Activating Transcription Factor 3, a Useful Marker for Regenerative Response after Nerve Root Injury

Activating transcription factor 3 (ATF3) is induced in various tissues in response to stress. In this experiment, ATF3 expression was studied in adult rats subjected either to a dorsal or ventral root avulsion (VRA; L4-6), or sciatic nerve transection (SNT). Post-operative survival times varied between 1.5 h and 3 weeks. In additional experiments an avulsed ventral root was directly replanted to the spinal cord. Dorsal root ganglias (DRGs) from humans exposed to traumatic dorsal root avulsions were also examined. After SNT ATF3 immunoreactivity (ATF3 IR) was detected in a few DRG neurons already 6 h after the lesion. After 24 h the number had clearly increased and still at 3 weeks DRG neurons remained labeled. In the ventral horn, ATF3 IR in motoneurons (MN) was first detected 24 h after the SNT, and still 3 weeks post-operatively lesioned MN showed ATF3 labeling. After a VRA many spinal MN showed ATF3 IR already after 3 h, and after 6 h all MN were labeled. At 3 weeks a majority of the lesioned MN had died, but all the remaining ones were labeled. When an avulsed ventral root was directly replanted, MN survived and were still labeled at 5 weeks. In DRG, a few neurons were labeled already at 1.5 h after a dorsal root avulsion. At 24 h the number had increased but still only a minority of the neurons were labeled. At 3 days the number of labeled neurons was reduced, and a further reduction was at hand at 7 days and 3 weeks. In parallel, in humans, 3 days after a traumatic dorsal root avulsion, only a few DRG neurons showed ATF3 IR. At 6 weeks no labeled neurons could be detected. These facts imply that ATF3 response to axotomy involves a distance-dependent mechanism. ATF3 also appears to be a useful and reliable neuronal marker of nerve lesions even in humans. In addition, ATF3 up-regulation in both motor and sensory neurons seems to be linked to regenerative competence

On Acute Gene Expression Changes after Ventral Root Replantation

Replantation of avulsed spinal ventral roots has been show to enable significant and useful regrowth of motor axons in both experimental animals and in human clinical cases, making up an interesting exception to the rule of unsuccessful neuronal regeneration in central nervous system. Compared to avulsion without repair, ventral root replantation seems to rescue lesioned motoneurons from death. In this study we have analyzed the acute response to ventral root avulsion and replantation in adult rats with gene arrays combined with cluster analysis of gene ontology search terms. The data show significant differences between rats subjected to ventral replantation compared to avulsion only. Even though number of genes related to cell death is similar in the two models after 24 h, we observed a significantly larger number of genes related to neurite growth and development in the rats treated with ventral root replantation, possibly reflecting the neuroregenerative capacity in the replantation model. In addition, an acute inflammatory response was observed after avulsion, while effects on genes related to synaptic transmission were much more pronounced after replantation than after avulsion alone. These data indicate that the axonal regenerative response from replantation is initiated at an earlier stage than the possible differences in terms of neuron survival. We conclude that this type of analysis may facilitate the comparison of the acute response in two types of injury

Karolinska Institutet 200-Year Anniversary. Symposium on Traumatic Injuries in the Nervous System: Injuries to the Spinal Cord and Peripheral Nervous System – Injuries and Repair, Pain Problems, Lesions to Brachial Plexus

The Karolinska Institutet 200-year anniversary symposium on injuries to the spinal cord and peripheral nervous system gathered expertise in the spinal cord, spinal nerve, and peripheral nerve injury field spanning from molecular prerequisites for nerve regeneration to clinical methods in nerve repair and rehabilitation. The topics presented at the meeting covered findings on adult neural stem cells that when transplanted to the hypoglossal nucleus in the rat could integrate with its host and promote neuron survival. Studies on vascularization after intraspinal replantation of ventral nerve roots and microarray studies in ventral root replantation as a tool for mapping of biological patterns typical for neuronal regeneration were discussed. Different immune molecules in neurons and glia and their very specific roles in synapse plasticity after injury were presented. Novel strategies in repair of injured peripheral nerves with ethyl-cyanoacrylate adhesive showed functional recovery comparable to that of conventional epineural sutures. Various aspects on surgical techniques which are available to improve function of the limb, once the nerve regeneration after brachial plexus lesions and repair has reached its limit were presented. Moreover, neurogenic pain after amputation and its treatment with mirror therapy were shown to be followed by dramatic decrease in phantom limb pain. Finally clinical experiences on surgical techniques to repair avulsed spinal nerve root and the motoric as well as sensoric regain of function were presented

