Be there on time: Spatial-temporal regularities guide young children's attention in dynamic environments.

Children's ability to benefit from spatiotemporal regularities to detect goal-relevant targets was tested in a dynamic, extended context. Young adults and children (from a low-deprivation area school in the United Kingdom; N = 80; 5-6 years; 39 female; ethics approval did not permit individual-level race/ethnicity surveying) completed a dynamic visual-search task. Targets and distractors faded in and out of a display over seconds. Half of the targets appeared at predictable times and locations. Search performance in children was poorer overall. Nevertheless, they benefitted equivalently from spatiotemporal regularities, detecting more predictable than unpredictable targets. Children's benefits from predictions correlated positively with their attention. The study brings ecological validity to the study of attentional guidance in children, revealing striking behavioral benefits of dynamic experience-based predictions.

Chronic histological changes in deceased donor kidneys at implantation do not predict graft survival: a single-centre retrospective analysis.

The use of preimplantation kidney biopsies (PIKBs) to aid deceased donor kidney utilization decisions is controversial. Outcomes of transplants that had been biopsied after the decision had been made to implant were analysed, in order to determine the association between chronic histological changes at implantation and graft outcomes. A retrospective analysis of transplants between the year range 2006-2015 was performed. Karpinski scores on biopsies were collected, and graft outcomes were analysed using univariate and multivariable techniques. Also, Karpinski scores from single and dual kidney transplants from older donors were examined to determine if knowledge of the score preoperatively would have altered utilization. Four hundred and eight single kidneys were transplanted. Although kidneys with scores >4 had lower 1- and 3-year median (IQR) estimated glomerular filtration rates (eGFRs) than those scoring 0-4 (51 (37-66) vs. 35 (26-52) ml/min/1.73 m2 , P < 0.001, and 52 (34-64) vs. 35 (24-52) ml/min/1.73 m2 , P < 0.001, respectively), there was no significant association between Karpinski score and death-censored graft survival on univariate or multivariable analyses. The utilization analysis (75 single and 25 dual kidney transplant recipients) suggested that systematic use of PIKBs would have resulted in 29% fewer patients being transplanted. This analysis does not support the systematic use of PIKBs to determine deceased donor kidney utilization.

Graft dysfunction in chronic antibody-mediated rejection correlates with B-cell-dependent indirect antidonor alloresponses and autocrine regulation of interferon-γ production by Th1 cells.

Chronic antibody-mediated rejection, a common cause of renal transplant failure, has a variable clinical phenotype. Understanding why some with chronic antibody-mediated rejection progress slowly may help develop more effective therapies. B lymphocytes act as antigen-presenting cells for in vitro indirect antidonor interferon-γ production in chronic antibody-mediated rejection, but many patients retain the ability to regulate these responses. Here we test whether particular patterns of T and B cell antidonor response associate with the variability of graft dysfunction in chronic antibody-mediated rejection. Our results confirm that dynamic changes in indirect antidonor CD4+ T-cell responses correlate with changes in estimated glomerular filtration rates, independent of other factors. Graft dysfunction progressed rapidly in patients who developed unregulated B-cell-driven interferon-γ production. However, conversion to a regulated or nonreactive pattern, which could be achieved by optimization of immunosuppression, associated with stabilization of graft function. Functional regulation by B cells appeared to activate an interleukin-10 autocrine pathway in CD4+ T cells that, in turn, impacted on antigen-specific responses. Thus, our data significantly enhance the understanding of graft dysfunction associated with chronic antibody-mediated rejection and provide the foundation for strategies to prolong renal allograft survival, based on regulation of interferon-γ production.

How Do Executive Functions Influence Children's Reasoning About Counterintuitive Concepts in Mathematics and Science?

