Elsevier

Brain Research

Volume 1685, 15 April 2018, Pages 91-104
Brain Research

Research report
MicroRNA let-7c-5p improves neurological outcomes in a murine model of traumatic brain injury by suppressing neuroinflammation and regulating microglial activation

https://doi.org/10.1016/j.brainres.2018.01.032Get rights and content

Highlights

  • The expression of let-7c-5p was decreased in traumatic foci after TBI and in activated primary microglia.

  • Overexpression of let-7c-5p attenuates microglia-mediated neuroinflammation after TBI.

  • Overexpression of let-7c-5p protects TBI via promoting microglia M2 polarization and inhibiting M1 polarization.

  • In a murine model of TBI, cleaved PKC-δ mediates effect of caspase-3 which is the target of let-7c-5p.

Abstract

MicroRNAs (miRNAs) are a class of non-coding small RNAs that regulate the expression of target genes. They derive from pre-miRNAs that are enzymatically processed by dicer to ∼22 nucleotide mature miRNAs. Members of the pre-miRNA lethal-7 (let-7) are known to regulate cell proliferation and apoptosis. Here, we showed that the level of let-7c-5p, a key member of the let-7 family, was rapidly reduced in the traumatically injured foci in brains of adult C57BL/6J mice and gradually recovered to the pre-injury level 14 days after traumatic brain injury (TBI) induction. This finding led us to test whether upregulating let-7c-5p in murine cerebral tissue by intracerebroventricular injection (ICV) of let-7c-5p mimic could improve the outcomes of mice subjected to controlled cortical impact (CCI). We found that let-7c-5p overexpression attenuated TBI-induced neurological dysfunction and brain edema. The improvements were attributed to let-7c-5p-mediated inhibiting neuroinflammation and attenuation of microglia/macrophage activation, both inhibiting M1 polarization and enhancing M2 polarization. In vitro experiments, we observed that let-7c-5p was decreased in primary microglia activated by LPS treatment or oxygen/glucose deprivation (OGD). Transfection of let-7c-5p mimic suppressed the release of inflammatory mediators in cultured activated primary microglia. In addition, the expressions of caspase-3, a let-7c-5p putative target gene, and the PKC-δ which mediates effect of caspase-3 were inhibited by let-7c-5p in a murine model of TBI. Taken together, these results define the biological activities of cerebral let-7c-5p and delineate its therapeutic potential for improving the neurological outcome of TBI.

Introduction

Traumatic brain injury (TBI) is one of the leading causes of death and disability worldwide, especially among young children and adults (Dong et al., 2016). Among the complex pathological changes of TBI, neuro-inflammation known as an important secondary injury mechanism exerts either detrimental or beneficial roles in the process of central nervous system (CNS) damage caused by TBI (Hellewell et al., 2016). However, sustained and excessive inflammation can exacerbate neuronal apoptosis after TBI (Xu et al., 2017). Neuro-inflammation is propagated by microglia and infiltrated macrophage principally via upregulating and releasing of soluble cellular constituents in the injured brain (Cornelius et al., 2017, Loane and Kumar, 2016). Microglia, which are among the first responders to CNS injuries, and infiltrated macrophage have been shown to be highly plastic involving either a “classically activated” M1-phenotype or the “alternatively activated” M2-phenotype (Loane and Kumar, 2016, Turtzo et al., 2014). The M1 polarization causes neuronal dysfunction, cell death and impairment of axon regrowth by inducing pro-inflammatory mediators and reducing neurotrophic factors. In contrast, M2 phenotype involved in anti-inflammatory response removes cellular debris and necrotic tissues through phagocytosis and releases trophic factors to promote CNS repair and preserve tissue integrity (Truettner et al., 2017). Altering the M2/M1 ratio which has been reported has promising effects in TBI (Yao et al., 2017). Therefore, strategies targeting the control of activation status of microglia/macrophages might have therapeutic implications after TBI.

MicroRNAs (miRNAs), which are a class of small non-coding RNAs that bind to the untranslated regions of target mRNAs to induce transcript degradation and inhibit translation, are critical regulators of immune signaling in CNS (Cardoso et al., 2016). MiRNAs have been extensively implicated in altering the M2/M1 ratio secondary to variety of brain injury including TBI (Jadhav et al., 2014). Let-7c which is one of the most abundant and highly conserved miRNAs, has been implicated in the macrophage polarization (Essandoh et al., 2016, Banerjee et al., 2013). Although overexpressing let-7c-5p may be involved in protecting against ischemic stroke by suppressing microglia activation (Ni et al., 2015, Tan et al., 2014), the role of let-7c-5p in TBI is unknown. To explore the function and mechanism of let-7c-5p in TBI, we focus on studying its effects on neurological outcome and relationship with pathological changes including ameliorating neuroinflammation and altering the microglia/macrophage polarization status.

Section snippets

Altered let-7c-5p expression in the traumatic foci after TBI and intracerebroventricular infusion of let-7c-5p oligomers

We detected let-7c-5p expression levels in the traumatic foci, which were defined as the impacted area with a diameter of 3 mm including injured cerebral cortex and injured hippocampus, at different time-points from 0 h to 14 d post-injury as measured by RT-PCR (Fig. 1). The expression of let-7c-5p in the injury group was progressively reduced after injury, reached a plateau on day 5 post-injury then gradually increased to baseline at 14 d post-injury. Compared with the mimic ctl group, the

Discussion

In the present study, we demonstrated for the first time the anti-inflammatory and immunomodulatory properties of let-7c-5p in mice post-TBI. The main findings were that intraventricular administration of let-7c-5p mimic (10 min post-trauma) effectively attenuated cerebral edema and improved neurological dysfunction induced by CCI in a mouse model. Overexpression of let-7c-5p decreased expressions of pro-inflammatory cytokines while simultaneously increased release of anti-inflammatory

Institutional approvals

All animal protocols were approved by the Animal Care and Use Committee of Tianjin Medical University and conformed to the standards set by the National Institutes of Health.

Animals

Adult male C57BL/6J mice (6–10-week-old) were purchased from the Chinese Academy of Military Science (Beijing, China). Animals were housed in 320 * 215 * 170 mm cages (RTSL2015886230, Rital company, China) in groups of four in a temperature (22 °C ± 2 °C) and humidity (60% ± 12%) animal facility with 1 week to acclimate. A

Acknowledgments

This work was supported by National Natural Science Foundation of China State Key Program (grant nos. 81330029) and National Natural Science Foundation of China research (grants nos. 81271361 and 81271359).

Author contributions

Jingfang Lv, Jianning Zhang and Jingfei Dong designed the experiments. Jingfang Lv, Yong Zeng, Yu Qian and Jianning Zhang performed the experiments. Jingfang Lv and Yong Zeng analyzed the data. Jingfang Lv, Jingfei Dong and Jianning Zhang wrote the manuscript.

Conflict of interest disclosures

All authors claim no conflict of interest.

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