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BACKGROUND: Gene ontology (GO) enrichment analysis is frequently undertaken during exploration of various -omics data sets. Despite the wide array of tools available to biologists to perform this analysis, meaningful visualisation of the overrepresented GO in a manner which is easy to interpret is still lacking. RESULTS: Monash Gene Ontology (MonaGO) is a novel web-based visualisation system that provides an intuitive, interactive and responsive interface for performing GO enrichment analysis and visualising the results. MonaGO supports gene lists as well as GO terms as inputs. Visualisation results can be exported as high-resolution images or restored in new sessions, allowing reproducibility of the analysis. An extensive comparison between MonaGO and 11 state-of-the-art GO enrichment visualisation tools based on 9 features revealed that MonaGO is a unique platform that simultaneously allows interactive visualisation within one single output page, directly accessible through a web browser with customisable display options. CONCLUSION: MonaGO combines dynamic clustering and interactive visualisation as well as customisation options to assist biologists in obtaining meaningful representation of overrepresented GO terms, producing simplified outputs in an unbiased manner. MonaGO will facilitate the interpretation of GO analysis and will assist the biologists into the representation of the results.
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Software , Análise por Conglomerados , Ontologia Genética , Probabilidade , Reprodutibilidade dos TestesRESUMO
In January 2019, a new plant-derived purified cannabidiol preparation, approved by the US Food and Drug Administration, became commercially available for patients ≥2 years old with Lennox-Gastaut syndrome or Dravet syndrome. Among our patients who were prescribed the new cannabidiol formulation, we observed several cases of thrombocytopenia and therefore embarked on this study. We conducted a single-center systematic chart review of all pediatric patients (<21 years old) who were prescribed cannabidiol from January to August 2019. We evaluated salient features of the patients' epilepsy syndrome, age, concurrent medications, and surveillance laboratory results before and after cannabidiol initiation. Among 87 patients, nine (10%) developed thrombocytopenia (platelet nadir range = 17 000-108 000) following initiation of cannabidiol. Each of these nine children was on combination therapy of cannabidiol with valproic acid. Whereas no children on cannabidiol without valproic acid (0/57) developed thrombocytopenia, nine of 23 treated with combination valproic acid and cannabidiol developed platelets < 110 000/µL (P < .0001). We report a novel and clinically important side effect of thrombocytopenia in one-third of patients treated concurrently with cannabidiol and valproic acid. If this finding is confirmed, clinicians should perform close monitoring for thrombocytopenia when adding cannabidiol to a regimen that includes valproic acid.
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Anticonvulsivantes/uso terapêutico , Canabidiol/uso terapêutico , Epilepsia Resistente a Medicamentos/tratamento farmacológico , Epilepsias Mioclônicas/tratamento farmacológico , Síndrome de Lennox-Gastaut/tratamento farmacológico , Trombocitopenia/epidemiologia , Ácido Valproico/uso terapêutico , Adolescente , Criança , Pré-Escolar , Quimioterapia Combinada , Epilepsia/tratamento farmacológico , Feminino , Humanos , Lactente , Masculino , Adulto JovemRESUMO
BACKGROUND: A strong focus of the post-genomic era is mining of the non-coding regulatory genome in order to unravel the function of regulatory elements that coordinate gene expression (Nat 489:57-74, 2012; Nat 507:462-70, 2014; Nat 507:455-61, 2014; Nat 518:317-30, 2015). Whole-genome approaches based on next-generation sequencing (NGS) have provided insight into the genomic location of regulatory elements throughout different cell types, organs and organisms. These technologies are now widespread and commonly used in laboratories from various fields of research. This highlights the need for fast and user-friendly software tools dedicated to extracting cis-regulatory information contained in these regulatory regions; for instance transcription factor binding site (TFBS) composition. Ideally, such tools should not require prior programming knowledge to ensure they are accessible for all users. RESULTS: We present TrawlerWeb, a web-based version of the Trawler_standalone tool (Nat Methods 4:563-5, 2007; Nat Protoc 5:323-34, 2010), to allow for the identification of enriched motifs in DNA sequences obtained from next-generation sequencing experiments in order to predict their TFBS composition. TrawlerWeb is designed for online queries with standard options common to web-based motif discovery tools. In addition, TrawlerWeb provides three unique new features: 1) TrawlerWeb allows the input of BED files directly generated from NGS experiments, 2) it automatically generates an input-matched biologically relevant background, and 3) it displays resulting conservation scores for each instance of the motif found in the input sequences, which assists the researcher in prioritising the motifs to validate experimentally. Finally, to date, this web-based version of Trawler_standalone remains the fastest online de novo motif discovery tool compared to other popular web-based software, while generating predictions with high accuracy. CONCLUSIONS: TrawlerWeb provides users with a fast, simple and easy-to-use web interface for de novo motif discovery. This will assist in rapidly analysing NGS datasets that are now being routinely generated. TrawlerWeb is freely available and accessible at: http://trawler.erc.monash.edu.au .
