MicroRNA-494 promotes apoptosis and extracellular matrix degradation in degenerative human nucleus pulposus cells
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Liang Kang1,*, Cao Yang1,*, Yu Song1,*, Kangcheng Zhao1, Wei Liu2, Wenbin Hua1, Kun Wang1, Ji Tu1, Shuai Li1, Huipeng Yin1, Yukun Zhang1
1Department of Orthopaedics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China
2Department of Orthopaedics, First Hospital of Wuhan, Wuhan 430022, China
*These authors contributed equally to this work
Yukun Zhang, email: email@example.com
Keywords: intervertebral disc degeneration, nucleus pulposus, miR-494, SOX9, methylation
Received: December 12, 2016 Accepted: February 20, 2017 Published: March 02, 2017
Purpose: This study investigated the expression and function of the microRNA-494 in intervertebral disc degeneration (IDD).
Results: MicroRNA-494 expression was upregulated during IDD progression; its overexpression increased the expression of ECM catabolic factors such as matrix metalloproteinase and A disintegrin and metalloproteinase with thrombospondin motif in NP cells while decreasing that of anabolic genes such as type II collagen and aggrecan; it also induced the apoptosis of NP cells, as determined by flow cytometry. These effects were reversed by microRNA-494 inhibitor treatment. SOX9 was identified as a target of negative regulation by microRNA-494. Promoter hypomethylation and NF-κB activation were associated with microRNA-494 upregulation in IDD.
Materials and Methods: MicroRNA-494 expression in degenerative nucleus pulposus (NP) tissue was assessed by quantitative real-time PCR. The effect of microRNA-494 on extracellular matrix (ECM) metabolism and NP cell apoptosis was evaluated by transfection of microRNA-494 mimic or inhibitor. The regulation of SRY-related high mobility group box (SOX)9 expression by microRNA-494 was assessed with the luciferase reporter assay, and the methylation status of the microRNA-494 promoter was evaluated by methylation-specific PCR and bisulfite sequencing PCR. The role of activated nuclear factor (NF)-κB in the regulation of microRNA-494 expression was evaluated using specific inhibitors.
Conclusions: MicroRNA-494 promotes ECM degradation and apoptosis of degenerative human NP cells by directly targeting SOX9.
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