Many scientific and mathematical concepts are counterintuitive because they conflict with misleading perceptual cues or incorrect naive theories that we build from our everyday experiences of the world. Executive functions (EFs) influence mathematics and science achievement, and inhibitory control (IC), in particular, might facilitate counterintuitive reasoning. Stop & Think (S&T) is a computerised learning activity that trains IC skills. It has been found effective in improving primary children's mathematics and science academic performance in a large scale RCT trial (Palak et al., 2019; Wilkinson et al., Journal of Cognitive Enhancement, 4, 296-314, 2020). The current study aimed to investigate the role of EFs and the moderating effects of S&T training on counterintuitive mathematics and science reasoning. A sample of 372 children in school Years 3 (7- to 8-year-olds) and 5 (9- to 10-year-olds) were allocated to S&T, active control or teaching as usual conditions, and completed tasks assessing verbal and visuospatial working memory (WM), IC, IQ, and counterintuitive reasoning, before and after training. Cross-sectional associations between counterintuitive reasoning and EF were found in Year 5 children, with evidence of a specific role of verbal WM. The intervention benefited counterintuitive reasoning in Year 3 children only and EF measures were not found to predict which children would most benefit from the intervention. Combined with previous research, these results suggest that individual differences in EF play a lesser role in counterintuitive reasoning in younger children, while older children show a greater association between EFs and counterintuitive reasoning and are able to apply the strategies developed during the S&T training to mathematics and science subjects. This work contributes to understanding why specifically the S&T intervention is effective. This work was preregistered with the ISRCTN registry (TRN: 54726482) on 10/10/2017. Supplementary Information: The online version contains supplementary material available at 10.1007/s41465-023-00271-0.

Exploiting the aggregation propensity of beta-lactamases to design inhibitors that induce enzyme misfolding.

There is an arms race between beta-lactam antibiotics development and co-evolving beta-lactamases, which provide resistance by breaking down beta-lactam rings. We have observed that certain beta-lactamases tend to aggregate, which persists throughout their evolution under the selective pressure of antibiotics on their active sites. Interestingly, we find that existing beta-lactamase active site inhibitors can act as molecular chaperones, promoting the proper folding of these resistance factors. Therefore, we have created Pept-Ins, synthetic peptides designed to exploit the structural weaknesses of beta-lactamases by causing them to misfold into intracellular inclusion bodies. This approach restores sensitivity to a wide range of beta-lactam antibiotics in resistant clinical isolates, including those with Extended Spectrum variants that pose significant challenges in medical practice. Our findings suggest that targeted aggregation of resistance factors could offer a strategy for identifying molecules that aid in addressing the global antibiotic resistance crisis.

Lipids revert inert Abeta amyloid fibrils to neurotoxic protofibrils that affect learning in mice.

Although soluble oligomeric and protofibrillar assemblies of Abeta-amyloid peptide cause synaptotoxicity and potentially contribute to Alzheimer's disease (AD), the role of mature Abeta-fibrils in the amyloid plaques remains controversial. A widely held view in the field suggests that the fibrillization reaction proceeds 'forward' in a near-irreversible manner from the monomeric Abeta peptide through toxic protofibrillar intermediates, which subsequently mature into biologically inert amyloid fibrils that are found in plaques. Here, we show that natural lipids destabilize and rapidly resolubilize mature Abeta amyloid fibers. Interestingly, the equilibrium is not reversed toward monomeric Abeta but rather toward soluble amyloid protofibrils. We characterized these 'backward' Abeta protofibrils generated from mature Abeta fibers and compared them with previously identified 'forward' Abeta protofibrils obtained from the aggregation of fresh Abeta monomers. We find that backward protofibrils are biochemically and biophysically very similar to forward protofibrils: they consist of a wide range of molecular masses, are toxic to primary neurons and cause memory impairment and tau phosphorylation in mouse. In addition, they diffuse rapidly through the brain into areas relevant to AD. Our findings imply that amyloid plaques are potentially major sources of soluble toxic Abeta-aggregates that could readily be activated by exposure to biological lipids.

Manipulation of tissue factor-mediated basal PAR-2 signalling on macrophages determines sensitivity for IFNγ responsiveness and significantly modifies the phenotype of murine DTH.

