Early phrenic motor neuron loss and transient respiratory abnormalities following unilateral cervical spinal cord contusion

Contusion-type cervical spinal cord injury (SCI) is one of the most common forms of SCI observed in patients. In particular, injuries targeting the C3-C5 region affect the pool of phrenic motor neurons (PhMNs) that innervates the diaphragm, resulting in significant and often chronic respiratory dysfunction. Using a previously described rat model of unilateral midcervical C4 contusion with the Infinite Horizon Impactor, we have characterized the early time course of PhMN degeneration and consequent respiratory deficits following injury, as this knowledge is important for designing relevant treatment strategies targeting protection and plasticity of PhMN circuitry. PhMN loss (48% of the ipsilateral pool) occurred almost entirely during the first 24 h post-injury, resulting in persistent phrenic nerve axonal degeneration and denervation at the diaphragm neuromuscular junction (NMJ). Reduced diaphragm compound muscle action potential amplitudes following phrenic nerve stimulation were observed as early as the first day post-injury (30% of pre-injury maximum amplitude), with slow functional improvement over time that was associated with partial reinnervation at the diaphragm NMJ. Consistent with ipsilateral diaphragmatic compromise, the injury resulted in rapid, yet only transient, changes in overall ventilatory parameters measured via whole-body plethysmography, including increased respiratory rate, decreased tidal volume, and decreased peak inspiratory flow. Despite significant ipsilateral PhMN loss, the respiratory system has the capacity to quickly compensate for partially impaired hemidiaphragm function, suggesting that C4 hemicontusion in rats is a model of SCI that manifests subacute respiratory abnormalities. Collectively, these findings demonstrate significant and persistent diaphragm compromise in a clinically relevant model of midcervical contusion SCI; however, the therapeutic window for PhMN protection is restricted to early time points post-injury. On the contrary, preventing loss of innervation by PhMNs and/or inducing plasticity in spared PhMN axons at the diaphragm NMJ are relevant long-term targets

Dysregulation of Kv3.4 channels in dorsal root ganglia following spinal cord injury.

Spinal cord injury (SCI) patients develop chronic pain involving poorly understood central and peripheral mechanisms. Because dysregulation of the voltage-gated Kv3.4 channel has been implicated in the hyperexcitable state of dorsal root ganglion (DRG) neurons following direct injury of sensory nerves, we asked whether such a dysregulation also plays a role in SCI. Kv3.4 channels are expressed in DRG neurons, where they help regulate action potential (AP) repolarization in a manner that depends on the modulation of inactivation by protein kinase C (PKC)-dependent phosphorylation of the channel\u27s inactivation domain. Here, we report that, 2 weeks after cervical hemicontusion SCI, injured rats exhibit contralateral hypersensitivity to stimuli accompanied by accentuated repetitive spiking in putative DRG nociceptors. Also in these neurons at 1 week after laminectomy and SCI, Kv3.4 channel inactivation is impaired compared with naive nonsurgical controls. At 2-6 weeks after laminectomy, however, Kv3.4 channel inactivation returns to naive levels. Conversely, Kv3.4 currents at 2-6 weeks post-SCI are downregulated and remain slow-inactivating. Immunohistochemistry indicated that downregulation mainly resulted from decreased surface expression of the Kv3.4 channel, as whole-DRG-protein and single-cell mRNA transcript levels did not change. Furthermore, consistent with Kv3.4 channel dysregulation, PKC activation failed to shorten the AP duration of small-diameter DRG neurons. Finally, re-expressing synthetic Kv3.4 currents under dynamic clamp conditions dampened repetitive spiking in the neurons from SCI rats. These results suggest a novel peripheral mechanism of post-SCI pain sensitization implicating Kv3.4 channel dysregulation and potential Kv3.4-based therapeutic interventions

Overexpression of the astrocyte glutamate transporter GLT1 exacerbates phrenic motor neuron degeneration, diaphragm compromise, and forelimb motor dysfunction following cervical contusion spinal cord injury.

