口腔医学研究

口腔医学研究 ›› 2021, Vol. 37 ›› Issue (10): 900-905.DOI: 10.13701/j.cnki.kqyxyj.2021.10.008

• 牙周病学研究 • 上一篇    下一篇

T型钙通道对人牙囊细胞成骨分化的影响

杨晶晶1,2, 左东川3, 谢沂航1, 蔡欣1, 袁小平1,2, 曾锦1,2*   

  1. 1.西南医科大学附属口腔医院正畸科 四川 泸州 646000;
    2.西南医科大学口颌面修复重建和再生实验室 四川 泸州 646000;
    3.西南医科大学电生理学教育部重点实验室,心血管医学研究所 四川 泸州 646000
  • 收稿日期:2021-05-10 出版日期:2021-10-28 发布日期:2021-10-18
  • 通讯作者: *曾锦,E-mail:zengjin@swmu.edu.cn
  • 作者简介:杨晶晶(1995~ ),女,四川达州人,硕士,医师,研究方向:口腔正畸学。
  • 基金资助:
    泸州市-西南医科大学联合项目(编号:2020LZXNYDJ05)四川省泸州市科技局应用基础研究项目(编号:2019-JYJ-55)

Effect of T-type Calcium Channel on Osteogenic Differentiation of Human Dental Follicle Cells

YANG Jingjing1,2, ZUO Dongchuan3, XIE Yihang1, CAI Xin1, YUAN Xiaoping1,2, ZENG Jin1,2*   

  1. 1. Department of Orthodontics, the Affiliated Stomatology Hospital of Southwest Medical University, Luzhou 646000, China;
    2. Laboratory of Oral and Maxillofacial Reconstruction and Regeneration, Southwest Medical University, Luzhou 646000, China;
    3. Institute of Cardiovascular Medicine, Key Laboratory of Electrophysiology of Ministry of Education, Southwest Medical University, Luzhou 646000, China
  • Received:2021-05-10 Online:2021-10-28 Published:2021-10-18

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摘要： 目的:探讨Ca2+对人牙囊细胞(hDFCs)成骨分化的影响及其分子机制。方法:分离培养hDFCs,免疫荧光染色及流式细胞术检测hDFCs的来源。反转录聚合酶链反应(RT-PCR)检测L型钙通道、T型钙通道、三磷酸肌醇受体以及雷诺定受体在hDFCs的表达及-成骨分化相关基因 RUNX 相关转录因子 2(runt-related transcription factor 2,RUNX2)、骨钙素(osteocalcin,OCN)的表达。茜素红染色分别检测移除细胞外Ca2+、细胞内Ca2+以及应用电压门控钙通道及Ca2+/CaMK Ⅱ信号通路特异阻断剂后,hDFCs的成骨水平。结果:移除细胞外Ca2+或细胞内Ca2+后,hDFCs成骨能力显著降低。RT-PCR结果显示,hDFCs表达L型钙通道、T型钙通道、三磷酸肌醇受体以及雷诺定受体。L型钙通道阻断剂Nifedipine对细胞成骨能力没有影响,而T型钙通道阻断剂Amlioride能够明显抑制细胞的成骨水平。提高细胞外Ca2+的浓度(5 mmol/L)能够促进hDFCs成骨分化,应用CaMK Ⅱ特异性阻断剂KN-93(10 μmol/L)后,Ca2+对hDFCs成骨分化的促进作用明显受到抑制。结论:T型钙通道介导的钙离子内流参与调控hDFCs的成骨分化。

关键词: 人牙囊细胞, 钙离子, T型钙通道, 成骨分化

Abstract: Objective: To investigate the effect of Ca2+ on osteogenic differentiation of human dental follicle cells (hDFCs) and to explore its molecular mechanism. Methods: hDFCs were isolated and cultured. Its source was verified by flow cytometry and immunofluorescence staining. The gene expression of L-type calcium channel, T-type calcium channel, inositol triphosphate receptor, and ryanodine receptor of hDFCs and mRNA expression of osteogenic differentiation related gene were detected by reverse transcription polymerase chain reaction (RT-PCR). Alizarin red staining was used to evaluate the formation of calcium nodules after the removal of extracellular Ca2+, intracellular Ca2+, and the application of voltage-gated calcium channel specific blockers. Results: The osteogenic ability of hDFCs was significantly decreased after removal of extracellular Ca2+ or intracellular Ca2+. RT-PCR results showed that hDFCs expressed L-type calcium channel, T-type calcium channel, inositol triphosphate receptor, and ryanodine receptor. Nifedipine, an L-type calcium channel blocker, had no effect on the osteogenic mineralization of the cells, while amlioride, a T-type calcium channel blockers significantly inhibited the osteogenic mineralization of the cells. Elevation of extracellular Ca2+ concentration (5 mmol/L) significantly promoted osteogenic differentiation of hDFCs, and the effect was inhibited by application of KN-93 (10 μmol/L, a Ca2+/CaMK Ⅱ signaling pathway specific blocker). Conclusion: T-type calcium channel mediated calcium influx is involved in the osteogenic differentiation of hDFCs.

Key words: human dental follicle cells, Ca2+, T-type calcium channel, osteogenic differentiation