A study of the educational and business experiences of the St. Patrick High School graduates, Watertown, Massachusetts.

Thesis (M.Ed.)--Boston Universit

AI is a viable alternative to high throughput screening: a 318-target study

: High throughput screening (HTS) is routinely used to identify bioactive small molecules. This requires physical compounds, which limits coverage of accessible chemical space. Computational approaches combined with vast on-demand chemical libraries can access far greater chemical space, provided that the predictive accuracy is sufficient to identify useful molecules. Through the largest and most diverse virtual HTS campaign reported to date, comprising 318 individual projects, we demonstrate that our AtomNet® convolutional neural network successfully finds novel hits across every major therapeutic area and protein class. We address historical limitations of computational screening by demonstrating success for target proteins without known binders, high-quality X-ray crystal structures, or manual cherry-picking of compounds. We show that the molecules selected by the AtomNet® model are novel drug-like scaffolds rather than minor modifications to known bioactive compounds. Our empirical results suggest that computational methods can substantially replace HTS as the first step of small-molecule drug discovery

Safety and efficacy of fluoxetine on functional outcome after acute stroke (AFFINITY): a randomised, double-blind, placebo-controlled trial

Background Trials of fluoxetine for recovery after stroke report conflicting results. The Assessment oF FluoxetINe In sTroke recoverY (AFFINITY) trial aimed to show if daily oral fluoxetine for 6 months after stroke improves functional outcome in an ethnically diverse population. Methods AFFINITY was a randomised, parallel-group, double-blind, placebo-controlled trial done in 43 hospital stroke units in Australia (n=29), New Zealand (four), and Vietnam (ten). Eligible patients were adults (aged ≥18 years) with a clinical diagnosis of acute stroke in the previous 2–15 days, brain imaging consistent with ischaemic or haemorrhagic stroke, and a persisting neurological deficit that produced a modified Rankin Scale (mRS) score of 1 or more. Patients were randomly assigned 1:1 via a web-based system using a minimisation algorithm to once daily, oral fluoxetine 20 mg capsules or matching placebo for 6 months. Patients, carers, investigators, and outcome assessors were masked to the treatment allocation. The primary outcome was functional status, measured by the mRS, at 6 months. The primary analysis was an ordinal logistic regression of the mRS at 6 months, adjusted for minimisation variables. Primary and safety analyses were done according to the patient's treatment allocation. The trial is registered with the Australian New Zealand Clinical Trials Registry, ACTRN12611000774921. Findings Between Jan 11, 2013, and June 30, 2019, 1280 patients were recruited in Australia (n=532), New Zealand (n=42), and Vietnam (n=706), of whom 642 were randomly assigned to fluoxetine and 638 were randomly assigned to placebo. Mean duration of trial treatment was 167 days (SD 48·1). At 6 months, mRS data were available in 624 (97%) patients in the fluoxetine group and 632 (99%) in the placebo group. The distribution of mRS categories was similar in the fluoxetine and placebo groups (adjusted common odds ratio 0·94, 95% CI 0·76–1·15; p=0·53). Compared with patients in the placebo group, patients in the fluoxetine group had more falls (20 [3%] vs seven [1%]; p=0·018), bone fractures (19 [3%] vs six [1%]; p=0·014), and epileptic seizures (ten [2%] vs two [<1%]; p=0·038) at 6 months. Interpretation Oral fluoxetine 20 mg daily for 6 months after acute stroke did not improve functional outcome and increased the risk of falls, bone fractures, and epileptic seizures. These results do not support the use of fluoxetine to improve functional outcome after stroke

