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Central anticholinergic syndrome (CAS) presents with central and/or peripheral neurological symptoms after exposure to anticholinergic medication. Pain management is a challenge for patients who have CAS and concurrent cancer-related pain. We present a patient who became lethargic and unresponsive after receiving butorphanol, with persistent symptoms despite administration of an opioid antagonist. This case report is the first to suggest a causal relationship between opioids and CAS without the presence of confounding anticholinergic medications. CAS should be considered for patients who develop neurological symptoms after opioid exposure and have an incomplete response to an opioid antagonist. Key words: central anticholinergic syndrome, butorphanol, opioids, cancer-related pain, anticholinergic toxicity, case report Abbreviations: CAS - Central Anticholinergic Syndrome; IV – intravenous; IM – intramuscular; ED - Emergency Department; WHO - World Health Organization; MRI - Magnetic Resonance Imaging; MED - Morphine Equivalent Dosing; ICU - Intensive Care Unit Citation: Choi A, Bui T, Pan EY, Ozcan M. Is central anticholinergic syndrome linked to opioid use for cervical cancer pain? Anaesth. pain intensive care 2022;26(6):826−830; DOI: 10.35975/apic.v26i6.2068

The gingiva is the target site for some topical drugs, but the permeability of human gingiva has not been systematically evaluated. Pigs are a common animal model for in vitro membrane transport studies. The objectives of this study were to: (a) determine the permeability coefficients of freshly excised human gingiva using model permeants, (b) compare the permeability coefficients of fresh human gingiva with those of fresh porcine gingiva, (c) evaluate the effect of freezing duration on the permeability of porcine gingiva, and (d) compare the permeability coefficients of fresh and cadaver (frozen) human gingiva. A goal was to examine the feasibility of using porcine gingiva as a surrogate for human gingiva. The potential of using frozen tissues in permeability studies of gingiva was also examined. Fresh and frozen porcine gingiva, fresh human gingiva, and frozen cadaver human gingiva were compared in the transport study with model polar and lipophilic permeants. The fresh porcine and human tissues showed similarities in the “permeability coefficient vs. octanol–water distribution coefficient” relationship. The porcine gingiva had a lower permeability than that of the human, with a moderate correlation between the permeability of the fresh porcine and fresh human tissues. The permeability of the porcine tissues for the model polar permeants increased significantly after the tissues were frozen in storage. Moreover, the frozen human cadaver tissue could not be utilized due to the high and indiscriminating permeability of the tissue for the permeants and large tissue sample-to-sample variabilities.

Background: Shivering is a common adverse effect of achieving and maintaining normothermia in neurocritical care patients. We compared the burden of shivering and shivering-related interventions between a novel trans-nasal temperature modulating device(tnTMD) and surface cooling temperature modulating devices(sTMDs) during the first 24 hours of targeted normothermia in mechanically ventilated febrile neurocritical care patients. Methods: This is a case: control study controlling for factors that impact shiver burden: age, sex, body surface area. All patients underwent transnasal cooling (COOLSTAT, KeyTech, Inc) as part of an ongoing multicenter clinical trial(NCT03360656). Patients undergoing treatment with sTMDs were selected from consecutively treated patients during the same time-period. Data collected included: core body temperature (every 2 hours), bedside shivering assessment scale(BSAS) score (every 2 hours), and administration of anti-shivering medication for BSAS>1.Time to normothermia(37.5 C(C*hr) were compared between groups using student’s t-test for mean differences. Proportion of patients requiring interventions as well as number of interventions per patient were compared using Chi-Square test. Significance was determined based on a P value < 0.05. Results: There were 10 tnTMD patients and 30 sTMD patients included in the analysis (mean age: 62+/-4, 30% women, BSA = 1.97+/-0.25). There were no differences between groups in temperature at cooling initiation (tnTMD: 38.5+/-0.2 C vs sTMD: 38.7+/-0.5 C, P=0.3), time to 37.5 (tnTMD:: -0.4 +/- 1.13 C*hr vs. sTMD median (IQR): -0.57 +/- 0.58 C*hr, P=0.67). The number of tnTMD patients who received pharmacologic shivering interventions was lower than the controls (20%vs.67%,p=0.01). tnTMD patients also had fewer shivering interventions per patient (0 (range: 0-3)vs.4(range: 0 – 23 ), p

SF1: TGF-β1 promotes Twist1-induced epithelial dissemination without loss of adherens junction protein expression; SF2: Prkd1 inhibitors kb-NB142-70 and Gö-6976 block Twist1- induced dissemination dose-dependently; SF3: Dissemination defects in Twist1-On organoids resulting from Prkd1 inactivation; SF4: Twist1-induced epithelial cells express Prkd1 and maintain β-catenin expression; SF5: Limited toxicity observed in response to drug treatment of Twist1-induced organoids; SF6: Supporting data for Figure 6: SF7: Molecular model for Twist1-induced epithelial dissemination; ST1: List of inhibitors tested in Figure 1; ST2: List of antibodies used in the study; ST3: Phosphoantibody microarray data. Supplementary movie legends.

Dissemination is an essential early step in metastasis but its molecular basis remains incompletely understood. To define the essential targetable effectors of this process, we developed a 3D mammary epithelial culture model, in which dissemination is induced by overexpression of the transcription factor Twist1. Transcriptomic analysis and ChIP-PCR together demonstrated that protein kinase D1 (Prkd1) is a direct transcriptional target of Twist1 and is not expressed in the normal mammary epithelium. Pharmacologic and genetic inhibition of Prkd1 in the Twist1-induced dissemination model demonstrated that Prkd1 was required for cells to initiate extracellular matrix (ECM)–directed protrusions, release from the epithelium, and migrate through the ECM. Antibody-based protein profiling revealed that Prkd1 induced broad phosphorylation changes, including an inactivating phosphorylation of β-catenin and two microtubule depolymerizing phosphorylations of Tau, potentially explaining the release of cell–cell contacts and persistent activation of Prkd1. In patients with breast cancer, TWIST1 and PRKD1 expression correlated with metastatic recurrence, particularly in basal breast cancer. Prkd1 knockdown was sufficient to block dissemination of both murine and human mammary tumor organoids. Finally, Prkd1 knockdown in vivo blocked primary tumor invasion and distant metastasis in a mouse model of basal breast cancer. Collectively, these data identify Prkd1 as a novel and targetable signaling node downstream of Twist1 that is required for epithelial invasion and dissemination.Significance:Twist1 is a known regulator of metastatic cell behaviors but not directly targetable. This study provides a molecular explanation for how Twist1-induced dissemination works and demonstrates that it can be targeted.

This file contains the full blot images of all Western blots.

Early inhibition of Prkd1 blocks epithelial invasion into ECM and loss of cell-cell adhesion.

Late inhibition of Prkd1 after onset of dissemination prevents disseminated cell migration and prevents de novo dissemination.

BACKGROUND: After subarachnoid hemorrhage (SAH), early brain injury (EBI) and delayed cerebral ischemia (DCI) lead to poor outcomes. Discovery of biomarkers indicative of disease severity and predictive of DCI is important. We tested whether leucine-rich alpha-2-glycoprotein 1 (LRG1) is a marker of severity, DCI, and functional outcomes after SAH. METHODS: We performed untargeted proteomics using mass spectrometry in plasma samples collected at < 48 h of SAH in two independent discovery cohorts (n = 27 and n = 45) and identified LRG1 as a biomarker for DCI. To validate our findings, we used enzyme-linked immunosorbent assay and confirmed this finding in an internal validation cohort of plasma from 72 study participants with SAH (22 DCI and 50 non-DCI). Further, we investigated the relationship between LRG1 and markers of EBI, DCI, and poor functional outcomes (quantified by the modified Rankin Scale). We also measured cerebrospinal fluid (CSF) levels of LRG1 and investigated its relationship to EBI, DCI, and clinical outcomes. RESULTS: Untargeted proteomics revealed higher plasma LRG1 levels across EBI severity and DCI in both discovery cohorts. In the validation cohort, the levels of LRG1 were higher in the DCI group compared with the non-DCI group (mean (SD): 95 [44] vs. 72 [38] pg/ml, p < 0.05, Student’s t-test) and in study participants who proceeded to have poor functional outcomes (84 [39.3] vs. 72 [43.2] pg/ml, p < 0.05). Elevated plasma LRG1 levels were also associated with markers of EBI. However, CSF levels of LRG1 were not associated with EBI severity or the occurrence of DCI. CONCLUSIONS: Plasma LRG1 is a biomarker for EBI, DCI, and functional outcomes after SAH. Further studies to elucidate the role of LRG1 in the pathophysiology of SAH are needed.

Aneurysmal subarachnoid hemorrhage (aSAH) causes a robust inflammatory response which leads worse brain injury and poor outcomes. We investigated if stimulation of nicotinic acetylcholine α7 receptors (α7-AChR) (receptors shown to have anti-inflammatory effects) would reduce inflammation and improve outcomes. To investigate the level of peripheral inflammation after aSAH, inflammatory markers were measured in plasma samples collected in a cohort of aSAH patients. To study the effect of α7-AChR stimulation, SAH was induced in adult mice which were then treated with a α7-AChR agonist, galantamine, or vehicle. A battery of motor and cognitive tests were performed 24 h after subarachnoid hemorrhage. Mice were euthanized and tissue collected for analysis of markers of inflammation or activation of α7-AChR-mediated transduction cascades. A separate cohort of mice was allowed to survive for 28 days to assess long-term neurological deficits and histological outcome. Microglia cell culture subjected to hemoglobin toxicity was used to assess the effects of α7-AChR agonism. Analysis of eighty-two patient plasma samples confirmed enhanced systemic inflammation after aSAH. α7-AChR agonism reduced neuroinflammation at 24 h after SAH in male and female mice, which was associated with improved outcomes. This coincided with JAK2/STAT3 and IRAK-M activity modulations and a robust improvement in neurological/cognitive status that was effectively reversed by interfering with various components of these signaling pathways. Pharmacologic inhibition partially reversed the α7-AChR agonist’s benefits, supporting α7-AChR as a target of the agonist’s therapeutic effect. The cell culture experiment showed that α7-AChR agonism is directly beneficial to microglia. Our results demonstrate that activation of α7-AChR represents an attractive target for treatment of SAH. Our findings suggest that α7-AChR agonists, and specifically galantamine, might provide therapeutic benefit to aSAH patients.

ObjectiveTo determine whether machine learning (ML) algorithms can improve the prediction of delayed cerebral ischemia (DCI) and functional outcomes after subarachnoid hemorrhage (SAH).MethodsML models and standard models (SMs) were trained to predict DCI and functional outcomes with data collected within 3 days of admission. Functional outcomes at discharge and at 3 months were quantified using the modified Rankin Scale (mRS) for neurologic disability (dichotomized as good [mRS ≤ 3] vs poor [mRS ≥ 4] outcomes). Concurrently, clinicians prospectively prognosticated 3-month outcomes of patients. The performance of ML, SMs, and clinicians were retrospectively compared.ResultsDCI status, discharge, and 3-month outcomes were available for 399, 393, and 240 participants, respectively. Prospective clinician (an attending, a fellow, and a nurse) prognostication of 3-month outcomes was available for 90 participants. ML models yielded predictions with the following area under the receiver operating characteristic curve (AUC) scores: 0.75 ± 0.07 (95% confidence interval [CI] 0.64–0.84) for DCI, 0.85 ± 0.05 (95% CI 0.75–0.92) for discharge outcome, and 0.89 ± 0.03 (95% CI 0.81–0.94) for 3-month outcome. ML outperformed SMs, improving AUC by 0.20 (95% CI −0.02 to 0.4) for DCI, by 0.07 ± 0.03 (95% CI −0.0018 to 0.14) for discharge outcomes, and by 0.14 (95% CI 0.03–0.24) for 3-month outcomes and matched physician's performance in predicting 3-month outcomes.ConclusionML models significantly outperform SMs in predicting DCI and functional outcomes and has the potential to improve SAH management.

Increased soluble endoglin (sENG) has been observed in human brain arteriovenous malformations (bAVMs). In addition, the overexpression of sENG in concurrence with vascular endothelial growth factor (VEGF)-A has been shown to induce dysplastic vessel formation in mouse brains. However, the underlying mechanism of sENG-induced vascular malformations is not clear. The evidence suggests the role of sENG as a pro-inflammatory modulator, and increased microglial accumulation and inflammation have been observed in bAVMs. Therefore, we hypothesized that microglia mediate sENG-induced inflammation and endothelial cell (EC) dysfunction in bAVMs. In this study, we confirmed that the presence of sENG along with VEGF-A overexpression induced dysplastic vessel formation. Remarkably, we observed increased microglial activation around dysplastic vessels with the expression of NLRP3, an inflammasome marker. We found that sENG increased the gene expression of VEGF-A, pro-inflammatory cytokines/inflammasome mediators (TNF-α, IL-6, NLRP3, ASC, Caspase-1, and IL-1β), and proteolytic enzyme (MMP-9) in BV2 microglia. The conditioned media from sENG-treated BV2 (BV2-sENG-CM) significantly increased levels of angiogenic factors (Notch-1 and TGFβ) and pERK1/2 in ECs but it decreased the level of IL-17RD, an anti-angiogenic mediator. Finally, the BV2-sENG-CM significantly increased EC migration and tube formation. Together, our study demonstrates that sENG provokes microglia to express angiogenic/inflammatory molecules which may be involved in EC dysfunction. Our study corroborates the contribution of microglia to the pathology of sENG-associated vascular malformations.

We prospectively studied SARS-CoV-2 transmission at schools in an era of Variants of Concern (VoCs), offering all close contacts serial viral asymptomatic testing up to 14 days. Of 229 school close contacts, 3 tested positive (1.3%), of which 2 were detected through asymptomatic testing. Most secondary transmission (90%) occurred in households. Routine asymptomatic testing of close contacts should be examined in the context of local testing rates, preventive measures, programmatic costs, and health impacts of asymptomatic transmission.

COVID-19 is an ongoing pandemic with a devastating impact on public health. Acute neurological symptoms have been reported after a COVID-19 diagnosis, however, the long-term neurological symptoms including pain is not well established. Using a prospective registry of hospitalized COVID-19 patients, we assessed pain and neurological function (including functional, cognitive and psychiatric assessments) of several hospitalized patients at 3 months. Our main finding is that 60% of the patients report pain symptoms. 71% of the patients still experienced neurological symptoms at 3 months and the most common symptoms being fatigue (42%) and PTSD (25%). Cognitive symptoms were found in 12%. Our preliminary findings suggests the importance of investigating long-term outcomes and rationalizes the need for further studies investigating the neurologic outcomes and symptoms of pain after COVID-19.

Objective Although COVID-19 is a respiratory disease, all organs can be affected including the brain. To date, specific investigations of brain injury markers (BIM) and endothelial injury markers (EIM) have been limited. Additionally, a male bias in disease severity and mortality after COVID-19 is evident globally. Sex differences in the immune response to COVID-19 may mediate this disparity. We investigated BIM, EIM and inflammatory cytokine/chemokine (CC) levels after COVID-19 and in across sexes. Methods Plasma samples from 57 subjects at Results Three BIMs: MAP2, NSE and S100B, two EIMs: sICAM1 and sVCAM1 and seven CCs: GRO IL10, sCD40L, IP10, IL1Ra, MCP1 and TNFα were significantly (p p Conclusion The acute elevation of BIMs, CCs, and EIMs and the robust associations among them at COVID-19 hospitalization are suggestive of brain and endothelial injury. Higher BIM and inflammatory markers in men additionally suggest that men are more susceptible to the risk compared to women.

Summary Organoid model systems recapitulate key features of mammalian tissues and enable high throughput experiments. However, the impact of these experiments may be limited by manual, non-standardized, static or qualitative phenotypic analysis. OrgDyn is an open-source and modular pipeline to quantify organoid shape dynamics using a combination of feature- and model-based approaches on time series of 2D organoid contour images. Our pipeline consists of (i) geometrical and signal processing feature extraction, (ii) dimensionality reduction to differentiate dynamical paths, (iii) time series clustering to identify coherent groups of organoids and (iv) dynamical modeling using point distribution models to explain temporal shape variation. OrgDyn can characterize, cluster and model differences among unique dynamical paths that define diverse final shapes, thus enabling quantitative analysis of the molecular basis of tissue development and disease. Availability and Implementation https://github.com/zakih/organoidDynamics (BSD 3-Clause License). Supplementary information Supplementary data are available at Bioinformatics online.

Although precision medicine has revolutionized the treatment for other solid tumors, comprehensive genome profiling of hepatocellular carcinoma (HCC) has demonstrated that there are few oncogenic mutational drivers. HCC arises nearly universally in the context of co-morbid hepatitis, driven by hepatitis C virus or hepatitis B virus (HBV). Previous work has highlighted the effects of HBV protein X (HBx) on signaling and proliferation. However, it remains unclear how these signaling changes interact with the stress imposed by HBV replication. We hypothesize that HBV replication and maintenance of HBV proteins like HBx creates specific vulnerabilities that can be uncovered and exploited. We performed whole-genome clustered regularly interspaced short palindromic repeat interference (CRISPRi) screening in two engineered, patient-derived HBV+ cell models, Hep3B and SNU-368, with cellular proliferation as the readout. A conditional induction of HBx was used to overexpress HBx in each cell model for the whole genome CRISPRi screen. Genes where HBx expression was significantly associated with a more negative dependency score were classified as HBV differential dependencies. 37 genes were identified as shared HBV-induced dependencies across both cell lines. Of these, 16 were also differentially expressed in HCC specimens with ongoing HBV replication vs. non-HBV expressing tumors collected in The Cancer Genome Atlas (TCGA), suggesting in vivo selective pressure to alter the expression of these genes. We focused on Zinc Fingers and Homeoboxes 2 (ZHX2) and methionine adenosyltransferase 2A (MAT2A). ZHX2 was identified as an HCC tumor suppressor, and data from TCGA showed differential gene expression of ZHX2 in HBV+HCC. Validation of ZHX2 sgRNA knockdown confirmed that ZHX2 knockdown preferentially reduced cell proliferation in the presence of HBx, consistent with an HBV-induced dependency. ZHX2 also can activate the HIF pathway; we identified alterations of HIF targets with ZHX2 knockdown and HBx expression. Similarly, MAT2A knockdown in HBV+HCC cell lines showed decreased cell proliferation with HBx expression. MAT2A is an emerging therapeutic target, with sensitivity to MAT2A inhibitors conferred by deletions in methylthioadenosine phosphorylase (MTAP), seen in 3% of HCC. Ongoing investigations include alteration of splicing and cell cycle changes with depletion of MAT2A in the presence of HBx. With small molecule inhibitors of MAT2A in clinical development, this finding may be therapeutically actionable. Thus, using functional genomics to interrogate HBV-associated gene dependencies has illuminated the biology of HCC and identified candidate therapeutic targets for pre-clinical validation. Citation Format: Rigney E Turnham, Huat Chye Lim, Lucille Ferret, Katherine Lo, LeeAnn Wang, Alex Choi, John D Gordan. Uncovering hepatitis B virus-induced signaling changes in hepatocellular carcinoma [abstract]. In: Proceedings of the AACR Special Conference: Advances in the Pathogenesis and Molecular Therapies of Liver Cancer; 2022 May 5-8; Boston, MA. Philadelphia (PA): AACR; Clin Cancer Res 2022;28(17_Suppl):Abstract nr PO023.

Background: In a disease that has only existed for 18 months, it is difficult to be fully informed of the long-term sequelae of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection. Evidence is growing that most organ systems can be affected by the virus, causing severe disabilities in survivors. The extent of the aftermath will declare itself over the next 5–10 years, but it is likely to be substantial with profound socio-economic impact on society.Methods: This is an international multi-center, prospective long-term follow-up study of patients who developed severe coronavirus disease-2019 (COVID-19) and were admitted to Intensive Care Units (ICUs). The study will be conducted at international tertiary hospitals. Patients will be monitored from time of ICU discharge up to 24 months. Information will be collected on demographics, co-existing illnesses before ICU admission, severity of illness during ICU admission and post-ICU quality of life as well as organ dysfunction and recovery. Statistical analysis will consist of patient trajectories over time for the key variables of quality of life and organ function. Using latent class analysis, we will determine if there are distinct patterns of patients in terms of recovery. Multivariable regression analyses will be used to examine associations between baseline characteristics and severity variables upon admission and discharge in the ICU, and how these impact outcomes at all follow-up time points up to 2 years.Ethics and Dissemination: The core study team and local principal investigators will ensure that the study adheres to all relevant national and local regulations, and that the necessary approvals are in place before a site may enroll patients.Clinical Trial Registration:anzctr.org.au: ACTRN12620000799954.

ObjectiveTo investigate brain injury markers (BIM), endothelial injury markers (EIM) and cytokine/chemokine (CC) markers of systemic inflammation in coronavirus disease 2019 (COVID-19) and across sex.MethodsPlasma samples from 57 subjects at ResultsThree BIMs: MAP2, NSE and S100B, two EIMs: sICAM1 and sVCAM1 and seven CCs: GRO IL10, sCD40L, IP10, IL1Ra, MCP1 and TNFα were significantly (pConclusionThe acute elevation of BIMs, CCs, and EIMs and the robust associations among them at COVID-19 hospitalization suggest that brain injury is mediated by endotheliopathy and inflammation. Higher BIM and inflammatory markers in men additionally suggest that men are more susceptible to the risk compared to women.

Aneurysmal subarachnoid hemorrhage (aSAH) causes a robust inflammatory response which leads worse brain injury and poor outcomes. We investigated if stimulation of nicotinic acetylcholine α(7) receptors (α(7)-AChR) (receptors shown to have anti-inflammatory effects) would reduce inflammation and improve outcomes. To investigate the level of peripheral inflammation after aSAH, inflammatory markers were measured in plasma samples collected in a cohort of aSAH patients. To study the effect of α(7)-AChR stimulation, SAH was induced in adult mice which were then treated with a α(7)-AChR agonist, galantamine, or vehicle. A battery of motor and cognitive tests were performed 24 h after subarachnoid hemorrhage. Mice were euthanized and tissue collected for analysis of markers of inflammation or activation of α(7)-AChR-mediated transduction cascades. A separate cohort of mice was allowed to survive for 28 days to assess long-term neurological deficits and histological outcome. Microglia cell culture subjected to hemoglobin toxicity was used to assess the effects of α(7)-AChR agonism. Analysis of eighty-two patient plasma samples confirmed enhanced systemic inflammation after aSAH. α(7)-AChR agonism reduced neuroinflammation at 24 h after SAH in male and female mice, which was associated with improved outcomes. This coincided with JAK2/STAT3 and IRAK-M activity modulations and a robust improvement in neurological/cognitive status that was effectively reversed by interfering with various components of these signaling pathways. Pharmacologic inhibition partially reversed the α(7)-AChR agonist’s benefits, supporting α(7)-AChR as a target of the agonist’s therapeutic effect. The cell culture experiment showed that α(7)-AChR agonism is directly beneficial to microglia. Our results demonstrate that activation of α(7)-AChR represents an attractive target for treatment of SAH. Our findings suggest that α(7)-AChR agonists, and specifically galantamine, might provide therapeutic benefit to aSAH patients. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13311-021-01052-3.

COVID-19 is an ongoing pandemic with a devastating impact on public health. Acute neurological symptoms have been reported after a COVID-19 diagnosis, however there is no data available on the long-term neurological symptoms. Using a prospective registry of hospitalized COVID-19 patients, we assessed the neurological assessments (including functional, cognitive and psychiatric assessments) of several hospitalized patients at 3 months. Our main finding is that 71% of the patients still experienced neurological symptoms at 3 months and the most common symptoms being fatigue (42%) and PTSD (29%). 64% of the patients report pain symptoms we well. Cognitive symptoms were found in 12%. Our preliminary findings suggests the importance of investigating long-term and rationalizes the need for further studies investigating the neurologic outcomes after COVID-19.

Background Electronic nicotine delivery systems (ENDSs), including electronic cigarettes (e-cigarettes), are rapidly gaining popularity. The aim of this study was to use poison centre data to describe epidemiological trends in ENDS-related exposures. Methods We conducted an observational case series study using records containing both coded fields and free-text narratives from the British Columbia Drug and Poison Information Centre for all calls involving exposure to ENDS received from 2012 to 2017. We described trends in exposures and exposed people, as well as clinical effects. Results A total of 243 calls were recorded for 186 unique exposures to ENDS devices, e-juice, e-cigarette cartridges and other associated paraphernalia over the study period. Calls related to ENDS exposures increased nearly sixfold between 2013 and 2014 and did not decline subsequently. Exposures were most frequently documented in children aged 4 years or less (81 [43.5%]), with 58 (31.0%) in 1- and 2-year-olds. Seventy-two exposures (89%) in children aged 4 years or less were due to accidental ingestion, whereas adults aged 25 years or more called the poison centre following ENDS malfunctions (7 [23%], spills (4 [13%]) and exposure to e-juice mistaken for other substances (4 [13%]). Of the 186 exposed people, 87 (46.8%) reported symptoms. Interpretation British Columbia experienced a sixfold increase in ENDS-related calls to the provincial poison centre between 2012 and 2017, driven by ingestions in young children. Regulatory approaches aimed at minimizing children's access to ENDS, clear labelling of nicotine concentration, and packaging that reduces the likelihood of spills, product confusion and malfunction should be considered.

The gut microbiome is fundamental in neurogenesis processes. Alterations in microbial constituents promote inflammation and immunosuppression. Recently, in immune-oncology, specific microbial taxa have been described to enhance the effects of therapeutic modalities. However, the effects of microbial dysbiosis on glioma are still unknown. The aim of this study was to explore the effects of glioma development and Temozolomide (TMZ) on fecal microbiome in mice and humans. C57BL/6 mice were implanted with GL261/Sham and given TMZ/Saline. Fecal samples were collected longitudinally and analyzed by 16S rRNA sequencing. Fecal samples were collected from healthy controls as well as glioma patients at diagnosis, before and after chemoradiation. Compared to healthy controls, mice and glioma patients demonstrated significant differences in beta diversity, Firmicutes/Bacteroides (F/B) ratio, and increase of Verrucomicrobia phylum and Akkermansia genus. These changes were not observed following TMZ in mice. TMZ treatment in the non-tumor bearing mouse-model diminished the F/B ratio, increase Muribaculaceae family and decrease Ruminococcaceae family. Nevertheless, there were no changes in Verrucomicrobia/Akkermansia. Glioma development leads to gut dysbiosis in a mouse-model, which was not observed in the setting of TMZ. These findings seem translational to humans and warrant further study.

Subarachnoid hemorrhage (SAH) results in severe neuronal dysfunction and degeneration. Since the nicotinic acetylcholine α(7) receptors (α(7)-AChR) are involved in neuronal function and survival, we investigated if stimulation of α(7)-AChR would promote neuronal survival and improve behavioral outcome following SAH in mice. Male mice subjected to SAH were treated with either galantamine (α(7)-AChR agonist) or vehicle. Neurobehavioral testing was performed 24 hours after SAH, and mice were euthanized for analysis of neuronal cell death or a cell survival (PI3K/Akt) signaling pathway. Neuron cell cultures were subjected to hemoglobin toxicity to assess the direct effects of α(7)-AChR agonism independent of other cells. Treatment with the α(7)-AChR agonist promoted neuronal survival and improved functional outcomes 24 hours post-SAH. The improved outcomes corresponded with increased PI3K/Akt activity. Antagonism of α(7)-AChR or PI3K effectively reversed galantamine’s beneficial effects. Tissue from α(7)-AChR knockout mice confirmed α(7)-AChR’s role in neuronal survival after SAH. Data from the neuronal cell culture experiment supported a direct effect of α(7)-AChR agonism in promoting cell survival. Our findings indicate that α(7)-AChR is a therapeutic target following SAH which can promote neuronal survival, thereby improving neurobehavioral outcome. Thus, the clinically relevant α(7)-AChR agonist, galantamine, might be a potential candidate for human use to improve outcome after SAH.

Corticosteroids (CSs) are used frequently in the neurocritical care unit mainly for their antiinflammatory and immunosuppressive effects. Despite their broad use, limited evidence exists for their efficacy in diseases confronted in the neurocritical care setting. There are considerable safety concerns associated with administering these drugs and should be limited to specific conditions in which their benefits outweigh the risks. The application of CSs in neurologic diseases, range from traumatic head and spinal cord injuries to central nervous system infections. Based on animal studies, it is speculated that the benefit of CSs therapy in brain and spinal cord, include neuroprotection from free radicals, specifically when given at a higher supraphysiologic doses. Regardless of these advantages and promising results in animal studies, clinical trials have failed to show a significant benefit of CSs administration on neurologic outcomes or mortality in patients with head and acute spinal injuries. This article reviews various chemical structures between natural and synthetic steroids, discuss its pharmacokinetic and pharmacodynamic profiles, and describe their use in clinical practice.

Background Plasma-based resuscitation improves outcomes in trauma patients with hemorrhagic shock, while large-animal and limited clinical data suggest that it also improves outcomes and is neuroprotective in the setting of combined hemorrhage and traumatic brain injury. However, the choice of initial resuscitation fluid, including the role of plasma, is unclear for patients after isolated traumatic brain injury. Methods We reviewed adult trauma patients admitted from January 2011 to July 2015 with isolated traumatic brain injury. “Early plasma” was defined as transfusion of plasma within 4 hours. Purposeful multiple logistic regression modeling was performed to analyze the relationship of early plasma and inhospital survival. After testing for interaction, subgroup analysis was performed based on the pattern of brain injury on initial head computed tomography: epidural hematoma, intraparenchymal contusion, subarachnoid hemorrhage, subdural hematoma, or multifocal intracranial hemorrhage. Results Of the 633 isolated traumatic brain injury patients included, 178 (28%) who received early plasma were injured more severely coagulopathic, hypoperfused, and hypotensive on admission. Survival was similar in the early plasma versus no early plasma groups (78% vs 84%, P = .08). After adjustment for covariates, early plasma was not associated with improved survival (odds ratio 1.18, 95% confidence interval 0.71–1.96). On subgroup analysis, multifocal intracranial hemorrhage was the largest subgroup with 242 patients. Of these, 61 (25%) received plasma within 4 hours. Within-group logistic regression analysis with adjustment for covariates found that early plasma was associated with improved survival (odds ratio 3.34, 95% confidence interval 1.20–9.35). Conclusion Although early plasma transfusion was not associated with improved in-hospital survival for all isolated traumatic brain injury patients, early plasma was associated with increased in-hospital survival in those with multifocal intracranial hemorrhage.

INTRODUCTION: traumatic brain injury (TBI) is associated with secondary injury to the central nervous system (CNS) via inflammatory mechanisms. The combination of polytrauma and TBI (TBI) further exacerbates the inflammatory response to injury, however combined injury phenomena has not been thoroughly studied. In this study, we examined the inflammatory differences between patients with TBI versus patients with polytrauma but no TBI (polytrauma). We hypothesize that patients with TBI have a heightened early inflammatory response compared to polytrauma. METHODS: We conducted a single center retrospective study of a cohort of patients with polytrauma who were enrolled in the PROPPR study. These patients had blood samples prospectively collected at eight time points in the first 3 days of admission. Using radiological data to determine TBI, our polytrauma cohort was dichotomized into TBI (n=30) or polytrauma (n=54). Inflammatory biomarkers were measured using ELISA. Data across time were compared for TBI vs. polytrauma groups using Wilcoxon rank sum test. Network analysis techniques were used to systematically characterize the inflammatory responses at admission. RESULTS: Patients with TBI (51.6%) had a higher 30-day mortality compared to polytrauma (16.9%) (p-value

Background Inflammatory mechanism has been implicated in delayed cerebral ischemia (DCI) and poor functional outcomes after subarachnoid hemorrhage (SAH). Identification of cytokine patterns associated with inflammation in acute SAH will provide insights into underlying biological processes of DCI and poor outcomes that may be amenable to interventions. Methods Serum samples were collected from a prospective cohort of 60 patients with acute non-traumatic SAH at four time periods ( Results Of the 60 patients enrolled in the study, 14 (23.3%) developed DCI and 16 (26.7%) had poor functional outcomes at 3 months. DCI was associated with increased levels of PDGF-ABBB and CCL5 and decreased levels of IP-10 and MIP-1α. Poor functional outcome was associated with increased levels of IL-6 and MCP-1α. Network analysis identified distinct cytokine clusters associated with DCI and functional outcomes. Conclusions Serum cytokine patterns in early SAH are associated with poor functional outcomes and DCI. The significant cytokines primarily modulate the inflammatory response. This supports earlier SAH studies linking inflammation and poor outcomes. In particular, this study identifies novel cytokine patterns over time that may indicate impending DCI.

Inhibitors targeting KRAS(G12C), a mutant form of the guanosine triphosphatase (GTPase) KRAS, are a promising new class of oncogene-specific therapeutics for the treatment of tumors driven by the mutant protein. These inhibitors react with the mutant cysteine residue by binding covalently to the switch-II pocket (S-IIP) that is present only in the inactive guanosine diphosphate (GDP)–bound form of KRAS(G12C), sparing the wild-type protein. We used a genome-scale CRISPR interference (CRISPRi) functional genomics platform to systematically identify genetic interactions with a KRAS(G12C) inhibitor in cellular models of KRAS(G12C) mutant lung and pancreatic cancer. Our data revealed genes that were selectively essential in this oncogenic driver–limited cell state, meaning that their loss enhanced cellular susceptibility to direct KRAS(G12C) inhibition. We termed such genes “collateral dependencies” (CDs) and identified two classes of combination therapies targeting these CDs that increased KRAS(G12C) target engagement or blocked residual survival pathways in cells and in vivo. From our findings, we propose a framework for assessing genetic dependencies induced by oncogene inhibition.

Traumatic brain injury (TBI) is the largest non-genetic, non-aging related risk factor for Alzheimer's disease (AD). We report here that TBI induces tau acetylation (ac-tau) at sites acetylated also in human AD brain. This is mediated by S-nitrosylated-GAPDH, which simultaneously inactivates Sirtuin1 deacetylase and activates p300/CBP acetyltransferase, increasing neuronal ac-tau. Subsequent tau mislocalization causes neurodegeneration and neurobehavioral impairment, and ac-tau accumulates in the blood. Blocking GAPDH S-nitrosylation, inhibiting p300/CBP, or stimulating Sirtuin1 all protect mice from neurodegeneration, neurobehavioral impairment, and blood and brain accumulation of ac-tau after TBI. Ac-tau is thus a therapeutic target and potential blood biomarker of TBI that may represent pathologic convergence between TBI and AD. Increased ac-tau in human AD brain is further augmented in AD patients with history of TBI, and patients receiving the p300/CBP inhibitors salsalate or diflunisal exhibit decreased incidence of AD and clinically diagnosed TBI.

Preventing secondary brain injury after neurological insults is one of the primary goals of the neurocritical care unit. Our understanding of the roles of intracranial pressure (ICP) and cerebral edema in managing patients in the neurocritical care units is still evolving. Recent clinical trials examining the monitoring and treatment of elevated ICP have influenced the way we think about intracranial hypertension. Additionally, new methods of monitoring ICP, new physiologic surrogates derived from ICP measurements, and evolving technology to measure cerebral edema are currently being studied and tested for clinical efficacy. In this article, we will discuss both traditional and novel methods of monitoring ICP and cerebral edema.

Background and Purpose Hemicraniectomy is a decompressive surgery used to remove a large bone flap to allow edematous brain tissue to bulge extracranially. However, early indicators of the decompressive effects of hemicraniectomy are unclear. We investigated whether reduction of midline shift following hemicraniectomy is associated with improved consciousness and survival in patients with malignant middle cerebral artery infarctions. Methods We studied 70 patients with malignant middle cerebral artery infarctions (MMI) who underwent hemicraniectomies. Midline shift was measured preoperatively and postoperatively using computed tomography (CT). Consciousness level was evaluated using the Glasgow Coma Scale on postoperative day 1. Patient survival was assessed six months after stroke onset. Results The median time interval between preoperative and postoperative CT was 8.3 hours (interquartile range, 6.1–10.2 hours). Reduction in midline shift was associated with higher postoperative Glasgow Coma Scale scores (P

Hepatocellular carcinoma (HCC) is the second leading cause of cancer death worldwide. Advanced HCC has proven particularly difficult to treat because of a scarcity of clear genetic drivers of cancer progression; thus, there are currently no predictive markers that guide HCC therapy. HCC arises in the context of co-morbid hepatitis due to hepatitis B virus (HBV), hepatitis C (HCV) or fatty liver disease. We hypothesize that protein-protein interactions (PPIs) between viral proteins and HCC genes may contribute to tumor initiation and maintenance. In order to characterize these PPIs, we performed affinity purification - mass spectrometry (APMS), defining 145 HBV/host PPIs including known and novel interacting partners. We next used a network propagation algorithm to identify host genes and protein complexes that were preferentially mutated in the absence of HBV infection. HBV is a small DNA virus, with 4 genes of which only one has enzymatic activity, raising a question as to how HBV interaction modifies host behavior. Using AP-MS of host proteins, we found that the HBV X protein (HBx) remodels multiple host protein complexes through direct interaction. These physical effects on complex components result in distinct biochemical behavior from the CRL4 E3 ubiquitin ligase complex as well as the phosphatase PP2A, as determined through global phosphoproteomics and ubiquitin analysis. We show that this remodeling driven by HBx substantially changes cellular protein turnover and downstream signaling dynamics. We followed this up with assessments of cellular viability and proliferation in response to pharmacological inhibition or CRISPRi-based knockdown of HBx effectors. Our data support a model where HBV proteins alter the components and behavior of key regulatory protein complexes in the cell, altering tumor behavior and raising the possibility of precision therapeutics for HCC. Citation Format: John D. Gordan, Adriana Pitea, Manon Eckhardt, Gwendolyn Jang, Rigney E. Turnham, Alex L M. Choi, John Von Dollen, Huat C. Lim, Elizabeth F. Thayer, R. Katie Kelley, Danielle L. Swaney, Wei Zhang, Fabian J. Theis, Trey Ideker, Nevan J. Krogan. Hepatitis B virus remodels host protein interaction networks to generate distinct cellular dependencies [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 4891.

Dissemination is an essential early step in metastasis but its molecular basis remains incompletely understood. To define the essential targetable effectors of this process, we developed a 3D mammary epithelial culture model, in which dissemination is induced by overexpression of the transcription factor Twist1. Transcriptomic analysis and ChIP-PCR together demonstrated that protein kinase D1 (Prkd1) is a direct transcriptional target of Twist1 and is not expressed in the normal mammary epithelium. Pharmacologic and genetic inhibition of Prkd1 in the Twist1-induced dissemination model demonstrated that Prkd1 was required for cells to initiate extracellular matrix (ECM)–directed protrusions, release from the epithelium, and migrate through the ECM. Antibody-based protein profiling revealed that Prkd1 induced broad phosphorylation changes, including an inactivating phosphorylation of β-catenin and two microtubule depolymerizing phosphorylations of Tau, potentially explaining the release of cell–cell contacts and persistent activation of Prkd1. In patients with breast cancer, TWIST1 and PRKD1 expression correlated with metastatic recurrence, particularly in basal breast cancer. Prkd1 knockdown was sufficient to block dissemination of both murine and human mammary tumor organoids. Finally, Prkd1 knockdown in vivo blocked primary tumor invasion and distant metastasis in a mouse model of basal breast cancer. Collectively, these data identify Prkd1 as a novel and targetable signaling node downstream of Twist1 that is required for epithelial invasion and dissemination. Significance: Twist1 is a known regulator of metastatic cell behaviors but not directly targetable. This study provides a molecular explanation for how Twist1-induced dissemination works and demonstrates that it can be targeted.

Survivors of cerebral aneurysm rupture are at risk for significant morbidity and neurological deficits. Much of this is related to the effects of blood in the subarachnoid space which induces an inflammatory cascade with numerous downstream consequences. Recent clinical trials have not been able to reduce the toxic effects of free hemoglobin or improve clinical outcome. One reason for this may be the inability to identify patients at high risk for neurologic decline. Recently, haptoglobin genotype has been identified as a pertinent factor in diabetes, sickle cell, and cardiovascular disease, with the Hp 2-2 genotype contributing to increased complications. Haptoglobin is a protein synthesized by the liver that binds free hemoglobin following red blood cell lysis, and in doing so, prevents hemoglobin induced toxicity and facilitates clearance. Clinical studies in patients with subarachnoid hemorrhage indicate that Hp 2-2 patients may be a high-risk group for hemorrhage related complications and poor outcome. We review the relevance of haptoglobin in subarachnoid hemorrhage and discuss the effects of genotype and expression levels on the known mechanisms of early brain injury (EBI) and cerebral ischemia after aneurysm rupture. A better understanding of haptoglobin and its role in preventing hemoglobin related toxicity should lead to novel therapeutic avenues.

Background: Infection with HIV predisposes patients to a myriad of neurologic disorders, including cerebrovascular disease. The pathophysiology is likely multifactorial, with proposed mechanisms including infectious vasculitis, HIV-induced endothelial dysfunction and adverse effects of combination antiretroviral therapy (cART). Epidemiologic data on clinically evident cerebral vasculopathy in HIV-infected adults is scarce, even though stroke hospitalizations are rising in this patient population. Methods: A total of 6,298 HIV-infected adults (San Francisco General Hospital, 2000-2013) were screened to generate a cohort of patients with dedicated neuroimaging of the intra- and extracranial cerebral vasculature. We extracted information regarding the extent of HIV disease (including serial viral load and CD4 counts), cardiovascular disease risk factors and exposure to cART (cross-referenced with pharmacy records) and performed multivariate logistic regression analysis to identify predictors of vasculopathy. Results: Of 144 patients, 55 patients (38.2%) had radiographic evidence of cerebral vasculopathy. Twenty (13.9%) had a vasculopathy characterized by vessel dolichoectasia and intracranial aneurysm formation. Thirty-five patients (24.3%) had intra- and or extracranial stenosis/occlusion. cART use (OR 2.27, 95% CI 1.03-5) and tobacco abuse (OR 2.35, 95% CI 1.04-5.25) were independently associated with the development of any vasculopathy, whereas cART use was also an independent risk factor for the stenosis/occlusion subtype specifically (OR 2.87, 95% CI 1.11-7.45). Conclusions: There was a high frequency of cerebral arterial disease in this neuroimaging cohort of HIV/AIDS patients. A history of cART use and a history of tobacco abuse were independent risk factors for vasculopathy, though these findings should be confirmed with large-scale prospective studies.

