MiR-199b-5p inhibits osteogenic differentiation in ligamentum flavum cells by targeting JAG1 and modulating the Notch signalling pathway.

Qiang Qi,Zhaoqing Guo,Weishi Li,Yan Zeng,Dongwei Fan,Zhongqiang Chen,Chuiguo Sun,Xiaochen Qu

doi:10.1111/jcmm.13047

Qiang Qi, Zhaoqing Guo + Show 6 more

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https://doi.org/10.1111/jcmm.13047

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Abstract

Ossification of the ligamentum flavum (OLF) is a pathology almost only reported in East Asian countries. The leading cause of OLF is thoracic spinal canal stenosis and myelopathy. In this study, the role of miR‐199b‐5p and jagged 1 (JAG1) in primary ligamentum flavum cell osteogenesis was examined. MiR‐199b‐5p was found to be down‐regulated during osteogenic differentiation in ligamentum flavum cells, while miR‐199b‐5p overexpression inhibited osteogenic differentiation. In addition, JAG1 was found to be up‐regulated during osteogenic differentiation in ligamentum flavum cells, while JAG1 knockdown via RNA interference caused an inhibition of Notch signalling and osteogenic differentiation. Moreover, target prediction analysis and dual luciferase reporter assays supported the notion that JAG1 was a direct target of miR‐199b‐5p, with miR‐199b‐5p found to down‐regulate both JAG1 and Notch. Further, JAG1 knockdown was demonstrated to block the effect of miR‐199b‐5p inhibition. These findings imply that miR‐199b‐5p performs an inhibitory role in osteogenic differentiation in ligamentum flavum cells by potentially targeting JAG1 and influencing the Notch signalling pathway.

Highlights

OLF is a pathology almost exclusively reported in Eastern Asian countries
Our results implied that miR-199b-5p, which is down-regulated during osteogenic differentiation in ligamentum flavum cells, inhibits the differentiation process by targeting jagged 1 (JAG1) and affecting the Notch signalling pathway
JAG1 mRNA expression levels increased at day 7 and remained elevated to day 17 (Fig. 3A), while protein levels were continually increased to day 17 (Fig. 3B). These results showed an inverse trend relative to miR-199b-5p levels, suggesting that JAG1 is post-transcriptionally regulated during osteogenic differentiation in ligamentum flavum cells

Summary

Introduction

OLF is a pathology almost exclusively reported in Eastern Asian countries. OLF primarily occurs in the thoracolumbar spine and is the main cause of thoracic spinal canal stenosis and myelopathy [1, 2]. In the past several years, many studies have examined OLF development and progression at both histopathological and cellular levels. While these studies identified potential contributing factors, such as mechanical [3,4,5,6], metabolic [7, 8], degenerative [9] and genetic factors [10, 11], OLF development and progression continues to be inadequately understood. While various signalling pathways have been connected with the regulation of osteogenic differentiation in general, some have been associated with OSL pathogenesis [6, 7, 13,14,15,16]. The Notch signalling pathway, which influences proliferation, differentiation and mineralization in osteoblasts, has been found to encourage osteogenic differentiation in ligamentum flavum cells [17, 18]

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