A conserved molecular logic for neurogenesis to gliogenesis switch in the cerebral cortex.

During development, neural stem cells in the cerebral cortex, also known as radial glial cells (RGCs), generate excitatory neurons, followed by production of cortical macroglia and inhibitory neurons that migrate to the olfactory bulb (OB). Understanding the mechanisms for this lineage switch is fundamental for unraveling how proper numbers of diverse neuronal and glial cell types are controlled. We and others recently showed that Sonic Hedgehog (Shh) signaling promotes the cortical RGC lineage switch to generate cortical oligodendrocytes and OB interneurons. During this process, cortical RGCs generate intermediate progenitor cells that express critical gliogenesis genes Ascl1, Egfr, and Olig2. The increased Ascl1 expression and appearance of Egfr+ and Olig2+ cortical progenitors are concurrent with the switch from excitatory neurogenesis to gliogenesis and OB interneuron neurogenesis in the cortex. While Shh signaling promotes Olig2 expression in the developing spinal cord, the exact mechanism for this transcriptional regulation is not known. Furthermore, the transcriptional regulation of Olig2 and Egfr has not been explored. Here, we show that in cortical progenitor cells, multiple regulatory programs, including Pax6 and Gli3, prevent precocious expression of Olig2, a gene essential for production of cortical oligodendrocytes and astrocytes. We identify multiple enhancers that control Olig2 expression in cortical progenitors and show that the mechanisms for regulating Olig2 expression are conserved between the mouse and human. Our study reveals evolutionarily conserved regulatory logic controlling the lineage switch of cortical neural stem cells.

Cell-type-specific 3D epigenomes in the developing human cortex.

Lineage-specific epigenomic changes during human corticogenesis have been difficult to study owing to challenges with sample availability and tissue heterogeneity. For example, previous studies using single-cell RNA sequencing identified at least 9 major cell types and up to 26 distinct subtypes in the dorsal cortex alone1,2. Here we characterize cell-type-specific cis-regulatory chromatin interactions, open chromatin peaks, and transcriptomes for radial glia, intermediate progenitor cells, excitatory neurons, and interneurons isolated from mid-gestational samples of the human cortex. We show that chromatin interactions underlie several aspects of gene regulation, with transposable elements and disease-associated variants enriched at distal interacting regions in a cell-type-specific manner. In addition, promoters with increased levels of chromatin interactivity-termed super-interactive promoters-are enriched for lineage-specific genes, suggesting that interactions at these loci contribute to the fine-tuning of transcription. Finally, we develop CRISPRview, a technique that integrates immunostaining, CRISPR interference, RNAscope, and image analysis to validate cell-type-specific cis-regulatory elements in heterogeneous populations of primary cells. Our findings provide insights into cell-type-specific gene expression patterns in the developing human cortex and advance our understanding of gene regulation and lineage specification during this crucial developmental window.

Key subphenotypes of bipolar disorder are differentially associated with polygenic liabilities for bipolar disorder, schizophrenia, and major depressive disorder.

Bipolar disorder (BD) features heterogenous clinical presentation and course of illness. It remains unclear how subphenotypes associate with genetic loadings of BD and related psychiatric disorders. We investigated associations between the subphenotypes and polygenic risk scores (PRS) for BD, schizophrenia, and major depressive disorder (MDD) in two BD cohorts from Sweden (N = 5180) and the UK (N = 2577). Participants were assessed through interviews and medical records for inter-episode remission, psychotic features during mood episodes, global assessment of functioning (GAF, function and symptom burden dimensions), and comorbid anxiety disorders. Meta-analyses based on both cohorts showed that inter-episode remission and GAF-function were positively correlated with BD-PRS but negatively correlated with schizophrenia-PRS (SCZ-PRS) and MDD-PRS. Moreover, BD-PRS was negatively, and MDD-PRS positively, associated with the risk of comorbid anxiety disorders. Finally, SCZ-PRS was positively associated with psychotic symptoms during mood episodes. Assuming a higher PRS of certain psychiatric disorders in cases with a positive family history, we further tested the associations between subphenotypes in index BD people and occurrence of BD, schizophrenia, or MDD in their relatives using Swedish national registries. BD patients with a relative diagnosed with BD had: (1) higher GAF and lower risk of comorbid anxiety than those with a relative diagnosed with schizophrenia or MDD, (2) lower risk of psychotic symptoms than those with a relative diagnosed with schizophrenia. Our findings shed light on the genetic underpinnings of the heterogeneity in clinical manifestations and course of illness in BD, which ultimately provide insights for developing personalized approaches to the diagnosis and treatment.