Expression of Semaphorins, Neuropilins, VEGF, and Tenascins in Rat and Human Primary Sensory Neurons after a Dorsal Root Injury

Dorsal root injury is a situation not expected to be followed by a strong regenerative growth, or growth of the injured axon into the central nervous system of the spinal cord, if the central axon of the dorsal root is injured but of strong regeneration if subjected to injury to the peripherally projecting axons. The clinical consequence of axonal injury is loss of sensation and may also lead to neuropathic pain. In this study, we have used in situ hybridization to examine the distribution of mRNAs for the neural guidance molecules semaphorin 3A (SEMA3A), semaphorin 3F (SEMA3F), and semaphorin 4F (SEMA4F), their receptors neuropilin 1 (NP1) and neuropilin 2 (NP2) but also for the neuropilin ligand vascular endothelial growth factor (VEGF) and Tenascin J1, an extracellular matrix molecule involved in axonal guidance, in rat dorsal root ganglia (DRG) after a unilateral dorsal rhizotomy (DRT) or sciatic nerve transcetion (SNT). The studied survival times were 1-365 days. The different forms of mRNAs were unevenly distributed between the different size classes of sensory nerve cells. The results show that mRNA for SEMA3A was diminished after trauma to the sensory nerve roots in rats. The SEMA3A receptor NP1, and SEMA3F receptor NP2, was significantly upregulated in the DRG neurons after DRT and SNT. SEMA4F was upregulated after a SNT. The expression of mRNA for VEGF in DRG neurons after DRT showed a significant upregulation that was high even a year after the injuries. These data suggest a role for the semaphorins, neuropilins, VEGF, and J1 in the reactions after dorsal root lesions

Editorial: Karolinska Institutet 200-Year Anniversary Symposium on Injuries to the Spinal Cord and Peripheral Nervous System : An Update on Recent Advances in Regenerative Neuroscience

The present E-book consists of original articles and reviews published in our Research Topic on injuries to the spinal cord and peripheral nerves and presents a wide array of novel findings and in depth discussions on topics within the field of nerve injury and repair. Our aim with this Research Topic is to bring together knowledge spanning from basic laboratory studies to clinical findings and strategies within the field of spinal cord and nerve injury and repair. We hope this publication will provide a basis for accelerated knowledge exchange within the field and hopefully a subsequent increase in research efforts and collaborations

Diffuse Axonal Injury in the Rat Brain : Axonal Injury and Oligodendrocyte Activity Following Rotational Injury

Traumatic brain injury (TBI) commonly results in primary diffuse axonal injury (DAI) and associated secondary injuries that evolve through a cascade of pathological mechanisms. We aim at assessing how myelin and oligodendrocytes react to head angular-acceleration-induced TBI in a previously described model. This model induces axonal injuries visible by amyloid precursor protein (APP) expression, predominantly in the corpus callosum and its borders. Brain tissue from a total of 27 adult rats was collected at 24 h, 72 h and 7 d post-injury. Coronal sections were prepared for immunohistochemistry and RNAscope(R) to investigate DAI and myelin changes (APP, MBP, Rip), oligodendrocyte lineage cell loss (Olig2), oligodendrocyte progenitor cells (OPCs) (NG2, PDGFRa) and neuronal stress (HSP70, ATF3). Oligodendrocytes and OPCs numbers (expressed as percentage of positive cells out of total number of cells) were measured in areas with high APP expression. Results showed non-statistically significant trends with a decrease in oligodendrocyte lineage cells and an increase in OPCs. Levels of myelination were mostly unaltered, although Rip expression differed significantly between sham and injured animals in the frontal brain. Neuronal stress markers were induced at the dorsal cortex and habenular nuclei. We conclude that rotational injury induces DAI and neuronal stress in specific areas. We noticed indications of oligodendrocyte death and regeneration without statistically significant changes at the timepoints measured, despite indications of axonal injuries and neuronal stress. This might suggest that oligodendrocytes are robust enough to withstand this kind of trauma, knowledge important for the understanding of thresholds for cell injury and post-traumatic recovery potential