Genomic investigations of unexplained acute hepatitis in children

Since its first identification in Scotland, over 1,000 cases of unexplained paediatric hepatitis in children have been reported worldwide, including 278 cases in the UK1. Here we report an investigation of 38 cases, 66 age-matched immunocompetent controls and 21 immunocompromised comparator participants, using a combination of genomic, transcriptomic, proteomic and immunohistochemical methods. We detected high levels of adeno-associated virus 2 (AAV2) DNA in the liver, blood, plasma or stool from 27 of 28 cases. We found low levels of adenovirus (HAdV) and human herpesvirus 6B (HHV-6B) in 23 of 31 and 16 of 23, respectively, of the cases tested. By contrast, AAV2 was infrequently detected and at low titre in the blood or the liver from control children with HAdV, even when profoundly immunosuppressed. AAV2, HAdV and HHV-6 phylogeny excluded the emergence of novel strains in cases. Histological analyses of explanted livers showed enrichment for T cells and B lineage cells. Proteomic comparison of liver tissue from cases and healthy controls identified increased expression of HLA class 2, immunoglobulin variable regions and complement proteins. HAdV and AAV2 proteins were not detected in the livers. Instead, we identified AAV2 DNA complexes reflecting both HAdV-mediated and HHV-6B-mediated replication. We hypothesize that high levels of abnormal AAV2 replication products aided by HAdV and, in severe cases, HHV-6B may have triggered immune-mediated hepatic disease in genetically and immunologically predisposed children

Effect of angiotensin-converting enzyme inhibitor and angiotensin receptor blocker initiation on organ support-free days in patients hospitalized with COVID-19

IMPORTANCE Overactivation of the renin-angiotensin system (RAS) may contribute to poor clinical outcomes in patients with COVID-19. Objective To determine whether angiotensin-converting enzyme (ACE) inhibitor or angiotensin receptor blocker (ARB) initiation improves outcomes in patients hospitalized for COVID-19. DESIGN, SETTING, AND PARTICIPANTS In an ongoing, adaptive platform randomized clinical trial, 721 critically ill and 58 non–critically ill hospitalized adults were randomized to receive an RAS inhibitor or control between March 16, 2021, and February 25, 2022, at 69 sites in 7 countries (final follow-up on June 1, 2022). INTERVENTIONS Patients were randomized to receive open-label initiation of an ACE inhibitor (n = 257), ARB (n = 248), ARB in combination with DMX-200 (a chemokine receptor-2 inhibitor; n = 10), or no RAS inhibitor (control; n = 264) for up to 10 days. MAIN OUTCOMES AND MEASURES The primary outcome was organ support–free days, a composite of hospital survival and days alive without cardiovascular or respiratory organ support through 21 days. The primary analysis was a bayesian cumulative logistic model. Odds ratios (ORs) greater than 1 represent improved outcomes. RESULTS On February 25, 2022, enrollment was discontinued due to safety concerns. Among 679 critically ill patients with available primary outcome data, the median age was 56 years and 239 participants (35.2%) were women. Median (IQR) organ support–free days among critically ill patients was 10 (–1 to 16) in the ACE inhibitor group (n = 231), 8 (–1 to 17) in the ARB group (n = 217), and 12 (0 to 17) in the control group (n = 231) (median adjusted odds ratios of 0.77 [95% bayesian credible interval, 0.58-1.06] for improvement for ACE inhibitor and 0.76 [95% credible interval, 0.56-1.05] for ARB compared with control). The posterior probabilities that ACE inhibitors and ARBs worsened organ support–free days compared with control were 94.9% and 95.4%, respectively. Hospital survival occurred in 166 of 231 critically ill participants (71.9%) in the ACE inhibitor group, 152 of 217 (70.0%) in the ARB group, and 182 of 231 (78.8%) in the control group (posterior probabilities that ACE inhibitor and ARB worsened hospital survival compared with control were 95.3% and 98.1%, respectively). CONCLUSIONS AND RELEVANCE In this trial, among critically ill adults with COVID-19, initiation of an ACE inhibitor or ARB did not improve, and likely worsened, clinical outcomes. TRIAL REGISTRATION ClinicalTrials.gov Identifier: NCT0273570

Enhanced Anxiety and Olfactory Microglial Activation in Early-Stage Familial Alzheimer’s Disease Mouse Model

