Secondary Injury Mechanisms Research Articles

Matrix metalloproteinase-9 regulates the blood brain barrier via the hedgehog pathway in a rat model of traumatic brain injury

The mechanisms of secondary brain injury after traumatic brain injury (TBI) are complex and are the result of multiple factors. Protecting the blood-brain barrier (BBB) and ameliorating cerebral edema are two key factors for improving the prognosis of TBI patients. The BBB is regulated by the hedgehog pathway through Scube2 and Shh protein. Matrix metalloproteinase-9 (MMP-9) influences the transport system and enzyme system of vascular endothelial cells, possibly via the hedgehog pathway. The present study aimed to investigate the role and mechanism of MMP-9 in TBI via the hedgehog pathway. Eighty male Sprague-Dawley rats were used to establish a murine model of TBI. Subsequently, the effect of SB-3CT—a specific inhibitor of MMP-9—was assessed via Western blotting, real-time PCR, immunofluorescence, apoptotic assays, and neurological scoring. The results showed that, compared with those of the sham-operation group, the mRNA and protein levels of MMP-9 were significantly increased after TBI, while the expressions of Scube2 and Shh were decreased. Application of SB-3CT at 24 h after TBI significantly reduced neuronal apoptosis and BBB permeability, while increasing expressions of Scube2 and Shh. In conclusion, these findings demonstrate an influence of TBI-induced MMP-9 upregulation in the induction of post-traumatic nerve and BBB injury, which may be partially mediated by Scube2 and Shh via the hedgehog pathway.

Neuro-Inflammation in Pediatric Traumatic Brain Injury-from Mechanisms to Inflammatory Networks.

Compared to traumatic brain injury (TBI) in the adult population, pediatric TBI has received less research attention, despite its potential long-term impact on the lives of many children around the world. After numerous clinical trials and preclinical research studies examining various secondary mechanisms of injury, no definitive treatment has been found for pediatric TBIs of any severity. With the advent of high-throughput and high-resolution molecular biology and imaging techniques, inflammation has become an appealing target, due to its mixed effects on outcome, depending on the time point examined. In this review, we outline key mechanisms of inflammation, the contribution and interactions of the peripheral and CNS-based immune cells, and highlight knowledge gaps pertaining to inflammation in pediatric TBI. We also introduce the application of network analysis to leverage growing multivariate and non-linear inflammation data sets with the goal to gain a more comprehensive view of inflammation and develop prognostic and treatment tools in pediatric TBI.

Unlocking the paradoxical endogenous stem cell response after spinal cord injury.

Nearly a century ago, the concept of the secondary injury in spinal cord trauma was first proposed to explain the complex cascade of molecular and cellular events leading to widespread neuronal and glial cell death after trauma. In recent years, it has been established that the ependymal region of the adult mammalian spinal cord contains a population of multipotent neural stem/progenitor cells (NSPCs) that are activated after spinal cord injury (SCI) and likely play a key role in endogenous repair and regeneration. How these cells respond to the various components of the secondary injury remains poorly understood. Emerging evidence suggests that many of the biochemical components of the secondary injury cascade which have classically been viewed as deleterious to host neuronal and glial cells may paradoxically trigger NSPC activation, proliferation, and differentiation thus challenging our current understanding of secondary injury mechanisms in SCI. Herein, we highlight new findings describing the response of endogenous NSPCs to spinal cord trauma, redefining the secondary mechanisms of SCI through the lens of the endogenous population of stem/progenitor cells. Moreover, we outline how these insights can fuel novel stem cell-based therapeutic strategies to repair the injured spinal cord.

Decellularized brain matrix enhances macrophage polarization and functional improvements in rat spinal cord injury

