中国组织工程研究 ›› 2020, Vol. 24 ›› Issue (1): 130-135.doi: 10.3969/j.issn.2095-4344.1845

• 干细胞基础实验 basic experiments of stem cells • 上一篇    下一篇

Asxl1在炎性微环境中对成骨细胞增殖分化的作用

徐慧君,史东梅,张  咪,吴赛璇,董  明,陆  颖,牛卫东   

  1. 大连医科大学口腔医学院牙体牙髓科,辽宁省大连市  116044
  • 收稿日期:2019-02-25 修回日期:2019-03-05 接受日期:2019-04-19 出版日期:2020-01-08 发布日期:2019-12-13
  • 通讯作者: 牛卫东,博士,教授,博士生导师,大连医科大学口腔医学院牙体牙髓科,辽宁省大连市 116044
  • 作者简介:徐慧君,女,1994年生,安徽省亳州市人,汉族,医师,大连医科大学在读硕士,主要从事牙体牙髓研究。 共同第一作者:史东梅,女,1990年生,山东省聊城市人,汉族,医师,大连医科大学在读硕士,主要从事牙体牙髓研究。
  • 基金资助:
    国家自然科学基金青年科学基金项目(81700962)

Effect of sex combing protein 1 on proliferation and differentiation of osteoblasts in inflammatory microenvironment#br#

Xu Huijun, Shi Dongmei, Zhang Mi, Wu Saixuan, Dong Ming, Lu Ying, Niu Weidong   

  1. Department of Endodontics, School of Stomatology, Dalian Medical University, Dalian 116044, Liaoning Province, China
  • Received:2019-02-25 Revised:2019-03-05 Accepted:2019-04-19 Online:2020-01-08 Published:2019-12-13
  • Contact: Niu Weidong, MD, Professor, Doctoral supervisor, Department of Endodontics, School of Stomatology, Dalian Medical University, Dalian 116044, Liaoning Province, China
  • About author:Xu Huijun, Master candidate, Physician, Department of Endodontics, School of Stomatology, Dalian Medical University, Dalian 116044, Liaoning Province, China Shi Dongmei, Master candidate, Physician, Department of Endodontics, School of Stomatology, Dalian Medical University, Dalian 116044, Liaoning Province, China Xu Huijun and Shi Dongmei contributed equally to this work
  • Supported by:
    the National Natural Science Foundation of China (Youth Project), No. 81700962

摘要:

文题释义:

Asxl1:是与果蝇Asx基因同源的3种基因中的一种类型,该基因属于表观遗传调控蛋白Trithorax家族(TrxG)和Polycomb家族(PcG)的增强子,位于染色体上,编码核蛋白,对于维持基因的同源性及不同基因位点表达的稳定性起决定性作用。

成骨细胞:该细胞起源于多功能的骨髓间充质干细胞,参与骨形成过程,是主要的功能性细胞。该细胞可形成新骨,主要通过促进细胞外骨基质的合成、分泌和矿化而实现。ALP和RUNX2是该细胞的主要标志性因子,前者可作为成骨早期分化的指标,后者是成骨细胞分化的特异性转录因子。成骨前体细胞系MC3T3-E1可作为体外研究成骨细胞增殖分化的良好模型。

背景:研究表明Asxl1的缺失可导致骨质发育不全、骨质缺损类疾病的发生,但目前在根尖周炎环境下该因子与骨破坏之间的关系暂无相关报道。

目的:探讨炎性微环境下Asxl1对成骨细胞增殖分化的影响。

方法:实验选用脂多糖刺激MC3T3-E1细胞建立体外炎性微环境,通过CCK-8实验筛取脂多糖最佳质量浓度和最佳作用时间,然后用20 mg/L脂多糖刺激MC3T3-E1细胞24 h,免疫荧光检测Asxl1的蛋白表达水平,Real Time-PCR检测Asxl1 mRNA的表达水平。为进一步验证Asxl1基因在炎性微环境中影响成骨细胞的增殖与分化,脂多糖刺激形成炎性微环境后转染Asxl1-SiRNA 24 h,采用CCK-8检测细胞增殖活性,Real Time-PCR检测Asxl1及成骨相关基因ALP和RUNX2 mRNA的表达水平。

结果与结论:①脂多糖刺激MC3T3-E1细胞后,Asxl1蛋白和mRNA表达水平呈降低趋势;②脂多糖刺激MC3T3-E1细胞后,转染Asxl1-SiRNA 24 h,细胞增殖活性下降趋势明显,Asxl1基因及成骨相关基因ALP和RUNX2 mRNA的表达水平明显降低;③结果提示,Asxl1可能通过参与炎性反应过程,影响成骨细胞的增殖与分化,进而参与骨破坏进程。

ORCID: 0000-0002-7601-4535(徐慧君)

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: MC3T3-E1细胞, Asxl1, 根尖周炎, 骨破坏, 基因转染, 脂多糖, 成骨分化, 国家自然科学基金

Abstract:

BACKGROUND: Studies have shown that the loss of sex combing protein 1 (Asxl1) can lead to the occurrence of bone dysplasia and bone defects, but the relationship between this factor and bone destruction in the microenvironment of apical periodontitis has not been reported.

OBJECTIVE: To study the effect of Asxl1 on proliferation and differentiation of osteoblasts in an inflammatory microenvironment.

METHODS: MC3T3-E1 cells were excited by lipopolysaccharide to establish an in vitro inflammatory microenvironment. The best concentration and optimal action time of lipopolysaccharide were screened by cell counting kit-8 test. MC3T3-E1 cells were then stimulated with 20 mg/L lipopolysaccharide for 24 hours. The expression levels of Asxl1 protein and mRNA were detected by immunofluorescence and real-time PCR, respectively. After lipopolysaccharide stimulated the formation of inflammatory microenvironment, Asxl1-Si RNA was transfected for 24 hours, cell counting kit-8 was applied to detect the activity of cell proliferation, and real-time PCR was used to detect the expression levels of Asxl1 and osteogenic related genes ALP and RUNX2 mRNA.

RESULTS AND CONCLUSION: After lipopolysaccharides stimulation of MC3T3-E1 cells, the expression levels of Asxl1 protein and mRNA were decreased. Under the inflammatory microenvironment, the proliferation activity of MC3T3-E1 cells transfected with Asxl1-Si RNA for 24 hours was significantly decreased, and the expression levels of Asxl1, ALP and RUNX2 mRNA were markedly decreased. These findings indicate that Asxl1 may influence the proliferation and differentiation of osteoblasts by involvement in the process of inflammatory reaction, thereby participating in bone destruction.

Key words: MC3T3-E1 cells, Asxl1, periapical periodontitis, bone destruction, gene transfection, lipopolysaccharide, osteogenic differentiation, National Natural Science Foundation of China

中图分类号: