Functional Effects of Epilepsy Associated KCNT1 Mutations Suggest Pathogenesis via Aberrant Inhibitory Neuronal Activity.

KCNT1 (K+ channel subfamily T member 1) is a sodium-activated potassium channel highly expressed in the nervous system which regulates neuronal excitability by contributing to the resting membrane potential and hyperpolarisation following a train of action potentials. Gain of function mutations in the KCNT1 gene are the cause of neurological disorders associated with different forms of epilepsy. To gain insights into the underlying pathobiology we investigated the functional effects of 9 recently published KCNT1 mutations, 4 previously studied KCNT1 mutations, and one previously unpublished KCNT1 variant of unknown significance. We analysed the properties of KCNT1 potassium currents and attempted to find a correlation between the changes in KCNT1 characteristics due to the mutations and severity of the neurological disorder they cause. KCNT1 mutations identified in patients with epilepsy were introduced into the full length human KCNT1 cDNA using quick-change site-directed mutagenesis protocol. Electrophysiological properties of different KCNT1 constructs were investigated using a heterologous expression system (HEK293T cells) and patch clamping. All mutations studied, except T314A, increased the amplitude of KCNT1 currents, and some mutations shifted the voltage dependence of KCNT1 open probability, increasing the proportion of channels open at the resting membrane potential. The T314A mutation did not affect KCNT1 current amplitude but abolished its voltage dependence. We observed a positive correlation between the severity of the neurological disorder and the KCNT1 channel open probability at resting membrane potential. This suggests that gain of function KCNT1 mutations cause epilepsy by increasing resting potassium conductance and suppressing the activity of inhibitory neurons. A reduction in action potential firing in inhibitory neurons due to excessively high resting potassium conductance leads to disinhibition of neural circuits, hyperexcitability and seizures.

A post-occupancy study of ventilation effectiveness from high-resolution CO2 monitoring at live theatre events to mitigate airborne transmission of SARS-CoV-2.

Mass-gathering events were closed around the world in 2020 to minimise the spread of the SARS-CoV-2 virus. Emerging research on the transmission of SARS-CoV-2 emphasised the importance of sufficient ventilation. This paper presents the results of an indoor air quality (IAQ) monitoring study over 82 events in seven mechanically ventilated auditoria to support the UK government Events Research Programme. Indoor carbon dioxide concentration was measured at high resolution before, during, and after occupancy to allow for assessment of the ventilation systems. Generally, good indoor air quality was measured in all auditoria, with average IAQ found to be excellent or very good for 70% of spaces. In some auditoria, spatial variation in IAQ was identified, indicating poor mixing of the air. In addition, surface and air samples were taken and analysed for the presence of bacteria by culture and SARS-CoV-2 using RT-qPCR in one venue. SARS-CoV-2 RNA was detected on a small number of surfaces at very low copy numbers, which are unlikely to pose an infection risk. Under the ventilation strategies and occupancy levels investigated, it is likely that most theatres pose a low risk of long-range transmission of COVID-19.

Narcissism, national narcissism, COVID-19 conspiracy belief, and social media use as predictors of compliance with COVID-19 public health guidelines.

Understanding health belief models, and the variables that influence adherence to public health measures imposed by local governments and international health bodies, is crucial to slowing down the spread of the virus that causes COVID-19. Conspiracy theories about the virus have quickly spread on social media and have been linked to reluctance to comply with COVID-19 regulations. Personality traits such as narcissism and collective national narcissism have also been associated with the way we perceive severity and susceptibility to the disease. To examine this further, participants (N = 183) completed an online questionnaire measuring belief in COVID-19 conspiracies, trait narcissism, national narcissism, and social media usage. A model containing these variables was able to significantly predict adherence to COVID-19 preventative health behaviours, with higher levels of COVID-19 conspiracy belief, narcissism, and social media usage all contributing to reduced adherence to recommended COVID-19 health behaviours. The findings suggest conspiracy beliefs, narcissism, and social media play a key role in adherence to behaviours orientated towards stopping the spread of COVID-19. Governments and social media companies need to demonstrate greater awareness of the negative effects of conspiracy theories spread through social media, in addition to awareness of how these effects may be greater in more narcissistic individuals.