Spinal cord injury (SCI) is a devastating lesion lacking effective treatment options currently available in clinics. The inflammatory process exacerbates the extent of the lesion through a secondary injury mechanism, where proinflammatory classically activated macrophages (M1) are prevalent at the lesion site. However, the polarized alternatively activated anti-inflammatory macrophages (M2) are known to play an important role in wound healing and regeneration following SCI. Herein, we introduce porcine brain decellularized extracellular matrix (dECM) to modulate the macrophages in the injured spinal cord. The hydrogels with collagen and dECM at various dECM concentrations (1, 5, and 8 mg/ml) were used to cultivate primary macrophages and neurons. The dECM hydrogels were shown to promote the polarization of macrophages toward M2 phase and the neurite outgrowth of cortical and hippocampal neurons. When the dECM hydrogels were applied to rat SCI models, the proportion of M1 and M2 macrophages in the injured spinal cord was substantially altered. When received dECM concetration of 5 mg/ml, the expression of molecules associated with M2 (CD206, arginase1, and IL-10) was significantly increased. Consistently, the population of total macrophages and cavity area were substantially reduced in the dECM-treated groups. As a result, the locomotor functions of injured spinal cord, as assessed by BBB and ladder scoring, were significantly improved. Collectively, the porcine brain dECM with optimal concentration promotes functional recovery in SCI models through the activation of M2 macrophages, suggesting the promising use of the engineered hydrogels in the treatment of acute SCI. Statement of significanceSpinal cord injury (SCI) is a devastating lesion, lacking effective treatment options currently available in clinics. Here we delineated that the treatment of injured spinal cord with porcine brain decellularized matrix-based hydrogels for the first time, and could modulate the macrophage polarization and the ultimate functional recovery. When appropriate formulations were applied to a contused spinal cord model in rats, the decellularized matrix hydrogels shifted the macrophages to polarize to pro-regenerative M2 phenotype, decreased the size of lesion cavity, and finally promoted the locomotor functions until 8 weeks following the injury. We consider this work can significantly augment the matrix(biomaterial)-based therapeutic options, as an alternative to drug or cell-free approaches, for the treatment of acute injury of spinal cord.

NDP-MSH binding melanocortin-1 receptor ameliorates neuroinflammation and BBB disruption through CREB/Nr4a1/NF-\u03baB pathway after intracerebral hemorrhage in mice

BackgroundNeuroinflammation and blood-brain barrier (BBB) disruption are two vital mechanisms of secondary brain injury following intracerebral hemorrhage (ICH). Recently, melanocortin-1 receptor (Mc1r) activation by Nle4-D-Phe7-α-MSH (NDP-MSH) was shown to play a neuroprotective role in an experimental autoimmune encephalomyelitis (EAE) mouse model. This study aimed to investigate whether NDP-MSH could alleviate neuroinflammation and BBB disruption after experimental ICH, as well as the potential mechanisms of its neuroprotective roles.MethodsTwo hundred and eighteen male C57BL/6 mice were subjected to autologous blood-injection ICH model. NDP-MSH, an agonist of Mc1r, was administered intraperitoneally injected at 1 h after ICH insult. To further explore the related protective mechanisms, Mc1r small interfering RNA (Mc1r siRNA) and nuclear receptor subfamily 4 group A member 1 (Nr4a1) siRNA were administered via intracerebroventricular (i.c.v) injection before ICH induction. Neurological test, BBB permeability, brain water content, immunofluorescence staining, and Western blot analysis were implemented.ResultsThe Expression of Mc1r was significantly increased after ICH. Mc1r was mainly expressed in microglia, astrocytes, and endothelial cells following ICH. Treatment with NDP-MSH remarkably improved neurological function and reduced BBB disruption, brain water content, and the number of microglia in the peri-hematoma tissue after ICH. Meanwhile, the administration of NDP-MSH significantly reduced the expression of p-NF-κB p65, IL-1β, TNF-α, and MMP-9 and increased the expression of p-CREB, Nr4a1, ZO-1, occludin, and Lama5. Inversely, the knockdown of Mc1r or Nr4a1 abolished the neuroprotective effects of NDP-MSH.ConclusionsTaken together, NDP-MSH binding Mc1r attenuated neuroinflammation and BBB disruption and improved neurological deficits, at least in part through CREB/Nr4a1/NF-κB pathway after ICH.

SCING—Spinal Cord Injury Neuroprotection with Glyburide: a pilot, open-label, multicentre, prospective evaluation of oral glyburide in patients with acute traumatic spinal cord injury in the USA

IntroductionAcute traumatic spinal cord injury (tSCI) is a devastating neurological disorder with no pharmacological neuroprotective strategy proven effective to date. Progressive haemorrhagic necrosis (PHN) represents an increasingly well-characterised mechanism of...

Monitoring of Protein Biomarkers of Inflammation in Human Traumatic Brain Injury Using Microdialysis and Proximity Extension Assay Technology in Neurointensive Care.

