Lack of evidence for decreased protein stability in the 2397 (Met) haplotype of the leucine rich repeat kinase 2 protein implicated in Parkinson’s disease

Missense mutations in the leucine rich repeat kinase 2 (LRRK2) gene are the leading genetic cause of autosomal dominant familial Parkinson’s disease. We previously reported that two mutations within the ROC domain, namely R1441C and A1442P, exhibit increased protein degradation leading to lowered steady state LRRK2 protein levels in HEK293 cells. More recently, the common WD40 domain LRRK2 haplotype, Met2397, which is a risk factor for Crohn’s disease, has been shown to lower steady state protein levels in HEK293 cells. In view of recent evidence implicating LRRK2 and inflame-mation in PD, we investigated the effects of Met2397 on LRRK2 expression, and compared them to the Thr2397 variant and other LRRK2 mutants. In this study, we transfected HEK293 cells with plasmid constructs encoding the different LRRK2 variants, and analyzed the resulting protein levels by Western blot and flow cytometry. Here we found that both the Met2397 and Thr2397 haplotypes yield similar levels of LRRK2 protein expression and do not appear to impact cell viability in HEK293 cells, compared to other LRRK mutants. Thus, we have concluded that the Met2397 haplotype is unlikely to play a role in LRRK2 mediated or idiopathic PD

Precipitating or prohibiting factor: Coaches’ perceptions of their role and actions in anti-doping

Coaches are frequently cited as potentially precipitating or preventing athletes’ engagement in doping. However, little is known about coaches’ perspectives. Therefore, the purpose of this study was to examine coaches’ perceptions of their role and actions in athletes’ anti-doping behaviour. Twenty-three coaches (M=17, F =6) working with performance athletes in Scotland participated in semi-structured interviews where topics related to doping and anti-doping were discussed. Thematic analysis, guided by Schön’s [1] role frame and reflective conversation concepts, was used to develop themes. Analysis led to the development of four internal role frame themes: clean sport value, approach to preparation and performance, responsibility to athletes, and knowledge; and five boundary role frame themes: Scottish/British sporting culture, potential for benefit, prevalence of doping and testing, clarity of responsibilities and consequences, and beyond coaches’ control. The coaches’ role frame supported an anti-doping stance, however, it also presented a risk and was insufficient to ensure action. Analysis of coaches’ reflective conversations revealed the issues set by the coaches differed and influenced subsequent actions and evaluations

Uncovering protein function: from classification to complexes

Almost all interactions and reactions that occur in living organisms involve proteins. The various biological roles of proteins include, but are not limited to, signal transduction, gene transcription, cell death, immune function, structural support, and catalysis of all the chemical reactions that enable organisms to survive. The varied roles of proteins have led to them being dubbed 'the workhorses of all living organisms'. This article discusses the functions of proteins and how protein function is studied in a laboratory setting. In this article, we begin by examining the functions of protein domains, followed by a discussion of some of the major classes of proteins based on their function. We consider protein binding in detail, which is central to protein function. We then examine how protein function can be altered through various mechanisms including post-translational modification, and changes to environment, oligomerisation and mutations. Finally, we consider a handful of the techniques employed in the laboratory to understand and measure the function of proteins

Relationships between selective neuronal loss and microglial activation after ischaemic stroke in man.