Protein arginine methyl transferase 1- and Jumonji C domain-containing protein 6-dependent arginine methylation regulate hepatocyte nuclear factor 4 alpha expression and hepatocyte proliferation in mice.

Alcohol is a well-established risk factor for hepatocellular carcinoma (HCC), but the mechanisms by which it promotes liver cancer are not well understood. Several studies have shown that cellular protein arginine methylation is inhibited by alcohol. Arginine methylation is controlled by the reciprocal activity of protein arginine methyltransferases, primarily protein arginine methyl transferase 1 (PRMT1), and a demethylase Jumonji C domain-containing protein 6 (JMJD6). The aim of this study was to explore the role of arginine methylation changes in alcohol pathogenesis. We found that PRMT1 activity is inhibited in livers of mice fed with alcohol compared to pair-fed mice. Using hepatocyte-specific PRMT1 knockout mice, we identified that loss of PRMT1 results in enhanced hepatocyte proliferation and a 33% increase in liver size. This increased hepatocyte proliferation was associated with reduced expression of hepatocyte nuclear factor 4 alpha (Hnf4α), an important regulator of liver tumorigenesis. We found that PRMT1 regulates Hnf4α expression directly through arginine methylation at the (Hnf4α) promoter. In the absence of PRMT1, JMJD6 can demethylate the Hnf4α promoter and suppress its expression. We were able to restore Hnf4α expression and abolish the increase in hepatocyte proliferation by knockdown of JMJD6 in PRMT1 knockout mice. Knockdown of JMJD6 in alcohol-fed mice similarly increased Hnf4α expression. We then examined whether loss of arginine methylation might play a role in alcohol-associated liver cancers. We examined 25 human HCC specimens and found a strong correlation (R = 0.8; P < 0.01) between arginine methylation levels and Hnf4α expression in these specimens, suggesting that the above mechanism is relevant in patients. CONCLUSION: Taken together, these data suggest that PRMT1 inhibition, such as induced by alcohol, may result in epigenetic changes leading to loss of Hnf4α. This effect may contribute to alcohol's ability to promote liver tumors. (Hepatology 2018;67:1109-1126).

Arginine methylation regulates c-Myc-dependent transcription by altering promoter recruitment of the acetyltransferase p300.

Protein arginine methyltransferase 1 (PRMT1) is an essential enzyme controlling about 85% of the total cellular arginine methylation in proteins. We have shown previously that PRMT1 is an important regulator of innate immune responses and that it is required for M2 macrophage differentiation. c-Myc is a transcription factor that is critical in regulating cell proliferation and also regulates the M2 transcriptional program in macrophages. Here, we sought to determine whether c-Myc in myeloid cells is regulated by PRMT1-dependent arginine methylation. We found that PRMT1 activity was necessary for c-Myc binding to the acetyltransferase p300. PRMT1 inhibition decreased p300 recruitment to c-Myc target promoters and increased histone deacetylase 1 (HDAC1) recruitment, thereby decreasing transcription at these sites. Moreover, PRMT1 inhibition blocked c-Myc-mediated induction of several of its target genes, including peroxisome proliferator-activated receptor γ (PPARG) and mannose receptor C-type 1 (MRC1), suggesting that PRMT1 is necessary for c-Myc function in M2 macrophage differentiation. Of note, in primary human blood monocytes, p300-c-Myc binding was strongly correlated with PRMT1 expression, and in liver sections, PRMT1, c-Myc, and M2 macrophage levels were strongly correlated with each other. Both PRMT1 levels and M2 macrophage numbers were significantly lower in livers from individuals with a history of spontaneous bacterial peritonitis, known to have defective cellular immunity. In conclusion, our findings demonstrate that PRMT1 is an important regulator of c-Myc function in myeloid cells. PRMT1 loss in individuals with cirrhosis may contribute to their immune defects.

Airborne asbestos exposures associated with the installation and removal of roofing products.

