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Vibrio vulnificuscauses life threatening infections dependent upon the effectors released from the Multifunctional-Autoprocessing Repeats-In-Toxin (MARTX) toxin. The Makes Caterpillars Floppy-like (MCF) cysteine protease effector is activated by host ADP ribosylation factors (ARFs), although the targets of processing activity were unknown. In this study we show MCF bindsRas-related proteins inbrain (Rab) GTPases at the same interface occupied by ARFs and then cleaves and/or degrades 24 distinct members of the Rab GTPases family. The cleavage occurs in the C-terminal tails of Rabs. We determine the crystal structure of MCF as a swapped dimer revealing the open, activated state of MCF and then use structure prediction algorithms to show that structural composition, rather than sequence or localization, determine Rabs selected as MCF proteolytic targets. Once cleaved, Rabs become dispersed in cells to drive organelle damage and cell death to promote pathogenesis of these rapidly fatal infections.

Figure S1. AKT and ERK activity following increasing doses of BGJ398, GDC-0941, BYL719 and BKM120 in AN3CA and JHUEM2 cells.

Supplementary Figure Legends 1-4.

Seizures are classically described as an epiphenomenon of hyperexcitability and hypersynchronicity across brain regions. However, this view is insufficient to explain the complex, dynamic evolution of focal-onset seizures in the brain. Recent studies have proposed mechanisms involving an evolution of excitability driven specifically by a spatiotemporally progressing seizure wavefront. These mechanisms attempt to align the abnormal propagation of neural activity with well-known neurobiological parameters, such as excitation-inhibition balance and neuronal connectivity patterns. We describe a direct test of these mechanisms by performing real-time,in vivoinvestigations of excitability in the acutely epileptic state and during seizure propagation. We used all-optical interrogation to test single-neuronal and local-circuit excitability in the epileptic brain. We demonstrate a surprising paradox during the acutely epileptic state, wherein the brain becomes susceptible to large synchronous inputs, yet single-cell excitability is largely maintained at baseline levels. At a finer scale, excitability of neurons at the single-cell level is related to their distance from the seizure wavefront. Local circuit excitability is increased in the distal penumbra but, crucially, we find inhibition in close proximity to the seizure wavefront. This is in contrast with previously suggested notions of widespread inhibition outside the direct area of action during a focal-onset seizure. These experimental results provide the first direct,in vivoevidence for the precise spatial scale over which single-cell excitability dynamics evolve during seizure propagation, providing support for local inhibitory restraint of seizure propagation.

Traumatic brain injury (TBI) is associated with chronic psychiatric complications and increased risk for development of neurodegenerative pathology. Aged individuals account for most TBI-related hospitalizations and deaths. Nonetheless, neurobiological mechanisms that underlie worsened functional outcomes after TBI in the elderly remain unclear. Therefore, this study aimed to identify pathways that govern differential responses to TBI with age. Here, adult (2 months of age) and aged (16–18 months of age) male C57BL/6 mice were subjected to diffuse brain injury (midline fluid percussion), and cognition, gliosis, and neuroinflammation were determined 7 or 30 d postinjury (dpi). Cognitive impairment was evident 7 dpi, independent of age. There was enhanced morphologic restructuring of microglia and astrocytes 7 dpi in the cortex and hippocampus of aged mice compared with adults. Transcriptional analysis revealed robust age-dependent amplification of cytokine/chemokine, complement, innate immune, and interferon-associated inflammatory gene expression in the cortex 7 dpi. Ingenuity pathway analysis of the transcriptional data showed that type I interferon (IFN) signaling was significantly enhanced in the aged brain after TBI compared with adults. Age prolonged inflammatory signaling and microgliosis 30 dpi with an increased presence of rod microglia. Based on these results, a STING (stimulator of interferon genes) agonist, DMXAA, was used to determine whether augmenting IFN signaling worsened cortical inflammation and gliosis after TBI. DMXAA-treated Adult-TBI mice showed comparable expression of myriad genes that were overexpressed in the cortex of Aged-TBI mice, includingIrf7,Clec7a,Cxcl10, andCcl5. Overall, diffuse TBI promoted amplified IFN signaling in aged mice, resulting in extended inflammation and gliosis.SIGNIFICANCE STATEMENTElderly individuals are at higher risk of complications following traumatic brain injury (TBI). Individuals >70 years old have the highest rates of TBI-related hospitalization, neurodegenerative pathology, and death. Although inflammation has been linked with poor outcomes in aging, the specific biological pathways driving worsened outcomes after TBI in aging remain undefined. In this study, we identify amplified interferon-associated inflammation and gliosis in aged mice following TBI that was associated with persistent inflammatory gene expression and microglial morphologic diversity 30 dpi. STING (stimulator of interferon genes) agonist DMXAA was used to demonstrate a causal link between augmented interferon signaling and worsened neuroinflammation after TBI. Therefore, interferon signaling may represent a therapeutic target to reduce inflammation-associated complications following TBI.

