中国组织工程研究

中国组织工程研究 ›› 2021, Vol. 25 ›› Issue (10): 1512-1516.doi: 10.3969/j.issn.2095-4344.3032

• 组织工程软骨材料Tissue-engineered cartilage • 上一篇    下一篇

脱细胞软骨细胞外基质对小鼠巨噬细胞系表型的调控

李珺琦1，2，田广招2，陈明学2，王  皓2，刘舒云2，眭  翔2，黄靖香2，李  明3，郭全义2   

  1. 长治医学院，1基础医学部，3解剖教研室，山西省长治市   046000；2中国人民解放军总医院第一医学中心骨科研究所，骨科再生医学北京市重点实验室，全军骨科战创伤重点实验室，北京市   100853
  • 收稿日期:2020-03-28 修回日期:2020-04-02 接受日期:2020-05-13 出版日期:2021-04-08 发布日期:2020-12-17
  • 通讯作者: 郭全义，博士，主任医师，教授，博士生导师，中国人民解放军总医院第一医学中心骨科研究所，骨科再生医学北京市重点实验室，全军骨科战创伤重点实验室，北京市 100853
  • 作者简介:李珺琦，女，1997年生，黑龙江省齐齐哈尔市人，汉族，长治医学院本科在读。
  • 基金资助:
    国家重点研发计划项目(2018YFC1105900) ；国家自然科学基金委员会资助项目(81772319)

Regulatory effect of acellular cartilage extracellular matrix on phenotype of mouse macrophage line

Li Junqi1, 2, Tian Guangzhao2, Chen Mingxue2,  Wang Hao2,  Liu Shuyun2,  Sui Xiang2,  Huang Jingxiang2,  Li Ming3,  Guo Quanyi2   

  1. 1Department of Basic Medicine, 3Department of Anatomy, Changzhi Medical College, Changzhi 046000, Shanxi Province, China; 2Institute of Orthopedics of First Medical Center of Chinese PLA General Hospital, Beijing Key Laboratory of Orthopedic Regenerative Medicine, Key Laboratory of Military Orthopedic Warfare Trauma, Beijing 100853, China 
  • Received:2020-03-28 Revised:2020-04-02 Accepted:2020-05-13 Online:2021-04-08 Published:2020-12-17
  • Contact: Guo Quanyi, MD, Chief physician, Professor, Doctoral supervisor, Institute of Orthopedics of First Medical Center of Chinese PLA General Hospital, Beijing Key Laboratory of Orthopedic Regenerative Medicine, Key Laboratory of Military Orthopedic Warfare Trauma, Beijing 100853, China
  • About author:Li Junqi, Department of Basic Medicine, Changzhi Medical College, Changzhi 046000, Shanxi Province, China; Institute of Orthopedics of First Medical Center of Chinese PLA General Hospital, Beijing Key Laboratory of Orthopedic Regenerative Medicine, Key Laboratory of Military Orthopedic Warfare Trauma, Beijing 100853, China
  • Supported by:
    the National Key Research and Development Project, No. 2018YFC1105900 ; the National Natural Science Foundation of China, No. 81772319

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摘要：

文题释义：
细胞外基质：是由各种组织和细胞合成并分泌到胞外、分布在细胞表面或细胞之间的大分子, 主要是一些多糖和蛋白聚糖等构成的复杂网架结构，有支持并连接组织结构、调节组织的发生和细胞的生理活动。
巨噬细胞：是天然免疫系统中重要组成部分，能消灭侵入机体的细菌，吞噬异物颗粒，消除体内衰老、损伤的细胞和变性的细胞间质、杀伤肿瘤细胞，并参与免疫反应。

背景：前期研究证实脱细胞软骨细胞外基质可有效促进关节软骨缺损修复再生，具有良好的修复效果，但是其促再生机制尚未阐明。
目的：探究猪来源脱细胞软骨细胞外基质支架对小鼠巨噬细胞系表型极化的调控。
方法：以猪膝关节软骨为原料，通过湿法粉碎、差速离心、冷冻干燥的方法制备脱细胞软骨细胞外基质支架。将小鼠RAW264.7巨噬细胞系接种于脱细胞软骨细胞外基质支架，构建细胞-支架复合物，体外极化诱导4 d，采用扫描电镜观察细胞黏附情况，死/活细胞染色观察细胞生长情况，免疫荧光染色观察M1型巨噬细胞特征性表面标志物CD86和M2型巨噬细胞特征性表面标志物CD206的表达，进而明确极化诱导情况。
结果与结论：①扫描电镜显示，复合物中的巨噬细胞为原来的圆形或类圆形，沿着支架的管状排列，广泛分布在支架表面及内部结构中；②死/活细胞染色显示，支架上附着的大部分细胞为活细胞，仅有极少数死细胞；③免疫荧光染色显示，复合物中的M2型巨噬细胞标志物CD206呈80%的高阳性表达，M1型巨噬细胞标志物CD86的阳性表达甚少；④结果表明，猪来源脱细胞软骨细胞外基质支架可诱导极化M0型巨噬细胞为修复性M2型巨噬细胞。

https://orcid.org/0000-0003-3457-4099 (李珺琦) 

中国组织工程研究杂志出版内容重点：生物材料；骨生物材料; 口腔生物材料; 纳米材料; 缓释材料; 材料相容性；组织工程

关键词: 骨, 软骨, 再生, 细胞外基质, 巨噬细胞, 极化, 组织工程, 免疫

Abstract: BACKGROUND: Previous studies have confirmed that the extracellular matrix of acellular cartilage can effectively promote the repair and regeneration of articular cartilage defects, and has a good repair effect, but its mechanism of promoting regeneration has not been elucidated.
OBJECTIVE: To investigate the regulation of phenotypic polarization of mouse macrophage cell line by extracellular matrix scaffolds derived from pigs.
METHODS: Using pig knee articular cartilage as raw material, the decellularized cartilage extracellular matrix scaffold was prepared by wet grinding, differential centrifugation, and freeze drying. The mouse RAW264.7 macrophage cell line was inoculated on the decellularized cartilage extracellular matrix scaffold to construct a cell-scaffold complex, which induced polarization in vitro for 4 days. Cell adhesion was observed using scanning electron microscopy, and cell growth was observed with dead/viable cells. Immunofluorescence staining was used to observe the expression of CD86, a characteristic surface marker of M1-type macrophages, and CD206, a characteristic surface marker of M2-type macrophages, and to clarify the polarization induction.
RESULTS AND CONCLUSION: (1) Scanning electron microscopy showed that the macrophages in the complex were originally round or quasi round, arranged along the tube of the scaffold, and widely distributed on the surface and internal structure of the scaffold. (2) Dead/live cell staining showed that most of the cells attached to the scaffold were living cells, with only a few dead cells. (3) Immunofluorescence staining showed that the M2 type macrophage marker CD206 in the complex was 80% highly expressed. The positive expression of CD86 the marker of M1 type macrophages was scarce. (4) The results show that the porcine-derived acellular cartilage extracellular matrix scaffold can induce polarized M0 type macrophages to repair M2 type macrophages.


Key words: bone, cartilage, regeneration, extracellular matrix, macrophages, polarization, tissue engineering, immune

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