Asbestos-containing roofing products were widely used throughout the 20th century, and certain products are still used in limited quantities today. Roofing products are generally considered non-friable and are not expected to release appreciable amounts of airborne asbestos fibers; however, despite the variety of roofing products that have contained asbestos over time, there are no comprehensive analyses of the exposure data associated with these products in the published literature. The objective of this study was to analyze the available data and characterize asbestos exposures associated with the installation, removal, and replacement of built-up roofing (BUR), felts, flashings, shingles, coatings, cements, and mastics under a variety of work practices. Published and unpublished literature that contained the following information was included in the analysis: (1) airborne fiber concentrations determined by PCM; (2) a description of the product(s) used; and (3) a description of the task(s) performed. More than 800 personal air samples from 12 studies performed between 1982 and 2010 were identified which fit the inclusion criteria. The findings indicate that short-term and full-shift exposures from the use of asbestos-containing roofing products were typically well below applicable occupational exposure limits. Additionally, the cumulative exposures associated with roofing work would be well below published chrysotile no-observed-adverse-effect-levels (NOAELs) for asbestos-related diseases.

COVID-19 risk mitigation in reopening mass cultural events: population-based observational study for the UK Events Research Programme in Liverpool City Region.

OBJECTIVES: To understand severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) transmission risks, perceived risks and the feasibility of risk mitigations from experimental mass cultural events before coronavirus disease 2019 (COVID-19) restrictions were lifted. DESIGN: Prospective, population-wide observational study. SETTING: Four events (two nightclubs, an outdoor music festival and a business conference) open to Liverpool City Region UK residents, requiring a negative lateral flow test (LFT) within the 36 h before the event, but not requiring social distancing or face-coverings. PARTICIPANTS: A total of 12,256 individuals attending one or more events between 28 April and 2 May 2021. MAIN OUTCOME MEASURES: SARS-CoV-2 infections detected using audience self-swabbed (5-7 days post-event) polymerase chain reaction (PCR) tests, with viral genomic analysis of cases, plus linked National Health Service COVID-19 testing data. Audience experiences were gathered via questionnaires, focus groups and social media. Indoor CO2 concentrations were monitored. RESULTS: A total of 12 PCR-positive cases (likely 4 index, 8 primary or secondary), 10 from the nightclubs. Two further cases had positive LFTs but no PCR. A total of 11,896 (97.1%) participants with scanned tickets were matched to a negative pre-event LFT: 4972 (40.6%) returned a PCR within a week. CO2 concentrations showed areas for improving ventilation at the nightclubs. Population infection rates were low, yet with a concurrent outbreak of >50 linked cases around a local swimming pool without equivalent risk mitigations. Audience anxiety was low and enjoyment high. CONCLUSIONS: We observed minor SARS-CoV-2 transmission and low perceived risks around events when prevalence was low and risk mitigations prominent. Partnership between audiences, event organisers and public health services, supported by information systems with real-time linked data, can improve health security for mass cultural events.

Measurement and rapid assessment of indoor air quality at mass gathering events to assess ventilation performance and reduce aerosol transmission of SARS-CoV-2.

To assess risk factors for COVID-19 transmission and address the closure of mass gathering events since March 2020, the UK Government ran the Events Research Programme (ERP), following which it reopened live events in sports, music, and culture in July 2021. We report the rapid post-occupancy evaluation of Indoor Air Quality (IAQ) and associated long-range airborne transmission risk conducted in the Environmental Study of the ERP. Ten large venues around the UK were monitored with CO2 sensors at a high spatial and temporal resolution during 90 events. An IAQ Index based on CO2 concentration was developed, and all monitored spaces were classified in bands from A to G based on their average and maximum CO2 concentrations from all events. High resolution monitoring and the IAQ Index depicted the overall state of ventilation at live events, and allowed identification of issues with ventilation effectiveness and distribution, and of spaces with poor ventilation and the settings in which long-range airborne transmission risk may be increased. In numerous settings, CO2 concentrations were found to follow patterns relating to event management and specific occupancy of spaces around the venues. Good ventilation was observed in 90% of spaces monitored for given occupancies. Practical applications: High-resolution monitoring of indoor CO2 concentrations is necessary to detect the spatial variation of indoor air quality (IAQ) in large mass gathering event venues. The paper summarises COVID-19 ventilation guidance for buildings and defines a methodology for measurement and rapid assessment of IAQ during occupancy at live events that can be implemented by venue managers. Comparisons of the CO2 concentrations measured during the events identified the spaces at high risk of long-range transmission of airborne pathogens. Building operators should be mindful of the ventilation strategies used relative to the total occupancy in different spaces and the occupant's activities.