Background: Tissue factor (TF) generates proteases that can signal through PAR-1 and PAR-2. We have previously demonstrated PAR-1 signalling primes innate myeloid cells to be exquisitely sensitive to interferon-gamma (IFNγ). In this work we explored how TF mediated PAR-2 signalling modulated responsiveness to IFNγ and investigated the interplay between PAR-1/-2 signalling on macrophages. Methodology: We characterised how TF through PAR-2 influenced IFNγ sensitivity in vitro using PCR and flow cytometry. and how it influenced oxazolone-induced delayed type hypersensitivity (DTH) responses in vivo. We investigated how basal signalling through PAR-2 influenced PAR-1 signalling using a combination of TF-inhibitors and PAR-1 &-2 agonists and antagonists. Finally, we investigated whether this system could be targeted therapeutically using 3-mercaptopropionyl-F-Cha-Cha-RKPNDK (3-MP), which has actions on both PAR-1 and -2. Results: TF delivered a basal signal through PAR-2 that upregulated SOCS3 expression and blunted M1 polarisation after IFNγ stimulation, opposing the priming achieved by signalling through PAR-1. PAR-1 and -2 agonists or antagonists could be used in combination to modify this basal signal in vitro and in vivo. 3-MP, by virtue of its PAR-2 agonist properties was superior to agents with only PAR-1 antagonist properties at reducing M1 polarisation induced by IFNγ and suppressing DTH. Tethering a myristoyl electrostatic switch almost completely abolished the DTH response. Conclusions: TF-mediated signalling through PARs-1 and -2 act in a homeostatic way to determine how myeloid cells respond to IFNγ. 3-MP, an agent that simultaneously inhibits PAR-1 whilst delivering a PAR-2 signal, can almost completely abolish immune responses dependent on M1 polarisation, particularly if potency is enhanced by targeting to cell membranes; this has potential therapeutic potential in multiple diseases.

Antisocial behaviour and teacher-student relationship quality: The role of emotion-related abilities and callous-unemotional traits.

BACKGROUND: Childhood antisocial behaviour has been associated with poorer teacher-student relationship (TSR) quality. It is also well-established that youth with antisocial behaviour have a range of emotion-related deficits, yet the impact of these students' emotion-related abilities on the TSR is not understood. Furthermore, the addition of the Limited Prosocial Emotions Specifier in the Diagnostic and Statistical Manual of Mental Disorders (DSM-5) indicates that understanding the role of callous-unemotional (CU) traits for youth with antisocial behaviour problems is of particular importance. AIMS: The aim of this study was to investigate the association between antisocial behaviour difficulties and the TSR by examining the influence of emotion-related abilities and CU traits. SAMPLE: Twelve teachers from 10 primary schools provided anonymized information on 108 children aged 6-11 years. RESULTS: Antisocial behaviour was associated with higher teacher-student conflict (but not closeness) as well as higher emotion lability/negativity and lower emotion understanding/empathy. Emotion lability/negativity was associated with higher teacher-student conflict (but not closeness), and emotion understanding/empathy was associated with lower teacher-student conflict and higher closeness. CU traits were associated with higher teacher-student conflict and lower teacher-student closeness (controlling for antisocial behaviour more broadly). We found no evidence of a moderating effect of CU traits or emotion-related abilities on the association between antisocial behaviour and TSR quality. CONCLUSIONS: Interventions for behaviour difficulties should consider teacher-student relationships in the classroom. Strategies which aim to improve teacher-student closeness as well as reduce teacher-student conflict may be of particular value to students with high CU traits.

PAR-1 signaling on macrophages is required for effective in vivo delayed-type hypersensitivity responses.