A major portion of spinal cord injury (SCI) cases affect midcervical levels, the location of the phrenic motor neuron (PhMN) pool that innervates the diaphragm. While initial trauma is uncontrollable, a valuable opportunity exists in the hours to days following SCI for preventing PhMN loss and consequent respiratory dysfunction that occurs during secondary degeneration. One of the primary causes of secondary injury is excitotoxic cell death due to dysregulation of extracellular glutamate homeostasis. GLT1, mainly expressed by astrocytes, is responsible for the vast majority of functional uptake of extracellular glutamate in the CNS, particularly in spinal cord. We found that, in bacterial artificial chromosome-GLT1-enhanced green fluorescent protein reporter mice following unilateral midcervical (C4) contusion SCI, numbers of GLT1-expressing astrocytes in ventral horn and total intraspinal GLT1 protein expression were reduced soon after injury and the decrease persisted for ≥6 weeks. We used intraspinal delivery of adeno-associated virus type 8 (AAV8)-Gfa2 vector to rat cervical spinal cord ventral horn for targeting focal astrocyte GLT1 overexpression in areas of PhMN loss. Intraspinal delivery of AAV8-Gfa2-GLT1 resulted in transduction primarily of GFAP(+) astrocytes that persisted for ≥6 weeks postinjury, as well as increased intraspinal GLT1 protein expression. Surprisingly, we found that astrocyte-targeted GLT1 overexpression increased lesion size, PhMN loss, phrenic nerve axonal degeneration, and diaphragm neuromuscular junction denervation, and resulted in reduced functional diaphragm innervation as assessed by phrenic nerve-diaphragm compound muscle action potential recordings. These results demonstrate that GLT1 overexpression via intraspinal AAV-Gfa2-GLT1 delivery exacerbates neuronal damage and increases respiratory impairment following cervical SCI

Children’s Moral Emotion Attribution in the Happy Victimizer Task: The Role of Response Format

Previous research in the happy victimizer tradition indicated that preschool and early elementary-school children attribute positive emotions to the violator of a moral norm, whereas older children attribute negative moral emotions. Cognitive and motivational processes have been suggested as underlying this developmental shift. The current research investigated whether making the happy victimizer task less cognitively demanding, by providing children with alternative response formats, would increase children’s attribution of moral emotions and moral motivation. In Study 1, 93 4- to 7-year-old British children responded to the happy victimizer questions either in a normal condition (where they spontaneously pointed with a finger), a wait condition (where they had to wait before giving their answers), or an arrow condition (where they had to point with a paper arrow). In Study 2, 40 Spanish 4-year-old children responded in the happy victimizer task either in a normal or a wait condition. In both studies, participants’ attribution of moral emotions and moral motivation was significantly higher in the conditions with alternative response formats (wait, arrow) than in the normal condition. The role of cognitive abilities for emotion attribution in the happy victimizer task is discussed

Antimicrobial resistance among migrants in Europe: a systematic review and meta-analysis