The discovery of population differences in network community structure: new methods and applications to brain functional networks in schizophrenia.

The modular organization of the brain network can vary in two fundamental ways. The amount of inter- versus intra-modular connections between network nodes can be altered, or the community structure itself can be perturbed, in terms of which nodes belong to which modules (or communities). Alterations have previously been reported in modularity, which is a function of the proportion of intra-modular edges over all modules in the network. For example, we have reported that modularity is decreased in functional brain networks in schizophrenia: There are proportionally more inter-modular edges and fewer intra-modular edges. However, despite numerous and increasing studies of brain modular organization, it is not known how to test for differences in the community structure, i.e., the assignment of regional nodes to specific modules. Here, we introduce a method based on the normalized mutual information between pairs of modular networks to show that the community structure of the brain network is significantly altered in schizophrenia, using resting-state fMRI in 19 participants with childhood-onset schizophrenia and 20 healthy participants. We also develop tools to show which specific nodes (or brain regions) have significantly different modular communities between groups, a subset that includes right insular and perisylvian cortical regions. The methods that we propose are broadly applicable to other experimental contexts, both in neuroimaging and other areas of network science.

Efficacy and Safety Outcomes of Cataract Surgery in Survivors of Ebola Virus Disease: 12-Month Results From the PREVAIL VII Study.

Purpose: In survivors of Ebola virus disease (EVD), intraocular viral persistence raises questions about the timing and safety of cataract surgery. To the best of our knowledge, this is the first controlled study evaluating Ebola virus persistence and cataract surgery safety and outcomes in EVD survivors. Methods: Seropositive EVD survivors and seronegative controls with vision worse than 20/40 from cataract and without active intraocular inflammation were enrolled. Aqueous humor from survivors was tested with reverse transcription-polymerase chain reaction for Ebola viral RNA. Participants underwent manual small-incision cataract surgery and 1 year of follow-up examinations. Results: Twenty-two eyes of 22 survivors and 12 eyes of eight controls underwent cataract surgery. All of the aqueous samples tested negative for Ebola viral RNA. Median visual acuity improved from 20/200 at baseline to 20/25 at 1 year in survivors and from count fingers to 20/50 in controls (overall, P < 0.001; between groups, P = 0.07). After a 1-month course of topical corticosteroids, 55% of survivors and 67% of controls demonstrated at least 1+ anterior chamber cell. Twelve months after surgery, optical coherence tomography revealed a median increase in macular central subfield thickness of 42 µm compared with baseline (overall, P = 0.029; between groups, P = 0.995). Conclusions: EVD survivors and controls demonstrated significant visual improvement from cataract surgery. The persistence of intraocular inflammation highlights the importance of follow-up. The absence of detectable intraocular Ebola viral RNA provides guidance regarding the safety of eye surgery in Ebola survivors. Translational Relevance: These findings demonstrate the safety and efficacy of cataract surgery in Ebola survivors and will inform ocular surgery guidelines in this population.

Awareness of oral and genital human papillomavirus (HPV) infection in young adolescents prior to gender-neutral vaccination.

