| 摘要: |
| 目的 筛选椎间盘退行性变（IDD）组织中表达升高的miRNA，探究其在IDD中的作用及可能机制。方法 比较已发表的2个IDD miRNA表达谱测序数据集，筛选出表达升高最显著的miRNA——miRNA-603。收集10例IDD患者和10例腰椎骨折患者的椎间盘组织，取正常椎间盘组织分离培养髓核细胞。用qRT-PCR检测IDD组织和正常椎间盘组织中miRNA-603的表达。通过miRNA-603过表达和抑制实验探究miRNA-603在TNF-α诱导的髓核细胞炎症反应中的作用。利用生物信息学网站及已发表的IDD mRNA测序数据集，获得miRNA-603的靶基因山梨糖和SH3结构域包含蛋白3（SORBS3），并利用双荧光素酶报告基因实验验证。通过miRNA-603和SORBS3过表达实验探究miRNA-603和SORBS3在髓核细胞炎症反应中的功能。结果 miRNA-603在IDD组织中表达升高（P<0.01）。在髓核细胞中过表达miRNA-603可以促进IL-6和IL-1β的表达（P均<0.01）；抑制miRNA-603表达后，TNF-α刺激诱导的IL-6和IL-1β表达降低（P<0.01，P<0.05）。同时，cleaved caspase-1的蛋白表达量也降低（P<0.01）。生物信息学预测显示miRNA-603可以靶向调控SORBS3基因，IDD组织样本检测显示SORBS3低表达（P<0.01）。与单独过表达miRNA-603相比，同时过表达miRNA-603和SORBS3可抑制TNF-α诱导的髓核细胞中IL-6和IL-1β的表达（P均<0.01）。结论 人IDD组织中miRNA-603表达升高。miRNA-603可能通过靶向抑制SORBS3继而诱导髓核细胞炎症反应，推动IDD进程。 |
| 关键词: 微RNA-603 椎间盘退行性变 炎症 山梨糖和SH3结构域包含蛋白3 |
| DOI：10.16781/j.0258-879x.2021.01.0096 |
| 投稿时间：2020-01-04修订日期：2020-09-02 |
| 基金项目: |
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| microRNA-603 targeting sorbin and SH3 domain-containing protein 3 inducing inflammatory response in nucleus pulposus cells and affecting intervertebral disc degeneration |
| LI Li1,LI Jin-cheng2,GAO Shu-hua3,QIN qin1* |
(1. Department of Laboratory Medicine, Changhai Hospital, Naval Medical University(Second Military Medical University), Shanghai 200433, China;
2. Department of Clinical Laboratory, Ningbo Meikang Traditional Chinese Medicine Hospital, Ningbo 315100, Zhejiang, China;
3. Outpatient Department, General Hospital of PLA Eastern Theater Command, Nanjing 210006, Jiangsu, China
*Corresponding author) |
| Abstract: |
| Objective To screen microRNA (miRNA) with elevated expression in intervertebral disc degeneration (IDD) tissues, and to explore its role and possible mechanisms in IDD. Methods Two published IDD miRNA sequencing databases were compared to screen the most highly expressed miRNA-miRNA-603. Intervertebral disc tissues of 10 patients with IDD and 10 patients with lumbar spine fracture were collected, and the nucleus pulposus cells from normal intervertebral disc tissues were isolated and cultured. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to detect the expression of miRNA-603 in the IDD and normal intervertebral disc tissues. The role of miRNA-603 in tumor necrosis factor α (TNF-α)-induced inflammation of nucleus pulposus cells was explored using miRNA-603 mimics and inhibitor factor. Sorbin and SH3 domain-containing protein 3 (SORBS3), a target gene of miRNA-603, was found by using bioinformatics website and the published IDD miRNA sequencing databases, and was verified by dual-luciferase reporter assay. The function of miRNA-603 and SORBS3 in the inflammatory response of nucleus pulposus cells was explored by the overexpression of miRNA-603 and SORBS3. Results The expression of miRNA-603 increased significantly in IDD tissues (P<0.01). Overexpression of miRNA-603 could promote the expression of interleukin (IL)-6 and IL-1β in nucleus pulposus cells (both P<0.01); inhibition of the miRNA-603 expression could reduce the TNF-α-induced expression of IL-6 and IL-1β (P<0.01, P<0.05). At the same time, the expression of cleaved caspase-1 also decreased (P<0.01). Bioinformatics prediction showed that miRNA-603 could targetedly regulate SORBS3, and the expression of SORBS3 in IDD tissues was low (P<0.01). Compared with the overexpression of miRNA-603 alone, co-overexpression of miRNA-603 and SORBS3 inhibited the TNF-α-induced expression of IL-6 and IL-1β in nucleus pulposus cells (both P<0.01). Conclusion The expression of miRNA-603 increases in human IDD tissues. miRNA-603 can induce the inflammatory response of nucleus pulposus cells by targetedly inhibiting SORBS3, which may promote the IDD process. |
| Key words: microRNA-603 intervertebral disc degeneration inflammation sorbin and SH3 domain-containing protein 3 |
