Elsevier

Neuroscience

Volume 189, 25 August 2011, Pages 397-407
Neuroscience

Regeneration, Repair, and Developmental Neuroscience
Research Paper
C-jun phosphorylation contributes to down regulation of neuronal nitric oxide synthase protein and motoneurons death in injured spinal cords following root-avulsion of the brachial plexus

https://doi.org/10.1016/j.neuroscience.2011.04.070Get rights and content

Abstract

Previous studies have shown that c-jun and neuronal nitric oxide synthase (nNOS) are both induced in injured motoneurons, but their roles in motoneuron death remain unclear. We hypothesized that nNOS might be the downstream effector of c-jun N-terminal kinase (JNK)/c-jun in avulsion-induced motoneuron death. Here, we found that brachial root-avulsion induced a temporary increase in JNK activity and three- and four-fold increases in phospho-c-jun and c-jun, respectively; however, brachial root-avulsion caused a decrease in nNOS protein expression from 4 h to 14 days post-injury. At 14 days post-injury, almost all nNOS-positive motoneurons were co-localized with phospho-c-jun-positive motoneurons in ipsilateral ventral horns. The JNK inhibitor SP600125, applied immediately post-injury, resulted in an upregulation of nNOS protein both in injured spinal cords and motoneurons and caused a slight alleviation of motoneuron death by inhibiting c-jun phosphorylation at 14 days post-injury. Our results demonstrated that the JNK/c-jun signal transduction pathway is involved in root-avulsion. The inhibition of c-jun phosphorylation prevents nNOS levels from dropping below baseline levels in the spinal cord and partially alleviates motoneuron death following root-avulsion. Therefore, inhibiting c-jun phosphorylation or up-regulating the nNOS protein in injured spinal cords at the early stage might be used in the future as the molecular-target strategies to prevent the motoneurons degeneration in root-avulsion.

Highlights

▶Root-avulsion induces c-jun phosphorylation in spinal cord and inside motoneurons of adult rats. ▶c-jun phosphorylation inhibits nNOS activation and counts for motoneurons death after avulsion. ▶nNOS protein level down to the baseline in spinal cord is harmful for motoneurons to survive avulsion. ▶The phosphorylated c-jun and nNOS are colocalized in injured motoneurons after avulsion. ▶nNOS maybe one of the downstream molecules of c-jun phosphorylation in root-avulsion injury.

Section snippets

Animal surgery

Adult male Sprague–Dawley rats (200–250 g) were bought from the Laboratory Animal Center of Sun Yat-sen University. All procedures related to the care of animals were carried out according to the Chinese National Institutes of Health Guide for the Care and Use of Laboratory Animals and were approved by the Committee on the Use of Live Animals in Teaching and Research of Sun Yat-sen University. Animals were housed in environmentally enriched conditions under a 12 h light/dark cycle throughout

Root-avulsion increased the level of c-jun phosphorylation but decreased the level of nNOS in injured spinal cords

The activation of the JNK/c-jun pathway and nNOS in spinal cord tissues following root-avulsion were assessed by a JNK kinase assay and Western blotting from 4 h to 14 days post-injury (Fig. 1). The result of JNK kinase assay showed an obvious change in JNK activity, which increased approximately 1.3 fold from 4 to 12 h and then decreased from 1 to 14 days post-injury in injured spinal cords following root-avulsion (Fig. 1A). Statistical analysis showed that the JNK activity was significantly

Discussion

In the present study, we demonstrated the specific involvement of the JNK/c-jun signal transduction pathway in root-avulsion, and we found that phosphorylation of c-jun contributes to the avulsion-induced reduction of nNOS expression in the injured spinal cord within the first 14 days of root-avulsion. In addition, the present study showed an inverse relationship between phospho-c-jun and nNOS expression in avulsion-injured spinal cords of adult rats. Inhibition of c-jun phosphorylation

Conclusion

The present study provides evidence that JNK/c-jun signaling is involved in brachial root-avulsion. The phosphorylation of c-jun contributed to the avulsion-induced reduction in nNOS gene expression in injured spinal cords within the first 14 days following root-avulsion. The reduction of nNOS below baseline levels in whole injured spinal cords is harmful for injured motoneurons to survive the root-avulsion. Further studies should use siRNA to downregulate c-jun gene expression to determine

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    These authors contributed equally to this work.

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