MiR-100 suppresses inflammatory activation of microglia and neuronal apoptosis following spinal cord injury via TLR4/NF-κB pathway

X.-H. Li, N.-S. Fu, Z.-M. Xing

Department of Rehabilitation, Liaocheng People’s Hospital, Liaocheng, China. 604353954@qq.com

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• Orthopedics

OBJECTIVE: This study aims to ascertain the effect of miR-100 on inflammation, apoptosis and functional rehabilitation after spinal cord injury (SCI) and the potential mechanism.

MATERIALS AND METHODS: Firstly, microglia were extracted from 3-day-old neonatal rats and cultured for the purpose of inflammatory activation. Quantitative Real Time-Polymerase Chain Reaction (qRT-PCR) was conducted to detect the levels of miR-100, toll-like receptors 4 (TLR4) and nuclear factor-κB (NF-κB). Moreover, proteins expressions of I-κB and induced-nitric oxide synthase (iNOS) and apoptosis-related genes were measured by Western blotting. In addition, SCI model was established in rats. Expressions of inflammatory factors in SCI rats were determined by enzyme-linked immunosorbent assay (ELISA) assay. Expression levels of TLR4/NF-κB pathway and miR-100 were determined by qRT-PCR. Immunofluorescence was conducted to measure activated microglia. Hindlimbs motor function in SCI rats was estimated via BBB 21-point rating scale.

RESULTS: In activated microglia, miR-100 level decreased, while TLR4 and NF-κB levels increased. The protein level of I-κB decreased and iNOS increased. Transfection of miR-100 mimics reversed the above trends. Inflammatory factors were highly elevated in SCI rats and mRNA levels of the TLR4/NF-κB pathway and miR-100 were down-regulated, which were ameliorated in SCI rats overexpressing miR-100 in vivo. The amount of activated microglia was declined with the administration of miR-100 mimics compared with the untreated SCI rats. Furthermore, apoptosis-related proteins were down-regulated by miR-100 mimics injection. Motor function in the miR-100 mimics group was improved better than that in the SCI group.

CONCLUSIONS: MiR-100 alleviates inflammation of microglia and neuronal tissue apoptosis, and improves motor function following SCI via inhibiting the TLR4/NF-κB pathway.

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