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Sequenciamento de Nucleotídeos em Larga Escala/métodos , Análise de Sequência de DNA/métodos , Software , Animais , Sequência de Bases , Sítios de Ligação , Sequência Conservada , DNA/química , DNA/metabolismo , Humanos , Internet , Mesotelina , Camundongos , Motivos de Nucleotídeos , Ratos , Fatores de Transcrição/metabolismoAssuntos
Anticonvulsivantes/uso terapêutico , Convulsões/tratamento farmacológico , Administração Intranasal , Administração Retal , Anticonvulsivantes/farmacologia , Benzodiazepinas/farmacologia , Criança , Diazepam/farmacologia , Diazepam/uso terapêutico , Epilepsia/tratamento farmacológico , Humanos , Midazolam/farmacologia , Midazolam/uso terapêutico , Estados Unidos , United States Food and Drug AdministrationRESUMO
OBJECTIVE: We aimed to determine the diagnostic yield of continuous monitoring with video electroencephalography (cVEEG) for pediatric inpatients with paroxysmal vital sign changes (PVSCs), and to identify risk factors for the PVSCs being seizures, based on clinical information available before cVEEG initiation. We hypothesized that PVSCs without nonautonomic symptoms (NAS) were unlikely to be seizures, and also that patients' clinical characteristics would alter the risk of recording seizures. METHODS: We performed a single-center chart review of 324 cVEEG studies that were obtained for differential diagnosis of PVSCs. We examined the type of PVSCs that prompted cVEEG, associated NAS, and patient characteristics, and whether the target events or seizures were recorded. We performed logistic regression analyses to determine which patient and semiologic features altered the risk of the PVSCs being seizures, and which patient characteristics altered the risk of recording any seizures. RESULTS: Target PVSCs were recorded in 52% (N = 169). Seizures were recorded in 21% (N = 69) of the studies, often unrelated to the PVSCs (N = 39). When examining only PVSCs without NAS, only 4% (3/75) of studies obtained for apnea and 2.1% (1/48) of studies obtained for oxygen desaturation revealed the target events to be seizures. No studies recorded ictal hypertension (0/26), hypotension (0/16), or bradycardia (0/18). In univariate analysis, there was a decreased risk that the events were seizures when PVSCs lacked NAS (odds ratio [OR] 0.23, 95% confidence interval [CI] 0.08-0.65). The risk was increased when the patient had received an antiseizure medication (2.9, 1.3-6.5), the target PVSC was apnea (3.5, 1.5-8.5), and in particular, apnea with NAS (8.7, 3.7-20.8). In adjusted analyses, only apnea with associated NAS independently increased the risk of the PVSCs being seizures (7.7, 3.2-18.5). SIGNIFICANCE: PVSCs in the absence of NAS are rarely due to seizures. Ideally, cVEEG should be reserved for children with additional risk factors for seizures, beyond isolated PVSCs.