High-throughput CRISPRi and CRISPRa technologies in 3D genome regulation for neuropsychiatric diseases.

Advances in genomics have led to the identification of many risk loci with hundreds of genes and thousands of DNA variants associated with neuropsychiatric disorders. A significant barrier to understanding the genetic underpinnings of complex diseases is the lack of functional characterization of risk genes and variants in biological systems relevant to human health and connecting disease-associated variants to pathological phenotypes. Characterizing gene and DNA variant functions requires genetic perturbations followed by molecular and cellular assays of neurobiological phenotypes. However, generating null or mutant alleles is low throughput, making it impossible to characterize disease-associated variants in large quantities efficiently. CRISPR interference (CRISPRi) and CRISPR activation (CRISPRa) screens can be leveraged to dissect the biological consequences of the tested genes and variants in their native context. Nevertheless, testing non-coding variants associated with complex diseases remains non-trivial. In this review, we first discuss the current challenges of interpreting the function of the non-coding genome and approaches to prioritizing disease-associated variants in the context of the 3D epigenome. Second, we provide a brief overview of high-throughput CRISPRi and CRISPRa screening strategies applicable for characterizing non-coding sequences in appropriate biological systems. Lastly, we discuss the promising prospects of using CRISPR-based technologies to dissect DNA sequences associated with neuropsychiatric diseases.

Meeting report - Cell size and growth: from single cells to the tree of life.

In April 2022, The Company of Biologists hosted their first post-pandemic in-person Workshop at Buxted Park Country House in the Sussex countryside. The Workshop, entitled 'Cell size and growth: from single cells to the tree of life', gathered a small group of early-career and senior researchers with expertise in cell size spanning a broad range of organisms, including bacteria, yeast, animal cells, embryos and plants, and working in fields from cell biology to ecology and evolutionary biology. The programme made ample room for fruitful discussions and provided a much-needed opportunity to discuss the most recent findings relating to the regulation of cell size and growth, identify the emerging challenges for the field, and build a community after the pandemic.

Oncomicrobial Community Profiling Identifies Clinicomolecular and Prognostic Subtypes of Colorectal Cancer.

BACKGROUND & AIMS: Dysbiosis of gut microbiota is linked to the development of colorectal cancer (CRC). However, microbiota-based stratification of CRC tissue and how this relates to clinicomolecular characteristics and prognosis remains to be clarified. METHODS: Tumor and normal mucosa from 423 patients with stage I to IV CRC were profiled by bacterial 16S rRNA gene sequencing. Tumors were characterized for microsatellite instability (MSI), CpG island methylator phenotype (CIMP), APC, BRAF, KRAS, PIK3CA, FBXW7, SMAD4, and TP53 mutations, subsets for chromosome instability (CIN), mutation signatures, and consensus molecular subtypes (CMS). Microbial clusters were validated in an independent cohort of 293 stage II/III tumors. RESULTS: Tumors reproducibly stratified into 3 oncomicrobial community subtypes (OCSs) with distinguishing features: OCS1 (Fusobacterium/oral pathogens, proteolytic, 21%), right-sided, high-grade, MSI-high, CIMP-positive, CMS1, BRAF V600E, and FBXW7 mutated; OCS2 (Firmicutes/Bacteroidetes, saccharolytic, 44%), and OCS3 (Escherichia/Pseudescherichia/Shigella, fatty acid ß-oxidation, 35%) both left-sided and exhibiting CIN. OCS1 was associated with MSI-related mutation signatures (SBS15, SBS20, ID2, and ID7) and OCS2 and OCS3 with SBS18 related to damage by reactive oxygen species. Among stage II/III patients, OCS1 and OCS3 both had poorer overall survival compared with OCS2 for microsatellite stable tumors (multivariate hazard ratio [HR], 1.85; 95% confidence interval [CI], 1.15-2.99; P = .012; and HR, 1.52; 95% CI 1.01-2.29; P = .044, respectively) and left-sided tumors (multivariate HR, 2.66; 95% CI, 1.45-4.86; P = .002; and HR, 1.76; 95% CI, 1.03-3.02; P = .039, respectively). CONCLUSIONS: OCS classification stratified CRCs into 3 distinct subgroups with different clinicomolecular features and outcomes. Our findings provide a framework for a microbiota-based stratification of CRC to refine prognostication and to inform the development of microbiota-targeted interventions.