Background Estimated glomerular filtration rate (eGFR) is a key determinant of eligibility for many life-saving therapies in HFrEF. Recently, the Chronic Kidney Disease Epidemiology Collaboration (CKD-EPI) provided new equations based on creatinine (CKD-EPIcr), cystatin C (CKD-EPIcys) or both (CKD-EPIcyscr) that do not include race. These new equations may reclassify individuals, irrespective of race, from one eGFR category to another, with implications for eligibility for HFrEF treatments. Purpose To assess the difference between eGFR estimation using the 2021 CKD-EPIcyscr equation and the 2009 CKD-EPIcr and Modification of Diet in Renal Disease Study (MDRD)-4 equations which are still standard in many European laboratories. Methods We included patients from PARADIGM-HF with cystatin C and creatinine values available at the time of randomization. For each patient, baseline eGFRs were calculated using the 3 equations described. Our focus was on patients with chronic kidney disease (CKD) stages III–V. Results Overall, 1910 patients were eligible. Mean age was 67.3 (10.1) year and 385 (18.7%) were female. Using 2009 CKD-EPIcr, 779 patients were in CKD stages 3–5, of which 233 (30%) were reclassified to a better CKD stage (higher eGFR) with the 2021 CKD-EPIcyscr equation (Table 1). Similar reclassification was seen when comparing MDRD-4 with the 2021 CKD-EPIcyscr equation: 277 (33%) of 831 patients in CKD stages 3–5 were reclassified to a better CKD stage (Figure 1). Conclusions The 2021 CKD-EPIcyscr equation favourably reclassified CKD stage in a large percentage of patients with HFrEF and a low eGFR, potentially increasing the proportion of these patients considered eligible for guideline-recommended therapies. Funding Acknowledgement Type of funding sources: None.

Background and objective High resting heart rate (HR) associates with cardiovascular death (CVD) and heart failure hospitalisation (HFH) in patients with reduced ejection fraction (HFrEF), but data are sparse in patients with preserved (HFpEF) or mildly reduced (HFmrEF) ejection fraction. Empagliflozin reduced the risk of CVD and HFH in HFpEF in the EMPEROR-Preserved trial. This study analyses the influence of HR on outcomes in patients with left ventricular ejection fraction (LVEF) >40% in EMPEROR-Preserved and evaluates the effects of empagliflozin across HR categories. Methods Patients (n=5988) with HFpEF (LVEF >40%) were categorised to HR 75 bpm. The composite of CVD or HFH (primary outcome), first HFH, CVD, recurrent HFH and all-cause mortality were studied in the HR groups and in patients separated by sinus rhythm (SR) or atrial fibrillation (AF) and true HFpEF (EF ≥50%) or HFmrEF (EF 40–49%). Results Empagliflozin did not influence HR over time. At HR >75 bpm, the primary outcome (hazard ratio: 1.31, 1.13–1.52, p=0.0003), time to first HFH (hazard ratio: 1.25, 1.04–1.49, p=0.02), recurrent HFH (hazard ratio: 1.29, 1.05–1.60, p=0.02), CVD (hazard ratio: 1.49, 1.21–1.84, p=0.0001) and all-cause mortality (hazard ratio: 1.49, 1.28–1.73, p Conclusions HR in SR, but not in AF, predicts heart failure outcomes in HFpEF and HFmrEF, but the effects of empagliflozin were not modified by HR. Funding Acknowledgement Type of funding sources: Private company. Main funding source(s): Boehringer Ingelheim & Eli Lilly and Company Diabetes Alliance

Introduction Bilirubin is a recognized predictor of adverse outcomes in patients with heart failure and reduced ejection fraction (HFrEF), possibly because it is a marker of congestion. Alkaline phosphatase (ALP) is an enzyme produced in many tissues including the biliary ducts and elevated levels are also associated with congestion. Purpose To examine the prognostic value of ALP alone and in combination with bilirubin in patients with HFrEF. Methods The study population was ambulatory patients with HFrEF enrolled in 2 recent clinical trials with similar inclusion and exclusion criteria: ATMOSPHERE (derivation cohort) and PARADIGM-HF (validation). Cut points to define elevated bilirubin and alkaline phosphatase were >17mg/dL and >120 U/L respectively. The composite of cardiovascular death or HF hospitalization, its components, and all-cause death related to elevation of one, other or both of bilirubin and ALP was examined using Cox regression. Univariable and multivariable models with adjustment for other recognized prognostic variables including NT-proBNP were analyzed. Results Of 7016 patients with HFrEF enrolled in ATMOSPHERE, 6870 had a measurement of both bilirubin and ALP at baseline: mean age 63 years, 22% women, mean LVEF 28% and proportion NYHA class III/IV 37%. Bilirubin and ALP were both normal in 4810 (70.0%) patients, bilirubin was elevated in 1393 (20.3%), ALP was elevated in 360 (5.2%) and both were elevated in 307 (4.5%) patients. Patients with elevation of both ALP and bilirubin were older, had lower systolic blood pressure, higher heart rate, higher NT-pro BNP, more clinical features of congestion, more atrial fibrillation and a greater proportion were treated with diuretics and digoxin. The primary endpoint rates (per 100 person-years) were 10.4 (95% CI 9.9–11.0) when both markers were normal, 15.1 (13.9–16.4) when bilirubin was elevated, 12.4 (10.4–14.9) when alkaline phosphatase was elevated, and 25.6 (22.0–29.9) when both markers were elevated (Figure 1). The adjusted hazard ratios (95% CI) were (both biomarkers normal = reference): elevated bilirubin 1.19 (1.07–1.31), P=0.001; elevated ALP 1.03 (0.84–1.26), P=0.81; both elevated 1.45 (1.21–1.73), P Conclusions Elevation of ALP in combination with elevated bilirubin identifies a small group of patients at very high risk of adverse outcomes. This may reflect more significant congestion. ALP and bilirubin, inexpensive and routinely measured biochemical tests, are useful prognostic markers in patients with HFrEF. Funding Acknowledgement Type of funding sources: Private grant(s) and/or Sponsorship.