Modern ischaemic stroke management involves intravenous thrombolysis followed by mechanical thrombectomy, which allows markedly higher rates of recanalization and penumbral salvage than thrombolysis alone. However, <50% of treated patients eventually enjoy independent life. It is therefore important to identify complementary therapeutic targets. In rodent models, the salvaged penumbra is consistently affected by selective neuronal loss, which may hinder recovery by interfering with plastic processes, as well as by microglial activation, which may exacerbate neuronal death. However, whether the salvaged penumbra in man is similarly affected is still unclear. Here we determined whether these two processes affect the non-infarcted penumbra in man and, if so, whether they are inter-related. We prospectively recruited patients with (i) acute middle-cerebral artery stroke; (ii) penumbra present on CT perfusion obtained <4.5 h of stroke onset; and (iii) early neurological recovery as a marker of penumbral salvage. PET with 11C-flumazenil and 11C-PK11195, as well as MRI to map the final infarct, were obtained at predefined follow-up times. The presence of selective neuronal loss and microglial activation was determined voxel-wise within the MRI normal-appearing ipsilateral non-infarcted zone and surviving penumbra masks, and their inter-relationship was assessed both across and within patients. Dilated infarct contours were consistently excluded to control for partial volume effects. Across the 16 recruited patients, there was reduced 11C-flumazenil and increased 11C-PK11195 binding in the whole ipsilateral non-infarcted zone (P = 0.04 and 0.02, respectively). Within the non-infarcted penumbra, 11C-flumazenil was also reduced (P = 0.001), but without clear increase in 11C-PK11195 (P = 0.18). There was no significant correlation between 11C-flumazenil and 11C-PK11195 in either compartment. This mechanistic study provides direct evidence for the presence of both neuronal loss and microglial activation in the ipsilateral non-infarcted zone. Further, we demonstrate the presence of neuronal loss affecting the surviving penumbra, with no or only mild microglial activation, and no significant relationship between these two processes. Thus, microglial activation may not contribute to penumbral neuronal loss in man, and its presence in the ipsilateral hemisphere may merely reflect secondary remote degeneration. Selective neuronal loss in the surviving penumbra may represent a novel therapeutic target as an adjunct to penumbral salvage to further improve functional outcome. However, microglial activation may not stand as the primary therapeutic approach. Protecting the penumbra by acutely improving perfusion and oxygenation in conjunction with thrombectomy for example, may be a better approach. 11C-flumazenil PET would be useful to monitor the effects of such therapies

Process evaluation protocol for a cluster randomised controlled trial of an intervention to improve the mental health support and training available to secondary school teachers– the Wellbeing in Secondary Education (WISE) project

Background: Secondary school teachers have low levels of wellbeing and high levels of depression compared with the general population. Teachers are in a key position to support students, but poor mental health may be a barrier to doing so effectively. The Wellbeing in Secondary Education (WISE) project is a cluster randomised controlled trial (RCT) of an intervention to improve the mental health support and training available to secondary school teachers through delivery of the training package Mental Health First Aid and a staff peer support service. We will conduct a process evaluation as part of the WISE trial to support the interpretation of trial outcomes and refine intervention theory. The domains assessed will be: the extent to which the hypothesised mechanisms of change are activated; system level influences on these mechanisms; programme differentiation and usual practice; intervention implementation, including any adaptations; intervention acceptability; and intervention sustainability. Methods: Research questions will be addressed via quantitative and qualitative methods. All study schools (n = 25) will provide process evaluation data, with more detailed focus group, interview and observation data being collected from a subsample of case study schools (4 intervention and 4 control). Mechanisms of change, as outlined in a logic model, will be measured via teacher and student surveys and focus groups. School context will be explored via audits of school practice that relate to mental health and wellbeing, combined with stakeholder interviews and focus groups. Implementation of the training and peer support service will be assessed via training observations, training participant evaluation forms, focus groups with participants, interviews with trainers and peer support service users, and peer supporter logs recording help provided. Acceptability and sustainability will be examined via interviews with funders, head teachers, trainers and peer support services users, and focus groups with training participants. Discussion: The process evaluation embedded within the WISE cluster RCT will illuminate how and why the intervention was effective, ineffective or conferred iatrogenic effects. It will contribute to the refinement of the theory underpinning the intervention, and will help to inform any future implementation. Trial registration: International Standard Randomised Controlled Trial Number: ISRCTN95909211 registered on 24 March 2016. Keywords: Mental health, Wellbeing, Schools, Children, Adolescents, Teachers, Process evaluation, Cluster Randomised Controlled Tria