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Apneia/etiologia , Bradicardia/etiologia , Eletroencefalografia , Hipertensão/etiologia , Hipotensão/etiologia , Convulsões/complicações , Taquicardia/etiologia , Gravação em Vídeo , Sinais Vitais , Adolescente , Apneia/diagnóstico , Bradicardia/diagnóstico , Criança , Pré-Escolar , Estudos de Coortes , Feminino , Humanos , Hipertensão/diagnóstico , Hipotensão/diagnóstico , Lactente , Recém-Nascido , Masculino , Monitorização Fisiológica , Estudos Retrospectivos , Convulsões/diagnóstico , Taquicardia/diagnósticoRESUMO
STUDY OBJECTIVES: Cultural sleep practices and COVID-19 mitigation strategies vary worldwide. The sleep of infants and toddlers during the COVID-19 pandemic in the United States is understudied. METHODS: Caregivers of children aged < 3 years responded to a cross-sectional survey during 2020 (divided into quarters, with the year quarter 1 being largely prelockdown). We assessed the global effect of year quarter on parent-reported total sleep time (hours) and sleep onset latency (hours) using an analysis of variance. We used multivariable linear regression to assess the adjusted effect of year quarter on total sleep time, sleep onset latency, and parental frustration. We used logistic regression to assess the adjusted effect of year quarter on nap consistency. RESULTS: Of 594 children, the mean age was 18.5 ± 9.7 months; 52% were female. In the adjusted analyses, the reference categories were as follows: quarter 1 (year quarter), ≤ 6 months (age category), and < $25,000 (annual household income). Total sleep time was associated with age category (ages 12 to ≤ 24 months: ß = -2.86; P = .0004; ages 24 to ≤ 36 months: ß = -3.25; P < .0001) and maternal age (ß = -0.04; P = .05). Sleep onset latency was associated with year quarter (year quarter 3: ß = 0.16; P = .04), age category (ages 24 to ≤ 36 months: ß = 0.28; P < .0001), annual household income ($100,000-$150,000: ß = -0.15; P = .03; > $150,000: ß = -0.19; P = .01), and lack of room-sharing (ß = -0.09; P = .05). Parental frustration with sleep increased with age (all P < .05) and lack of room-sharing (P = .01). The effect of lack of room-sharing on nap consistency approached significance (adjusted odds ratio, 1.88; 95% confidence interval, 0.95-3.72). CONCLUSIONS: Social factors such as lower household income and room-sharing affected the sleep of U.S. infants and toddlers as opposed to the COVID-19 lockdown itself. CITATION: Gupta G, O'Brien LM, Dang LT, Shellhaas RA. Sleep of infants and toddlers during 12 months of the COVID-19 pandemic in the midwestern United States. J Clin Sleep Med. 2022;18(5):1225-1234.
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COVID-19 , Pré-Escolar , Controle de Doenças Transmissíveis , Estudos Transversais , Feminino , Humanos , Lactente , Masculino , Meio-Oeste dos Estados Unidos/epidemiologia , Pandemias , Sono , Estados Unidos/epidemiologiaRESUMO
BACKGROUND: Sleep comorbidities are common, and sometimes severe, for children with early-life epilepsies (ELEs). Yet, there is a paucity of data regarding the profile of these sleep disturbances and their complications. METHODS: Participants registered with the Rare Epilepsy Network (REN) were queried about sleep via online questionnaires. Descriptive statistics and logistic regression were performed. RESULTS: Median age of the 356 children was 56 months (interquartile range 30 to 99), 56% were female, and 53% (188/356) endorsed a sleep concern. Frequent nighttime awakenings (157 of 350; 45%), difficulty falling asleep (133 of 350; 38%), and very restless sleep (118 of 345; 34%) were most endorsed. Nocturnal seizures were associated with sleep concerns and were reported in 75% (268 of 356) of children. Of the children with nocturnal seizures, 56% (118 of 268) had sleep concerns. Of the children without nocturnal seizures, 43% (38 of 88) had sleep concerns. Sleep concerns were most common in dup15q syndrome (16 of 19; 84%). Children aged 4 to ≤10 years (adjusted odds ratio [aOR] 16.1; 95% confidence interval [CI] 2.0, 131.0) and 10 to <13 years (aOR 22.2; 95% CI 2.6, 188.6) had a greater odds of having a sleep concern compared with children aged ≤6 months. Female sex appeared protective for sleep concerns (aOR 0.6; 95% CI 0.4, 0.9). The association between sleep concerns and nocturnal seizures was weaker when adjusted for sex and age category in a logistic regression model. CONCLUSIONS: Reported sleep concerns are highly prevalent in children with ELEs and persist with age, in contrast to what is expected in healthy children. There may be unmet sleep-related clinical needs in children with ELEs.