Background Although multiple novel biomarkers have individually been shown to predict outcomes in patients with HFrEF, the value of these over and above conventional clinical and laboratory variables, plus natriuretic peptides, is uncertain. Purpose To test the incremental predictive value of 11 novel biomarkers added to a recent prognostic model 1 (PREDICT-HF) derived in PARADIGM-HF and validated in ATMOSPHERE and the Swedish heart failure registry. The PREDICT-HF model includes clinical variables, standard laboratory variables, and BNP or NT-proBNP. Methods 1559 participants enrolled in PARADIGM-HF had all 11 biomarkers of interest measured. These reflected different pathophysiological pathways: (i) myocyte injury (high sensitivity cardiac troponin T), (ii) cardiac remodelling and inflammation (growth stimulation expressed gene 2, growth differentiation factor-15 and galectin-3), (iii) extracellular matrix remodelling (matrix metalloproteinase-2, matrix metalloproteinase-9, tissue inhibitor of metalloproteinase-1), (iv) neurohormonal pathways (aldosterone) and (v) renal dysfunction and injury (cystatin C, kidney injury molecule-1 and urinary albumin to creatinine ratio). The incremental prognostic value of these biomarkers was evaluated using Harrell's C statistic. Results The mean age of participants studied was 67.3 (SD 9.9) years, 1254 (80%) were men and 1103 (71%) were in NYHA class II. During a median follow-up of 31 months, 197 patients died and 300 experienced the primary composite outcome (cardiovascular death or heart failure hospitalization). When each candidate biomarker (log unit) was added individually to the PREDICT-HF base model, GDF-15, ST2, TIMP1, cystatin C, hsTnT and UACR were independent predictors of all-cause mortality (Table 1). GDF-15, TIMP1, hs-TnT and cystatin C consistently increased the risk of both all-cause mortality and the primary outcome. Individuals who had all 4 biomarkers elevated (compared to none elevated) had the highest risk: HR for all-cause mortality 3.65 (2.01–6.64), p Conclusion Several novel biomarkers provide meaningful additional prognostic information in patients with HFrEF. A multimarker approach incorporating biomarkers reflecting different pathophysiological pathways added most information. This approach may be useful in refining risk and targeting treatment. Funding Acknowledgement Type of funding sources: Private grant(s) and/or Sponsorship. Main funding source(s): The PARADIGM-HF trial was funded by Novartis.J.J.V.M is supported by a British Heart Foundation Centre of Excellence Grant

Background Though current heart failure (HF) biomarkers are highly prognostic, systematically characterizing associations between circulating proteins and risk of subsequent events may improve clinical risk prediction and illuminate new biological pathways. Large-scale assays measuring thousands of proteins now enable unbiased proteomic investigation in clinical trials. Purpose To identify and replicate serum proteins associated with HF events in patients with chronic HF with reduced ejection fraction (HFrEF), and to develop and validate a proteomic risk score. Methods Serum levels of 4076 proteins were measured at baseline in the ATMOSPHERE (n=1261, 487 events over 6 years) and PARADIGM-HF (n=1257, 287 events over 4 years) trials of chronic HFrEF using a modified aptamer-based proteomics assay. Proteins associated with the primary endpoint, HF hospitalization or cardiovascular death, were identified in the ATMOSPHERE discovery cohort (false discovery rate Results We identified 377 serum proteins associated with the primary endpoint in ATMOSPHERE and replicated 167 in PARADIGM-HF. Prognostic proteins included known HF biomarkers Growth Differentiation Factor 15, NT-proBNP, and Angiopoietin-2, and also a previously unrecognized HF biomarker: Sushi, Von Willebrand Factor Type A, EGF And Pentraxin Domain Containing 1 (SVEP1) (HR 1.60 [95% CI 1.44–1.79] per standard deviation [SD], p=2x10–17) (Table 1). Proteins related to hepatic fibrosis, granulocyte adhesion, and inhibition of matrix metalloproteinases were over-represented. A 64-protein risk score derived in ATMOSPHERE predicted clinical events in PARADIGM-HF with greater discrimination (c-statistic 0.70) than the MAGGIC clinical score (c-statistic 0.61), NT-proBNP (c-statistic 0.65), or both (c-statistic 0.66) (Figure 1). Genetically predicted levels of NT-proBNP, WISP2, FSTL1, and CTSS were associated with the primary endpoint by Mendelian randomization. Conclusions We identify SVEP1, an extracellular matrix protein known to cause inflammation in vascular smooth muscle cells, as a previously unrecognized HF biomarker. A 64-protein score improved risk discrimination compared with NT-proBNP and may assist in identifying high-risk patients for clinical trials or disease management programs. Funding Acknowledgement Type of funding sources: Private company. Main funding source(s): The ATMOSPHERE and PARADIGM-HF trials were sponsored by Novartis

Background Anaemia is common in patients with HFrEF and is associated with poor clinical outcomes. Although they reduce rates of mortality and heart failure hospitalization, renin-angiotensin (RAS) blockers lower haemoglobin and may induce anaemia. Concomitant neprilysin inhibition might ameliorate this effect of RAS blockers. Purpose We investigated the effect of sacubitril/valsartan compared with enalapril on clinical outcomes, according to anaemia status, and on haemoglobin levels in PARADIGM-HF. Methods Patient characteristics and clinical outcomes were compared between patients with and without anaemia (defined as haemoglobin Results Of 8239 participants with a baseline haemoglobin measurement, 1677 (20.4%) were anaemic. Compared to those without anaemia, patients with anaemia had a more severe heart failure profile, worse kidney function, greater neurohormonal derangement and worse clinical outcomes. Sacubitril/valsartan, compared to enalapril, reduced the risk of the primary endpoint similarly in patients with anaemia (HR 0.84, 95% CI 0.71–1.00) and without anaemia (HR 0.78, 95% CI 0.71–0.87), p-value for interaction=0.478. Between baseline and 12 months, haemoglobin decreased by 1.5 (95% CI 1.7 to 1.2) g/L with sacubitril/valsartan compared with 2.3 (2.6 to 2.0) g/L with enalapril group: mean difference 0.8 (95% CI 0.5 to 1.2) g/L, p Conclusions Compared to enalapril, sacubitril/valsartan reduced mortality and hospitalization in HFrEF patients with and without anaemia. Haemoglobin decreased less with sacubitril/valsartan and the incidence of new anaemia was lower in the sacubitril/valsartan group compared with the enalapril group. Funding Acknowledgement Type of funding sources: Private company. Main funding source(s): PARADIGM-HF was funded by Novartis.

