De novo monoallelic Reelin missense variants act in a dominant-negative manner causing neuronal migration disorders.

Reelin (RELN) is a secreted glycoprotein essential for cerebral cortex development. In humans, recessive RELN variants cause cortical and cerebellar malformations, while heterozygous variants were associated to epilepsy, autism and mild cortical abnormalities. However, their functional effects remain unknown. We identified inherited and de novo RELN missense variants in heterozygous patients with neuronal migration disorders (NMDs) as diverse as pachygyria and polymicrogyria. We investigated in culture and in the developing mouse cerebral cortex how different variants impacted RELN function. Polymicrogyria-associated variants behaved as gain-of-function showing an enhanced ability to induce neuronal aggregation, while those linked to pachygyria as loss-of-function leading to defective neuronal aggregation/migration. The pachygyria-associated de novo heterozygous RELN variants acted as dominant-negative by preventing wild-type RELN secretion in culture, animal models and patients, thereby causing dominant NMDs. We demonstrated how mutant RELN proteins in vitro and in vivo predict cortical malformation phenotypes, providing valuable insights into the pathogenesis of such disorders.

Individualised human phenotype ontology gene panels improve clinical whole exome and genome sequencing analytical efficacy in a cohort of developmental and epileptic encephalopathies.

BACKGROUND: The majority of genetic epilepsies remain unsolved in terms of specific genotype. Phenotype-based genomic analyses have shown potential to strengthen genomic analysis in various ways, including improving analytical efficacy. METHODS: We have tested a standardised phenotyping method termed 'Phenomodels' for integrating deep-phenotyping information with our in-house developed clinical whole exome/genome sequencing analytical pipeline. Phenomodels includes a user-friendly epilepsy phenotyping template and an objective measure for selecting which template terms to include in individualised Human Phenotype Ontology (HPO) gene panels. In a pilot study of 38 previously solved cases of developmental and epileptic encephalopathies, we compared the sensitivity and specificity of the individualised HPO gene panels with the clinical epilepsy gene panel. RESULTS: The Phenomodels template showed high sensitivity for capturing relevant phenotypic information, where 37/38 individuals' HPO gene panels included the causative gene. The HPO gene panels also had far fewer variants to assess than the epilepsy gene panel. CONCLUSION: We have demonstrated a viable approach for incorporating standardised phenotype information into clinical genomic analyses, which may enable more efficient analysis.

Outcome at age 7 of epilepsy presenting in the first 2 years of life. A population-based study.

OBJECTIVE: Existing data suggest that epilepsy presenting in the first few years of life carries a worse prognosis than later onset. However, studies are few and methods differ, making interpretations of data uncertain. This study analyzes outcome at age 7 and potential prognostic factors in a well-characterized population-based cohort with epilepsy onset during the first 2 years of life. METHODS: An incidence cohort of 116 prospectively identified cases of epilepsy with seizure onset before age 2 years was described in Stödberg et al. (2020). Cases were originally retrieved from the Stockholm Incidence Registry of Epilepsy (SIRE), which registered all cases with a first unprovoked epileptic seizure from September 1, 2001, in Northern Stockholm. Data on treatment and outcome at age 7 years were collected from electronic medical records and through interviews with parents. Outcome and potential prognostic factors were analyzed with descriptive statistics and multivariable log binomial regression analysis. RESULTS: Eleven children (9.5%) died before age 7. Polytherapy was common. Epilepsy surgery was performed in two children. At age 7 years, 61 of 116 children (53%) had been seizure-free for the last 2 years or longer. Intellectual disability was diagnosed in 57 of 116 children (49%), autism spectrum disorder in 13 (11%), and cerebral palsy in 28 (24%). West syndrome had a similar seizure remission rate but a worse cognitive outcome. There was no difference in outcome between first and second year onset. Six predictors, including etiology, remained associated with two or more outcome variables after regression analysis. SIGNIFICANCE: About half of children with infantile-onset epilepsy will become seizure-free and half of them will have intellectual disability. Etiology was confirmed as a major independent predictor of outcome. Our study contributes to a more firm knowledge base when counseling parents of infants diagnosed with epilepsy.

Evidence for a Dual-Pathway, 2-Hit Genetic Model for Focal Cortical Dysplasia and Epilepsy.