Anxiety is a known comorbidity and risk factor for conversion to neuroinflammation-mediated dementia in patients with Alzheimer’s disease (AD). Here, we investigated if anxiety occurred as an early endophenotype of mutant familial AD (5 × FAD) male mice and the underlying neuroinflammatory mechanisms. We observed that compared to wildtype (WT) littermates, 5 × FAD mice showed enhanced anxiety at as early as 2 months old (mo). Interestingly, these 5 × FAD male mice had concomitantly increased mRNA levels of pro-inflammatory cytokines such as interleukin 1 beta (Il1b) and tumor necrosis factor (Tnf) in the olfactory bulb (OB) but not the frontal cortex (FC). Increased expression of Tnf in the OB was significantly correlated with the anxious behavior in the FAD but not WT mice. Furthermore, we found more prominent microglial activation and morphological changes in the OB of 2 mo 5 × FAD mice, while only microglial ramification was seen in the FC. To understand if neuroinflammatory changes in the FC could occur at a later stage, we studied 5~6 mo male mice and found that Il1b, interleukin 18 (Il18), and Tnf were upregulated in the FC at this older age. Furthermore, we observed that numbers of microglia and macrophage as well as microglial synaptic pruning, as indicated by phagocytosis of presynaptic component of vesicular glutamate transporter-2, were increased in the OB but not the FC of 5~6 mo 5 × FAD mice. Our findings demonstrated the OB as a more sensitive brain region than the cerebral cortex for microglia-mediated neuroinflammation in association with anxiety in FAD mice and supported the notion that the OB can be an early-stage biomarker in AD

Role of Mitochondrial Dynamics in Neuronal Development: Mechanism for Wolfram Syndrome

International audienceDeficiency of the protein Wolfram syndrome 1 (WFS1) is associated with multiple neurological and psychiatric abnormalities similar to those observed in pathologies showing alterations in mitochondrial dynamics. The aim of this study was to examine the hypothesis that WFS1 deficiency affects neuronal function via mitochondrial abnormalities. We show that down-regulation of WFS1 in neurons leads to dramatic changes in mitochondrial dynamics (inhibited mitochondrial fusion, altered mitochondrial trafficking, and augmented mitophagy), delaying neuronal development. WFS1 deficiency induces endoplasmic reticulum (ER) stress, leading to inositol 1,4,5-trisphosphate receptor (IP3R) dysfunction and disturbed cytosolic Ca2+ homeostasis, which, in turn, alters mitochondrial dynamics. Importantly, ER stress, impaired Ca2+ homeostasis, altered mitochondrial dynamics, and delayed neuronal development are causatively related events because interventions at all these levels improved the downstream processes. Our data shed light on the mechanisms of neuronal abnormalities in Wolfram syndrome and point out potential therapeutic targets. This work may have broader implications for understanding the role of mitochondrial dynamics in neuropsychiatric diseases

WFS1 deficiency decreases mitochondrial membrane potential and cytosolic ATP level.

<p>(A) Primary cortical neurons were transfected with control or <i>Wfs1</i> siRNA using the N-TER nanoparticle siRNA transfection system and stained with JC-10, which emits light from 525 nm to 590 nm, depending on mitochondrial membrane potential. Values shown are corrected by subtracting the values obtained in the presence of FCCP incubation (5 μM). The red to green fluorescence ratio demonstrated a slight but significant decrease in the <i>Wfs1</i> siRNA group. (B) Neurons were transfected with plasmids expressing scrambled shRNA or <i>Wfs1</i> shRNA, firefly luciferase construct containing NRF2 binding site, and <i>Renilla</i> luciferase. Firefly luciferase signal normalized to <i>Renilla</i> signal demonstrates no change in NRF2 activity. NRF2 overexpression-induced reporter activity was used as a positive control. (C) Neurons transfected with the ATP sensor ATeam and treated with 2-deoxyglucose (12 mM)/oligomycin (2.5 μM) or glutamate (2 mM) (both used as positive controls) show a decrease in relative cytosolic ATP levels. (D) Neurons were transfected with the ATP sensor ATeam and scrambled or <i>Wfs1</i> shRNAs. WFS1-deficient neurons show a lower cytosolic ATP level as compared to control. **<i>p</i> < 0.01 and ***<i>p</i> < 0.001 compared with respective control group. Underlying data is shown in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1002511#pbio.1002511.s001" target="_blank">S1 Data</a>.</p

WFS1 deficiency impairs mitochondrial dynamics.