Traumatic brain injury (TBI) is followed by secondary injury mechanisms strongly involving neuroinflammation. To monitor the complex inflammatory cascade in human TBI, we used cerebral microdialysis (MD) and multiplex proximity extension assay (PEA) technology and simultaneously measured levels of 92 protein biomarkers of inflammation in MD samples every three hours for five days in 10 patients with severe TBI under neurointensive care. One μL MD samples were incubated with paired oligonucleotide-conjugated antibodies binding to each protein, allowing quantification by real-time quantitative polymerase chain reaction. Sixty-nine proteins were suitable for statistical analysis. We found five different patterns with either early (<48 h; e.g., CCL20, IL6, LIF, CCL3), mid (48–96 h; e.g., CCL19, CXCL5, CXCL10, MMP1), late (>96 h; e.g., CD40, MCP2, MCP3), biphasic peaks (e.g., CXCL1, CXCL5, IL8) or stable (e.g., CCL4, DNER, VEGFA)/low trends. High protein levels were observed for e.g., CXCL1, CXCL10, MCP1, MCP2, IL8, while e.g., CCL28 and MCP4 were detected at low levels. Several proteins (CCL8, -19, -20, -23, CXCL1, -5, -6, -9, -11, CST5, DNER, Flt3L, and SIRT2) have not been studied previously in human TBI. Cross-correlation analysis revealed that LIF and CXCL5 may play a central role in the inflammatory cascade. This study provides a unique data set with individual temporal trends for potential inflammatory biomarkers in patients with TBI. We conclude that the combination of MD and PEA is a powerful tool to map the complex inflammatory cascade in the injured human brain. The technique offers new possibilities of protein profiling of complex secondary injury pathways.

34. Timing of surgery for thoracolumbar spine trauma: patients without neurological injury

BACKGROUND CONTEXT Over 100,000 thoracolumbar fractures occur annually in North America, with presence of concurrent neurological injury associated with greater long-term morbidity and unemployment. Previous investigations document heterogeneity in timing-outcome studies for thoracolumbar fracture, with most studies failing to differentiate patients with neurological injury from those without. PURPOSE This study analyzes the effect of time from hospital admission to spinal fusion on mortality, in-hospital complications, length of stay (LOS), and hospital charges in a large population of thoracolumbar fracture patients with neurological injury. STUDY DESIGN/SETTING Retrospective cohort study of de-identified National Inpatient Sample data from 2004-2014. PATIENT SAMPLE Nonelective cases containing ICD-9-CM diagnosis codes for closed thoracic/lumbar spinal fracture with concurrent neurological injury (806.20-806.29, 806.4) and procedure codes for primary thoracolumbar/lumbosacral fusion (81.04-81.08). Patients with diagnosis codes for any open, cervical, or sacral spinal fractures or any neurological injury of a non-thoracolumbar region were excluded. OUTCOME MEASURES In-hospital mortality and complications were primary measures. Inpatient surgical complication subtypes included all complications, cardiac complications, intraoperative hemorrhage or hematoma formation, respiratory complications, and postoperative infections. In 19,136 patients analyzed, patients undergoing surgery within 72 hours (n=12,845) had the lowest odds of in-hospital complications (OR=0.834; 95% CI, 0.697-0.999), cardiac complications (OR=0.573; 95% CI, 0.350-0.937), respiratory complications (OR=0.442; 95% CI, 0.282-0.691), and postoperative infection (OR=0.573; 95% CI, 0.366-0.899). No significant differences in outcomes were observed between same-day surgery (n=4,724) and 1-2-day delay (n=8,121). A ≥7-day delay in thoracolumbar fusion (n=2,002) was associated with the greatest odds of intraoperative hemorrhage or hematoma formation (OR=2.471; 95% CI, 1.351-4.517), respiratory complications (OR=1.909; 95% CI, 1.125-3.241), and postoperative infection (OR=3.559; 95% CI, 2.162-5.859). Greater than 7-day delay to fusion was also associated with the highest total and postoperative LOS and total charges (all p METHODS Classification of time from hospital admission to fusion: Same-day, 1-2-day delay, 3-6-day delay, and ≥7-day delay. To reduce data confounding by patient health and stability status, validated injury severity scoring algorithm from the International Classification of Diseases Program for Injury Categorization was used. Survey-design-weighted logistic regressions were performed to assess the effect of surgical timing on the primary inpatient outcomes, controlling for age, sex, fusion approach, and multi-organ-system injury severity score. Using the same controls, survey-design-weighted linear regressions were performed to assess the effect of surgical timing on secondary inpatient outcomes. RESULTS In 19,136 patients analyzed, patients undergoing surgery within 72 hours (n=12,845) had the lowest odds of in-hospital complications (OR=0.834; 95% CI, 0.697-0.999), cardiac complications (OR=0.573; 95% CI, 0.350-0.937), respiratory complications (OR=0.442; 95% CI, 0.282-0.691), and postoperative infection (OR=0.573; 95% CI, 0.366-0.899). No significant differences in outcomes were observed between same-day surgery and 1-2-day delay. A ≥7-day delay in thoracolumbar fusion was associated with the greatest odds of intraoperative hemorrhage/hematoma (OR=2.471; 95% CI, 1.351-4.517), respiratory complications (OR=1.909; 95% CI, 1.125-3.241), and postoperative infection (OR=3.559; 95% CI, 2.162-5.859). Greater than 7-day delay to fusion was also associated with the highest total and postoperative LOS and total charges (all p CONCLUSIONS While timing of spinal fusion following thoracolumbar fracture with neurological injury was not associated with mortality, patients who underwent surgery in ≤3 days after admission experienced fewer in-hospital complications, including hemorrhage, hematoma, respiratory complications, and postoperative infection. These benefits may be due to secondary injury mechanism avoidance and earlier mobilization. FDA DEVICE/DRUG STATUS This abstract does not discuss or include any applicable devices or drugs.