Utilizing Precursor Ion Connectivity of Different Charge States to Improve Peptide and Protein Identification in MS/MS Analysis.

Tandem mass spectrometry (MS/MS) has become a key method for the structural analysis of biomolecules such as peptides and proteins. A pervasive problem in MS/MS analyses, especially for top-down proteomics, is the occurrence of chimeric spectra, when two or more precursor ions are co-isolated and fragmented, thus leading to complex MS/MS spectra that are populated with fragment ions originating from different precursor ions. This type of convoluted data typically results in low sequence database search scores due to the vast number of mixed-source fragment ions, of which only a fraction originates from a specific precursor ion. Herein, we present a novel workflow that deconvolutes the data of chimeric MS/MS spectra, improving the protein search scores and sequence coverages in database searching and thus providing a more confident peptide and protein identification. Previously misidentified proteins or proteins with insignificant search scores can be correctly and significantly identified following the presented data acquisition and analysis workflow with search scores increasing by a factor of 3-4 for smaller precursor ions (peptides) and >6 for larger precursor ions such as intact ubiquitin and cytochrome C.

Supermarket/hypermarket opportunistic screening for atrial fibrillation (SHOPS-AF) using sensors embedded in the handles of supermarket trolleys: A feasibility study.

BACKGROUND: Atrial fibrillation (AF) increases the risk of death, stroke, heart failure, cognitive decline, and healthcare costs but is often asymptomatic and undiagnosed. There is currently no national screening program for AF. The advent of validated hand-held devices allows AF to be detected in non-healthcare settings, enabling screening to be undertaken within the community. METHOD AND RESULTS: In this novel observational study, we embedded a MyDiagnostick single lead ECG sensor into the handles of shopping trolleys in four supermarkets in the Northwest of England: 2155 participants were recruited. Of these, 231 participants either activated the sensor or had an irregular pulse, suggesting AF. Some participants agreed to use the sensor but refused to provide their contact details, or consent to pulse assessment. In addition, some data were missing, resulting in 203 participants being included in the final analyses. Fifty-nine participants (mean age 73.6 years, 43% female) were confirmed or suspected of having AF; 20 were known to have AF and 39 were previously undiagnosed. There was no evidence of AF in 115 participants and the remaining 46 recordings were non-diagnostic, mainly due to artefact. Men and older participants were significantly more likely to have newly diagnosed AF. Due to the number of non-diagnostic ECGs (n = 46), we completed three levels of analyses, excluding all non-diagnostic ECGs, assuming all non-diagnostic ECGs were masking AF, and assuming all non-diagnostic ECGs were not AF. Based on the results of the three analyses, the sensor's sensitivity (95% CI) ranged from 0.70 to 0.93; specificity from 0.15 to 0.97; positive predictive values (PPV) and negative predictive values (NPV) ranged from 0.24 to 0.56 and 0.55 to 1.00, respectively. These values should be interpreted with caution, as the ideal reference standard on 1934 participants was imperfect. CONCLUSION: The study demonstrates that the public will engage with AF screening undertaken as part of their daily routines using hand-held devices. Sensors can play a key role in identifying asymptomatic patients in this way, but the technology must be further developed to reduce the quantity of non-diagnostic ECGs.