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Epilepsia/complicações , Transtornos do Sono-Vigília/diagnóstico , Transtornos do Sono-Vigília/epidemiologia , Fatores Etários , Criança , Pré-Escolar , Estudos de Coortes , Feminino , Humanos , Masculino , Pais , Fatores de Risco , Inquéritos e QuestionáriosRESUMO
Background: Pathogenic variants in SCN1B, the gene encoding voltage-gated sodium channel b1/b1B subunits are associated with a spectrum of epileptic disorders. This study describes a child with early myoclonic encephalopathy and a compound heterozygous variant in the SCN1B gene (p.Arg85Cys and c.3G>C/p.Met1), along with the child's clinical response to anti-seizure medications (ASMs) and the ketogenic diet. We reviewed the current clinical literature pertinent to SCN1B-related epilepsy. Methods: We described the evaluation and management of a patient with SCN1B-related developmental and epileptic encephalopathy (DEE). We used the Medline and Pubmed databases to review the various neurological manifestations associated with SCN1B genetic variants, and summarize the functional studies performed on SCN1B variants. Results: We identified 20 families and six individuals (including the index case described herein) reported to have SCN1B-related epilepsy. Individuals with monoallelic pathogenic variants in SCN1B often present with genetic epilepsy with febrile seizures plus (GEFS+), while those with biallelic pathogenic variants may present with developmental and epileptic encephalopathy (DEE). Individuals with DEE present with seizures of various semiologies (commonly myoclonic seizures) and status epilepticus at early infancy and are treated with various antiseizure medications. In our index case, adjunctive fenfluramine was started at 8 months of age at 0.2 mg/kg/day with gradual incremental increases to the final dose of 0.7 mg/kg/day over 5 weeks. Fenfluramine was effective in the treatment of seizures, resulting in a 50% reduction in myoclonic seizures, status epilepticus, and generalized tonic-clonic seizures, as well as a 70−90% reduction in focal seizures, with no significant adverse effects. Following the initiation of fenfluramine at eight months of age, there was also a 50% reduction in the rate of hospitalizations. Conclusions: SCN1B pathogenic variants cause epilepsy and neurodevelopmental impairment with variable expressivity and incomplete penetrance. The severity of disease is associated with the zygosity of the pathogenic variants. Biallelic variants in SCN1B can result in early myoclonic encephalopathy, and adjunctive treatment with fenfluramine may be an effective treatment for SCN1B-related DEE. Further research on the efficacy and safety of using newer ASMs, such as fenfluramine in patients under the age of 2 years is needed.
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Genetic diseases involving overactivation of the mechanistic target of rapamycin (mTOR) pathway, so-called "mTORopathies," often manifest with malformations of cortical development (MCDs), epilepsy, and cognitive impairment. How mTOR pathway hyperactivation results in abnormal human cortical development is poorly understood. To study the effect of mTOR hyperactivity on early stages of cortical development, we focused on Pretzel Syndrome (polyhydramnios, megalencephaly, symptomatic epilepsy; PMSE syndrome), a rare mTORopathy caused by homozygous germline mutations in the STRADA gene. We developed a human cortical organoid (hCO) model of PMSE and examined morphology and size for the first 2 weeks of organoid growth, and cell type composition at weeks 2, 8, and 12 of differentiation. In the second week, PMSE hCOs enlarged more rapidly than controls and displayed an abnormal Wnt pathway-dependent increase in neural rosette structures. PMSE hCOs also exhibited delayed neurogenesis, decreased subventricular zone progenitors, increased proliferation and cell death, and an abnormal architecture of primary cilia. At week 8, PMSE hCOs had fewer deep layer neurons. By week 12, neurogenesis recovered in PMSE organoids, but they displayed increased outer radial glia, a cell type thought to contribute to the expansion of the human cerebral cortex. Together, these findings suggest that megalencephaly in PMSE arises from the expansion of neural stem cells in early corticogenesis and potentially also from increased outer radial glial at later gestational stages. The delayed neuronal differentiation in PMSE organoids demonstrates the important role the mTOR pathway plays in the maintenance and expansion of the stem cell pool.