Stroke : Highlights of Selected Articles

<i>Stroke</i> : Highlights of Selected Articles

Intracerebral Hemorrhage: A Brief Evidence-Based Review of Common Etiologies, Mechanisms of Secondary Injury, and Medical and Surgical Management

AbstractIntracerebral hemorrhage (ICH) accounts for only 10 to 15% of all strokes but remains a significant cause of morbidity and mortality. Despite lengthy stays in critical care units, only one-half of those experiencing an ICH survive after 30 days, and those who do are often left with considerable disability. Treatment has traditionally focused on minimizing the hemorrhage expansion and reducing clot volume through both medical and surgical means. Management of ICH is a complex and multidisciplinary process. This review will discuss a few common etiologies, explore the pathophysiology of secondary neuronal injury after ICH, review the basics of ICH imaging with computed tomography and magnetic resonance imaging, and highlight latest practices in medical and surgical management. Secondary injury mechanisms such as perihematomal edema and disordered cerebral autoregulation are discussed as potential targets for new treatment modalities. Emergent treatment in the “golden hour” after ictus provides a template of measures to adopt from initial contact with emergency medical services, to the emergency department, and thereafter, triage to the intensive care unit. Medical management including blood pressure control, hemostasis, and coagulopathy reversal are discussed and evidence from trials such as INTERACT 2, ATACH 2, and ANNEXA-4 are given a clinical context. Surgical management including intracranial pressure monitoring, surgical evacuation with open craniotomy, and minimally invasive approaches such as stereotactic-guided aspiration and thrombolysis, ultrasound-induced thrombolysis, image-guided stereotactic endoscopic aspiration, and stereotactic ICH underwater blood aspiration are enumerated. The outcomes and relevance of STICH, MISTIE, and CLEAR trials to present surgical care are elaborated. The review summarizes the current guidelines for the treatment of ICH and the latest literature in the field they are based upon. It aims to provide a concise article beneficial to the emergency physicians and neurointensivists/neuroanesthesiologists.

The Role of Temporal Lobectomy as a Part of Surgical Resuscitation in Patients with Severe Traumatic Brain Injury.

Background:Traumatic brain injuries (TBIs) are serious morbidity and mortality risk for especially in the young population. Primary and secondary injury mechanisms may cause cerebral edema and intracranial hypertension. The target point of the TBI treatment is lowering the intracranial pressure medically or surgically if indicated.Methods:The files of the patients with severe brain injury admitted between January 2015 and December 2017 were reviewed retrospectively. Patients who underwent decompression surgery due to severe brain injury ([The Glasgow Coma Scale [GCS] score] <8) and additional temporal lobectomy were included in the study group.Results:Ten patients were included in the study during the 3 years. All the patients were suffering from blunt severe TBI. Traumatic etiology was vehicle traffic accident in six cases, nonvehicle traffic accident in two cases, and falling from height in two cases. All the cases suffered from blunt trauma. The admission GCS of the patients was 4–7 (mean = 5.5). Right-sided decompression surgery and lobectomy were performed for seven patients and left-sided in three cases. The postoperational survival was 60%. All the survivors were functionally independent with mild cognitive disturbances.Conclusion:Temporal lobectomy might be added to the surgery to apply all the interventions available in combat with progressively increasing intracerebral pressure as a part of surgical resuscitation.