To ensure that the science, technology, engineering, and mathematics (STEM) workforce of the future is inclusive of diverse peoples and ideas, there is a need to ensure that STEM outreach activities that encourage students into STEM education and careers are inclusive of and provide equity for people from diverse racial/ethnic, socioeconomic, and academic backgrounds. Since 2004, the National Indigenous Science Education Program (NISEP), based at Macquarie University, Sydney, has been using best ethical and inclusion strategies to engage with Australian Aboriginal and Torres Strait Islander school students and their communities. This publication discusses the inclusion and equity strategies utilized by NISEP, including community consultation, student leadership and autonomy, volunteer mentors, and the respectful and best practice use of Indigenous knowledge. These factors have all contributed to NISEP's success in increasing Aboriginal and Torres Strait Islander students' aspiration toward and engagement with secondary and tertiary education.

Resume La phacoemulsification est la chirurgie la plus frequemment pratiquee dans le monde. Depuis quelques annees, cette chirurgie semble avoir atteint un plateau et qu’il n’y a plus d’innovations de rupture. Dans cet article, nous faisons le point sur les techniques alternatives, les dernieres innovations et les voies de recherche et developpement dans ce domaine.

Contains fulltext : 249977.pdf (Publisher’s version ) (Closed access) BACKGROUND: It is important to evaluate whether a new treatment for heart failure with reduced ejection fraction (HFrEF) provides additive benefit to background foundational treatments. As such, we aimed to evaluate the efficacy and safety of empagliflozin in patients with HFrEF in addition to baseline treatment with specific doses and combinations of disease-modifying therapies. METHODS: We performed a post-hoc analysis of the EMPEROR-Reduced randomised, double-blind, parallel-group trial, which took place in 520 centres (hospitals and medical clinics) in 20 countries in Asia, Australia, Europe, North America, and South America. Patients with New York Heart Association (NYHA) classification II-IV with an ejection fraction of 40% or less were randomly assigned (1:1) to receive the addition of either oral empagliflozin 10 mg per day or placebo to background therapy. The primary composite outcome was cardiovascular death and heart failure hospitalisation; the secondary outcome was total heart failure hospital admissions. An extended composite outcome consisted of inpatient and outpatient HFrEF events was also evaluated. Outcomes were analysed according to background use of angiotensin-converting enzyme (ACE) inhibitors, angiotensin II receptor blockers (ARBs) or angiotensin receptor neprilysin inhibitors (ARNIs), as well as β blockers and mineralocorticoid receptor antagonists (MRAs) at less than 50% or 50% or more of target doses and in various combinations. This study is registered with ClinicalTrials.gov, NCT03057977. FINDINGS: In this post-hoc analysis of 3730 patients (mean age 66·8 years [SD 11·0], 893 [23·9%] women; 1863 [49·9%] in the empagliflozin group, 1867 [50·1%] in the placebo group) assessed between March 6, 2017, and May 28, 2020, empagliflozin reduced the risk of the primary outcome (361 in 1863 participants in the empagliflozin group and 462 of 1867 in the placebo group; HR 0·75 [95% CI 0·65-0·86]) regardless of background therapy or its target doses for ACE inhibitors or ARBs at doses of less than 50% of the target dose (HR 0·85 [0·69-1·06]) and for doses of 50% or more of the target dose (HR 0·67 [0·52-0·88]; p(interaction)=0·18). A similar result was seen for β blockers at doses of less than 50% of the target dose (HR 0·66 [0·54-0·80]) and for doses of 50% or more of the target dose (HR 0·81 [0·66-1·00]; p(interaction)=0·15). Empagliflozin also reduced the risk of the primary outcome irrespective of background use of triple therapy with an ACE inhibitor, ARB, or ARNI plus β blocker plus MRA (given combination HR 0·73 [0·61-0·88]; not given combination HR 0·76 [0·62-0·94]; p(interaction)=0·77). Similar patterns of benefit were observed for the secondary and extended composite outcomes. Empagliflozin was well tolerated and rates of hypotension, symptomatic hypotension, and hyperkalaemia were similar across all subgroups. INTERPRETATION: Empagliflozin reduced serious heart failure outcomes across doses and combinations of disease-modifying therapies for HFrEF. Clinically, these data suggest that empagliflozin might be considered as a foundational therapy in patients with HFrEF regardless of their existing background therapy. FUNDING: Boehringer Ingelheim and Eli Lilly and Company.

The claustrum is highly interconnected with many structures in the brain, but the organizing principles governing its vast connectivity have yet to be fully explored. We investigated the defining characteristics and activity of single claustrum neurons, the nature of their relationship with the cortex, and their connectivity within the claustrum itself to gain a comprehensive view of claustral circuits. We show that the claustrum is composed of excitatory and inhibitory cell types that are connected through intraclaustral excitatory synapses, especially between neurons with disparate projection targets. Investigations of corticoclaustral innervation demonstrated that axons from the cortex localize to distinct dorsoventral modules depending on their region of origin.In vitrodual-color optogenetic mapping experiments revealed that individual claustrum neurons integrated inputs from more than one cortical region in a cell type- and projection target-specific manner. Integration in claustrum neurons was observed to be particularly common between areas of the frontal cortex and less so for sensory, motor, and association cortices. Finally, activity in claustrum axons in midline cortical areas recorded within vivotwo-photon calcium imaging showed responses to multimodal sensory stimuli. Our findings shed light on the organizing principles of claustrum circuits, demonstrating that individual claustrum neurons integrate cortical inputs and redistribute this information back to cortex after performing output target- and cell type-dependent local computations.