Delayed-type hypersensitivity (DTH) responses underpin chronic inflammation. Using a model of oxazolone-induced dermatitis and a combination of transgenic mice, adoptive cell transfer, and selective agonists/antagonists against protease activated receptors, we show that that PAR-1 signaling on macrophages by thrombin is required for effective granuloma formation. Using BM-derived macrophages (BMMs) in vitro, we show that thrombin signaling induced (a) downregulation of cell membrane reverse cholesterol transporter ABCA1 and (b) increased expression of IFNγ receptor and enhanced co-localization within increased areas of cholesterol-rich membrane microdomains. These two key phenotypic changes combined to make thrombin-primed BMMs sensitive to M1 polarization by 1000-fold less IFNγ, compared to resting BMMs. We confirm that changes in ABCA1 expression were directly responsible for the exquisite sensitivity to IFNγ in vitro and for the impact on granuloma formation in vivo. These data indicate that PAR-1 signaling plays a hitherto unrecognized and critical role in DTH responses.

Accurate prediction of DnaK-peptide binding via homology modelling and experimental data.

Molecular chaperones are essential elements of the protein quality control machinery that governs translocation and folding of nascent polypeptides, refolding and degradation of misfolded proteins, and activation of a wide range of client proteins. The prokaryotic heat-shock protein DnaK is the E. coli representative of the ubiquitous Hsp70 family, which specializes in the binding of exposed hydrophobic regions in unfolded polypeptides. Accurate prediction of DnaK binding sites in E. coli proteins is an essential prerequisite to understand the precise function of this chaperone and the properties of its substrate proteins. In order to map DnaK binding sites in protein sequences, we have developed an algorithm that combines sequence information from peptide binding experiments and structural parameters from homology modelling. We show that this combination significantly outperforms either single approach. The final predictor had a Matthews correlation coefficient (MCC) of 0.819 when assessed over the 144 tested peptide sequences to detect true positives and true negatives. To test the robustness of the learning set, we have conducted a simulated cross-validation, where we omit sequences from the learning sets and calculate the rate of repredicting them. This resulted in a surprisingly good MCC of 0.703. The algorithm was also able to perform equally well on a blind test set of binders and non-binders, of which there was no prior knowledge in the learning sets. The algorithm is freely available at http://limbo.vib.be.

Thermodynamic and Evolutionary Coupling between the Native and Amyloid State of Globular Proteins.

The amyloid-like aggregation propensity present in most globular proteins is generally considered to be a secondary side effect resulting from the requirements of protein stability. Here, we demonstrate, however, that mutations in the globular and amyloid state are thermodynamically correlated rather than simply associated. In addition, we show that the standard genetic code couples this structural correlation into a tight evolutionary relationship. We illustrate the extent of this evolutionary entanglement of amyloid propensity and globular protein stability. Suppressing a 600-Ma-conserved amyloidogenic segment in the p53 core domain fold is structurally feasible but requires 7-bp substitutions to concomitantly introduce two aggregation-suppressing and three stabilizing amino acid mutations. We speculate that, rather than being a corollary of protein evolution, it is equally plausible that positive selection for amyloid structure could have been a driver for the emergence of globular protein structure.

Domain-Specific Inhibitory Control Training to Improve Children's Learning of Counterintuitive Concepts in Mathematics and Science.

Evidence from cognitive neuroscience suggests that learning counterintuitive concepts in mathematics and science requires inhibitory control (IC). This prevents interference from misleading perceptual cues and naïve theories children have built from their experiences of the world. Here, we (1) investigate associations between IC, counterintuitive reasoning, and academic achievement and (2) evaluate a classroom-based computerised intervention, called Stop & Think, designed to embed IC training within the learning domain (i.e. mathematics and science content from the school curricula). Cross-sectional analyses of data from 627 children in Years 3 and 5 (7- to 10-year-olds) demonstrated that IC, measured on a Stroop-like task, was associated with counterintuitive reasoning and mathematics and science achievement. A subsample (n = 456) participated either in Stop & Think as a whole-class activity (teacher-led, STT) or using individual computers (pupil-led, STP), or had teaching as usual (TAU). For Year 3 children (but not Year 5), Stop & Think led to better counterintuitive reasoning (i.e. near transfer) in STT (p < .001, ηp 2 = .067) and STP (p < .01, ηp 2 = .041) compared to TAU. Achievement data was not available for Year 3 STP or Year 5 STT. For Year 3, STT led to better science achievement (i.e. far transfer) compared to TAU (p < .05, ηp 2 = .077). There was no transfer to the Stroop-like measure of IC. Overall, these findings support the idea that IC may contribute to counterintuitive reasoning and mathematics and science achievement. Further, we provide preliminary evidence of a domain-specific IC intervention with transferable benefits to academic achievement for Year 3 children.