INTRODUCTION: Oral human papillomavirus (HPV) and oropharyngeal cancer prevalence are increasing, particularly in men. Raising greater awareness of male HPV disease is perceived as an important intervention strategy. This study investigated the effectiveness of HPV education on adolescents' perception of HPV disease and the impact of HPV vaccination on their sexual health. METHODS: An HPV questionnaire was completed by 357 UK-based adolescents, aged 12-13 years. RESULTS: Most adolescents knew HPV causes cervical cancer and HPV vaccination prevents this. A minority acknowledged HPV causes other genital cancers, with under one-fifth knowing HPV causes genital warts. Adolescents' awareness of HPV transmission activities were limited. There was very poor awareness of oral HPV infection or HPV-induced oropharyngeal cancer. Half of the participants stated HPV vaccination reduced their concerns about sexually transmitted infection contraction. Over half the males said they may take more sexual risks following vaccination, while a similar proportion of females did not expect their partner to take more risks. CONCLUSIONS: Adolescents had little awareness of male HPV infection and the role HPV vaccination can play in preventing these diseases. With variable rates of HPV vaccination uptake in males reported worldwide, this study indicates that in the UK greater emphasis on male HPV disease within educational information is required, to raise better awareness of how HPV affects both genders. As both genders preferred to receive education via healthcare professionals, educating a wider range of healthcare professionals on oral HPV could help facilitate awareness of HPV's role in head and neck cancer.

An examination of the profile and journey of patients with mental illness in the emergency department.

OBJECTIVE: To examine the profile and journey of patients with mental illness in the emergency departments (ED) of a Local Health District in Australia. There is limited evidence around the patient profile and journey of people in who present to EDs with mental illness, and the aim of this study was to inform the development of a psychiatric emergency service model. METHODS: The study design was a retrospective descriptive analysis of routinely collected data of two hospital EDs. Retrospective health care data of patients who presented to the EDs between July 2016 and June 2017 was analysed using descriptive and inferential statistics. RESULTS: Mental health presentations (N = 4506) consisted of 3.44% of all presentations to the Emergency Department, mostly for anxiety and stress, comorbid alcohol and drug issues, and suicidality. Females were more likely to present, and significantly more likely to present multiple times (specifically for age groups 12-17 and over 65). Presenting issues were associated with age and gender for most diagnostic groups, but not with number of presentations, except for personality disorder. 24.83% were admitted to a mental health inpatient unit, 54.34% were referred to a mental health community team, 4.63% medically, and the remaining referred to alcohol and drug services, GPs, or NGOs. Length of stay was associated with gender (female), age (>65) and diagnosis (drug related and eating disorder). CONCLUSION: There is limited evidence around the patient profile and journey of people who present to EDs for mental health reasons, and this study contributes to this literature.

The Complex Holiday Calendar of 1902: Responses to the Coronation of Edward VII and the Growth of Edwardian Event Fatigue.

The coronation of Edward VII and events to mark the end of the South African War led to a series of public ceremonies and events in the United Kingdom that had a profound effect on attitudes linked to national occasions and public holidays. This article explores the circumstances surrounding the numerous local and national holidays of 1902. It considers the decision-making process linked to the declaration of a coronation double-bank holiday, which demonstrated the inadequacy of contemporary legislation. The public response to the postponement of the coronation, due to the king's contraction of appendicitis, led to a period of 'event fatigue' in response to further ceremonial events. This showcased how much the British people guarded their right to holiday time and how the coronation had become more synonymous with celebration than with royal ceremony. It also showcased the degree to which the British people had been politicized and were ready to defend what they saw as their rights, in rejection of deference and traditional authority.

Role of the von Hippel-Lindau tumour suppressor protein in the regulation of HIF-1alpha and its oxygen-regulated transactivation domains at high cell density.

Hypoxia-inducible factor-1alpha (HIF-1alpha) induction and associated transcription were investigated during high cell density, focusing on the negative regulator of HIF-1alpha expression, the von Hippel-Lindau (VHL) protein. In 293T and HeLa cells, HIF-1alpha protein levels and associated transcription were induced as cells approached confluence. To determine whether these changes were due to a deficit in nuclear VHL-mediated ubiquitination of HIF-1alpha at confluence, cells were stably transfected with VHL. Overexpression of VHL in 293T cells had no demonstrable effect on the induction and nuclear accumulation of HIF-1alpha during high cell density or associated transcription. Moreover, RCC cells stably transfected with full-length VHL failed to exhibit the cell-density-dependent induction of HIF-1alpha noted in other cell lines. Investigation of both N-terminal and C-terminal (aa 727-826) oxygen-regulated proline and asparagine hydroxylation of HIF-1alpha revealed that both are inhibited during high cell density, as determined by impaired capture of HIF-1alpha by VHL and enhanced C-terminal transactivation. Finally, cell-density-mediated induction of HIF-1alpha and GLUT1 in RCC cells could be completely reconstituted by mutations in VHL binding affinity, suggesting that cell-density dependent induction of HIF-1alpha and transactivation may underpin some of the deregulated gene expression observed in VHL disease.

