Anna V. Leonard

Neurogenic inflammation after traumatic brain injury and its potentiation of classical inflammation

BackgroundThe neuroinflammatory response following traumatic brain injury (TBI) is known to be a key secondary injury factor that can drive ongoing neuronal injury. Despite this, treatments that have targeted aspects of the inflammatory pathway have not shown significant efficacy in clinical trials.Main bodyWe suggest that this may be because classical inflammation only represents part of the story, with activation of neurogenic inflammation potentially one of the key initiating inflammatory events following TBI. Indeed, evidence suggests that the transient receptor potential cation channels (TRP channels), TRPV1 and TRPA1, are polymodal receptors that are activated by a variety of stimuli associated with TBI, including mechanical shear stress, leading to the release of neuropeptides such as substance P (SP). SP augments many aspects of the classical inflammatory response via activation of microglia and astrocytes, degranulation of mast cells, and promoting leukocyte migration. Furthermore, SP may initiate the earliest changes seen in blood-brain barrier (BBB) permeability, namely the increased transcellular transport of plasma proteins via activation of caveolae. This is in line with reports that alterations in transcellular transport are seen first following TBI, prior to decreases in expression of tight-junction proteins such as claudin-5 and occludin. Indeed, the receptor for SP, the tachykinin NK1 receptor, is found in caveolae and its activation following TBI may allow influx of albumin and other plasma proteins which directly augment the inflammatory response by activating astrocytes and microglia.ConclusionsAs such, the neurogenic inflammatory response can exacerbate classical inflammation via a positive feedback loop, with classical inflammatory mediators such as bradykinin and prostaglandins then further stimulating TRP receptors. Accordingly, complete inhibition of neuroinflammation following TBI may require the inhibition of both classical and neurogenic inflammatory pathways.

Kinin Receptor Antagonists as Potential Neuroprotective Agents in Central Nervous System Injury

Injury to the central nervous system initiates complex physiological, cellular and molecular processes that can result in neuronal cell death. Of interest to this review is the activation of the kinin family of neuropeptides, in particular bradykinin and substance P. These neuropeptides are known to have a potent pro-inflammatory role and can initiate neurogenic inflammation resulting in vasodilation, plasma extravasation and the subsequent development of edema. As inflammation and edema play an integral role in the progressive secondary injury that causes neurological deficits, this review critically examines kinin receptor antagonists as a potential neuroprotective intervention for acute brain injury, and more specifically, traumatic brain and spinal cord injury and stroke.

The neuroprotective activity of the amyloid precursor protein against traumatic brain injury is mediated via the heparin binding site in residues 96-110

We have previously shown that following traumatic brain injury (TBI) the presence of the amyloid precursor protein (APP) may be neuroprotective. APP knockout mice have increased neuronal death and worse cognitive and motor outcomes following TBI, which is rescued by treatment with exogenous sAPPα (the secreted ectodomain of APP generated by α‐secretase cleavage). Two neuroprotective regions were identified in sAPPα, the N and C‐terminal domains D1 and D6a/E2 respectively. As both D1 and D6a/E2 contain heparin binding activity it was hypothesized that this is responsible for the neuroprotective activity. In this study, we focused on the heparin binding site, encompassed by residues 96‐110 in D1, which has previously been shown to have neurotrophic properties. We found that treatment with APP96‐110 rescued motor and cognitive deficits in APP−/− mice following focal TBI. APP96‐110 also provided neuroprotection in Sprague–Dawley rats following diffuse TBI. Treatment with APP96‐110 significantly improved functional outcome as well as preserve histological cellular morphology in APP−/− mice following focal controlled cortical impact injury. Furthermore, following administration of APP96‐110 in rats after diffuse impact acceleration TBI, motor and cognitive outcomes were significantly improved and axonal injury reduced. These data define the heparin binding site in the D1 domain of sAPPα, represented by the sequence APP96‐110, as the neuroprotective site to confer neuroprotection following TBI.

Substance P Antagonists as a Novel Intervention for Brain Edema and Raised Intracranial Pressure

Increased intracranial pressure (ICP) following acute brain injury requires the accumulation of additional water in the intracranial vault. One source of such water is the vasculature, although the mechanisms associated with control of blood-brain barrier permeability are unclear. We have recently shown that acute brain injury, such as neurotrauma and stroke, results in perivascular accumulation of the neuropeptide, substance P. This accumulation is associated with increased blood-brain barrier permeability and formation of vasogenic edema. Administration of a substance P antagonist targeting the tachykinin NK1 receptor profoundly reduced the increased blood-brain barrier permeability and edema formation, and in small animal models of acute brain injury, improved functional outcome. In a large, ovine model of experimental traumatic brain injury, trauma resulted in a significant increase in ICP. Administration of an NK1 antagonist caused a profound reduction in post--traumatic ICP, with levels returning to normal within 4 h of drug administration. Substance P NK1 antagonists offer a novel therapeutic approach to the treatment of acute brain injury.