Nerve Tracing in Juvenile Rats: A Feasible Model for the Study of Brachial Plexus Birth Palsy and Cocontractions?

Brachial plexus birth injuries cause diminished motor function in the upper extremity. The most common sequel is internal rotation contracture. A number of these patients also suffer from cocontractions, preventing the use of an otherwise good passive range of motion in the shoulder. One theory behind the co-contracture problem is that injured nerve fibers grow into distal support tissue not corresponding to the proximal support tissue, resulting in reinnervation of the wrong muscle groups. To further elucidate this hypothesis, we used rat neonates to investigate a possible model for the study of cocontractions in brachial plexus birth injuries. Five-day-old rats were subjected to a crush injury to the C5–C6 roots. After a healing period of 4 weeks, the infraspinatus muscle was injected with Fluoro-Gold. A week later, the animals were perfused and spinal cords harvested and sectioned. Differences in the uptake of Fluoro-Gold and NeuN positive cells of between sides of the spinal cord were recorded. We found a larger amount of Fluoro-Gold positive cells on the uninjured side, while the injured side had positive cells dispersed over a longer area in the craniocaudal direction. Our findings indicate that the method can be used to trace Fluoro-Gold from muscle through a neuroma. Our results also indicate that a neuroma in continuity somewhat prevents the correct connection from being established between the motor neuron pool in the spinal cord and target muscle and that some neurons succumb to a crushing injury. We also present future research ideas

Hippocampal Expression of Cytochrome P450 1B1 in Penetrating Traumatic Brain Injury

Hippocampal dysfunction contributes to multiple traumatic brain injury sequala. Female rodents' outcome is superior to male which has been ascribed the neuroprotective sex hormones 17 beta-estradiol and progesterone. Cytochrome P450 1B1 (CYP1B1) is an oxidative enzyme influencing the neuroinflammatory response by creating inflammatory mediators and metabolizing neuroprotective 17 beta-estradiol and progesterone. In this study, we aimed to describe hippocampal CYP1B1 mRNA expression, protein presence of CYP1B1 and its key redox partner Cytochrome P450 reductase (CPR) in both sexes, as well as the effect of penetrating traumatic brain injury (pTBI). A total 64 adult Sprague Dawley rats divided by sex received pTBI or sham-surgery and were assigned survival times of 1-, 3-, 5- or 7 days. CYP1B1 mRNA was quantified using in-situ hybridization and immunohistochemistry performed to verify protein colocalization. CYP1B1 mRNA expression was present in all subregions but greatest in CA2 irrespective of sex, survival time or intervention. At 3-, 5- and 7 days post-injury, expression in CA2 was reduced in male rats subjected to pTBI compared to sham-surgery. Females subjected to pTBI instead exhibited increased expression in all CA subregions 3 days post-injury, the only time point expression in CA2 was greater in females than in males. Immunohistochemical analysis confirmed neuronal CYP1B1 protein in all hippocampal subregions, while CPR was limited to CA1 and CA2. CYP1B1 mRNA is constitutively expressed in both sexes. In response to pTBI, females displayed a more urgent but brief regulatory response than males. This indicates there may be sex-dependent differences in CYP1B1 activity, possibly influencing inflammation and neuroprotection in pTBI