Role of nuclear and cytoplasmic localization in the tumour-suppressor activity of the von Hippel-Lindau protein.

Mutations in the von Hippel-Lindau (VHL) tumour-suppressor gene result in several forms of cancer. In the present study, we investigated the role of VHL subcellular localization in its antitumour properties. We generated renal cell carcinoma (RCC) lines stably expressing either exclusively nuclear (RCC/NLS-VHL), cytoplasmic (RCC/NES-VHL) or nucleo-cytoplasmic shuttling (RCC/DeltaNES-VHL or RCC/VHL) forms of VHL and investigated several parameters linked to tumorigenesis and known to be dysregulated in VHL disease. Remarkably, although the expression of wild-type VHL is largely cytoplasmic, all of the antitumour properties of VHL tested could be reconstituted by expressing exclusively nuclear VHL.

Role of the C-terminal alpha-helical domain of the von Hippel-Lindau protein in its E3 ubiquitin ligase activity.

In the present study, the role of the C-terminal alpha-helical domain (amino acid (aa) 195-208) of the von Hippel-Lindau (VHL) tumour suppressor was investigated. Deletions of the VHL C-terminus up to the naturally occurring 195-Gln-Term resulted in hypoxia-inducible factor (HIF)-1alpha downregulation in renal cell carcinoma (RCC)4 cells during normoxia, suggesting that this domain is not an absolute requirement for the ubiquitination of HIF-1alpha. However, detailed investigation of the ubiquitin protein isopeptide ligase ubiquitin ligase properties of VHL revealed C-terminal deletions to cause a significant impairment of HIF-1alpha ubiquitination, which is shown to be due to a loss in high-affinity binding to the target substrate. When VHL regulation of both HIF-1alpha N- and C-terminal oxygen-dependent degradation domains (HIF-ODDD) was investigated, it was found that only ubiquitination of the C-terminal HIF-ODDD was affected by the deletion of the VHL C-terminus. When RCC4 cells expressing C-terminal truncations of VHL were exposed to graded hypoxia, differences in the induction of HIF-1alpha were observed in comparison with full-length VHL, with a shift in the maximal induction of HIF-1alpha to a higher oxygen tension. These changes were accompanied by increased glucose transporter 1 expression, p300 CH1 domain binding and HIF-mediated reporter activity. We have thus defined a role for the C-terminal alpha-helical domain of VHL in the regulation of HIF-1alpha.

Role of CYP2E1 activity in endoplasmic reticulum ubiquitination, proteasome association, and the unfolded protein response.

In an experimental model of liver cirrhosis, marked increases in ER proteasome content in rat livers were observed 5 h after acute i.p. injection of the hepatotoxicant CCl4. To confirm the role of CYP2E1 in mediating protein misfolding/damage in the ER via its metabolism of CCl4, 293T cells stably transfected with human CYP2E1 were exposed to CCl4 and cell ER fractions assessed for ubiquitination. Increases in ER ubiquitin conjugates were noted in CYP2E1/293T cells treated with CCl4 and not in controls, suggesting these effects are CYP2E1 specific. Finally, the role of CYP2E1 in ER homeostasis was investigated by examining the unfolded protein response (UPR). When exposed to CCl4, CYP2E1/293T cells but not 293T or CYP1A2/293T cells showed rapid induction of the UPR-inducible ER chaperone BiP. Collectively, the data presented suggest that CYP2E1 is capable of inducing significant ER protein damage and stress via its catalytic activation of pro-oxidants.