Du Ketao, He Xuguang, Deng Jiaqin
Department of Rehabilitation, The First Affiliated Hospital, Jinan University, Guangzhou, Guangdong, China.
Department of Rehabilitation, Chenzhou No. 1 People's Hospital, Chenzhou, Hunan, China.
Arch Med Sci. 2018 Apr 6;17(2):500-513. doi: 10.5114/aoms.2018.74950. eCollection 2021.
A variety of inflammatory mediators are produced by the degenerative human intervertebral disc (IVD) tissues spontaneously, suggesting their role in the development of intervertebral disc degeneration (IDD). Our present study was designed to investigate the regulatory effect of microRNA-16 (miR-16) on the lipopolysaccharide (LPS)-induced inflammatory response in nucleus pulposus (NP) cells of the IVD.
NP cells were treated with LPS to induce inflammatory responses. The expression of miRNA and genes was determined by qRT-PCR. Western blot and an ELISA kit were used to detect the proteins and protein expression, respectively. A dual luciferase reporter assay was applied to identify the correlation between a miRNA and a gene, and to test nuclear factor-κB (NF-κB) activity.
The results suggested that miR-16 positively regulated the mRNA and protein expression of extracellular matrix (ECM) genes (including and ) in NP cells, while it was negatively correlated with ECM degrading enzymes (including , , , ) and related genes of nitric oxide (NO) reaction. Further studies revealed that miR-16 could oppositely regulate NF-κB and MAPK pathways by directly mediating their upstream gene .
Our study suggested that, in NP cells of the IVD, miR-16 could exert inhibitory effects on the LPS-induced inflammatory response through NF-κB and MAPK pathways by directly mediating TAB3. In this way, miR-16 would play a protective role against LPS-induced IDD and inflammation. Therefore, miR-16 may be a novel therapeutic target for the inhibit of the ECM in the IVD.
退变的人类椎间盘（IVD）组织可自发产生多种炎症介质，提示其在椎间盘退变（IDD）发展过程中的作用。我们目前的研究旨在探讨微小RNA-16（miR-16）对IVD髓核（NP）细胞中脂多糖（LPS）诱导的炎症反应的调节作用。
用LPS处理NP细胞以诱导炎症反应。通过qRT-PCR测定miRNA和基因的表达。分别使用蛋白质印迹法和ELISA试剂盒检测蛋白质及蛋白质表达。应用双荧光素酶报告基因检测法来鉴定miRNA与基因之间的相关性，并检测核因子κB（NF-κB）活性。
结果表明，miR-16正向调节NP细胞中细胞外基质（ECM）基因（包括 和 ）的mRNA和蛋白质表达，而它与ECM降解酶（包括 、 、 、 ）以及一氧化氮（NO）反应的相关基因呈负相关。进一步研究表明，miR-16可通过直接介导其上游基因 来反向调节NF-κB和丝裂原活化蛋白激酶（MAPK）信号通路。
我们的研究表明，在IVD的NP细胞中，miR-16可通过直接介导TAB3，经由NF-κB和MAPK信号通路对LPS诱导的炎症反应发挥抑制作用。通过这种方式，miR-16将对LPS诱导的IDD和炎症起到保护作用。因此，miR-16可能是抑制IVD中ECM的一个新的治疗靶点。