Perihematomal Edema After Spontaneous Intracerebral Hemorrhage.

Perihematomal Edema After Spontaneous Intracerebral Hemorrhage.

A hydrogel engineered to deliver minocycline locally to the injured cervical spinal cord protects respiratory neural circuitry and preserves diaphragm function

We tested a biomaterial-based approach to preserve the critical phrenic motor circuitry that controls diaphragm function by locally delivering minocycline hydrochloride (MH) following cervical spinal cord injury (SCI). MH is a clinically-available antibiotic and anti-inflammatory drug that targets a broad range of secondary injury mechanisms via its anti-inflammatory, anti-oxidant and anti-apoptotic properties. However, MH is only neuroprotective at high concentrations that cannot be achieved by systemic administration, which limits its clinical efficacy. We have developed a hydrogel-based MH delivery system that can be injected into the intrathecal space for local delivery of high concentrations of MH, without damaging spinal cord tissue. Implantation of MH hydrogel after unilateral level-C4/5 contusion SCI robustly preserved diaphragm function, as assessed by in vivo recordings of compound muscle action potential (CMAP) and electromyography (EMG) amplitudes. MH hydrogel also decreased lesion size and degeneration of cervical motor neuron somata, demonstrating its central neuroprotective effects within the injured cervical spinal cord. Furthermore, MH hydrogel significantly preserved diaphragm innervation by the axons of phrenic motor neurons (PhMNs), as assessed by both detailed neuromuscular junction (NMJ) morphological analysis and retrograde PhMN labeling from the diaphragm using cholera toxin B (CTB). In conclusion, our findings demonstrate that local MH hydrogel delivery to the injured cervical spinal cord is effective in preserving respiratory function after SCI by protecting the important neural circuitry that controls diaphragm activation.

Acute Spinal Cord Injury: A Systematic Review Investigating miRNA Families Involved.

Acute traumatic spinal cord injury (SCI) involves primary and secondary injury mechanisms. The primary mechanism is related to the initial traumatic damage caused by the damaging impact and this damage is irreversible. Secondary mechanisms, which begin as early as a few minutes after the initial trauma, include processes such as spinal cord ischemia, cellular excitotoxicity, ionic dysregulation, and free radical-mediated peroxidation. SCI is featured by different forms of injury, investigating the pathology and degree of clinical diagnosis and treatment strategies, the animal models that have allowed us to better understand this entity and, finally, the role of new diagnostic and prognostic tools such as miRNA could improve our ability to manage this pathological entity. Autopsy could benefit from improvements in miRNA research: the specificity and sensitivity of miRNAs could help physicians in determining the cause of death, besides the time of death.

A New Vision for Therapeutic Hypothermia in the Era of Targeted Temperature Management: A Speculative Synthesis.

Three decades of animal studies have reproducibly shown that hypothermia is profoundly cerebroprotective during or after a central nervous system (CNS) insult. The success of hypothermia in preclinical acute brain injury has not only fostered continued interest in research on the classic secondary injury mechanisms that are prevented or blunted by hypothermia but has also sparked a surge of new interest in elucidating beneficial signaling molecules that are increased by cooling. Ironically, while research into cold-induced neuroprotection is enjoying newfound interest in chronic neurodegenerative disease, conversely, the scope of the utility of therapeutic hypothermia (TH) across the field of acute brain injury is somewhat controversial and remains to be fully defined. This has led to the era of Targeted Temperature Management, which emphasizes a wider range of temperatures (33–36°C) showing benefit in acute brain injury. In this comprehensive review, we focus on our current understandings of the novel neuroprotective mechanisms activated by TH, and discuss the critical importance of developmental age germane to its clinical efficacy. We review emerging data on four cold stress hormones and three cold shock proteins that have generated new interest in hypothermia in the field of CNS injury, to create a framework for new frontiers in TH research. We make the case that further elucidation of novel cold responsive pathways might lead to major breakthroughs in the treatment of acute brain injury, chronic neurological diseases, and have broad potential implications for medicines of the distant future, including scenarios such as the prevention of adverse effects of long-duration spaceflight, among others. Finally, we introduce several new phrases that readily summarize the essence of the major concepts outlined by this review—namely, Ultramild Hypothermia, the “Responsivity of Cold Stress Pathways,” and “Hypothermia in a Syringe.”