Background and Purpose— Neurological deterioration (ND) is a devastating complication after intracerebral hemorrhage but little is known about time course and predictors. Methods— We performed a retrospective cohort study of placebo patients in intracerebral hemorrhage trials. We performed computed tomographic scans within 3 hours of symptoms and at 24 and 72 hours; and clinical evaluations at baseline, 1-hour, and days 1, 2, 3, and 15. Timing of ND was predefined as follows: hyperacute (within 1 hour), acute (1–24 hours), subacute (1–3 days), and delayed (3–15 days). Results— We enrolled 376 patients and 176 (47%) had ND within 15 days. In multivariate analyses of ND by category, hyperacute ND was associated with hematoma expansion (odds ratio [OR], 3.6; 95% confidence interval [CI], 1.7–7.6) and baseline intracerebral hemorrhage volume (OR, 1.04 per mL; 95% CI 1.02–1.06); acute ND with hematoma expansion (OR, 7.59; 95% CI, 3.91–14.74), baseline intracerebral hemorrhage volume (OR, 1.02 per mL; 95% CI, 1.01–1.04), admission Glasgow Coma Scale (OR, 0.77 per point; 95% CI, 0.65–0.91), and interventricular hemorrhage (OR, 2.14; 95% CI, 1.05–4.35); subacute ND with 72-hour edema (OR, 1.03 per mL; 95% CI, 1.02–1.05) and fever (OR, 2.49; 95% CI, 1.01–6.14); and delayed ND with age (OR, 1.11 per year; 95% CI, 1.04–1.18), troponin (OR, 4.30 per point; 95% CI, 1.71–10.77), and infections (OR, 3.69; 95% CI, 1.11–12.23). Patients with ND had worse 90-day modified Rankin scores (5 versus 3; P Conclusions— ND occurs frequently and predicts poor outcomes. Our results implicate hematoma expansion and interventricular hemorrhage in early ND, and cerebral edema, fever, and medical complications in later ND.

The mechanism of early neurological deterioration (END) in patients with stroke remains unclear. We assessed the relationship between nocturnal oxygen desaturation (NOD) in the stroke unit (SU) and END, especially occurring at nighttime, following acute stroke.A retrospective analysis was performed on a total of 276 patients with ischemic stroke who were admitted to the SU between July 2013 and June 2015. The oxygen desaturation index was calculated from pulse oximetry data sampled every 1 minute during 9 hours on the first night (10:00 PM to 7:00 AM) after admission, and NOD was defined as oxygen desaturation index ≥ 5 events/h. END was defined as an increase of ≥ 2 points from the baseline National Institutes of Health Stroke Scale during 7 days after onset. We compared clinical characteristics and NOD between patients with and without END.Among the included patients (mean age 69.2; male 55.4%), 42 patients (15.2%) experienced END. The proportion of NOD was significantly greater in the END group (45.2% versus 12.8%,NOD in the SU was associated with END, especially during nighttime, after ischemic stroke. This suggests that treatment of sleep-disordered breathing could be a modifiable factor to possibly reduce the risk of neurological worsening among acute stroke patients.

SUMMARYAlthough macrophages are armed with potent anti-bacterial functions, Mycobacterium tuberculosis (Mtb) replicates inside these innate immune cells. Determinants of macrophage-intrinsic bacterial control, and the Mtb strategies to overcome them are poorly understood. To further study these processes, we used a systematic affinity tag purification mass spectrometry (AP-MS) approach to identify 187 Mtb-human protein-protein interactions (PPIs) involving 34 secreted Mtb proteins. This interaction map revealed two new factors involved in Mtb pathogenesis - the secreted Mtb protein, LpqN, and its binding partner, the human ubiquitin ligase CBL. We discovered that an lpqN Mtb mutant is attenuated in macrophages, but growth is restored when CBL is removed. Conversely, Cbl-/- macrophages are resistant to viral infection, indicating that CBL regulates cell-intrinsic polarization between anti-bacterial and anti-viral immunity. Collectively, these findings illustrate the utility of this Mtb-human PPI map as a resource for developing a deeper understanding of the intricate interactions between Mtb and its host.

Although macrophages are armed with potent antibacterial functions, Mycobacterium tuberculosis (Mtb) replicates inside these innate immune cells. Determinants of macrophage intrinsic bacterial control, and the Mtb strategies to overcome them, are poorly understood. To further study these processes, we used an affinity tag purification mass spectrometry (AP-MS) approach to identify 187 Mtb-human protein-protein interactions (PPIs) involving 34 secreted Mtb proteins. This interaction map revealed two factors involved in Mtb pathogenesis - the secreted Mtb protein, LpqN, and its binding partner, the human ubiquitin ligase CBL. We discovered that an lpqN Mtb mutant is attenuated in macrophages, but growth is restored when CBL is removed. Conversely, Cbl(−/−) macrophages are resistant to viral infection, indicating that CBL regulates cell-intrinsic polarization between antibacterial and antiviral immunity. Collectively, these findings illustrate the utility of this Mtb-human PPI map for developing a deeper understanding of the intricate interactions between Mtb and its host.

Background. The gut is a major reservoir for human immunodeficiency virus (HIV) in patients receiving antiretroviral therapy (ART). We hypothesized that distinct immune environments within the gut may support varying levels of HIV.Methods. In 8 HIV-1-positive adults who were receiving ART and had CD4 + T cell counts of >200 cells/µL and plasma viral loads of