Mid-infrared Doppler-free saturation absorption spectroscopy of the Q branch of CH4ν3 = 1 band using a rapid-scanning continuous-wave optical parametric oscillator.

We have developed a mid-infrared Doppler-free saturation absorption spectroscopy apparatus that employs a commercial continuous-wave optical parametric oscillator (CW OPO), complemented by a home-built automation and wavelength scanning system. Here, we report a comprehensive spectral scan of the Q branch transitions of the ν3 = 1 band of methane (CH4) with an average linewidth (FWHM) of 4.5 MHz. The absolute frequency calibration was achieved using previously reported transition frequencies determined using optical frequency combs, while a Fabry-Perot etalon was used for the relative frequency calibration. We report 15 transitions with improved accuracies of 1.13 MHz (3.76 × 10-5 cm-1).

Methods of identifying surgical Necrotizing Enterocolitis-a systematic review and meta-analysis.

BACKGROUND: Current data suggests potential benefit of earlier surgery for necrotizing enterocolitis (NEC) however this requires accurate prognostication early in the disease course. This study aims to identify and determine the effectiveness of previously reported methods or tests for the identification of surgical NEC. METHODS: Systematic review and meta-analysis with registration on PROSPERO including articles describing a method of identifying surgical NEC. Outcomes of interest were effectiveness and repeatability of index test. RESULTS: Of the 190 full-text articles screened, 90 studies were included which contained 114 methods of identifying surgical NEC in 9546 infants. Of these methods, 44 were a scoring system, 37 a single biomarker, 24 an imaging method, and 9 an invasive method. Sensitivity and specificity ranged from 12.8-100% to 13-100%, respectively. Some methods (9.6%) provided insufficient methods for repeatability within clinical practice or research. Meta-analyses were possible for only 2 methods, the metabolic derangement 7 score and abdominal ultrasound. CONCLUSIONS: A range of methods for identifying surgical NEC have been identified with varying overall performance and uncertainties about reproducibility and superiority of any method. External validation in large multicentre datasets should allow direct comparison of accuracy and prospective study should evaluate impact on clinical outcomes. IMPACT: Earlier identification of need for surgery in necrotizing enterocolitis (NEC) has the potential to improve the unfavourable outcomes in this condition. As such, many methods have been developed and reported to allow earlier identification of surgical NEC. This study is the first synthesis of the literature which identifies previously reported methods and the effectiveness of these. Many methods, including scoring systems and biomarkers, appear effective for prognostication in NEC and external validation is now required in multicentre datasets prior to clinical utility.

Perioperative Dexamethasone in Diabetic Patients: A Systematic Review and Meta-Analysis of Randomized, Placebo-Controlled Trials.

BACKGROUND: The perioperative use of dexamethasone in diabetic patients remains controversial due to concerns related to infection and adverse events. This study aimed to determine whether clinical evidence supports withholding dexamethasone in diabetic patients due to concern for infection risk. We hypothesized that there is no difference in infectious outcomes between dexamethasone-treated patients and controls. METHODS: A literature search was performed on November 22, 2022 to identify randomized, placebo-controlled trials investigating short-course (<72 hours), perioperative dexamethasone that explicitly included diabetic patients and measured at least 1 clinical outcome. Pertinent studies were independently searched in PubMed, Embase, and Cochrane. Authors for all identified studies were contacted with the aim of performing quantitative subgroup analyses of diabetic patients. The primary end point was surgical site infection and the secondary end point was a composite of adverse events. Qualitative remarks were reported based on the total available data and a quality assessment tool. Meta-analyses were performed using inverse variance with random effects. Heterogeneity was assessed via standard χ2 and I2 tests. RESULTS: Sixteen unique studies were included, 5 of which were analyzed quantitatively. Of the 2592 diabetic patients, 2344 (1184 randomized to dexamethasone and 1160 to placebo) were analyzed in at least 1 quantitative outcome. Quantitative analysis showed that the use of perioperative dexamethasone had no effect on the risk of surgical site infections (log odds ratio [LOR], -0.10, 95%; 95% confidence interval [CI], -0.64 to 0.44) while significantly reducing the risk of composite adverse events (LOR, -0.33; 95% CI, -0.62 to -0.05). Qualitative analysis reinforced these findings, demonstrating noninferior to superior results across all clinical outcomes. There was high heterogeneity between the included studies. CONCLUSIONS: Current evidence suggests perioperative dexamethasone may be given to diabetic patients without increasing the risk of infectious complications. Prospective investigations aimed at optimizing dose, frequency, and timing are needed, as well as studies aimed explicitly at exploring the use of dexamethasone in patients with poorly controlled diabetes.

