中国组织工程研究

中国组织工程研究 ›› 2022, Vol. 26 ›› Issue (20): 3164-3172.doi: 10.12307/2022.615

• 口腔组织构建 oral tissue construction • 上一篇    下一篇

浓缩生长因子与表皮生长因子干预口腔黏膜等效细胞的增殖和衰老

李腾艳1，2，聂敏海1，2，刘旭倩1，2   

  1. 1西南医科大学附属口腔医院牙周黏膜病科，四川省泸州市 646000；2西南医科大学口颌面修复重建和再生实验室，四川省泸州市 646000
  • 收稿日期:2021-04-06 接受日期:2021-06-09 出版日期:2022-07-18 发布日期:2022-01-19
  • 通讯作者: 刘旭倩，博士，副教授，西南医科大学附属口腔医院牙周黏膜病科，四川省泸州市 646000；西南医科大学口颌面修复重建和再生实验室，四川省泸州市 646000
  • 作者简介:李腾艳，女，1993年生，四川省叙永县人，汉族，硕士，医师，主要从事口腔内科学研究。
  • 基金资助:
    四川省科技厅科技计划项目(2020YJ0387)，项目负责人：刘旭倩；四川省泸州市人民政府-西南医科大学科技战略合作重点项目(2019LZXNYDZ10)，项目负责人：刘旭倩

Effects of concentrated growth factor combined with epidermal growth factor on the proliferation and aging of oral mucosa equivalents

Li Tengyan1, 2, Nie Minhai1, 2, Liu Xuqian1, 2   

  1. 1Department of Periodontal Mucosal Diseases, Affiliated Hospital of Stomatology, Southwest Medical University, Luzhou 646000, Sichuan Province, China; 2Laboratory of Oral and Maxillofacial Reconstruction and Regeneration, Southwest Medical University, Luzhou 646000, Sichuan Province, China
  • Received:2021-04-06 Accepted:2021-06-09 Online:2022-07-18 Published:2022-01-19
  • Contact: Liu Xuqian, MD, Associate professor, Department of Periodontal Mucosal Diseases, Affiliated Hospital of Stomatology, Southwest Medical University, Luzhou 646000, Sichuan Province, China; Laboratory of Oral and Maxillofacial Reconstruction and Regeneration, Southwest Medical University, Luzhou 646000, Sichuan Province, China
  • About author:Li Tengyan, Master, Physician, Department of Periodontal Mucosal Diseases, Affiliated Hospital of Stomatology, Southwest Medical University, Luzhou 646000, Sichuan Province, China; Laboratory of Oral and Maxillofacial Reconstruction and Regeneration, Southwest Medical University, Luzhou 646000, Sichuan Province, China
  • Supported by:
    the Science and Technology Planning Project of Sichuan Provincial Department of Science and Technology, No. 2020YJ0387 (to LXQ); Luzhou Municipal People’s Government-Southwest Medical University Science and Technology Strategic Cooperation Key Project, No. 2019LZXNYDZ10 (to LXQ)

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摘要：

文题释义：
浓缩生长因子(concentrate growth factors，CGF)：是第3代血小板制品，利用静脉血通过差速离心程序制备，其中含有多种生长因子及纤维蛋白。
表皮生长因子(epidermal growth factor，EGF)：是一种广泛存在于人或其它动物体内的小分子多肽，极微量即能强烈刺激细胞生长、抑制衰老基因的出现，延缓表皮细胞衰老。表皮生长因子通过与细胞表面的受体表皮生长因子受体(EGFR)结合而起作用。