Views and experience of breastfeeding in public: A qualitative systematic review

Breastfeeding rates in many Global North countries are low. Qualitative research highlights that breastfeeding in public is a particular challenge, despite mothers often having the legal right to do so. To identify barriers and facilitators, we systematically searched the qualitative research from Organisation for Economic Co‐operation and Development countries relating to breastfeeding in public spaces from 2007 to 2021. Data were analysed using the Thematic Synthesis technique. The review was registered with PROSPERO (registration number: CRD42017081504). Database searching identified 3570 unique records. In total, 74 papers, theses, or book chapters, relating to 71 studies, were included, accounting for over 17,000 mothers. Overall, data quality was high. Our analysis identified that five core factors influenced mothers' thought processes and their breastfeeding in public behaviour: legal system; structural (in)equality; knowledge; beliefs and the social environment. Macro‐level factors relating to legislation and inequality urgently require redress if breastfeeding rates are to be increased. Widespread culture change is also required to enhance knowledge, change hostile beliefs and thus the social environment in which mother/infant dyads exist. In particular, the sexualisation of breasts, disgust narratives and lack of exposure among observers to baby‐led infant feeding patterns resulted in beliefs which created a stigmatising environment. In this context, many mothers felt unable to breastfeed in public; those who breastfed outside the home were usually highly self‐aware, attempting to reduce their exposure to conflict. Evidence‐based theoretically informed interventions to remove barriers to breastfeeding in public are urgently required

An intervention to improve teacher well-being support and training to support students in UK high schools (the WISE study): A cluster randomised controlled trial.

Funder: Public Health WalesFunder: Bristol City CouncilFunder: Public Health EnglandBACKGROUND: Teachers are at heightened risk of poor mental health and well-being, which is likely to impact on the support they provide to students, and student outcomes. We conducted a cluster randomised controlled trial, to test whether an intervention to improve mental health support and training for high school teachers led to improved mental health and well-being for teachers and students, compared to usual practice. We also conducted a cost evaluation of the intervention. METHODS AND FINDINGS: The intervention comprised (i) Mental Health First Aid training for teachers to support students; (ii) a mental health awareness session; and (iii) a confidential staff peer support service. In total 25 mainstream, non-fee-paying secondary schools stratified by geographical area and free school meal entitlement were randomly allocated to intervention (n = 12) or control group (n = 13) after collection of baseline measures. We analysed data using mixed-effects repeated measures models in the intention-to-treat population, adjusted for stratification variables, sex, and years of experience. The primary outcome was teacher well-being (Warwick-Edinburgh Mental Well-being Scale). Secondary outcomes were teacher depression, absence, and presenteeism, and student well-being, mental health difficulties, attendance, and attainment. Follow-up was at months 12 (T1) and 24 (T2). We collected process data to test the logic model underpinning the intervention, to aid interpretation of the findings. A total of 1,722 teachers were included in the primary analysis. Teacher well-being did not differ between groups at T2 (intervention mean well-being score 47.5, control group mean well-being score 48.4, adjusted mean difference -0.90, 95% CI -2.07 to 0.27, p = 0.130). The only effect on secondary outcomes was higher teacher-reported absence among the intervention group at T2 (intervention group median number of days absent 0, control group median number of days absent 0, ratio of geometric means 1.04, 95% CI 1.00 to 1.09, p = 0.042). Process measures indicated little change in perceived mental health support, quality of relationships, and work-related stress. The average cost of the intervention was £9,103 per school. The study's main limitations were a lack of blinding of research participants and the self-report nature of the outcome measures. CONCLUSIONS: In this study, we observed no improvements to teacher or student mental health following the intervention, possibly due to a lack of impact on key drivers of poor mental health within the school environment. Future research should focus on structural and cultural changes to the school environment, which may be more effective at improving teacher and student mental health and well-being. TRIAL REGISTRATION: www.isrctn.com ISRCTN95909211

Dental attendance and behavioural pathways to adult oral health inequalities.