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Epilepsia , Megalencefalia , Córtex Cerebral , Epilepsia/genética , Feminino , Humanos , Megalencefalia/genética , Neurogênese , Organoides/metabolismo , GravidezRESUMO
mTORopathies are a heterogeneous group of neurological disorders characterized by malformations of cortical development (MCD), enhanced cellular mechanistic target of rapamycin (mTOR) signaling, and epilepsy that results from mutations in mTOR pathway regulatory genes. Homozygous mutations (del exon 9-13) in the pseudokinase STE20-related kinase adaptor alpha (STRAD-α; STRADA), an mTOR modulator, are associated with Pretzel Syndrome (PS), a neurodevelopmental disorder within the Old Order Mennonite Community characterized by megalencephaly, intellectual disability, and intractable epilepsy. To study the cellular mechanisms of STRADA loss, we generated CRISPR-edited Strada mouse N2a cells, a germline mouse Strada knockout (KO-/-) strain, and induced pluripotent stem cell (iPSC)-derived neurons from PS individuals harboring the STRADA founder mutation. Strada KO in vitro leads to enhanced mTOR signaling and iPSC-derived neurons from PS individuals exhibit enhanced cell size and mTOR signaling activation, as well as subtle alterations in electrical firing properties e.g., increased input resistance, a more depolarized resting membrane potential, and decreased threshold for action potential (AP) generation. Strada-/- mice exhibit high rates of perinatal mortality and out of more than 100 litters yielding both WT and heterozygous pups, only eight Strada-/- animals survived past P5. Strada-/- mice are hypotonic and tremulous. Histopathological examination (n = 5 mice) revealed normal gross brain organization and lamination but all had ventriculomegaly. Ectopic neurons were seen in all five Strada-/- brains within the subcortical white matter mirroring what is observed in human PS brain tissue. These distinct experimental platforms demonstrate that STRADA modulates mTOR signaling and is a key regulator of cell size, neuronal excitability, and cortical lamination.
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For both disease and basic science research, loss-of-function (LOF) mutations are vitally important. Herein, we provide a simple stream-lined protocol for generating LOF iPSC lines that circumvents the technical challenges of traditional gene-editing and cloning of established iPSC lines by combining the introduction of the CRISPR vector concurrently with episomal reprogramming plasmids into fibroblasts. Our experiments have produced nearly even numbers of all 3 genotypes in autosomal genes. In addition, we provide a detailed approach for maintaining and genotyping 96-well plates of iPSC clones.
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Dravet syndrome (DS) is a severe developmental and epileptic encephalopathy with a high incidence of sudden unexpected death in epilepsy (SUDEP). Most DS patients carry de novo variants in SCN1A, resulting in Nav1.1 haploinsufficiency. Because SCN1A is expressed in heart and in brain, we proposed that cardiac arrhythmia contributes to SUDEP in DS. We generated DS patient and control induced pluripotent stem cell-derived cardiac myocytes (iPSC-CMs). We observed increased sodium current (INa) and spontaneous contraction rates in DS patient iPSC-CMs versus controls. For the subject with the largest increase in INa, cardiac abnormalities were revealed upon clinical evaluation. Generation of a CRISPR gene-edited heterozygous SCN1A deletion in control iPSCs increased INa density in iPSC-CMs similar to that seen in patient cells. Thus, the high risk of SUDEP in DS may result from a predisposition to cardiac arrhythmias in addition to seizures, reflecting expression of SCN1A in heart and brain.