Dynamic protein changes in the perihaemorrhagic zone of Surgically Treated Intracerebral Haemorrhage Patients

The secondary injury cascades exacerbating the initial brain injury following intracerebral haemorrhage (ICH) are incompletely understood. We used dual microdialysis (MD) catheters placed in the perihaemorrhagic zone (PHZ) and in seemingly normal cortex (SNX) at time of surgical ICH evacuation in ten patients (range 26–70 years). Routine interstitial MD markers (including glucose and the lactate/pyruvate ratio) were analysed and remaining microdialysate was analysed by two-dimensional gel electrophoresis (2-DE) and nano-liquid chromatography tandem mass spectrometry (nLC-MS/MS). Two time intervals were analysed; median 2–10 hours post-surgery (time A) and median 68–76 hours post-ICH onset (time B). Using 2-DE, we quantified 232 ± 31 different protein spots. Two proteins differed between the MD catheters at time A, and 12 proteins at time B (p < 0.05). Thirteen proteins were significantly altered between time A and time B in the SNX and seven proteins in the PHZ, respectively. Using nLC-MS/MS ca 800 proteins were identified out of which 76 were present in all samples. At time A one protein was upregulated and two downregulated, and at time B, seven proteins were upregulated, and four downregulated in the PHZ compared to the SNX. Microdialysis-based proteomics is feasible for study of secondary injury mechanisms and discovery of biomarkers after ICH.

Old age increases microglial senescence, exacerbates secondary neuroinflammation, and worsens neurological outcomes after acute traumatic brain injury in mice

After traumatic brain injury (TBI), individuals aged over 65 years show increased mortality and worse functional outcomes compared with younger persons. As neuroinflammation is a key pathobiological mechanism of secondary injury after TBI, we examined how aging affects post-traumatic microglial responses and functional outcomes. Young (3-month-old) and aged (18-month-old) male C57Bl/6 mice were subjected to moderate-level controlled cortical impact or sham surgery, and neurological function was evaluated. At 72 hours after injury, brain, blood, and spleen leukocyte counts were assessed ex vivo using flow cytometry. Aged mice demonstrated more severe deficits in forelimb grip strength, balance and motor coordination, spontaneous locomotor activity, and anxiety-like behavior. These animals also exhibited more robust microglial proliferation and significantly higher numbers of brain-infiltrating leukocytes. Microglia in aged mice showed impairments in phagocytic activity and higher production of interleukin-1β (IL-1β). Infiltrating myeloid cells in aged TBI mice also had deficits in phagocytosis but showed diminished proinflammatory cytokine production and greater reactive oxygen species production. Expression of several senescence markers (Bcl-2, p16ink4a, p21cip1a, lipofuscin, and H2AX [pS139]) was increased with age and/or TBI in both microglia and injured cortex. Although there was no difference in the number of circulating blood neutrophils as a function of age, young mice exhibited more pronounced TBI-induced splenomegaly and splenic myeloid cell expansion. Thus, worse post-traumatic behavioral outcomes in aged animals are associated with exaggerated microglial responses, increased leukocyte invasion, and upregulation of senescence markers.

The role of microglial inflammasome activation in pyroptotic cell death following penetrating traumatic brain injury

