MicroRNA-204-5p targets SOX11 to regulate the inflammatory response in spinal cord injury

J.-S. Feng, J.-D. Sun, X.-D. Wang, C.-H. Fu, L.-L. Gan, R. Ma

Intensive Care Unit, Gansu Provincial Hospital, Lanzhou, China. fjsh1982jd@163.com

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• Orthopedics

OBJECTIVE: The purpose of this study was to investigate whether microRNA-204-5p can regulate the inflammatory response of spinal cord injury (SCI) by targeting SOX11.
PATIENTS AND METHODS: Quantitative Real Time-Polymerase Chain Reaction (qRT-PCR) was used to detect the expression of microRNA-204-5p in patients with SCI. The mouse SCI model was established to detect the recovery of the grip strength of the upper and lower limbs. Then, the expression of microRNA-204-5p in these mice with SCI was detected by qRT-PCR, and the levels of the inflammatory factors Toll-like receptor 4 (TLR4) and iNOS were examined by Western blot. Subsequently, microRNA- 204-5p was overexpressed in the mouse SCI model using lentivirus, and the changes in mouse grip strength and the inflammatory factor levels were observed. SOX11 was then searched as the target gene of microRNA-204-5p through bioinformatics analysis, and its expression in patients or mice with SCI was examined using qRT-PCR. SOX11 expression was again detected after the overexpression or knockdown of microRNA-204-5p in cells. The binding of microRNA-204-5p to SOX11 was verified by dual-luciferase reporting assay. After microRNA-204-5p and SOX11 were co-overexpressed in cells, the levels of TLR4 and iNOS were analyzed. Furthermore, the changes in the grip strength were observed in mice with SCI after simultaneous up-regulation of microRNA-204-5p and SOX11.
RESULTS: Micro-204-5p level was conspicuously decreased in the population with SCI. And the SCI mouse model showed that the upper and lower limb strength conspicuously decreased and began to recover after 7 days. During the seven days, microRNA-204-5p level in the SCI mice decreased with time, while the levels of the inflammatory cytokines TLR4 and iNOS conspicuously increased. After microRNA-204-5p was overexpressed in SCI mice, their upper and lower limb strength was conspicuously restored, while the levels of TLR4 and iNOS were also remarkably decreased. The bioinformatics analysis revealed that there exist some binding sites between microRNA-204-5p and SOX11, and we found that SOX11 expression was conspicuously enhanced in the plasma of the SCI patients. Meanwhile, the SOX11 level in SCI mice was also conspicuously increased, and it was time-dependent. The expression of SOX11 was decreased after the upregulation of microRNA-204-5p, while the opposite result was observed after the downregulation of microRNA-204-5p. In addition, the result of the dual-luciferase reporter gene assay revealed that microRNA-204-5p could bind to SOX11 in a targeted manner. Meanwhile, the up-regulation of SOX11 was partially relieved by the inhibitory effect of microRNA-204-5p on TLR4 and iNOS. Moreover, the simultaneous overexpression of SOX11 and microRNA-204-5p partially reversed the impact of the up-regulated microRNA-204-5p alone on the recovery of the upper and lower limb strength in SCI mice.
CONCLUSIONS: The low expression of microRNA-204-5p in patients with SCI can affect the levels of the inflammatory cytokines TLR4 and iNOS and improve SCI by targeting SOX11.

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