BACKGROUND: While inequalities in oral health are documented, little is known about the extent to which they are attributable to potentially modifiable factors. We examined the role of behavioural and dental attendance pathways in explaining oral health inequalities among adults in England, Wales and Northern Ireland. METHODS: Using nationally representative data, we analysed inequalities in self-rated oral health and number of natural teeth. Highest educational attainment, equivalised household income and occupational social class were used to derive a latent socioeconomic position (SEP) variable. Pathways were dental attendance and behaviours (smoking and oral hygiene). We used structural equation modelling to test the hypothesis that SEP influences oral health directly and also indirectly via dental attendance and behavioural pathways. RESULTS: Lower SEP was directly associated with fewer natural teeth and worse self-rated oral health (standardised path coefficients, -0.21 (SE=0.01) and -0.10 (SE=0.01), respectively). We also found significant indirect effects via behavioural factors for both outcomes and via dental attendance primarily for self-rated oral health. While the standardised parameters of total effects were similar between the two outcomes, for number of teeth, the estimated effect of SEP was mostly direct while for self-rated oral health, it was almost equally split between direct and indirect effects. CONCLUSION: Reducing inequalities in dental attendance and health behaviours is necessary but not sufficient to tackle socioeconomic inequalities in oral health

Hydrogen activation by [NiFe]-hydrogenases

Hydrogenase-1 (Hyd-1) from Escherichia coli is a membrane-bound enzyme that catalyses the reversible oxidation of molecular H2 The active site contains one Fe and one Ni atom and several conserved amino acids including an arginine (Arg(509)), which interacts with two conserved aspartate residues (Asp(118) and Asp(574)) forming an outer shell canopy over the metals. There is also a highly conserved glutamate (Glu(28)) positioned on the opposite side of the active site to the canopy. The mechanism of hydrogen activation has been dissected by site-directed mutagenesis to identify the catalytic base responsible for splitting molecular hydrogen and possible proton transfer pathways to/from the active site. Previous reported attempts to mutate residues in the canopy were unsuccessful, leading to an assumption of a purely structural role. Recent discoveries, however, suggest a catalytic requirement, for example replacing the arginine with lysine (R509K) leaves the structure virtually unchanged, but catalytic activity falls by more than 100-fold. Variants containing amino acid substitutions at either or both, aspartates retain significant activity. We now propose a new mechanism: heterolytic H2 cleavage is via a mechanism akin to that of a frustrated Lewis pair (FLP), where H2 is polarized by simultaneous binding to the metal(s) (the acid) and a nitrogen from Arg(509) (the base)

A new primary dental care service compared with standard care for child and family to reduce the re-occurrence of childhood dental caries (Dental RECUR): study protocol for a randomised controlled trial

Background: In England and Scotland, dental extraction is the single highest cause of planned admission to the hospital for children under 11 years. Traditional dental services have had limited success in reducing this disease burden. Interventions based on motivational interviewing have been shown to impact positively dental health behaviours and could facilitate the prevention of re-occurrence of dental caries in this high-risk population. The objective of the study is to evaluate whether a new, dental nurse-led service, delivered using a brief negotiated interview based on motivational interviewing, is a more cost-effective service than treatment as usual, in reducing the re-occurrence of dental decay in young children with previous dental extractions. Methods/Design: This 2-year, two-arm, multicentre, randomised controlled trial will include 224 child participants, initially aged 5 to 7 years, who are scheduled to have one or more primary teeth extracted for dental caries under general anaesthesia (GA), relative analgesia (RA: inhalation sedation) or local anaesthesia (LA). The trial will be conducted in University Dental Hospitals, Secondary Care Centres or other providers of dental extraction services across the United Kingdom. The intervention will include a brief negotiated interview (based on the principles of motivational interviewing) delivered between enrolment and 6 weeks post-extraction, followed by directed prevention in primary dental care. Participants will be followed up for 2 years. The main outcome measure will be the dental caries experienced by 2 years post-enrolment at the level of dentine involvement on any tooth in either dentition, which had been caries-free at the baseline assessment. Discussion: The participants are a hard-to-reach group in which secondary prevention is a challenge. Lack of engagement with dental care makes the children and their families scheduled for extraction particularly difficult to recruit to an RCT. Variations in service delivery between sites have also added to the challenges in implementing the Dental RECUR protocol during the recruitment phase. Trial registration: ISRCTN24958829 (date of registration: 27 September 2013), Current protocol version: 5.0