<p>(A) Primary cortical neurons were transfected with the photoconvertible mitochondrially targeted construct mito-Kikume-Green and scrambled shRNA or <i>Wfs1</i> shRNA. Selected mitochondria were irradiated using a 405-nm laser line, thereby converting mito-Kikume-Green into mito-Kikume-Red. Fusion events between mito-Kikume-Green and photoactivated mito-Kikume-Red mitochondria are visible when mitochondria become yellow after mixing of the contents of the red and green mitochondria. (B–E) In primary cortical neurons, <i>Wfs1</i> shRNA significantly decreases fusion rate (B) and mitochondrial length (C). These parameters are restored by overexpression of wild-type (wt) WFS1 but not by P724L WFS1, a mutant found in Wolfram syndrome. Similar changes are observed in cortical neurons isolated from <i>Wfs1</i>^-/- mice. The lower fusion rate (D) and reduced mitochondrial length (E) in <i>Wfs</i>1^-/- neurons is restored by wt WFS1 overexpression, but <i>Wfs1</i> shRNA has no effect on these parameters. (F–H) Primary cortical neurons were transfected with mitochondrially targeted Keima (which changes its excitation spectrum under acidic conditions) and scrambled shRNA or <i>Wfs1</i> shRNA (F). The number of autolysosomes containing mitochondria increases in <i>Wfs1</i>-silenced neurons (G) and in neurons isolated from <i>Wfs1</i>^-/- mice (H). (I–K) Representative images of mitochondrial morphology and density in the axons of scrambled- and <i>Wfs1</i>-shRNA transfected neurons (I). The density of axonal mitochondria is reduced in <i>Wfs1</i>-silenced neurons (J) and in neurons isolated from <i>Wfs1</i>^-/- mice (K). This parameter is restored by wt WFS1 overexpression, but <i>Wfs1</i> shRNA has no effect. *<i>p</i> < 0.05, **<i>p</i> < 0.01, and ***<i>p</i> < 0.001 compared with respective control groups. Underlying data is shown in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1002511#pbio.1002511.s001" target="_blank">S1 Data</a>.</p

WFS1 deficiency leads to impaired neuronal development.

<p>Primary cortical neurons were transfected with the neuronal marker pAAV-hSyn-DsRed1 and scrambled shRNA or <i>Wfs1</i> shRNA at DIV (day in vitro) 1, and neuronal morphology was assessed at different time points. (A) Cell morphology at different stages of development. (B) Morphological analysis demonstrating the retarded development of WFS1-deficient neurons. (C) Examples of reconstructed control and WFS1-deficient neurons at different DIV. (D–F) WFS1 deficiency retards growth of the longest axon (D) and growth of the axonal tree (E) and decreases the number of axon tips (F). (G) Survival of WFS1-deficient neurons is decreased when compared with control neurons (<i>n</i> = 61–62 individual dishes from 17 independent sister cultures at DIV 6–11). (H) Visualisation of synapses (red) using an antibody targeted against the post-synaptic marker PSD-95 in neurons transfected with GFP (green). The right panel shows a zoomed image. (I) WFS1-deficiency decreases synaptic density at late stages. *<i>p</i> < 0.05, **<i>p</i> < 0.01, and ***<i>p</i> < 0.001 compared with respective scrambled shRNA-treated groups. Underlying data is shown in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.1002511#pbio.1002511.s001" target="_blank">S1 Data</a>.</p