BACKGROUND AND OBJECTIVES: The 2-hit model of genetic disease is well established in cancer, yet has only recently been reported to cause brain malformations associated with epilepsy. Pathogenic germline and somatic variants in genes in the mechanistic target of rapamycin (mTOR) pathway have been implicated in several malformations of cortical development. We investigated the 2-hit model by performing genetic analysis and searching for germline and somatic variants in genes in the mTOR and related pathways. METHODS: We searched for germline and somatic pathogenic variants in 2 brothers with drug-resistant focal epilepsy and surgically resected focal cortical dysplasia (FCD) type IIA. Exome sequencing was performed on blood- and brain-derived DNA to identify pathogenic variants, which were validated by droplet digital PCR. In vitro functional assays of a somatic variant were performed. RESULTS: Exome analysis revealed a novel, maternally inherited, germline pathogenic truncation variant (c.48delG; p.Ser17Alafs*70) in NPRL3 in both brothers. NPRL3 is a known FCD gene that encodes a negative regulator of the mTOR pathway. Somatic variant calling in brain-derived DNA from both brothers revealed a low allele fraction somatic variant (c.338C>T; p.Ala113Val) in the WNT2 gene in 1 brother, confirmed by droplet digital PCR. In vitro functional studies suggested a loss of WNT2 function as a consequence of this variant. A second somatic variant has not yet been found in the other brother. DISCUSSION: We identify a pathogenic germline mTOR pathway variant (NPRL3) and a somatic variant (WNT2) in the intersecting WNT signaling pathway, potentially implicating the WNT2 gene in FCD and supporting a dual-pathway 2-hit model. If confirmed in other cases, this would extend the 2-hit model to pathogenic variants in different genes in critical, intersecting pathways in a malformation of cortical development. Detection of low allele fraction somatic second hits is challenging but promises to unravel the molecular architecture of FCDs.

"I was 15 when I started doing drugs with my dad": Victimization, Social Determinants of Health, and Criminogenic Risk Among Women Opioid Intervention Court Participants.

The presence and severity of childhood and adult victimization increase the likelihood of substance use disorder (SUD), crimes, antisocial behaviors, arrests, convictions, and medical and psychiatric disorders among women more than men. These problems are compounded by the impact of social determinants of health (SDH) challenges, which include predisposition to the understudied, dramatic increase in opioid dependence among women. This study examined victimization, related SDH challenges, gender-based criminogenic risk factors for female participants, and public health opportunities to address these problems. We recruited women from the first national Opioid Intervention Court, a fast-track SUD treatment response to rapidly increasing overdose deaths. We present a consensual qualitative research analysis of 24 women Opioid Intervention Court participants (among 31 interviewed) who reported childhood, adolescent, and/or adult victimization experiences in the context of substance use and recovery, mental health symptoms, heath behaviors, and justice-involved trajectories. We iteratively established codes and overarching themes. Six primary themes emerged: child or adolescent abuse as triggers for drug use; impact of combined child or adolescent abuse with loss or witnessing abuse; adult abduction or assault; trajectory from lifetime abuse, substance use, and criminal and antisocial behaviors to sobriety; role of friends and family support in recovery; and role of treatment and opioid court in recovery, which we related to SDH, gender-based criminogenic factors, and public health. These experiences put participants at risk of further physical and mental health disorders, yet indicate potential strategies. Findings support future studies examining strategies where courts and health systems could collaboratively address SDH with women Opioid Intervention Court participants.

A Model for Sleep Apnea Management in Underserved Patient Populations.

INTRODUCTION: Obstructive sleep apnea (OSA) is a common condition in the United States that is strongly linked to metabolic disease, cardiovascular disease, and increased mortality. Uninsured populations experience sleep health disparities, including delayed recognition, diagnosis, and treatment of OSA due to barriers accessing and affording care. Partnerships between primary care clinics and sleep medicine specialists for sleep apnea management have the potential to increase screening, testing, and treatment among underserved populations. Here, we present an integrated and cost-effective model that is easier to navigate for patients while maintaining high quality care. METHODS: We designed and implemented a specialty sleep clinic at Shade Tree Clinic, Vanderbilt's student-run, free primary care clinic. Patients with signs and symptoms of OSA were identified at primary care appointments and screened using the STOP-BANG questionnaire. Clinic visits took place over telehealth with a medical student and sleep specialist. Patients were diagnosed using a home sleep test, and if indicated, were prescribed and given a CPAP device for treatment. CPAP adherence was monitored using a cloud-based remote monitoring system. RESULTS: From December 2020 through August 2021, we hosted 6 telehealth Sleep Clinics, seeing a total of 28 patients across these visits. We have received a total of 37 referrals and have coordinated sleep evaluations and diagnostic testing for 18 of these patients so far. Prior to initiation of the sleep clinic, there were 17 patients on our primary care panel at Shade Tree with a diagnosis of OSA. These patients were using donated equipment and many had been lost to follow-up or had broken parts. We were able to replace 10 of these patient's CPAP devices and plan to replace the remaining seven. CONCLUSIONS: We have created a model of integrated specialty care that is efficient and cost-effective. This paradigm can be replicated for the many specialties that are typically overlooked and undertreated when working with uninsured patients. As awareness of this sleep medicine program becomes more widespread at Shade Tree Clinic, we anticipate reaching more primary care patients with signs and symptoms of sleep apnea through student education, cost-effective diagnostics, and partnership with sleep specialists.