BackgroundTraumatic brain injury remains a significant cause of death and disability in the USA. Currently, there are no effective therapies to mitigate disability except for surgical interventions necessitating a need for continued research into uncovering novel therapeutic targets. In a recent study, we used a rodent model of penetrating traumatic brain injury known as penetrating ballistic-like brain injury (PBBI) to examine the role of innate immunity in post-traumatic secondary injury mechanisms. We previously reported that the inflammasome, a multiprotein complex composed of apoptosis-associated speck-like protein containing card and caspase-1, plays a role in secondary cell death mechanisms after PBBI, including inflammatory cell death (pyroptosis).MethodsIn the current study, we used flow cytometry analysis to evaluate activated microglia and CD11b-positive leukocytes after PBBI and assessed inflammasome activation and pyroptosis of specific cellular populations. Sprague-Dawley male rats underwent PBBI or sham-operated procedures and ipsilateral cortical regions processed for flow cytometry and cellular analysis. Flow cytometry results were compared using one-way ANOVA followed by Tukey’s multiple comparisons.ResultsAt 48 h following PBBI, there was an increase in activated microglia and infiltrating leukocytes compared to sham controls that were associated with increased caspase-1 activity. Using a florescent probe to identify caspase-1 activity and a fluorescent assay to determine cell viability, evidence for pyroptosis in CD11b+ cells was also determined. Finally, while post-traumatic treatment with an anti-ASC antibody had no effect on the number of activated microglia and infiltrating leukocytes, antibody treatment decreased caspase-1 activity in both resident microglia and infiltrating leukocytes and reduced pyroptotic CD11b+ cell death.ConclusionsThese results provide evidence for inflammasome activation in microglia and infiltrating leukocytes after penetrating traumatic brain injury and a role for pyroptotic cell death in the pathophysiology. In addition to inhibiting neuronal cell death, therapeutic treatments targeting inflammasome activation may also provide beneficial effects by reducing the potentially detrimental consequences of activated microglia and infiltrating CD11b+ leukocytes following penetrating traumatic brain injury.

Facile fabrication of flexible glutamate biosensor using direct writing of platinum nanoparticle-based nanocomposite ink

Glutamate excitotoxicity is a pathology in which excessive glutamate can cause neuronal damage and degeneration. It has also been linked to secondary injury mechanisms in traumatic spinal cord injury. Conventional bioanalytical techniques used to characterize glutamate levels in vivo, such as microdialysis, have low spatiotemporal resolution, which has impeded our understanding of this dynamic event. In this study, we present an amperometric biosensor fabricated using a simple direct ink writing technique for the purpose of in vivo glutamate monitoring. The biosensor is fabricated by immobilizing glutamate oxidase on nanocomposite electrodes made of platinum nanoparticles, multi-walled carbon nanotubes, and a conductive polymer on a flexible substrate. The sensor is designed to measure extracellular dynamics of glutamate and other potential biomarkers during a traumatic spinal cord injury event. Here we demonstrate good sensitivity and selectivity of these rapidly prototyped implantable biosensors that can be inserted into a spinal cord and measure extracellular glutamate concentration. We show that our biosensors exhibit good flexibility, linear range, repeatability, and stability that are suitable for future in vivo evaluation.

Circular RNA Expression Alteration and Bioinformatics Analysis in Rats After Traumatic Spinal Cord Injury.

Spinal cord injury (SCI) is mostly caused by trauma. As primary mechanical injury is unavoidable in SCI, a focus on the pathophysiology and underlying molecular mechanisms of SCI-induced secondary injury is necessary to develop promising treatments for SCI patients. Circular RNAs (circRNAs) are associated with various diseases. Nevertheless, studies to date have not yet determined the functional roles of circRNAs in traumatic SCI. We examined circRNA expression profiles in the contused spinal cords of rats using microarray and quantitative reverse transcription-PCR (qRT-PCR) then predict their potential roles in post-SCI pathophysiology with bioinformatics. We found a total of 1676 differentially expressed circRNAs (fold change ≥ 2.0; P < 0.05) in spinal cord 3 days after contusion using circRNA microarray; 1261 circRNAs were significantly downregulated, whereas the remaining 415 were significantly upregulated. Then, five selected circRNAs, namely, rno_circRNA_005342, rno_circRNA_015513, rno_circRNA_002948, rno_circRNA_006096, and rno_circRNA_013017 were all significantly downregulated in the SCI group after verification by qRT-PCR, demonstrating a similar expression pattern in both microarray and PCR data. The next section of the study was concerned with the prediction of circRNA/miRNA/mRNA interactions using bioinformatics analysis. In the final part of the study, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes analyses indicated carbohydrate metabolic process was one of the most significant enrichments and meaningful terms after GO analysis, and the top two signaling pathways affected by the circRNAs-miRNAs axes were the AMP-activated protein kinase signaling pathway and the peroxisome related pathway. In summary, this study showed an altered circRNA expression pattern that may be involved in physiological and pathological processes in rats after traumatic SCI, providing deep insights into numerous possibilities for SCI treatment targets by regulating circRNAs.