Transcriptional network orchestrating regional patterning of cortical progenitors.

We uncovered a transcription factor (TF) network that regulates cortical regional patterning in radial glial stem cells. Screening the expression of hundreds of TFs in the developing mouse cortex identified 38 TFs that are expressed in gradients in the ventricular zone (VZ). We tested whether their cortical expression was altered in mutant mice with known patterning defects (Emx2, Nr2f1, and Pax6), which enabled us to define a cortical regionalization TF network (CRTFN). To identify genomic programming underlying this network, we performed TF ChIP-seq and chromatin-looping conformation to identify enhancer-gene interactions. To map enhancers involved in regional patterning of cortical progenitors, we performed assays for epigenomic marks and DNA accessibility in VZ cells purified from wild-type and patterning mutant mice. This integrated approach has identified a CRTFN and VZ enhancers involved in cortical regional patterning in the mouse.

Global perspectives on heart disease rehabilitation and secondary prevention: a scientific statement from the Association of Cardiovascular Nursing and Allied Professions, European Association of Preventive Cardiology, and International Council of Cardiovascular Prevention and Rehabilitation.

Cardiovascular disease is a leading cause of death, morbidity, disability, and reduced health-related quality of life, as well as economic burden worldwide, with some 80% of disease burden occurring in the low- and middle-income country (LMIC) settings. With increasing numbers of people living longer with symptomatic disease, the effectiveness and accessibility of secondary preventative and rehabilitative health services have never been more important. Whilst LMICs experience the highest prevalence and mortality rates, the global approach to secondary prevention and cardiac rehabilitation, which mitigates this burden, has traditionally been driven from clinical guidelines emanating from high-income settings. This state-of-the art review provides a contemporary global perspective on cardiac rehabilitation and secondary prevention, contrasting the challenges of and opportunities for high vs. lower income settings. Actionable solutions to overcome system, clinician, programme, and patient level barriers to cardiac rehabilitation access in LMICs are provided.

Dexamethasone-Associated Hyperglycemia is Not Associated With Infectious Complications After Total Joint Arthroplasty in Diabetic Patients.

BACKGROUND: Postoperative infection is a devastating complication of total joint arthroplasty (TJA). Perioperative use of dexamethasone in patients who have diabetes mellitus (DM) remains controversial due to concern for increased infection risk. This study aimed to evaluate the association between dexamethasone and infection risk among patients who have DM undergoing TJA. METHODS: This was a retrospective cohort study conducted on adult patients who underwent primary, elective total knee arthroplasty (TKA) or total hip arthroplasty (THA) between January 2016 and December 2021 using a large national database. We identified 110,568 TJA patients (TKA: 66.6%; THA: 33.4%), 31.0% (34,298) of which had DM. Patients who received perioperative dexamethasone were compared to those who did not. The primary end points were the 90-day risk of postoperative periprosthetic joint infection, surgical site infection (SSI), and other non-SSI (urinary tract infection, pneumonia, sepsis). RESULTS: When modeling the association between dexamethasone exposure and study outcomes while accounting for the interaction between dexamethasone and morning blood glucose levels, dexamethasone administration conferred no increased odds of postoperative periprosthetic joint infection nor SSI in diabetics. However, dexamethasone significantly lowered the adjusted odds of other postoperative infections in diabetic patients (TKA: adjusted odds ratio = 09, 95% confidence interval = 0.8 to 1.0, P = .030; THA: adjusted odds ratio = 0.7, 95% confidence interval = 0.6 to 0.9, P = .001); specifically in patients with morning blood glucose levels between 110 to 248 mg/dL in TKA and ≤ 172 mg/dL in THA. CONCLUSIONS: This study provides strong evidence against withholding dexamethasone in diabetic patients undergoing TJA based on concern for infection. Instead, short-course perioperative dexamethasone reduced infection risk in select patients. The narrative surrounding dexamethasone should shift away from questions about whether dexamethasone is appropriate for diabetic patients, and instead focus on how best to optimize its use.