The brains of higher organisms are composed of anatomically and functionally distinct regions performing specialised tasks; but regions do not operate in isolation. Orchestration of complex behaviours requires communication between brain regions, but how neural activity dynamics are organised to facilitate reliable transmission is not well understood. We studied this process directly by generating neural activity that propagates between brain regions and drives behaviour, allowing us to assess how populations of neurons in sensory cortex cooperate to transmit information. We achieved this by imaging two hierarchically organised and densely interconnected regions, the primary and secondary somatosensory cortex (S1 and S2) in mice while performing two-photon photostimulation of S1 neurons and assigning behavioural salience to the photostimulation. We found that the probability of perception is determined not only by the strength of the photostimulation signal, but also by the variability of S1 neural activity. Therefore, maximising the signal-to-noise ratio of the stimulus representation in cortex relative to the noise or variability in cortex is critical to facilitate activity propagation and perception. Further, we show that propagated, behaviourally salient activity elicits balanced, persistent, and generalised activation of the downstream region. Hence, our work adds to existing understanding of cortical function by identifying how population activity is formatted to ensure robust transmission of information, allowing specialised brain regions to communicate and coordinate behaviour.

The accurate control of magnetic fields is a cornerstone of multiple emerging quantum technologies. These technologies often require passive high permeability magnetic shielding and internal active field-generating coils to create their own bespoke magnetic field landscape. However, magnetic fields generated by coils are distorted by high permeability shielding, preventing the accurate and efficient generation of the desired field environment. Here, we design a cylindrical four-layer magnetic shield with an interior hybrid active-passive coil system that is explicitly optimised to include the electromagnetic coupling between the active and passive components. We use a combination of analytical methods and numerical simulations to determine the shield parameters - geometry, thickness, and access hole positions - to maximise the passive shielding efficiency and minimise the shield-induced Johnson noise and weight. Then, we apply an analytical formulation of the magnetic field, which accounts for the interaction with the magnetic shield, to design nine orthogonal hybrid active-passive field-generating coils inside the shield. The coils will be manufactured on thin low-via flex-PCBs near the shield's interior surface and generate three uniform fields and six gradient fields that deviate by less than 0.4% and 2%, respectively, over an internal cylindrical region extending over half the diameter and length of the innermost shield layer. These hybrid active-passive coils can accurately remove deviations in the background field or generate various complex magnetic field landscapes. Consequently, the hybrid shield provides an ideal platform for miniaturising and commercialising quantum technologies that require precisely-controlled magnetic fields within a low-noise environment.

Objectives/AimsThe claustrum is a sheet-like bilateral brain region whose function remains unknown. We comprehensively examined cases of human claustrum lesions to assess their support for various hypotheses of function. To do so we searched the following terms on PubMed and Scopus: ‘claustrum AND (lesion OR contusion OR in- jury OR trauma)’.ResultsIn total our search uncovered 103 cases, which were then screened for confirmed damage to the claustrum by neuroimaging. Thirty-eight individual cases and 14 cohort studies were included. Our results suggest human claustral lesions do not selectively impair a singular domain. Instead, they argue that human claustrum may have a more general function. The high incidence of seizures following claustral lesions suggests this may involve maintenance of excitation-inhibition balance.ConclusionsThe wide range of symptoms observed following claustral lesions do not provide compelling evidence to support prominent current theories of claustrum function such as spatial navigation, attentional allocation, cognition, multisensory integration, and salience computation. Conversely, the lesions studies support the hypothesis that the claustrum regulates cortical excitability.

Recent advances combining two-photon calcium imaging and two-photon optogenetics with digital holography now allow us to read and write neural activity in vivo at cellular resolution with millisecond temporal precision. Such “all-optical” techniques enable experimenters to probe the impact of functionally defined neurons on neural circuit function and behavioural output with new levels of precision. This protocol describes the experimental strategy and workflow for successful completion of typical all-optical interrogation experiments in awake, behaving head-fixed mice. We describe modular procedures for the setup and calibration of an all-optical system, the preparation of an indicator and opsin-expressing and task-performing animal, the characterization of functional and photostimulation responses and the design and implementation of an all-optical experiment. We discuss optimizations for efficiently selecting and targeting neuronal ensembles for photostimulation sequences, as well as generating photostimulation response maps from the imaging data that can be used to examine the impact of photostimulation on the local circuit. We demonstrate the utility of this strategy using all-optical experiments in three different brain areas – barrel cortex, visual cortex and hippocampus – using different experimental setups. This approach can in principle be adapted to any brain area for all-optical interrogation experiments to probe functional connectivity in neural circuits and for investigating the relationship between neural circuit activity and behaviour.

Recent advances combining two-photon calcium imaging and two-photon optogenetics with computer-generated holography now allow us to read and write the activity of large populations of neurons in vivo at cellular resolution and with high temporal resolution. Such 'all-optical' techniques enable experimenters to probe the effects of functionally defined neurons on neural circuit function and behavioral output with new levels of precision. This greatly increases flexibility, resolution, targeting specificity and throughput compared with alternative approaches based on electrophysiology and/or one-photon optogenetics and can interrogate larger and more densely labeled populations of neurons than current voltage imaging-based implementations. This protocol describes the experimental workflow for all-optical interrogation experiments in awake, behaving head-fixed mice. We describe modular procedures for the setup and calibration of an all-optical system (~3 h), the preparation of an indicator and opsin-expressing and task-performing animal (~3-6 weeks), the characterization of functional and photostimulation responses (~2 h per field of view) and the design and implementation of an all-optical experiment (achievable within the timescale of a normal behavioral experiment; ~3-5 h per field of view). We discuss optimizations for efficiently selecting and targeting neuronal ensembles for photostimulation sequences, as well as generating photostimulation response maps from the imaging data that can be used to examine the impact of photostimulation on the local circuit. We demonstrate the utility of this strategy in three brain areas by using different experimental setups. This approach can in principle be adapted to any brain area to probe functional connectivity in neural circuits and investigate the relationship between neural circuit activity and behavior.

