xinjiang Medical University, Urumqi 830063, Xinjiang Uygur Autonomous Region, China
  • Supported by:
    Science Foundation of Xinjiang Uygur Autonomous Region, No. 2019D01C231 (to LSH)

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摘要：

文题释义：
人牙周膜干细胞：是来源于牙周膜组织中的间充质干细胞，具有自我更新和多向分化潜能，是牙周组织修复的重要细胞群，其可以分化为成骨细胞、成脂细胞和成软骨细胞等，也能够形成牙周膜样结构。人牙周膜干细胞可以重建口腔中被牙周疾病损伤的牙周膜组织，并有望通过成骨向分化潜能促进牙周围骨组织缺损的再生修复。
巨噬细胞炎性蛋白1α：又称为趋化因子配体3(CCL3)，可由B细胞、中性粒细胞等多种细胞产生，属于CC趋化因子家族，对单核巨噬细胞、T淋巴细胞、B淋巴细胞等具有趋化、活化作用，并可穿透血管内皮细胞，结合特异性受体，刺激炎性因子产生，发挥促炎作用，在口腔疾病中具有重要作用。
Notch信号通路：是高度保守的特定信号通路，决定细胞代谢和命运，在细胞增殖、生长发育和细胞生物学行为中发挥重要作用。Notch信号通路由Notch1、Notch2、Notch3、Notch4、Jagged1、Jagged2、Dll1、Dll3、Dll4配体和受体激活发生作用，相邻细胞裂解，Notch蛋白移位于细胞内，从而调控Notch通路。Notch作为关键的信号通路在牙周组织再生领域研究中备受关注。

背景：目前大量研究集中于炎症因子对人牙周膜干细胞的影响，然而关于趋化因子巨噬细胞炎性蛋白1α(macrophage inflammatory protein-1α，MIP-1α)在人牙周膜干细胞中的表达，国内外相关文献报道较少。
目的：探究不同质量浓度MIP-1α对人牙周膜干细胞增殖、凋亡及迁移的影响及与Notch信号通路的相关性。
方法：将12-25岁患者(均签署知情同意书)拔下的健康正畸减数前磨牙，通过组织块法联合酶消化法分离培养人牙周膜干细胞。将第3代对数生长期人牙周膜干细胞分为对照组和1，10，100 mg/L MIP-1α组，干预第1，3，5，7天采用CCK-8法检测细胞增殖能力，干预48 h采用流式检测细胞凋亡能力，Transwell小室检测细胞迁移能力，ELISA检测分泌肿瘤坏死因子α水平，qRT-PCR、Western blot检测Notch1、Hes1 mRNA和蛋白表达量。
结果与结论：①1 mg/L MIP-1α对人牙周膜干细胞生物学行为无明显影响，10 mg/L MIP-1α显著促进人牙周膜干细胞的增殖及迁移，而100 mg/L MIP-1α显著促进人牙周膜干细胞的凋亡，抑制人牙周膜干细胞的增殖和迁移；②1 mg/L MIP-1α对人牙周膜干细胞中Notch1及Hes1蛋白表达无明显影响；10 mg/L MIP-1α对人牙周膜干细胞中Notch1蛋白表达无显著影响，但可下调细胞中Hes1蛋白表达水平，100 mg/L MIP-1α可上调细胞中Notch1及Hes1蛋白表达水平；③1 mg/L MIP-1α组人牙周膜干细胞分泌的肿瘤坏死因子α水平与对照组无显著差异；10 mg/L MIP-1α促进肿瘤坏死因子α分泌；100 mg/L MIP-1α抑制肿瘤坏死因子α分泌；④结果表明：10 mg/L MIP-1α促进人牙周膜干细胞增殖、迁移及肿瘤坏死因子α分泌，其机制可能与下调Notch信号通路相关；100 mg/L MIP-1α抑制人牙周膜干细胞增殖、迁移及肿瘤坏死因子α分泌，促进人牙周膜干细胞凋亡，其机制可能与上调Notch信号通路有关。

https://orcid.org/0000-0002-1538-5818 (王钊鑫)；https://orcid.org/0000-0001-5190-4163 (李淑慧) 

中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程

关键词: 人牙周膜干细胞, 巨噬细胞炎性蛋白1α, Notch信号通路, 增殖, 迁移, 凋亡

Abstract: BACKGROUND: A large number of studies focus on the effect of inflammatory factors on human periodontal ligament stem cells; however, there are few reports on the expression of chemokine macrophage inflammatory protein-1α in human periodontal ligament stem cells at home and abroad.  
OBJECTIVE: To investigate the effects of different mass concentrations of macrophage inflammatory protein 1α on the proliferation, apoptosis, and migration of human periodontal ligament stem cells and the correlation of Notch signaling pathway.
METHODS:  Healthy orthodontic reduced premolars were extracted from patients aged 12-25 years (all signed informed consent), and human periodontal ligament stem cells were isolated and cultured by tissue block method combined with enzymatic digestion. Passage 3 human periodontal ligament stem cells at logarithmic growth stage were divided into control group, 1, 10, 100 mg/L macrophage inflammatory protein-1α groups. Cell proliferation was detected by CCK-8 assay at 1, 3, 5, and 7 days. At 48 hours, the apoptotic ability of each group was detected by flow cytometry. Cell migration ability of each group was detected by Transwell chamber test. ELISA was used to detect the level of secreted tumor necrosis factor α. qRT-PCR and western blot assay were used to detect the mRNA and protein expression levels of Notch1 and Hes1.  
RESULTS AND CONCLUSION: (1) 1 mg/L macrophage inflammatory protein-1α had no significant effect on the biological behavior of human periodontal ligament stem cells. 10 mg/L macrophage inflammatory protein-1α significantly promoted the proliferation and migration of human periodontal ligament stem cells, and 100 mg/L macrophage inflammatory protein-1α significantly promoted the apoptosis of human periodontal ligament stem cells, inhibited the proliferation and migration of human periodontal ligament stem cells. (2) 1 mg/L macrophage inflammatory protein-1α had no significant effect on the expression of Notch1 and Hes1 proteins in human periodontal ligament stem cells. 10 mg/L macrophage inflammatory protein-1α had no significant effect on the expression of Notch1 protein in human periodontal ligament stem cells, but could down-regulate the expression level of Hes1 protein in the cells. 100 mg/L macrophage inflammatory protein-1α could up-regulate the expression of Notch1 and Hes1 protein in cells. (3) The level of tumor necrosis factor α secreted by  human periodontal ligament stem cells of the 1 mg/L macrophage inflammatory protein-1α group was not significantly different from that of the control group. 10 mg/L macrophage inflammatory protein-1α promoted the secretion of tumor necrosis factor α; 100 mg/L macrophage inflammatory protein-1α inhibited the secretion of tumor necrosis factor α. (4) These results exhibited that 10 mg/L macrophage inflammatory protein-1α promoted the proliferation, migration and tumor necrosis factor α secretion of human periodontal ligament stem cells, and the mechanism might be related to the down-regulation of Notch signaling pathway. 100 mg/L macrophage inflammatory protein-1α inhibited the proliferation and migration of human periodontal ligament stem cells and the secretion of tumor necrosis factor α, and promoted the apoptosis of human periodontal ligament stem cells. The mechanism may be related to the up-regulation of Notch signaling pathway.

Key words: human periodontal ligament stem cell, macrophage inflammatory protein-1α, Notch signaling pathway, proliferation, migration, apoptosis

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