| 丹酚酸A调控miR-940与miR-576-5p促进退变终板软骨细胞修复的作用研究 |
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投稿时间：2022-01-21
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| 作者 | Author | 单位 | Address | E-Mail |
| 展嘉文 |
ZHAN Jia-wen |
中国中医科学院望京医院运动医学中心, 北京 100102
中医正骨技术北京市重点实验室, 北京 100102 |
Sport Medicine center, Wangjing Hospital, China Academy of Chinese Meidical Science, Beijing 100102, China
Beijing Key Laboratory of Manipulative Technique Beijing 100102, Chiese |
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| 王尚全 |
WANG Shang-quan |
中国中医科学院望京医院运动医学中心, 北京 100102
中医正骨技术北京市重点实验室, 北京 100102 |
Sport Medicine center, Wangjing Hospital, China Academy of Chinese Meidical Science, Beijing 100102, China
Beijing Key Laboratory of Manipulative Technique Beijing 100102, Chiese |
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| 陈明 |
CHEN Ming |
中国中医科学院望京医院运动医学中心, 北京 100102
中医正骨技术北京市重点实验室, 北京 100102 |
Sport Medicine center, Wangjing Hospital, China Academy of Chinese Meidical Science, Beijing 100102, China
Beijing Key Laboratory of Manipulative Technique Beijing 100102, Chiese |
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| 孙凯 |
SUN Kai |
中医正骨技术北京市重点实验室, 北京 100102
中国中医科学院望京医院脊柱二科, 北京 100102 |
Beijing Key Laboratory of Manipulative Technique Beijing 100102, Chiese
The Second Department of Spine, Wangjing Hospital, China Academy of Chinese Medical Science, Beijing 100102, China |
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| 于杰 |
YU Jie |
中医正骨技术北京市重点实验室, 北京 100102
中国中医科学院望京医院脊柱二科, 北京 100102 |
Beijing Key Laboratory of Manipulative Technique Beijing 100102, Chiese
The Second Department of Spine, Wangjing Hospital, China Academy of Chinese Medical Science, Beijing 100102, China |
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| 李玲慧 |
LI Ling-hui |
中国中医科学院望京医院运动医学中心, 北京 100102
中医正骨技术北京市重点实验室, 北京 100102 |
Sport Medicine center, Wangjing Hospital, China Academy of Chinese Meidical Science, Beijing 100102, China
Beijing Key Laboratory of Manipulative Technique Beijing 100102, Chiese |
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| 魏戌 |
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中国中医科学院附属望京医院学术发展处, 北京 100102 |
Department of Academic Development, Wangjing Hospital, China Academy of Chinese Medical Science, Beijing 100102, China |
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| 孙武 |
SUN Wu |
中国中医科学院望京医院脊柱二科, 北京 100102 |
The Second Department of Spine, Wangjing Hospital, China Academy of Chinese Medical Science, Beijing 100102, China |
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| 陈忻 |
CHEN Xin |
中国中医科学院望京医院脊柱二科, 北京 100102 |
The Second Department of Spine, Wangjing Hospital, China Academy of Chinese Medical Science, Beijing 100102, China |
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| 蔡楚豪 |
CAI Chu-hao |
中国中医科学院望京医院运动医学中心, 北京 100102 |
Sport Medicine center, Wangjing Hospital, China Academy of Chinese Meidical Science, Beijing 100102, China |
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| 张伟业 |
ZHANG Wei-ye |
中国中医科学院望京医院运动医学中心, 北京 100102 |
Sport Medicine center, Wangjing Hospital, China Academy of Chinese Meidical Science, Beijing 100102, China |
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| 韩涛 |
HAN Tao |
中国中医科学院望京医院运动医学中心, 北京 100102
中医正骨技术北京市重点实验室, 北京 100102 |
Sport Medicine center, Wangjing Hospital, China Academy of Chinese Meidical Science, Beijing 100102, China
Beijing Key Laboratory of Manipulative Technique Beijing 100102, Chiese |
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| 尹煜辉 |
YIN Yu-hui |
中国中医科学院望京医院运动医学中心, 北京 100102 |
Sport Medicine center, Wangjing Hospital, China Academy of Chinese Meidical Science, Beijing 100102, China |
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| 唐彬 |
TANG Bin |
中国中医科学院望京医院脊柱二科, 北京 100102 |
The Second Department of Spine, Wangjing Hospital, China Academy of Chinese Medical Science, Beijing 100102, China |
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| 朱立国 |
ZHU Li-guo |
中医正骨技术北京市重点实验室, 北京 100102
中国中医科学院望京医院脊柱二科, 北京 100102 |
Beijing Key Laboratory of Manipulative Technique Beijing 100102, Chiese