Hybrid magnetic shields with both active field generating components and high-permeability magnetic shielding are increasingly needed for a variety of technologies and experiments that require precision-controlled magnetic field environments. However, the fields generated by the active components interact with the passive magnetic shield, distorting the desired field profiles. Consequently, optimization of the active components needed to generate user-specified target fields must include coupling to the high-permeability passive components. Here, we consider the optimization of planar active systems, on which an arbitrary static current flows, coupled to a closed high-permeability cylindrical shield. We modify the Green's function for the magnetic vector potential to match boundary conditions on the shield's interior surface, enabling us to construct an inverse optimization problem to design planar coils that generate user-specified magnetic fields inside high-permeability shields. We validate our methodology by designing two bi-planar hybrid active--passive systems, which generate a constant transverse field, $\mathbf{B}=\mathbf{\hat{x}}$, and a linear field gradient, $\mathbf{B}=(-x~\mathbf{\hat{x}}-y~\mathbf{\hat{y}}+2z~\mathbf{\hat{z}})$, respectively. For both systems, the inverse-optimized magnetic field profiles agree well with forward numerical simulations. Our design methodology is accurate and flexible, facilitating the miniaturization of high-performance hybrid magnetic field generating technologies with strict design constraints and spatial limitations., Comment: 24 pages, 10 figures

The reuse of water in a range of potable and non-potable applications is an important factor in the augmentation of water supply and in improving water security and productivity worldwide. A key hindrance to the reuse of water is the cost of compliance testing and process validation associated with ensuring that pathogen and chemicals in the feedwater are removed to a level that ensures no acute or chronic health and/or environmental effects. The critical control point (CCP) approach is well established and widely adopted by water utilities to provide an operational and risk management framework for the removal of pathogens in the treatment system. The application of a CCP approach to barriers in a treatment system for the removal of chemicals is presented. The application exemplar is to a small community wastewater treatment system that aims to produce potable quality water from a secondary treated wastewater effluent, however, the concepts presented are generic. The example used seven treatment barriers, five of which were designed and operated as CCP barriers for pathogens. The work demonstrates a method and risk management framework by which three of the seven barriers could also include a CCP approach for the removal of chemicals. Analogous to a CCP approach for pathogens, the potential is to reduce the use of chemical analysis as a routine determinant of performance criteria. The operational deployment of a CCP approach for chemicals was augmented with the development of a decision tree encompassing the classification of chemicals and the total removal credits across the treatment train in terms of the mechanistic removal of chemicals for each barrier. Validation of the approach is shown for an activated sludge, ozone and reverse osmosis barrier.

Endometrial cancer is the most commonly diagnosed gynaecological malignancy. Unfortunately, 15-20% of women demonstrate persistent or recurrent tumours that are refractory to current chemotherapies. We previously identified activating mutations in fibroblast growth factor receptor 2 (FGFR2) in 12% (stage I/II) to 17% (stage III/IV) endometrioid ECs and found that these mutations are associated with shorter progression-free and cancer-specific survival. Although FGFR inhibitors are undergoing clinical trials for treatment of several cancer types, little is known about the mechanism by which they induce cell death. We show that treatment with BGJ398, AZD4547 and PD173074 causes mitochondrial depolarization, cytochrome c release and impaired mitochondrial respiration in two FGFR2-mutant EC cell lines (AN3CA and JHUEM2). Despite this mitochondrial dysfunction, we were unable to detect caspase activation following FGFR inhibition; in addition, the pan-caspase inhibitor Z-VAD-FMK was unable to prevent cell death, suggesting that the cell death is caspase-independent. Furthermore, while FGFR inhibition led to an increase in LC3 puncta, treatment with bafilomycin did not further increase lipidated LC3, suggesting that FGFR inhibition led to a block in autophagosome degradation. We confirmed that cell death is mitochondrial-dependent as it can be blocked by overexpression of Bcl-2 and/or Bcl-XL. Importantly, we show that combining FGFR inhibitors with the BH3 mimetics ABT737/ABT263 markedly increased cell death in vitro and is more effective than BGJ398 alone in vivo, where it leads to marked tumour regression. This work may have implications for the design of clinical trials to treat a wide range of patients with FGFR-dependent malignancies.

BackgroundEndometrial cancer (EC) is a major gynecological cancer with increasing incidence. It comprised of four molecular subtypes with differing etiology, prognoses, and response to chemotherapy. In the future, clinical trials testing new single agents or combination therapies will be targeted to the molecular subtype most likely to respond. Pre-clinical models that faithfully represent the molecular subtypes of EC are urgently needed, we sought to develop and characterize a panel of novel EC patient-derived xenograft (PDX) models.MethodsHere, we report whole exome or whole genome sequencing of 11 PDX models and the matched primary tumor. Analysis of multiple PDX lineages and passages was performed to study tumor heterogeneity across lineages and/or passages. Based on recent reports of frequent defects in the homologous recombination (HR) pathway in EC, we assessed mutational signatures and HR deficiency scores and correlated these with in vivo responses to the PARP inhibitor (PARPi) talazoparib in six PDXs representing the different molecular subtypes of EC.ResultsPDX models were successfully generated from all four molecular subtypes of EC and uterine carcinosarcomas, and they recapitulated morphology and the molecular landscape of primary tumors without major genomic drift. We also observed a wide range of inter-tumor and intra-tumor heterogeneity, well captured by different PDX lineages, which could lead to different treatment responses. An in vivo response to talazoparib was detected in two p53mut models consistent with stable disease, however both lacked the HR deficiency genomic signature.ConclusionsEC PDX models represent the four molecular subtypes of disease and can capture intra-tumoral heterogeneity of the original primary tumor. PDXs of the p53mut molecular subtype showed sensitivity to PARPi, however, deeper and more durable responses will likely require combination of PARPi with other agents.