BACKGROUND: Rates of antimicrobial resistance (AMR) are rising globally and there is concern that increased migration is contributing to the burden of antibiotic resistance in Europe. However, the effect of migration on the burden of AMR in Europe has not yet been comprehensively examined. Therefore, we did a systematic review and meta-analysis to identify and synthesise data for AMR carriage or infection in migrants to Europe to examine differences in patterns of AMR across migrant groups and in different settings. METHODS: For this systematic review and meta-analysis, we searched MEDLINE, Embase, PubMed, and Scopus with no language restrictions from Jan 1, 2000, to Jan 18, 2017, for primary data from observational studies reporting antibacterial resistance in common bacterial pathogens among migrants to 21 European Union-15 and European Economic Area countries. To be eligible for inclusion, studies had to report data on carriage or infection with laboratory-confirmed antibiotic-resistant organisms in migrant populations. We extracted data from eligible studies and assessed quality using piloted, standardised forms. We did not examine drug resistance in tuberculosis and excluded articles solely reporting on this parameter. We also excluded articles in which migrant status was determined by ethnicity, country of birth of participants' parents, or was not defined, and articles in which data were not disaggregated by migrant status. Outcomes were carriage of or infection with antibiotic-resistant organisms. We used random-effects models to calculate the pooled prevalence of each outcome. The study protocol is registered with PROSPERO, number CRD42016043681. FINDINGS: We identified 2274 articles, of which 23 observational studies reporting on antibiotic resistance in 2319 migrants were included. The pooled prevalence of any AMR carriage or AMR infection in migrants was 25·4% (95% CI 19·1-31·8; I2 =98%), including meticillin-resistant Staphylococcus aureus (7·8%, 4·8-10·7; I2 =92%) and antibiotic-resistant Gram-negative bacteria (27·2%, 17·6-36·8; I2 =94%). The pooled prevalence of any AMR carriage or infection was higher in refugees and asylum seekers (33·0%, 18·3-47·6; I2 =98%) than in other migrant groups (6·6%, 1·8-11·3; I2 =92%). The pooled prevalence of antibiotic-resistant organisms was slightly higher in high-migrant community settings (33·1%, 11·1-55·1; I2 =96%) than in migrants in hospitals (24·3%, 16·1-32·6; I2 =98%). We did not find evidence of high rates of transmission of AMR from migrant to host populations. INTERPRETATION: Migrants are exposed to conditions favouring the emergence of drug resistance during transit and in host countries in Europe. Increased antibiotic resistance among refugees and asylum seekers and in high-migrant community settings (such as refugee camps and detention facilities) highlights the need for improved living conditions, access to health care, and initiatives to facilitate detection of and appropriate high-quality treatment for antibiotic-resistant infections during transit and in host countries. Protocols for the prevention and control of infection and for antibiotic surveillance need to be integrated in all aspects of health care, which should be accessible for all migrant groups, and should target determinants of AMR before, during, and after migration. FUNDING: UK National Institute for Health Research Imperial Biomedical Research Centre, Imperial College Healthcare Charity, the Wellcome Trust, and UK National Institute for Health Research Health Protection Research Unit in Healthcare-associated Infections and Antimictobial Resistance at Imperial College London

Surgical site infection after gastrointestinal surgery in high-income, middle-income, and low-income countries: a prospective, international, multicentre cohort study

Background: Surgical site infection (SSI) is one of the most common infections associated with health care, but its importance as a global health priority is not fully understood. We quantified the burden of SSI after gastrointestinal surgery in countries in all parts of the world. Methods: This international, prospective, multicentre cohort study included consecutive patients undergoing elective or emergency gastrointestinal resection within 2-week time periods at any health-care facility in any country. Countries with participating centres were stratified into high-income, middle-income, and low-income groups according to the UN's Human Development Index (HDI). Data variables from the GlobalSurg 1 study and other studies that have been found to affect the likelihood of SSI were entered into risk adjustment models. The primary outcome measure was the 30-day SSI incidence (defined by US Centers for Disease Control and Prevention criteria for superficial and deep incisional SSI). Relationships with explanatory variables were examined using Bayesian multilevel logistic regression models. This trial is registered with ClinicalTrials.gov, number NCT02662231. Findings: Between Jan 4, 2016, and July 31, 2016, 13 265 records were submitted for analysis. 12 539 patients from 343 hospitals in 66 countries were included. 7339 (58·5%) patient were from high-HDI countries (193 hospitals in 30 countries), 3918 (31·2%) patients were from middle-HDI countries (82 hospitals in 18 countries), and 1282 (10·2%) patients were from low-HDI countries (68 hospitals in 18 countries). In total, 1538 (12·3%) patients had SSI within 30 days of surgery. The incidence of SSI varied between countries with high (691 [9·4%] of 7339 patients), middle (549 [14·0%] of 3918 patients), and low (298 [23·2%] of 1282) HDI (p < 0·001). The highest SSI incidence in each HDI group was after dirty surgery (102 [17·8%] of 574 patients in high-HDI countries; 74 [31·4%] of 236 patients in middle-HDI countries; 72 [39·8%] of 181 patients in low-HDI countries). Following risk factor adjustment, patients in low-HDI countries were at greatest risk of SSI (adjusted odds ratio 1·60, 95% credible interval 1·05–2·37; p=0·030). 132 (21·6%) of 610 patients with an SSI and a microbiology culture result had an infection that was resistant to the prophylactic antibiotic used. Resistant infections were detected in 49 (16·6%) of 295 patients in high-HDI countries, in 37 (19·8%) of 187 patients in middle-HDI countries, and in 46 (35·9%) of 128 patients in low-HDI countries (p < 0·001). Interpretation: Countries with a low HDI carry a disproportionately greater burden of SSI than countries with a middle or high HDI and might have higher rates of antibiotic resistance. In view of WHO recommendations on SSI prevention that highlight the absence of high-quality interventional research, urgent, pragmatic, randomised trials based in LMICs are needed to assess measures aiming to reduce this preventable complication

