Elsevier

Neurobiology of Disease

Volume 51, March 2013, Pages 104-112
Neurobiology of Disease

Transgenic mice with high endogenous omega-3 fatty acids are protected from spinal cord injury

https://doi.org/10.1016/j.nbd.2012.10.021Get rights and content

Abstract

Omega-3 polyunsaturated fatty acids have been shown to have therapeutic potential in a variety of neurological disorders, including acute traumatic injury of the spinal cord. We addressed the question whether the neuroprotective effect of these compounds after spinal cord injury could also be seen when their level is raised in tissues prophylactically, prior to injury. In this study we used transgenic fat-1 mice to examine whether enriching spinal cord tissue in endogenous omega-3 polyunsaturated fatty acids has an effect on the outcome after compression spinal cord injury. The results demonstrate that after thoracic compression spinal cord injury, fat-1 mice display better locomotor recovery compared with the wild-type mice on a high omega-6 diet (high omega-6 polyunsaturated fatty acids in tissues), and wild-type mice on a normal diet (controls). This is associated with a significant increase in neuronal and oligodendrocyte survival and a decrease in non-phosphorylated neurofilament loss. The protection from spinal cord injury in fat-1 mice was also correlated with a reduction in microglia/macrophage activation and in pro-inflammatory mediators. In vitro experiments in dorsal root ganglia primary sensory neurons further demonstrated that a fat-1 tissue background confers robust neuroprotection against a combined mechanical stretch and hypoxic injury. In conclusion, our studies support the hypothesis that a raised omega-3 polyunsaturated fatty acid level and an altered tissue omega-6/omega-3 ratio prior to injury leads to a much improved outcome after spinal cord injury.

Introduction

Studies have found that omega-3 polyunsaturated fatty acids (PUFAs) can reverse excitotoxicity triggered by glutamate, decrease lipid peroxidation and modulate the inflammation triggered by central nervous system (CNS) injury. These observations led to the hypothesis that omega-3 PUFAs could act as neuroprotective agents in acute spinal cord injury (SCI). We have previously shown that treatment with omega-3 PUFAs after injury has a therapeutic potential in rat SCI models (Huang et al., 2007, King et al., 2006, Lim et al., 2010, Ward et al., 2010). There have been reports demonstrating the protective effect of pre-injury administration of omega-3 PUFAs in the prevention of the damaging consequences of traumatic brain injury (TBI) (Wu et al., 2004, Wu et al., 2007). Prophylactic treatment would also be important in SCI, especially in military operations, where personnel are at high risk of SCI during combat, or to limit the damage which may occur as a consequence of sports-related injury. It would be worth ascertaining whether a raised tissue level of omega-3 PUFA before injury could confer neuroprotection in SCI. The aim of this study was to assess the effect of enriching spinal cord tissue in endogenous omega-3 PUFAs on the response to compression SCI. Changes in the endogenous levels of PUFAs could be achieved through dietary supplementation, or alternatively through the use of the fat-1 transgenic mice. The fat-1 transgenic mouse carries the Caenorhabditis elegans fat-1 gene, which encodes an omega-3 fatty acid desaturase that converts omega-6 to omega-3 PUFA (Kang et al., 2004). When fat-1 mice and wild-type (WT) mice are maintained on a diet high in omega-6 PUFAs, fat-1 mice are capable of producing omega-3 PUFAs from the omega-6 type, leading to an abundance of omega-3 fatty acids and a low omega-6/omega-3 ratio in their tissues. In contrast, the WT mice exhibit an increased ratio of omega-6/omega-3 PUFAs. Therefore, by using these mice, we can produce two fatty acid profiles (high vs. low omega-6/omega-3 ratios) and study the impact of these changes on SCI.

We evaluated histological injury as well as functional motor recovery following compression SCI in three groups of mice, namely fat-1 mice on an omega-6 diet, WT mice on an omega-6 diet, and WT mice on a normal diet, as a control baseline. We also studied the impact of different omega-6/omega-3 ratios on inflammatory cytokines and chemokines after SCI. Finally, we also isolated dorsal root ganglion (DRG) primary sensory neurons from animals to investigate the effect of enhanced endogenous omega-3 PUFA on neuronal survival in vitro after a combined mechanical stretch and hypoxic injury.

Section snippets

Animals and diets

Fat-1 breeders on a C57BL/6 background were obtained from Dr. Jing X. Kang (Harvard Medical School). The F1 progeny were obtained by mating C57BL/6 X C3H fat-1 breeders with C57BL/6 WT mice from Charles River Laboratories (Margate, UK). Generations of heterozygous fat-1 mice were then mated with WT littermates to obtain WT and heterozygous fat-1 mice. The presence of the fat-1 gene was confirmed by genotyping and analysing the fatty acid composition of ear punches. Both the fat-1 and WT mice

Fatty acid profiles of the spinal cord before and after SCI

As indicated in Table 1, the expression of the fat-1 gene resulted in a significantly higher level of omega-3 docosapentaenoic acid (DPA) and docosahexaenoic acid (DHA) in the spinal cord of fat-1 mice, compared with the two WT groups. There was also a decrease in omega-6 DTA and AA in fat-1 mice, with marked effects found for DTA. WT mice on the high omega-6 diet had a higher level of omega-6 AA compared with WT mice on the control diet, whereas there were no significant differences in the

Discussion

The results presented here clearly show that the spinal cord damage induced by compression injury, in terms of both functional neurological manifestations and histopathological outcome, is significantly less severe in fat-1 mice than in the WT mice on a normal or omega-6 PUFA enriched diet. Two behavioural assessments (open-field and activity monitor analysis) showed better functional recovery in injured fat-1 mice compared with the two WT groups. The improved functional outcome in fat-1 mice

Conclusions

In summary, by using the fat-1 transgenic mouse model, we demonstrated that increased endogenous omega-3 PUFA levels and a decreased omega-6/omega-3 ratio prior to compression SCI lead to a significantly lesser impact of SCI, improved neurological outcome, decreased tissue damage and reduced inflammatory response. A robust resilience against both mechanical stretch and hypoxia was found in fat-1 mice. A significant increase in spinal cord omega-3 DPA and DHA in the fat-1 mice suggests that

Acknowledgment

This work was supported by Chang Gung Memorial Hospital, Taiwan (CMRPG360622). We gratefully acknowledge the assistance of Patrick Pallier, Laura Poddighe and Rasall Choudhury with preparatory staining for the Abercrombie correction.

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