Protection of Messenger RNA Vaccines Against Hospitalized Coronavirus Disease 2019 in Adults Over the First Year Following Authorization in the United States.

BACKGROUND: Coronavirus disease 2019 (COVID-19) messenger RNA (mRNA) vaccines were authorized in the United States in December 2020. Although vaccine effectiveness (VE) against mild infection declines markedly after several months, limited understanding exists on the long-term durability of protection against COVID-19-associated hospitalization. METHODS: Case-control analysis of adults (≥18 years) hospitalized at 21 hospitals in 18 states 11 March-15 December 2021, including COVID-19 case patients and reverse transcriptase-polymerase chain reaction-negative controls. We included adults who were unvaccinated or vaccinated with 2 doses of a mRNA vaccine before the date of illness onset. VE over time was assessed using logistic regression comparing odds of vaccination in cases versus controls, adjusting for confounders. Models included dichotomous time (<180 vs ≥180 days since dose 2) and continuous time modeled using restricted cubic splines. RESULTS: A total of 10 078 patients were included, 4906 cases (23% vaccinated) and 5172 controls (62% vaccinated). Median age was 60 years (interquartile range, 46-70), 56% were non-Hispanic White, and 81% had ≥1 medical condition. Among immunocompetent adults, VE <180 days was 90% (95% confidence interval [CI], 88-91) versus 82% (95% CI, 79-85) at ≥180 days (P < .001). VE declined for Pfizer-BioNTech (88% to 79%, P < .001) and Moderna (93% to 87%, P < .001) products, for younger adults (18-64 years) (91% to 87%, P = .005), and for adults ≥65 years of age (87% to 78%, P < .001). In models using restricted cubic splines, similar changes were observed. CONCLUSIONS: In a period largely predating Omicron variant circulation, effectiveness of 2 mRNA doses against COVID-19-associated hospitalization was largely sustained through 9 months.

Changing Severity and Epidemiology of Adults Hospitalized With Coronavirus Disease 2019 (COVID-19) in the United States After Introduction of COVID-19 Vaccines, March 2021-August 2022.

INTRODUCTION: Understanding the changing epidemiology of adults hospitalized with coronavirus disease 2019 (COVID-19) informs research priorities and public health policies. METHODS: Among adults (≥18 years) hospitalized with laboratory-confirmed, acute COVID-19 between 11 March 2021, and 31 August 2022 at 21 hospitals in 18 states, those hospitalized during the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron-predominant period (BA.1, BA.2, BA.4/BA.5) were compared to those from earlier Alpha- and Delta-predominant periods. Demographic characteristics, biomarkers within 24 hours of admission, and outcomes, including oxygen support and death, were assessed. RESULTS: Among 9825 patients, median (interquartile range [IQR]) age was 60 years (47-72), 47% were women, and 21% non-Hispanic Black. From the Alpha-predominant period (Mar-Jul 2021; N = 1312) to the Omicron BA.4/BA.5 sublineage-predominant period (Jun-Aug 2022; N = 1307): the percentage of patients who had ≥4 categories of underlying medical conditions increased from 11% to 21%; those vaccinated with at least a primary COVID-19 vaccine series increased from 7% to 67%; those ≥75 years old increased from 11% to 33%; those who did not receive any supplemental oxygen increased from 18% to 42%. Median (IQR) highest C-reactive protein and D-dimer concentration decreased from 42.0 mg/L (9.9-122.0) to 11.5 mg/L (2.7-42.8) and 3.1 mcg/mL (0.8-640.0) to 1.0 mcg/mL (0.5-2.2), respectively. In-hospital death peaked at 12% in the Delta-predominant period and declined to 4% during the BA.4/BA.5-predominant period. CONCLUSIONS: Compared to adults hospitalized during early COVID-19 variant periods, those hospitalized during Omicron-variant COVID-19 were older, had multiple co-morbidities, were more likely to be vaccinated, and less likely to experience severe respiratory disease, systemic inflammation, coagulopathy, and death.