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Canalopatias/patologia , Morte Súbita/patologia , Epilepsias Mioclônicas/patologia , Miócitos Cardíacos/patologia , Arritmias Cardíacas/genética , Arritmias Cardíacas/patologia , Sistemas CRISPR-Cas , Células Cultivadas , Canalopatias/genética , Criança , Pré-Escolar , Epilepsias Mioclônicas/genética , Feminino , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Células-Tronco Pluripotentes Induzidas/patologia , Masculino , Miócitos Cardíacos/metabolismo , Canal de Sódio Disparado por Voltagem NAV1.1/genéticaRESUMO
The mainstay of treatment of childhood epilepsy is to administer antiepileptic drugs (AEDs). This article provides an overview of the clinical approach to drug treatment of childhood epilepsy, focusing on general principles of therapy and properties of recently introduced medications. Initiation and cessation of therapy, adverse medication effects, drug interactions, indications for the various AEDs, and off-label use of AEDs are reviewed. The distinct challenges in treatment of epileptic spasms and neonatal seizures are addressed. Finally, ideas for the future of drug treatment of childhood epilepsy are presented, with particular attention to precision medicine.
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Anticonvulsivantes/uso terapêutico , Epilepsia/tratamento farmacológico , Convulsões/tratamento farmacológico , Anticonvulsivantes/efeitos adversos , Criança , Humanos , Recém-NascidoRESUMO
Specifically ablating genes in human induced pluripotent stem cells (iPSCs) allows for studies of gene function as well as disease mechanisms in disorders caused by loss-of-function (LOF) mutations. While techniques exist for engineering such lines, we have developed and rigorously validated a method of simultaneous iPSC reprogramming while generating CRISPR/Cas9-dependent insertions/deletions (indels). This approach allows for the efficient and rapid formation of genetic LOF human disease cell models with isogenic controls. The rate of mutagenized lines was strikingly consistent across experiments targeting four different human epileptic encephalopathy genes and a metabolic enzyme-encoding gene, and was more efficient and consistent than using CRISPR gene editing of established iPSC lines. The ability of our streamlined method to reproducibly generate heterozygous and homozygous LOF iPSC lines with passage-matched isogenic controls in a single step provides for the rapid development of LOF disease models with ideal control lines, even in the absence of patient tissue.
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Reprogramação Celular/genética , Edição de Genes , Células-Tronco Pluripotentes Induzidas/metabolismo , Mutação com Perda de Função/genética , Sistemas CRISPR-Cas/genética , Linhagem Celular , Heterogeneidade Genética , Instabilidade Genômica , Genótipo , Humanos , Mutação INDEL/genética , RNA Guia de Cinetoplastídeos/metabolismo , Reprodutibilidade dos TestesAssuntos
Epilepsia Resistente a Medicamentos , Epilepsia Generalizada , Fenitoína/farmacologia , Convulsões Febris , Subunidade beta-1 do Canal de Sódio Disparado por Voltagem/genética , Anticonvulsivantes , Pré-Escolar , Epilepsia Resistente a Medicamentos/tratamento farmacológico , Epilepsia Resistente a Medicamentos/genética , Epilepsia Resistente a Medicamentos/fisiopatologia , Eletroencefalografia , Epilepsia Generalizada/tratamento farmacológico , Epilepsia Generalizada/genética , Epilepsia Generalizada/fisiopatologia , Humanos , Masculino , Convulsões Febris/tratamento farmacológico , Convulsões Febris/genética , Convulsões Febris/fisiopatologiaRESUMO
Low-molecular-weight heparins, such as enoxaparin, are often used to treat thrombosis in infants. We present 4 infants with diffuse brain injury who developed cerebral venous sinus thrombosis or deep vein thrombosis and were treated with enoxaparin. These infants subsequently developed subdural hemorrhages, and enoxaparin was stopped. In 3 cases, the subdural hemorrhages were found on routine surveillance brain MRI, and in 1 case imaging was urgently obtained because of focal seizures. Two patients needed urgent neurosurgical intervention, and all subdural hemorrhages improved or resolved on follow-up imaging. Each infant developed severe neurologic deficits, probably from the coexisting diffuse brain injury rather than from the subdural hemorrhages themselves. The risk of intracranial hemorrhage from enoxaparin may be accentuated in patients with diffuse brain injury, and careful consideration should be given before treatment in this population.