背景：浓缩生长因子与表皮生长因子是近年来被广泛研究的生物学材料，经研究证实其可通过影响细胞生物学行为，进而促进组织修复与损伤愈合。
目的：探讨浓缩生长因子联合表皮生长因子对人牙龈成纤维细胞、上皮细胞、人正常黑色素细胞系正常人皮肤黑色素细胞增殖和衰老的影响。
方法：①浓缩生长因子/表皮生长因子与人牙龈成纤维细胞、上皮细胞、正常人皮肤黑色素细胞体外共培养，MTT分别筛选出浓缩生长因子、表皮生长因子、浓缩生长因子+表皮生长因子的最佳作用浓度；②实验分组：人牙龈成纤维细胞对照组、50% 浓缩生长因子组、10 μg/L表皮生长因子组、50%浓缩生长因子+10 μg/L表皮生长因子组；上皮细胞对照组、30% 浓缩生长因子组、10 μg/L表皮生长因子组、30% 浓缩生长因子+10 μg/L表皮生长因子组；正常人皮肤黑色素细胞对照组、30% 浓缩生长因子组、20 μg/L表皮生长因子组、30% 浓缩生长因子+10 μg/L表皮生长因子组、30% 浓缩生长因子+20 μg/L表皮生长因子组；③CCK-8检测细胞增殖，Transwell检测细胞迁移，Annexin V/PI双染色法检测细胞凋亡。研究通过西南医科大学附属口腔医院生物医学科学研究伦理委员会审查(申请受理号：20181008001)及患者和志愿者知情同意。
结果与结论：①MTT结果提示，浓缩生长因子对人牙龈成纤维细胞、上皮细胞、正常人皮肤黑色素细胞增殖影响的最佳质量浓度分别为50%，30%，30%；表皮生长因子对人牙龈成纤维细胞、上皮细胞、正常人皮肤黑色素细胞增殖影响的最佳质量浓度分别是10，10，20 μg/L；与既定最佳浓度浓缩生长因子协同作用，对3种细胞增殖影响最佳的表皮生长因子质量浓度为10 μg/L；②CCK-8、Transwell、Annexin V/PI双染色法结果显示，浓缩生长因子联合表皮生长因子可促进人牙龈成纤维细胞、上皮细胞、正常人皮肤黑色素细胞增殖、迁移及抑制细胞凋亡，作用效应呈现出表皮生长因子>浓缩生长因子 + 表皮生长因子>浓缩生长因子的趋势；③浓缩生长因子联合表皮生长因子促进细胞增殖、迁移和抑制细胞凋亡的效应较浓缩生长因子单独作用强，较表皮生长因子单独作用弱。

https://orcid.org/0000-0001-6662-2265 (李腾艳) 

中国组织工程研究杂志出版内容重点：组织构建；骨细胞；软骨细胞；细胞培养；成纤维细胞；血管内皮细胞；骨质疏松；组织工程

关键词: 浓缩生长因子, 表皮生长因子, 增殖, 迁移, 凋亡, 衰老, 口腔黏膜

Abstract: BACKGROUND: Concentrated growth factor (CGF) and epidermal growth factor (EGF) are biological materials that have been extensively studied in recent years. Studies have shown that they can affect the biological behavior of cells, thereby promoting tissue repair and injury healing.
OBJECTIVE: To explore the effects of CGF combined with EGF on the proliferation and aging of human gingival fibroblasts, epithelial cells, and human normal melanocytes.
METHODS: CGF/EGF was co-cultured with human gingival fibroblasts, epithelial cells and human normal melanocytes in vitro. The MTT assay was used to screen the optimal concentrations of CGF, EGF and their combination. Experiments were divided into: human gingival fibroblast control group, 50% CGF group, 10 μg/L EGF group, 50% CGF+10 μg/L EGF group; epithelial cell control group, 30% CGF group, 10 μg/L EGF, 30% CGF + 10 μg/L EGF group; human skin melanocyte control group, 30% CGF group, 20 μg/L EGF group, 30% CGF + 10 μg/L EGF group, 30% CGF + 20 μg/L EGF group. Cell counting kit-8 assay was used to detect the proliferation of cells, Transwell technology used to detect the migration of cells, and Annexin V/PI double staining method used to detect the apoptosis of cells. An ethical approval was obtained from the Biomedical Science Research Ethics Committee of the Affiliated Stomatological Hospital of Southwest Medical University (approval No. 20181008001) and written informed consents were obtained from patients and volunteers.
RESULTS AND CONCLUSION: The results of MTT assay suggested that the optimal concentrations of CGF on the cell proliferation were 50% for human gingival fibroblasts, 30% for epithelial cells and 30% for human normal melanocytes, while the optimal concentrations of EGF on the cell proliferation were 10 μg/L for human gingival fibroblasts, 10 μg/L for epithelial cells, and 20 μg/L for human normal melanocytes. Cell counting kit-8 assay, Transwel assay and Annexin V/PI double staining method showed that CGF combined with EGF could promote the proliferation and migration of human gingival fibroblasts, epithelial cells and human normal melanocytes and inhibit cell apoptosis. Their effects were ranked as follows: EGF > CGF + EGF > CGF. To conclude, CGF combined with EGF can promote cell proliferation and migration and inhibit cell apoptosis, and its effect is stronger than that of CGF alone but weaker than that of EGF alone.

Key words: concentrated growth factor, epidermal growth factor, proliferation, migration, apoptosis, aging, oral mucosa

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