Unilateral contusion SCI induced a loss of ventral horn motor neurons at the injury epicenter.

<p>Harvested tissue was stained with Cresyl violet and Eriochrome cyanine. At the level of the laminectomy, uninjured control animals (A) exhibited large motor neurons in the ventral horn (arrowheads). At six weeks post-injury, animals receiving unilateral C5 (B) or C6 (C) contusion SCI showed a loss of these motor neurons at the injury epicenter but not 1.0 mm caudal to the injury (D). The spread of ventral horn motor neuron loss was approximately 1.0 mm rostrally and 1.0 mm caudally (2.0 mm total) from the epicenter (E). Immunohistochemical analyses of the injury models were performed in laminae I–II and lamina III of the cervical spinal cord dorsal horn (F–G). Lam = laminectomy.</p

Enhanced cell proliferation, including proliferation of astrocytes, was evident after cervical contusion SCI.

<p>In the superficial laminae of the injured ipsilateral dorsal horn, we observed cells co-expressing GFAP and Ki67 (A), representing activated and proliferating astrocytes. In laminectomy control animals, little to no cell proliferation was evident (B). However, the number of Ki67-positive cells was significantly increased at two weeks (C) and six weeks (D) after injury on the ipsilateral side both at the level of and caudal to the injury (E). Additionally, at two weeks, there was a significant increase in proliferating cells contralaterally at the injury site (E). At both two and six weeks, a significant percentage of Ki67-positive cells were also either GFAP-positive or CD11b-positive (F). Compared to laminectomy (G), the intensity of CD11b expression in the superficial dorsal horn was greater in animals with C6 SCI at both two (H–I) and six (I) weeks after injury. This increase was significant at the injury site both ipsilaterally and contralaterally, as well as caudal to the injury on the contralateral side (I). * = p<0.05; ** = p<0.01; **** = p<0.0001.</p

Chronic neuronal activation in the dorsal horn resulted from cervical contusion SCI.

<p>ΔfosB staining was used to measure the extent of persistent neuronal activation in the spinal cord. Following injury, ΔfosB-positive nuclei (arrowheads) were evident in laminae I–II (A–C). At all regions, laminectomy control animals (A) showed little-to-no ΔfosB expression (D). Two weeks after C6 injury (B), a significant increase in numbers of ΔfosB expressing cells in the superficial laminae of the ipsilateral dorsal horn was observed at the injury epicenter and caudal to the injury (D). ΔfosB levels were further increased at all regions in animals sacrificed six weeks post-injury (C, D). * = p<0.05; ** = p<0.01; *** = p 0.001; **** = p<0.0001.</p

Astrocyte GLT1 promoter activity was reduced in injured BAC-GLT1-eGFP transgenic reporter mice.

<p>Following cervical contusion SCI, GLT1 expression/promoter activity, represented by eGFP-positive cells, was decreased in the superficial dorsal horn. In laminectomy animals, there were low levels of GFAP, although many of the cells that were GFAP-positive also expressed GLT1 (A–B). Following injury, however, an increase in GFAP was observed, but fewer of these activated astrocytes co-expressed GLT1 (C). Laminectomy control animals (D) had greater numbers of eGFP-positive cells compared to animals that received contusion injury two days, two weeks, or six weeks (E) prior to analysis. This decrease in GLT1 expression was observed at the injury epicenter in both the ipsilateral (F) and contralateral (G) superficial laminae as well as caudal to the injury on the ipsilateral side (H). No change in the number of eGFP-positive cells was found caudal to the injury on the contralateral side (I). * = p<0.05; ** = p<0.01; *** = p 0.001.</p