Differential Effects of 670 and 830 nm Red near Infrared Irradiation Therapy: A Comparative Study of Optic Nerve Injury, Retinal Degeneration, Traumatic Brain and Spinal Cord Injury

Red/near-infrared irradiation therapy (R/NIR-IT) delivered by laser or light-emitting diode (LED) has improved functional outcomes in a range of CNS injuries. However, translation of R/NIR-IT to the clinic for treatment of neurotrauma has been hampered by lack of comparative information regarding the degree of penetration of the delivered irradiation to the injury site and the optimal treatment parameters for different CNS injuries. We compared the treatment efficacy of R/NIR-IT at 670 nm and 830 nm, provided by narrow-band LED arrays adjusted to produce equal irradiance, in four in vivo rat models of CNS injury: partial optic nerve transection, light-induced retinal degeneration, traumatic brain injury (TBI) and spinal cord injury (SCI). The number of photons of 670 nm or 830 nm light reaching the SCI injury site was 6.6% and 11.3% of emitted light respectively. Treatment of rats with 670 nm R/NIR-IT following partial optic nerve transection significantly increased the number of visual responses at 7 days after injury (P≤0.05); 830 nm R/NIR-IT was partially effective. 670 nm R/NIR-IT also significantly reduced reactive species and both 670 nm and 830 nm R/NIR-IT reduced hydroxynonenal immunoreactivity (P≤0.05) in this model. Pre-treatment of light-induced retinal degeneration with 670 nm R/NIR-IT significantly reduced the number of Tunel+ cells and 8-hydroxyguanosine immunoreactivity (P≤0.05); outcomes in 830 nm R/NIR-IT treated animals were not significantly different to controls. Treatment of fluid-percussion TBI with 670 nm or 830 nm R/NIR-IT did not result in improvements in motor or sensory function or lesion size at 7 days (P>0.05). Similarly, treatment of contusive SCI with 670 nm or 830 nm R/NIR-IT did not result in significant improvements in functional recovery or reduced cyst size at 28 days (P>0.05). Outcomes from this comparative study indicate that it will be necessary to optimise delivery devices, wavelength, intensity and duration of R/NIR-IT individually for different CNS injury types.

A Substance P Antagonist Improves Outcome in Female Sprague Dawley Rats Following Diffuse Traumatic Brain Injury

Over the past decade it has become increasingly clear from work in experimental models that the secondary injury cascade following traumatic brain injury (TBI) may differ between males and females [1, 2], with females exhibiting a different temporal profile of edema [1] and neurodegeneration compared to males [2]. Furthermore significant neuroprotection was noted in male, but not female rodents treated with posttraumatic hypothermia [3], or a dopamine agonist [4]. This highlights the need to ensure that experimental treatments are equally efficacious in both genders before proceeding to clinical trials that are performed in mixed populations. Previous research in our laboratory found that substance P (SP) plays an integral role in the secondary injury cascade following TBI in males [5]. SP is a member of the tachykinin family of neuropeptides, with its release causing the development of neurogenic inflammation characterised by vasodilation, plasma extravasation, and tissue swelling [6]. These effects are principally mediated by the binding of SP to the NK1 tachykinin receptor [6]. Following impact-acceleration TBI in male rodents SP levels were found to increase. Furthermore, inhibition of SP, through treatment with an NK1 receptor antagonist reduced vasogenic edema formation and axonal injury, with a corresponding improvement in motor and cognitive outcome [5, 7]. However, it is not known whether SP plays a similar role in females following TBI. As such, the effects of an NK1 receptor antagonist, N-acetyl-L-tryptophan (NAT) on outcome following TBI in female rodents were investigated. Female Sprague Dawley rats were injured using the impactacceleration model of diffuse TBI, as previously described [5]. At 30 mins postinjury rats were treated intravenously with either 2.5 mg/kg of NAT or an equal volume of saline. Sham animals were surgically prepared but not injured. Motor deficits were assessed on rotarod as previously described [5]. For histological analysis, rats were transcardially perfused fixed, the brains removed and processed. Slides were labeled with APP (1:1,000), SP (1:2,000), or albumin (1:20,000). To objectively analyse levels of SP and albumin, Ruifrok and Johnston’s color deconvolution method was employed on 2 slides per animal to determine the amount of DAB and hence antigen that was present, as previously described [8]. Axonal injury was determined by counting the number of APP immunopositive lengths within the corpus callosum. For analysis of edema the specific gravity of the brain was determined by placing it in a Percoll density gradient, with this measurement then converted to% water as previously described [9]. All data were analyzed using a one or two-way ANOVA as appropriate, followed by Bonferonni t-tests using Graphpad Prism software. SP immunoreactivity was assessed postinjury to determine whether levels were increased, as previously reported in males (Figures 1A and B). Indeed, at 24 h postinjury increased SP immunoreactivity was observed, particularly apparent in perivascular nerve fibers and astrocytic processes in vehicle treated rats. In contrast low levels of SP immunoreactivity were observed in shams. Furthermore color deconvolution analysis demonstrated a significant increase in%DAB weight in vehicle treated rats, with 28.0 ± 3.6% compared to 16.6 ± 2.8% in sham controls (P < 0.01).