Effect of Optimized Immunosuppression (Including Rituximab) on Anti-Donor Alloresponses in Patients With Chronically Rejecting Renal Allografts.

RituxiCAN-C4 combined an open-labeled randomized controlled trial (RCT) in 7 UK centers to assess whether rituximab could stabilize kidney function in patients with chronic rejection, with an exploratory analysis of how B cell-depletion influenced T cell anti-donor responses relative to outcome. Between January 2007 and March 2015, 59 recruits were enrolled after screening, 23 of whom consented to the embedded RCT. Recruitment was halted when in a pre-specified per protocol interim analysis, the RCT was discovered to be significantly underpowered. This report therefore focuses on the exploratory analysis, in which we confirmed that when B cells promoted CD4+ anti-donor IFNγ production assessed by ELISPOT, this associated with inferior clinical outcome; these patterns were inhibited by optimized immunosuppression but not rituximab. B cell suppression of IFNγ production, which associated with number of transitional B cells and correlated with slower declines in kidney function was abolished by rituximab, which depleted transitional B cells for prolonged periods. We conclude that in this patient population, optimized immunosuppression but not rituximab promotes anti-donor alloresponses associated with favorable outcomes. Clinical Trial Registration: Registered with EudraCT (2006-002330-38) and www.ClinicalTrials.gov, identifier: NCT00476164.

Regression of Atherosclerosis in ApoE-/- Mice Via Modulation of Monocyte Recruitment and Phenotype, Induced by Weekly Dosing of a Novel "Cytotopic" Anti-Thrombin Without Prolonged Anticoagulation.

Background Anticoagulants induce atherosclerosis regression in animal models but exploiting this clinically is limited by bleeding events. Here we test a novel thrombin inhibitor, PTL060, comprising hirulog covalently linked to a synthetic myristoyl electrostatic switch to tether to cell membranes. Methods and Results ApoE-/- mice were fed chow or high-fat diets, before transplantation of congenic aortic segments or injection of PTL060, parental hirulog, control saline, or labeled CD11b positive cells. Aortic transplants from transgenic mice expressing anticoagulants on endothelium did not develop atherosclerosis. A single intravenous injection of PTL060, but not hirulog inhibited atheroma development by >50% compared with controls when assessed 4 weeks later. Mice had prolonged bleeding times for only one seventh of the time that PTL060 was biologically active. Repeated weekly injections of PTL060 but not hirulog caused regression of atheroma. We dissected 2 contributory mechanisms. First, the majority of CCR2+ (C-C chemokine receptor type 2+) monocytes recruited into plaques expressed CCR7 (C-C chemokine receptor type 7), ABCA1 (ATP-binding cassette transporter - 1), and interleukin-10 in PTL060 mice, a phenotype seen in <20% of CCR2+ recruits in controls. Second, after several doses, there was a significant reduction in monocyte recruits, the majority of which were CCR2-negative with a similar regression-associated phenotype. Regression equivalent to that induced by intravenous PTL060 was induced by adoptive transfer of CD11b+ cells pre-coated with PTL060. Conclusions Covalent linkage of a myristoyl electrostatic switch onto hirulog in PTL060 uncouples the pharmacodynamic effects on hemostasis and atherosclerosis, such that plaque regression, mediated predominantly via effects on monocytes, is accompanied by only transient anticoagulation.

Reverse engineering synthetic antiviral amyloids.