Understanding the Accuracy of Clinician Provided Estimated Discharge Dates.

Discharge planning is a vital tool in managing hospital capacity, which is essential for maintaining hospital throughput for surgical postoperative admissions. Early discharge planning has been effective in reducing length of stay and hospital readmissions. Between 2014 and 2017, Vanderbilt University Medical Center (VUMC) implemented a tool in the electronic health record (EHR) requiring providers to input the patient's estimated discharge date on each hospital day. We hypothesized discharge estimates would be more accurate, on average, for surgical patients compared to non-surgical patients because treatment plans are known in advance of surgical admissions. We also analyzed the data to identify factors associated with more accurate discharge estimates. In this retrospective observational study, via an analysis of covariance (ANCOVA) approach, we identified factors associated with more accurate discharge estimates for admitted adult patients at VUMC. The primary outcome was the difference between estimated and actual discharge date, and the primary exposure of interest was whether the patient underwent surgery while admitted to the hospital. A total of 304,802 date of discharge estimate entries from 68,587 inpatient encounters met inclusion criteria. After controlling for measured confounding, we found the discharge estimates were more precise as the difference between estimated and actual discharge date narrowed; for each additional day closer to discharge, prediction accuracy improved by .67 days (95% confidence interval [CI], 0.66 to 0.67; p < 0.001), on average. No difference was observed on the primary outcome in patients undergoing surgery compared with non-surgical treatment (0.02 days; 95% CI, 0.00 to 0.03; p = 0.111). Faculty members were found to perform best among all clinicians in predicting estimated discharge date with a 0.24-day better accuracy (95% CI, 0.20 to 0.27; p < 0.001), on average, than other staff. Weekend and holiday, specific clinical teams, staff types, and discharge dispositions were associated with the variability in estimated versus actual discharge date (p < 0.001). Given the widespread variation in current efforts to improve discharge planning and the recommended approach of assigning a discharge date early in the hospital stay, understanding provider estimated discharge dates is an important tool in hospital capacity management. While we did not determine a difference in discharge estimates among surgical and non-surgical patients, we found estimates were more accurate as discharge came nearer and identified notable trends in provider inputs and patient factors. Assessing factors that impact variability in discharge accuracy can allow hospitals to design targeted interventions to improve discharge planning and reduce unnecessary hospital days.

Intravenous immunoglobulin therapy in COVID-19-related encephalopathy.

OBJECTIVE: To report on efficacy and safety of intravenous immunoglobulin (IVIg) therapy in a case series of patients with COVID-19-related encephalopathy. METHODS: We retrospectively collected data on all patients with COVID-19 hospitalized at two Italian hospitals who developed encephalopathy during disease course and were treated with IVIg. RESULTS: Five patients (two females, mean age 66.8 years) developed encephalopathy after a mean of 12.6 days, since the onset of respiratory/constitutional symptoms related to COVID-19. Four patients suffered severe respiratory distress, three of which required invasive mechanical ventilation. Neurological manifestations included impaired consciousness, agitation, delirium, pyramidal and extrapyramidal signs. EEG demonstrated diffuse slowing in all patients. Brain MRI showed non-specific findings. CSF analysis revealed normal cell count and protein levels. In all subjects, RT-PCR for SARS-CoV-2 in CSF tested negative. IVIg at 0.4 g/kg/die was commenced 29.8 days (mean, range: 19-55 days) after encephalopathy onset, leading to complete electroclinical recovery in all patients, with an initial improvement of neuropsychiatric symptoms observed in 3.4 days (mean, range: 1-10 days). No adverse events related to IVIg were observed. CONCLUSIONS: Our preliminary findings suggest that IVIg may represent a safe and effective treatment for COVID-19-associated encephalopathy. Clinical efficacy may be driven by the anti-inflammatory action of IVIg, associated with its anti-cytokine qualities.

The SMC complex MukBEF recruits topoisomerase IV to the origin of replication region in live Escherichia coli.

