Elsevier

Brain Research Bulletin

Volume 140, June 2018, Pages 154-161
Brain Research Bulletin

Research report
Neuroprotective effects of metformin on traumatic brain injury in rats associated with NF-κB and MAPK signaling pathway

https://doi.org/10.1016/j.brainresbull.2018.04.008Get rights and content

Highlights

  • Metformin ameliorated neurological deficits, BBB disruption and brain edema after TBI.

  • Metformin reduced neuronal apoptosis and degeneration after TBI.

  • Metformin inhibited microglia-mediated inflammation after TBI.

  • Metformin suppressed NF-κB and MAPK activation after TBI.

Abstract

Traumatic brain injury (TBI) triggers a complex sequence of inflammatory responses that contribute to secondary injury. Metformin, a first-line drug used to treat type 2 diabetes, is reported to exhibit potent anti-inflammatory activity on diseases associated with the central nervous system (CNS). The aim of this study is to investigate the potential neuroprotective effects of metformin on acute brain injury after TBI and explore the underlying mechanisms. Male Sprague-Dawley (SD) rats were divided into four groups: sham group, TBI group, TBI + saline (NS) group and TBI + metformin group. A weight-dropping model was employed to induce TBI in rats. Modified neurological severity scores (mNSS) were employed to assess the short-term neurological deficits, neuronal degeneration and apoptosis in the brain tissues were assayed with Fluoro-Jade B and TUNEL staining, immunofluorescence was designed to investigate microglial activation. The mRNA and protein expression levels of pro-inflammatory cytokines such as necrosis factor-alpha (TNF-α), interleukin-beta (IL-1β) and nterleukin-6 (IL-6) were evaluated by real-time quantitative reverse transcriptase polymerase chain reaction (QPCR) and enzyme-linked immunosorbent assay (ELISA). Western blotting analysis was engaged to examine the expression of NF-κB p65 and phosphorylation of ERK1/2 and p38 MAPK. Our results showed that metformin significantly ameliorated neurological deficit, cerebral edema and neuronal apoptosis in rats following TBI. Moreover, metformin administration inhibited microglial activation and decreased the production of pro-inflammatory cytokines including TNF-α, IL-1β and IL-6. In addition, metformin inhibited the translocation of NF-κB p65 from cytoplasm into the nucleus, as well as the phosphorylation of ERK1/2 and p38 MAPK. This study suggests that metformin administration inhibits microglia activation-mediated inflammation via NF-κB and MAPK signaling pathway to improve neurobehavioral function following TBI, which provide a potential therapeutic benefit in treating brain injury.

Introduction

Traumatic brain injury (TBI) has been recognized a public health problem worldwide and is responsible for mortality and disability in children and young adults. It is a complicated pathophysiological process induced by traumatic mechanical forces and can be characterized with two broad phases: primary and secondary injury (Werner and Engelhard, 2007). Primary injury occurs at the moment of impact containing brain contusion, diffused axonal injury, intracranial hemorrhages, which results in focal necrotic cell death (LaPlaca et al., 2007). The secondary injury is a progressive pathological process including oxidative stress, inflammation, mitochondrial dysfunction, and apoptosis occur within minutes and may last from hours to days following the initial insult (Wang et al., 2015).

Microglia-mediated neuroinflammation is one of the hallmarks in secondary injury. In response to brain injury, microglia cells are rapidly activated and release inflammatory factors such as tumor necrosis factor (TNF), interleukin (IL) and interferon (IFN), which are believed to contribute to direct neuronal cell death and also induce vascular endothelial cells to express a variety of cell adhesion molecules and cell chemotaxis (Corrigan et al., 2016). Notably, pro-inflammatory cytokines released by microglia such as TNF-α, IL-1β and IL-6 play an important role in early events mediating blood–brain barrier (BBB) disruption and subsequent development of cerebral edema (Ramilo et al., 1990; Sun et al., 2013). It has been reported that inhibiting TBI-induced microglial activation and the subsequent inflammatory responses could improve the functional recovery in TBI patients (Kumar et al., 2017). Therefore, the inhibition of microglial activation could be a valuable therapeutic option for improving the recovery in TBI patients.

Metformin is a biguanide drug widely used in the treatment of type 2 diabetic patients as it reduces hepatic glucose production and improves insulin sensitivity (Schulte et al., 2014). Accumulating evidence showed that metformin exerts potent anti-inflammatory, anti-apoptotic and anti-oxidative properties on diseases associated with the central nervous system such as ischemic stroke, intracerebral hemorrhage and multiple sclerosis (Nath et al., 2009). It has been demonstrated that metformin protects rats from ischemia/reperfusion injuries by reducing oxidative stress (Abd-Elsameea et al., 2014). Metformin also protects the liver from ischemia reperfusion-induced injury by reducing post-ischemic inflammation (Cahova et al., 2015). However, there are few reports focused on the effects of metformin in TBI. Therefore, our present study aimed to investigate the effects and mechanisms of metformin on TBI-induced inflammation, shedding light on the potential therapeutic benefit for TBI.

Section snippets

Animals and treatment

Adult male Sprague-Dawley (SD) rats (300–350 g) were purchased from Animal Center of Chinese Academy of Sciences, Shanghai, China. The rats were housed in a temperature (23 ± 2 °C) and light (12 h light/dark cycle)-controlled room and supplied with food and water ad libitum. All procedures were approved by the Institutional Animal Care Committee and were in accordance with the guidelines of the National Institutes of Health on the care and use of animals.

We randomly divided the rats into four

Metformin reduced neurological deficits, brain water content and BBB breakdown after TBI

To determine the effects of metformin in neurological deficits after TBI, we performed mNSS test before rats were sacrificed in each group. At 24 h after TBI induction, the rats showed higher scores than the sham group, metformin treated group showed impaired neurological functions compared with the TBI + NS group. There were no significant differences between TBI and TBI + NS group (Fig. 1B).

Brain water content was evaluated to explore the effects of metformin treatment on TBI-induced brain

Discussion

In our present study, we investigated the anti-inflammatory effect of metformin in a rat model of TBI. The main findings were that acute (1 h post-trauma) metformin treatment reduced brain edema, ameliorated neuronal death and improved short-term neurological functions. Metformin suppressed activation of microglia and the expression of pro-inflammatory cytokines including TNF-α, IL-1β and IL-6 both in mRNA and protein levels. Additionally, metformin attenuated the translocation of NF-κB p65

Conflict of interest statement

The authors declare that they have no competing interests.

Acknowledgement

This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.

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