The Second Department of Spine, Wangjing Hospital, China Academy of Chinese Medical Science, Beijing 100102, China |
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| 期刊信息：《中国骨伤》2023年，第36卷，第10期，第982-989页 |
| DOI：10.12200/j.issn.1003-0034.2023.10.014 |
| 基金项目:国家自然科学基金项目(编号:81930118,81804120,81774330);国家中医药管理局中医药创新团队及人才支持计划项(编号:ZYYCXTD-C-202003);中国中医科学院基本科研业务费优秀青年科技人才(创新类)培养专项(编号:ZZ13-YQ-038) |
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中文摘要:
目的:观察丹酚酸A(salvianolic acid A,SAA)对退变终板软骨细胞(cartilaginous endplates cells,CEPCs)的干预作用,及其调控的潜在非编码RNA(micro-RNA,miRNA)作用靶点。
方法:从腰椎间盘手术标本中分离CEPCs,用不同浓度SAA(2、5、10 μM处理24、48、72 h,利用CCK-8检测细胞活性确定SAA的最适剂量和干预时间。通过阿利新蓝染色和对血小板反应蛋白解整合素金属肽酶-5(A disintegrin and metalloproteinase with thrombospondin-5,ADAMTS-5)、基质金属肽酶3(matrix metallopepridase 3,MMP-3)、Ⅱ型胶原a1(clollagen typeⅡ a1,Col2a1)的蛋白表达检测,分析SAA对IL-1β诱导的退变CEPCs的干预作用。进一步结合生信分析,以及实时荧光定量PCR(quantitative real-time,qRT-PCR)与Western blot检测,并应用miRNA模拟物(miR-mimics) 与抑制剂(miR-inhibitor),验证SAA对潜在靶miRNAs的调控作用。
结果:10 μM SAA处理48 h显著提高了CEPCs活性,增加了白细胞介素(interlenkin-1β,IL-1β)所抑制的糖胺聚糖积累与Col2a1表达,并降低了细胞基质中ADAMTS-5与MMP3表达(P<0.05)。通过生物信息分析筛选与差异基因表达检测,确定了SAA的潜在靶miRNAs为miR-940和miR-576-5p。进一步在SAA处理组中加入miR-940-mimic或miR-576-5p-mimic,与SAA处理组相比过表达miR-940或miR-576-5p后CEPCs基质中ADAMTS-5与MMP3表达显著升高,Col2a1表达显著降低。
结论:SAA能够提高CEPCs活性,改善退变CEPCs的基质成分表达,而SAA的调控作用与抑制miR-940和miR-576-5p表达有关,两者可能是SAA调节CEPCs退变的作用靶点。 |
| 【关键词】椎间盘 丹酚酸A 终板软骨细胞 细胞外基质 micro-RNA |
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| Salvianolic acid A contributes to cartilage endplate cell restoration by regulating miR-940 and miR-576-5p |
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ABSTRACT
Objective To investigate whether Salvianolic acid A (SAA) can restore cartilage endplate cell degeneration of intervertebral discs and to identify the mechanism via regulation of micro-RNA.
Methods Cartilage endplate cells were isolated from lumbar intervertebral disc surgical samples and were treated with serum containing a series of concentrations of SAA (2,5,and 10 μM) for 24,48,and 72 h to identify a proper dose and treatment time of SAA. The effect SAA on interlenkin-1β (IL-1β)-induced extracellular matrix degradation of cartilage endplate cells were analyzed by Alcian blue staining and assessment of the expression levels of ADAMTS-5,MMP3 and Col2a1. Further,the potential target miRNAs were preliminarily screened by micro-RNA sequencing combining qRT-PCR and Western blot,and then,the miRNAs mimics and inhibitors were used to verify the regulatory effect of SAA on potential target miRNAs.
Results The 10 μM SAA treatment for 48 h significantly enhanced the viability of cartilage endplate cells,and increased Col2a1 expression and glycosaminoglycan accumulation that were repressed by IL-1β,and reduced the effect of IL-1β on ADAMTS-5,and MMP3. Screening analysis based on micro-RNA sequencing and Venny analysis identified the downstream micro-RNAs,including miR-940 and miR-576-5p. Then,the miR-940-mimic or miR-576-5p-mimic were transfected into CEPCs. Compared with the SAA group,the expression of ADAMTS-5 and MMP3 increased significantly and the expression of COL2A1 obviously decreased after overexpression of miR-940 or miR-576-5p in CEPCs.
Conclusion Salvianolic acid A attenuated the IL-1β-induced extracellular matrix degradation of cartilage endplate cells by targeting regulate the miR-940 and the miR-576-5p. |
| KEY WORDS Intervertebral disc Salvianolic acid A Cartilaginous endplates cells Extracellular matrix Micro-RNA |
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| 引用本文，请按以下格式著录参考文献： |
| 中文格式: | 展嘉文,王尚全,陈明,孙凯,于杰,李玲慧,魏戌,孙武,陈忻,蔡楚豪,张伟业,韩涛,尹煜辉,唐彬,朱立国.丹酚酸A调控miR-940与miR-576-5p促进退变终板软骨细胞修复的作用研究[J].中国骨伤,2023,36(10):982~989 |
| 英文格式: | ZHAN Jia-wen,WANG Shang-quan,CHEN Ming,SUN Kai,YU Jie,LI Ling-hui,SUN Wu,CHEN Xin,CAI Chu-hao,ZHANG Wei-ye,HAN Tao,YIN Yu-hui,TANG Bin,ZHU Li-guo.Salvianolic acid A contributes to cartilage endplate cell restoration by regulating miR-940 and miR-576-5p[J].zhongguo gu shang / China J Orthop Trauma ,2023,36(10):982~989 |
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