MiR-223-5p inhibitor suppresses microglia inflammation and promotes Nrg-1 in rats of spinal cord injury

Y.-Z. Guan, C. Sun, H.-L. Wang, X.-L. Xia, F.-Z. Lu, J. Song, X.-S. Ma, J.-Y. Jiang

Department of Orthopedics, Huashan Hospital, Fudan University, Shanghai, China. mxs0356099@163.com

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• Orthopedics

OBJECTIVE: The aim of this study was to investigate the role of microRNA-233-5p (miR-233-5p) in spinal cord injury (SCI), and to explore the possible underlying mechanism.
MATERIALS AND METHODS: Microglia were first isolated from neonate rats and cultured in a suitable environment in vitro. Lipopolysaccharide (LPS) and interleukin-4 (IL-4) were used to activate microglia. The expressions of miR-223-5p, inducible nitric oxide synthase (iNOS) and arginase 1 (Arg-1) were measured by qRT-PCR, respectively. After transfection of miR-233-5p inhibitor, the expression levels of miR-223-5p, iNOS and Arg-1 in cells were detected as well. A moderate SCI model was successfully established in rats (10 g fallen on T10 spinal cord at the height of 5 cm). Subsequently, inflammation indexes at miR-223-5p peak moment were observed. Meanwhile, its neuro-protective effect at 28 days after SCI was estimated. Finally, Basso, Beattie, and Bresnahan (BBB) rating scale was applied to evaluate the hindlimb locomotor function of rats at 1, 3, 7, 14, 21, 28 days after SCI.
RESULTS: MiR-223-5p inhibitor significantly promoted M2 microglia expression and degenerated M1 microglia expression in vitro. SCI elevated the level of miR-223-5p in injured spinal cord tissues within one week, which reached a peak at 5 days after injury. Meanwhile, miR-223-5p inhibitor remarkably reduced the expressions of inflammatory factors, including interleukin-1 beta (IL-1β), interleukin-6 (IL-6) and tumor necrosis factor alpha (TNF-α) at 3 days after SCI, as well as increased neuregulin1 (NRG-1) expression. However, miR-223-5p inhibitor significantly declined the levels of apoptosis key enzyme-caspase-3 and glia reaction marker-glial fibrillary acidic protein (GFAP) at 7 and 28 days after SCI, respectively. As a result, BBB rating scale demonstrated that hindlimb locomotor function was significantly recovered in miR-223-5p injection group.
CONCLUSIONS: MiR-223-5p was up-regulated in M1 microglia, whereas down-regulated in M2 microglia. MiR-223-5p inhibitor could significantly increase M2 microglia expression, while decrease M1 microglia expression in vitro. In vivo, miR-223-5p inhibitor suppressed the inflammatory response and reinforced NRG-1 level to reduce glia reaction and neuron apoptosis. Thereby, its treatment promoted the hindlimb locomotor function of rats.

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