Identification of Host Factors Differentially Induced by Clinically Diverse Strains of Tick-Borne Encephalitis Virus.

Tick-borne encephalitis virus (TBEV) is an important human arthropod-borne virus that causes tick-borne encephalitis (TBE) in humans. TBEV acutely infects the central nervous system (CNS), leading to neurological symptoms of various severity. No therapeutics are currently available for TBEV-associated disease. Virus strains of various pathogenicity have been described, although the basis of their diverse clinical outcome remains undefined. Work with infectious TBEV requires high-level biocontainment, meaning model systems that can recapitulate the virus life cycle are highly sought. Here, we report the generation of a self-replicating, noninfectious TBEV replicon used to study properties of high (Hypr) and low (Vs) pathogenic TBEV isolates. Using a Spinach2 RNA aptamer and luciferase reporter system, we perform the first direct comparison of Hypr and Vs in cell culture. Infectious wild-type (WT) viruses and chimeras of the nonstructural proteins 3 (NS3) and 5 (NS5) were investigated in parallel to validate the replicon data. We show that Hypr replicates to higher levels than Vs in mammalian cells, but not in arthropod cells, and that the basis of these differences map to the NS5 region, encoding the methyltransferase and RNA polymerase. For both Hypr and Vs strains, NS5 and the viral genome localized to intracellular structures typical of positive-strand RNA viruses. Hypr was associated with significant activation of IRF-3, caspase-3, and caspase-8, while Vs activated Akt, affording protection against caspase-mediated apoptosis. Higher activation of stress-granule proteins TIAR and G3BPI were an additional early feature of Vs but not for Hypr. These findings highlight novel host cell responses driven by NS5 that may dictate the differential clinical characteristics of TBEV strains. This highlights the utility of the TBEV replicons for further virological characterization and antiviral drug screening. IMPORTANCE Tick-borne encephalitis virus (TBEV) is an emerging virus of the flavivirus family that is spread by ticks and causes neurological disease of various severity. No specific therapeutic treatments are available for TBE, and control in areas of endemicity is limited to vaccination. The pathology of TBEV ranges from mild to fatal, depending on the virus genotype. Characterization of TBEV isolates is challenging due to the requirement for high-containment facilities. Here, we described the construction of novel TBEV replicons that permit a molecular comparison of TBEV isolates of high and low pathogenicity.

Aberrant glycosylation of anti-SARS-CoV-2 spike IgG is a prothrombotic stimulus for platelets.

A subset of patients with coronavirus disease 2019 (COVID-19) become critically ill, suffering from severe respiratory problems and also increased rates of thrombosis. The causes of thrombosis in severely ill patients with COVID-19 are still emerging, but the coincidence of critical illness with the timing of the onset of adaptive immunity could implicate an excessive immune response. We hypothesized that platelets might be susceptible to activation by anti-severe acute respiratory syndrome coronavirus 2 (anti-SARS-CoV-2) antibodies and might contribute to thrombosis. We found that immune complexes containing recombinant SARS-CoV-2 spike protein and anti-spike immunoglobulin G enhanced platelet-mediated thrombosis on von Willebrand factor in vitro, but only when the glycosylation state of the Fc domain was modified to correspond with the aberrant glycosylation previously identified in patients with severe COVID-19. Furthermore, we found that activation was dependent on FcγRIIA, and we provide in vitro evidence that this pathogenic platelet activation can be counteracted by the therapeutic small molecules R406 (fostamatinib) and ibrutinib, which inhibit tyrosine kinases Syk and Btk, respectively, or by the P2Y12 antagonist cangrelor.