Magnetically-sensitive experiments and newly-developed quantum technologies with integrated high-permeability magnetic shields require increasing control of their magnetic field environment and reductions in size, weight, power and cost. However, magnetic fields generated by active components are distorted by high-permeability magnetic shielding, particularly when they are close to the shield's surface. Here, we present an efficient design methodology for creating desired static magnetic field profiles by using discrete coils electromagnetically-coupled to a cylindrical passive magnetic shield. We utilize a modified Green's function solution that accounts for the interior boundary conditions on a closed finite-length high-permeability cylindrical magnetic shield, and determine simplified expressions when a cylindrical coil approaches the interior surface of the shield. We use an analytic formulation of simple discrete building blocks to provide a complete discrete coil basis to generate any physically-attainable magnetic field inside the shield. We then use a genetic algorithm to find optimized discrete coil structures composed of this basis. We use our methodology to generate an improved linear axial gradient field, $\mathrm{d}B_z/\mathrm{d}z$, and transverse bias field, $B_x$. These optimized structures increase, by a factor of seven and three compared to the standard configurations, the volume in which the desired and achieved fields agree within $1\%$ accuracy, respectively. This coil design method can be used to optimize active--passive magnetic field shaping systems that are compact and simple to manufacture, enabling accurate magnetic field control in spatially-confined experiments at low cost., Comment: The authors M. Packer and P. J. Hobson have contributed equally to this work. 24 pages, 16 figures

Background Endometrial cancer (EC) is a major gynecological cancer with increasing incidence. It comprises four molecular subtypes with differing etiology, prognoses, and responses to chemotherapy. In the future, clinical trials testing new single agents or combination therapies will be targeted to the molecular subtype most likely to respond. As pre-clinical models that faithfully represent the molecular subtypes of EC are urgently needed, we sought to develop and characterize a panel of novel EC patient-derived xenograft (PDX) models. Methods Here, we report whole exome or whole genome sequencing of 11 PDX models and their matched primary tumor. Analysis of multiple PDX lineages and passages was performed to study tumor heterogeneity across lineages and/or passages. Based on recent reports of frequent defects in the homologous recombination (HR) pathway in EC, we assessed mutational signatures and HR deficiency scores and correlated these with in vivo responses to the PARP inhibitor (PARPi) talazoparib in six PDXs representing the copy number high/p53-mutant and mismatch-repair deficient molecular subtypes of EC. Results PDX models were successfully generated from grade 2/3 tumors, including three uterine carcinosarcomas. The models showed similar histomorphology to the primary tumors and represented all four molecular subtypes of EC, including five mismatch-repair deficient models. The different PDX lineages showed a wide range of inter-tumor and intra-tumor heterogeneity. However, for most PDX models, one arm recapitulated the molecular landscape of the primary tumor without major genomic drift. An in vivo response to talazoparib was detected in four copy number high models. Two models (carcinosarcomas) showed a response consistent with stable disease and two models (one copy number high serous EC and another carcinosarcoma) showed significant tumor growth inhibition, albeit one consistent with progressive disease; however, all lacked the HR deficiency genomic signature. Conclusions EC PDX models represent the four molecular subtypes of disease and can capture intra-tumor heterogeneity of the original primary tumor. PDXs of the copy number high molecular subtype showed sensitivity to PARPi; however, deeper and more durable responses will likely require combination of PARPi with other agents.

Magnetic shields that use both active and passive components to enable the generation of a tailored low-field environment are required for many applications in science, engineering, and medical imaging. Until now, accurate field nulling, or field generation, has only been possible over a small fraction of the overall volume of the shield. This is due to the interaction between the active field-generating components and the surrounding high-permeability passive shielding material. In this paper, we formulate the interaction between an arbitrary static current flow on a cylinder and an exterior closed high-permeability cylinder. We modify the Green's function for the magnetic vector potential and match boundary conditions on the shield's interior surface to calculate the total magnetic field generated by the system. We cast this formulation into an inverse optimization problem to design active--passive magnetic field shaping systems that accurately generate any physical static magnetic field in the interior of a closed cylindrical passive shield. We illustrate this method by designing hybrid systems that generate a range of magnetic field profiles to high accuracy over large interior volumes, and simulate them in real-world shields whose passive components have finite permeability, thickness, and axial entry holes. Our optimization procedure can be adapted to design active--passive magnetic field shaping systems that accurately generate any physical user-specified static magnetic field in the interior of a closed cylindrical shield of any length, enabling the development and miniaturization of systems that require accurate magnetic shielding and control., 22 pages, 9 figures, 1 table

We present an open-source synchronization software package, PackIO, that can record and generate voltage signals to enable complex experimental paradigms across multiple devices. This general purpose package is built on National Instruments data acquisition and generation hardware and has temporal precision up to the limit of the hardware. PackIO acts as a flexibly programmable master clock that can record experimental data (e.g. voltage traces), timing data (e.g. event times such as imaging frame times) while generating stimuli (e.g. voltage waveforms, voltage triggers to drive other devices, etc.). PackIO is particularly useful to record from and synchronize multiple devices, for example when simultaneously acquiring electrophysiology while generating and recording imaging timing data. Experimental control is easily enabled by an intuitive graphical user interface. We also release an open-source data visualisation and analysis tool, EphysViewer, written in MATLAB, as well as a module to import data into Python. These flexible and programmable tools allow experimenters to configure and set up customised input and output protocols in a synchronized fashion for controlling, recording, and analysing experiments.