Human amyloids have been shown to interact with viruses and interfere with viral replication. Based on this observation, we employed a synthetic biology approach in which we engineered virus-specific amyloids against influenza A and Zika proteins. Each amyloid shares a homologous aggregation-prone fragment with a specific viral target protein. For influenza we demonstrate that a designer amyloid against PB2 accumulates in influenza A-infected tissue in vivo. Moreover, this amyloid acts specifically against influenza A and its common PB2 polymorphisms, but not influenza B, which lacks the homologous fragment. Our model amyloid demonstrates that the sequence specificity of amyloid interactions has the capacity to tune amyloid-virus interactions while allowing for the flexibility to maintain activity on evolutionary diverging variants.

Aggregating sequences that occur in many proteins constitute weak spots of bacterial proteostasis.

Aggregation is a sequence-specific process, nucleated by short aggregation-prone regions (APRs) that can be exploited to induce aggregation of proteins containing the same APR. Here, we find that most APRs are unique within a proteome, but that a small minority of APRs occur in many proteins. When aggregation is nucleated in bacteria by such frequently occurring APRs, it leads to massive and lethal inclusion body formation containing a large number of proteins. Buildup of bacterial resistance against these peptides is slow. In addition, the approach is effective against drug-resistant clinical isolates of Escherichia coli and Acinetobacter baumannii, reducing bacterial load in a murine bladder infection model. Our results indicate that redundant APRs are weak points of bacterial protein homeostasis and that targeting these may be an attractive antibacterial strategy.

Domain swapping in p13suc1 results in formation of native-like, cytotoxic aggregates.

The field of protein aggregation has been occupied mainly with the study of beta-strand self-association that occurs as a result of misfolding and leads to the formation of toxic protein aggregates and amyloid fibers. However, some of these aggregates retain native-like structural and enzymatic properties suggesting mechanisms other than beta-strand assembly. p13suc1 is a small protein that can exist as a monomer or a domain-swapped dimer. Here, we show that, under native conditions, p13suc1 forms three-dimensional domain-swapped aggregates, and that these aggregates are cytotoxic. Thus, toxicity of protein aggregates is not only associated with beta-rich assemblies and amyloid fibers, involving non-native interactions, but it can be induced by oligomeric misassembly that maintains predominantly native-like interactions.

Fear appeals, engagement, and examination performance: The role of challenge and threat appraisals.

BACKGROUND: Fear appeals are persuasive messages that draw attention to the negative consequences (e.g., academic failure) that follow a particular course of action (e.g., not engaging in lessons) and how negative consequences can be avoided with an alternate course of action. Previous studies have shown that when fear appeals are appraised as threatening, they are related to lower examination performance. AIM: In this study, we examined how challenge, as well as threat, appraisals are indirectly related to performance on a mathematics examination through behavioural engagement. SAMPLE: A total of 579 students from two secondary schools. METHOD: Data were collected over four waves at approximately 3-month intervals. Behavioural engagement data were collected at T1 and T3 , fear appeal frequency and appraisal at T3 , and examination performance at T2 and T4 . RESULTS: A challenge appraisal of fear appeals predicted better examination performance through higher behavioural engagement whereas a threat appraisal of fear appeals predicted worse examination performance through lower behavioural engagement. CONCLUSION: The relationship between fear appeals and examination performance depended on their appraisal.

The structure of the transition state for folding of domain-swapped dimeric p13suc1.

suc1 has two native states, a monomer and a domain-swapped dimer, in which one molecule exchanges a beta strand with an identical partner. Thus, monomer and dimer have the same structures but are topologically distinct. Importantly, residues that exchange are part of the folding nucleus of the monomer and therefore forming these interactions in the dimer would be expected to incur a large entropic cost. Here we present the transition state for folding/unfolding of domain-swapped dimeric suc1 and compare it with its monomeric counterpart. The same overall structure is observed in the two transition states but the phi values are consistently higher for the domain-swapped dimer. Thus, a greater entropic penalty for bringing together the key interactions in the dimer is overcome by mobilizing more contacts in the transition state, thereby achieving a greater enthalpic gain.