UNLABELLED: The Escherichia coli structural maintenance of chromosome (SMC) complex, MukBEF, and topoisomerase IV (TopoIV) interact in vitro through a direct contact between the MukB dimerization hinge and the C-terminal domain of ParC, the catalytic subunit of TopoIV. The interaction stimulates catalysis by TopoIV in vitro. Using live-cell quantitative imaging, we show that MukBEF directs TopoIV to ori, with fluorescent fusions of ParC and ParE both forming cellular foci that colocalize with those formed by MukBEF throughout the cell cycle and in cells unable to initiate DNA replication. Removal of MukBEF leads to loss of fluorescent ParC/ParE foci. In the absence of functional TopoIV, MukBEF forms multiple foci that are distributed uniformly throughout the nucleoid, whereas multiple catenated oris cluster at midcell. Once functional TopoIV is restored, the decatenated oris segregate to positions that are largely coincident with the MukBEF foci, thereby providing support for a mechanism by which MukBEF acts in chromosome segregation by positioning newly replicated and decatenated oris. Additional evidence for such a mechanism comes from the observation that in TopoIV-positive (TopoIV(+)) cells, newly replicated oris segregate rapidly to the positions of MukBEF foci. Taken together, the data implicate MukBEF as a key component of the DNA segregation process by acting in concert with TopoIV to promote decatenation and positioning of newly replicated oris. IMPORTANCE: Mechanistic understanding of how newly replicated bacterial chromosomes are segregated prior to cell division is incomplete. In this work, we provide in vivo experimental support for the view that topoisomerase IV (TopoIV), which decatenates newly replicated sister duplexes as a prelude to successful segregation, is directed to the replication origin region of the Escherichia coli chromosome by the SMC (structural maintenance of chromosome) complex, MukBEF. We provide in vivo data that support the demonstration in vitro that the MukB interaction with TopoIV stimulates catalysis by TopoIV. Finally, we show that MukBEF directs the normal positioning of sister origins after their replication and during their segregation. Overall, the data support models in which the coordinate and sequential action of TopoIV and MukBEF plays an important role during bacterial chromosome segregation.

Encouraging responsible drinking... with a focus on the Northern Territory.

Alcohol consumption per capita in the Northern Territory is substantially higher, at 13.3 litres than the national figure of 9.02 litres (People's Alcohol Action Coalition 2012 and Department of Health and Ageing 2009a). As well as measures to reduce supply, continued efforts to reduce the demand for alcohol have been implemented, including the use of public awareness campaigns.

Suppression of homocysteine levels by vitamin B12 and folates: age and gender dependency in the Jackson Heart Study.

INTRODUCTION: To examine factors potentially contributing to premature cardiovascular disease mortality in African Americans (40% versus 20% all other populations), plasma homocysteine, serum vitamin B12 and folate levels were examined for African American participants in the Jackson Heart Study. METHODS: Of 5192 African American Jackson Heart Study participants (21-94 years), 5064 (mean age, 55 ± 13 years; 63% female) had homocysteine levels measured via fasting blood samples, with further assessments of participants' vitamin B12 (n = 1790) and folate (n = 1788) levels. Regression analyses were used to examine age, gender, vitamin B12 and folate with homocysteine levels. RESULTS: Homocysteine levels, a purported surrogate risk factor for cardiovascular disease, increased with age, were inversely proportional to folate and vitamin B12 levels (P < 0.001) and were higher for men of all ages. CONCLUSIONS: The results show that, as with other populations, age, gender, vitamin B12 and folate may predict homocysteine levels for African Americans. Diet may be an important predictive factor as well, given the relationships that were observed between plasma homocysteine and serum B vitamin levels.

Validity and calibration of food frequency questionnaires used with African-American adults in the Jackson Heart Study.

OBJECTIVE: To examine the relative validity of two food frequency questionnaires (FFQs) developed for use in investigating diet and disease relationships within the adult African-American population in the southern United States. DESIGN: Cross-sectional analyses of dietary nutrient intake data, comparing four 24-hour dietary recalls with an FFQ developed by the Lower Mississippi Delta Nutrition Intervention Research Initiative, and its shorter version adapted for use in the Jackson Heart Study. SUBJECTS: A representative subset of participants (n=499, aged 35 to 81 years) from the baseline Jackson Heart Study cohort (N=5,302) was selected for this study. Data collection took place between winter 2000 and spring 2004. STATISTICAL ANALYSES: Pearson's correlation coefficients (energy adjusted and de-attenuated) for 26 nutrients estimates from each of the FFQs, comparing them with the mean of four 24-hour dietary recalls. The ability of the FFQs to rank individuals based on nutrient intakes was compared to that of the mean of four 24-hour dietary recalls and attenuation coefficients were also calculated. RESULTS: Median nutrient intake estimates tended to be higher on the long and lower on the short FFQ compared to the median for the mean of four 24-hour dietary recalls. Energy adjusted and deattenuated correlations of FFQ intake estimates with recalls ranged from 0.20 for sodium to 0.70 for carbohydrate for the short FFQ and from 0.23 for polyunsaturated fat to 0.75 for dietary fiber and magnesium for the long. Attenuation coefficients for men on average were 0.42 for the short and 0.49 for the long FFQ. For women, these were 0.31 for the short and 0.42 for the long FFQ. CONCLUSIONS: Both FFQs appear to be reasonably valid for assessment of dietary intake of adult African Americans in the South. The Lower Mississippi Delta Nutrition Intervention Research Initiative FFQ exhibited higher intake estimates and stronger correlations with recalls than the Jackson Heart Study FFQ for most nutrients analyzed, more so for women than men.