The capabilities of a radio-frequency atomic magnetometer for object detection based on magnetic induction tomography are explored. The determination of object orientation is demonstrated by utilizing the measurement geometry. The self-compensation configuration of the atomic magnetometer is implemented to address the issue of saturation of the sensor response by the radio-frequency primary field that generates the object signature. Three methods of "covert" detection are investigated as a testbed for exploring the functionalities of this sensor, where (1) the operational frequency of the sensor is continuously changed, (2) the primary field has non-monochromatic frequency distribution, and (3) the sensor operates in the so-called spin maser mode. The results of the measurements are also discussed in terms of possible magnetic field communication.

Phacoemulsification is the most frequently performed surgery in the world. Over the past few years, this surgery seems to have reached a plateau with no further innovative breakthroughs. In this paper, we focus on alternatives techniques, the latest innovations, and the research and development pipeline in this field.

The human claustrum, a major hub of widespread neocortical connections, is a thin, bilateral sheet of gray matter located between the insular cortex and the striatum. The subplate is a largely transient cortical structure that contains some of the earliest generated neurons of the cerebral cortex and has important developmental functions to establish intra‐ and extracortical connections. In human and macaque some subplate cells undergo regulated cell death, but some remain as interstitial white matter cells. In mouse and rat brains a compact layer is formed, Layer 6b, and it remains underneath the cortex, adjacent to the white matter. Whether Layer 6b in rodents is homologous to primate subplate or interstitial white matter cells is still debated. Gene expression patterns, such as those of Nurr1/Nr4a2 , have suggested that the rodent subplate and the persistent subplate cells in Layer 6b and the claustrum might have similar origins. Moreover, the birthdates of the claustrum and Layer 6b are similarly precocious in mice. These observations prompted our speculations on the common developmental and evolutionary origin of the claustrum and the subplate. Here we systematically compare the currently available data on cytoarchitecture, evolutionary origin, gene expression, cell types, birthdates, neurogenesis, lineage and migration, circuit connectivity, and cell death of the neurons that contribute to the claustrum and subplate. Based on their similarities and differences we propose a partially common early evolutionary origin of the cells that become claustrum and subplate, a likely scenario that is shared in these cell populations across all amniotes.

The coronavirus disease 2019 (COVID-19) pandemic, caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has important implications for the safety of participants in clinical trials and the research staff caring for them and, consequently, for the trials themselves. Patients with heart failure may be at greater risk of infection with COVID-19 and the consequences might also be more serious, but they are also at risk of adverse outcomes if their clinical care is compromised. As physicians and clinical trialists, it is our responsibility to ensure safe and effective care is delivered to trial participants without affecting the integrity of the trial. The social contract with our patients demands no less. Many regulatory authorities from different world regions have issued guidance statements regarding the conduct of clinical trials during this COVID-19 crisis. However, international trials may benefit from expert guidance from a global panel of experts to supplement local advice and regulations, thereby enhancing the safety of participants and the integrity of the trial. Accordingly, the Heart Failure Association of the European Society of Cardiology on 21 and 22 March 2020 conducted web-based meetings with expert clinical trialists in Europe, North America, South America, Australia, and Asia. The main objectives of this Expert Position Paper are to highlight the challenges that this pandemic poses for the conduct of clinical trials in heart failure and to offer advice on how they might be overcome, with some practical examples. While this panel of experts are focused on heart failure clinical trials, these discussions and recommendations may apply to clinical trials in other therapeutic areas. © 2020 Published on behalf of the European Society of Cardiology. All rights reserved. © The Author(s) 2020.

Background Customary medicine of Australia’s Indigenous peoples draws upon knowledge developed through millennia of interaction with Australia’s unique flora and fauna. Many Indigenous Australians are interested in developing modern medicinal and commercial translations of traditional knowledge; however, barriers of trust and benefit sharing often thwart progress. Methods Using a participatory action research framework, university researchers collaborated with members of two Australian Indigenous communities to investigate selected medicinal plants and locally made bush products. A trusted community liaison facilitated the collaboration that was fostered through bilateral site visits. Material transfer and confidentiality agreements ensured that the plant materials were only used for the agreed purpose. Plain language written reports of the laboratory results were provided to the communities with follow up discussions. Results In case study 1, only some of the traditional uses for the raw plants were shared with the researchers. Deidentified plants were assessed for antioxidant and antimicrobial properties. In case study 2, the plant names, traditional uses, and preparation methods were shared with the aim of learning more about their plants, potential uses, and optimising their bush products. Literature reviews were conducted that also helped guide in vitro testing of the crude and solvent partitioned extracts. These differences reflected the community’s reasons for conducting the research and intellectual property considerations. In both cases, observed benefits included building trust and strengthening working relationships for ongoing collaboration, fostering enthusiasm for linking traditional and scientific knowledge, promoting cross-cultural learning about scientific methods and traditional medicine, maintaining the relevance of traditional knowledge in the modern world, and initiating community discussions around their bush medicine product development. Conclusions Community-driven scientific investigation of traditional medicinal knowledge can facilitate culturally meaningful outcomes, with potentially wide-reaching direct and indirect benefits. Community liaisons were invaluable for establishment of strong relationships and ensured that the research was culturally and locally appropriate. The need for clearer guidelines and regulation around community-driven biomedical research of their plants was identified. Australia would benefit from a user-friendly, open-source toolkit that promotes use of local traditional medicines, contains information about processes and protocols that communities and scientists could use to develop collaborative projects, and guides regulation and ethical commercialisation. Close consultation and collaboration with communities and researchers will be needed to ensure that such a toolkit is culturally appropriate and fit-for-purpose.