Rare copy number variation in posttraumatic stress disorder.

Posttraumatic stress disorder (PTSD) is a heritable (h2 = 24-71%) psychiatric illness. Copy number variation (CNV) is a form of rare genetic variation that has been implicated in the etiology of psychiatric disorders, but no large-scale investigation of CNV in PTSD has been performed. We present an association study of CNV burden and PTSD symptoms in a sample of 114,383 participants (13,036 cases and 101,347 controls) of European ancestry. CNVs were called using two calling algorithms and intersected to a consensus set. Quality control was performed to remove strong outlier samples. CNVs were examined for association with PTSD within each cohort using linear or logistic regression analysis adjusted for population structure and CNV quality metrics, then inverse variance weighted meta-analyzed across cohorts. We examined the genome-wide total span of CNVs, enrichment of CNVs within specified gene-sets, and CNVs overlapping individual genes and implicated neurodevelopmental regions. The total distance covered by deletions crossing over known neurodevelopmental CNV regions was significant (beta = 0.029, SE = 0.005, P = 6.3 × 10-8). The genome-wide neurodevelopmental CNV burden identified explains 0.034% of the variation in PTSD symptoms. The 15q11.2 BP1-BP2 microdeletion region was significantly associated with PTSD (beta = 0.0206, SE = 0.0056, P = 0.0002). No individual significant genes interrupted by CNV were identified. 22 gene pathways related to the function of the nervous system and brain were significant in pathway analysis (FDR q < 0.05), but these associations were not significant once NDD regions were removed. A larger sample size, better detection methods, and annotated resources of CNV are needed to explore this relationship further.

Robotic colorectal surgery in the emergent diverticulitis setting: is it safe? A review of large national database.

BACKGROUND: As robotic colorectal surgery continues to advance in conjunction with improved recovery protocols, we began implementing robotic surgery (RS) as an option for emergent diverticulitis surgery. Our hospital system utilizes the Da Vinci Xi system, and staff are required to undergo training, making emergent colorectal surgery a feasible option. However, it is essential to determine the safety with reproducibility of our experiences. METHODS: A de-identified retrospective review was performed of Intuitive's national database which obtained data from 262 facilities from January 2018 through December 2021. This identified over 22,000 emergent colorectal surgeries. Of those, over 2500 were performed for diverticulitis in which 126 were RS, 446 laparoscopic surgery (LS), and 1952 open surgery (OS). Clinical outcome metrics including conversion rates, anastomotic leaks, intensive care unit (ICU) admissions, length of stay, mortality, and readmissions were obtained. The cohort was defined by patients who were seen in the emergency department (ED) with diverticulitis and proceeded to have a sigmoid colectomy within 24 h of ED arrival. RESULTS: RS was associated with increased operating time (RS 262, LS 207, OS 182 min), but data has shown many benefits of emergent RS compared to OS. We identified significant decreases in ICU admission rates (OS 19.0%, RS 9.5%, p = 0.01) and anastomotic leak rates (OS 4.4%, RS 0.8%, p = 0.04), with borderline improvement in overall length of stay (OS 9.9, RS 8.9 days, p = 0.05). When compared with LS, RS showed many comparable results. However, RS witnessed a statistically significant improvement in anastomotic leak rates (LS 4.5%, RS 0.8%, p = 0.04). Importantly, there was a striking difference in conversion rates to OS. LS converted over 28.7% of cases to OS, whereas RS only converted 7.9%, p = 0.000005. CONCLUSION: Given these findings, RS is another MIS tool that could be a safe and feasible option for the acute management of emergent diverticulitis.