Substance P as a Mediator of Neurogenic Inflammation after Balloon Compression Induced Spinal Cord Injury

Although clinical spinal cord injury (SCI) occurs within a closed environment, most experimental models of SCI create an open injury. Such an open environment precludes the measurement of intrathecal pressure (ITP), whose increase after SCI has been linked to the development of greater tissue damage and functional deficits. Raised ITP may be potentiated by edema, which we have recently shown to be associated with substance P (SP) induced neurogenic inflammation in both traumatic brain injury and stroke. The present study investigates whether SP plays a similar role as a mediator of neurogenic inflammation after SCI. A closed balloon compression injury was induced at T10 in New Zealand white rabbits. Animals were thereafter assessed for blood spinal cord barrier (BSCB) permeability, edema, ITP, histological outcome, and functional outcome from 5 h to 2 weeks post-SCI. The balloon compression model produced significant increases in BSCB permeability, edema, and ITP along with significant functional deficits that persisted for 2 weeks. Histological assessment demonstrated decreased SP immunoreactivity in the injured spinal cord while NK1 receptor immunoreactivity initially increased before returning to sham levels. In addition, aquaporin 4 immunoreactivity increased early post-SCI, implicating this water channel in the development of edema after SCI. The changes described in the present study support a role for SP as a mediator of neurogenic inflammation after SCI.

Localization of the corticospinal tract within the porcine spinal cord: Implications for experimental modeling of traumatic spinal cord injury

Spinal cord injury (SCI) researchers have predominately utilized rodents for SCI modeling and experimentation. Unfortunately, a large number of novel therapies developed in rodent models have failed to demonstrate efficacy in human clinical trials which suggests that improved animal models are an important translational tool. Recently, porcine models of SCI have been identified as a valuable intermediary model for preclinical evaluation of promising therapies to aid clinical translation. However, the localization of the major spinal tracts in pigs has not yet been described. Given that significant differences exist in the location of the corticospinal tract (CST) between rodents and humans, determining its location in pigs will provide important information related to the translational potential of the porcine pre-clinical model of SCI. Thus, the goal of this study is to investigate the localization of the CST within the porcine spinal cord. Mature female domestic pigs (n=4, 60kg) received microinjections of fluorescent dextran tracers (Alexa Fluor, 10,000MW) into the primary motor cortex, using image-guided navigation (StealthStation®), to label the CST. At 5 weeks post-tracer injection animals were euthanized, the entire neuroaxis harvested and processed for histological examination. Serial sections of the brain and spinal cord were prepared and imaged using confocal microscopy to observe the location of the CST in pigs. Results demonstrate that the CST of pigs is located in the lateral white matter, signifying greater similarity to human anatomical structure compared to that of rodents. We conclude that the corticospinal tract in pigs demonstrates anatomical similarity to human, suggesting that the porcine model has importance as a translational intermediary pre-clinical model.

Reducing intrathecal pressure after traumatic spinal cord injury: a potential clinical target to promote tissue survival

Spinal cord injury (SCI) is an unexpected event that is both devastating and debilitating, resulting in not just motor and sensory loss, but also autonomic dysfunction of the bladder, bowel and sexual organs. Currently, there are no treatments available to improve outcome following SCI, leaving individuals with permanent and lifelong physical disability. Worldwide it is estimated that more than 500,000 people sustain a SCI each year, with average lifetime cost of paraplegia and quadriplegia estimated at

Changes in substance P and NK1 receptor immunohistochemistry following human spinal cord injury

5 million and