Titin mutations in iPS cells define sarcomere insufficiency as a cause of dilated cardiomyopathy

Human mutations that truncate the massive sarcomere protein titin [TTN-truncating variants (TTNtvs)] are the most common genetic cause for dilated cardiomyopathy (DCM), a major cause of heart failure and premature death. Here we show that cardiac microtissues engineered from human induced pluripotent stem (iPS) cells are a powerful system for evaluating the pathogenicity of titin gene variants. We found that certain missense mutations, like TTNtvs, diminish contractile performance and are pathogenic. By combining functional analyses with RNA sequencing, we explain why truncations in the A-band domain of TTN cause DCM, whereas truncations in the I band are better tolerated. Finally, we demonstrate that mutant titin protein in iPS cell-derived cardiomyocytes results in sarcomere insufficiency, impaired responses to mechanical and {beta}-adrenergic stress, and attenuated growth factor and cell signaling activation. Our findings indicate that titin mutations cause DCM by disrupting critical linkages between sarcomerogenesis and adaptive remodeling

Saliva Proteins of Vector Culicoides Modify Structure and Infectivity of Bluetongue Virus Particles

Bluetongue virus (BTV) and epizootic haemorrhagic disease virus (EHDV) are related orbiviruses, transmitted between their ruminant hosts primarily by certain haematophagous midge vectors (Culicoides spp.). The larger of the BTV outer-capsid proteins, ‘VP2’, can be cleaved by proteases (including trypsin or chymotrypsin), forming infectious subviral particles (ISVP) which have enhanced infectivity for adult Culicoides, or KC cells (a cell-line derived from C. sonorensis). We demonstrate that VP2 present on purified virus particles from 3 different BTV strains can also be cleaved by treatment with saliva from adult Culicoides. The saliva proteins from C. sonorensis (a competent BTV vector), cleaved BTV-VP2 more efficiently than those from C. nubeculosus (a less competent / non-vector species). Electrophoresis and mass spectrometry identified a trypsin-like protease in C. sonorensis saliva, which was significantly reduced or absent from C. nubeculosus saliva. Incubating purified BTV-1 with C. sonorensis saliva proteins also increased their infectivity for KC cells ∼10 fold, while infectivity for BHK cells was reduced by 2–6 fold. Treatment of an ‘eastern’ strain of EHDV-2 with saliva proteins of either C. sonorensis or C. nubeculosus cleaved VP2, but a ‘western’ strain of EHDV-2 remained unmodified. These results indicate that temperature, strain of virus and protein composition of Culicoides saliva (particularly its protease content which is dependent upon vector species), can all play a significant role in the efficiency of VP2 cleavage, influencing virus infectivity. Saliva of several other arthropod species has previously been shown to increase transmission, infectivity and virulence of certain arboviruses, by modulating and/or suppressing the mammalian immune response. The findings presented here, however, demonstrate a novel mechanism by which proteases in Culicoides saliva can also directly modify the orbivirus particle structure, leading to increased infectivity specifically for Culicoides cells and, in turn, efficiency of transmission to the insect vector

Dimethyl fumarate in patients admitted to hospital with COVID-19 (RECOVERY): a randomised, controlled, open-label, platform trial

Dimethyl fumarate (DMF) inhibits inflammasome-mediated inflammation and has been proposed as a treatment for patients hospitalised with COVID-19. This randomised, controlled, open-label platform trial (Randomised Evaluation of COVID-19 Therapy [RECOVERY]), is assessing multiple treatments in patients hospitalised for COVID-19 (NCT04381936, ISRCTN50189673). In this assessment of DMF performed at 27 UK hospitals, adults were randomly allocated (1:1) to either usual standard of care alone or usual standard of care plus DMF. The primary outcome was clinical status on day 5 measured on a seven-point ordinal scale. Secondary outcomes were time to sustained improvement in clinical status, time to discharge, day 5 peripheral blood oxygenation, day 5 C-reactive protein, and improvement in day 10 clinical status. Between 2 March 2021 and 18 November 2021, 713 patients were enroled in the DMF evaluation, of whom 356 were randomly allocated to receive usual care plus DMF, and 357 to usual care alone. 95% of patients received corticosteroids as part of routine care. There was no evidence of a beneficial effect of DMF on clinical status at day 5 (common odds ratio of unfavourable outcome 1.12; 95% CI 0.86-1.47; p = 0.40). There was no significant effect of DMF on any secondary outcome

A deletion mutation in bovine \u3ci\u3eSLC4A2\u3c/i\u3e is associated with osteopetrosis in Red Angus cattle

Background: Osteopetrosis is a skeletal disorder of humans and animals characterized by the formation of overly dense bones, resulting from a deficiency in the number and/or function of bone-resorbing osteoclast cells. In cattle, osteopetrosis can either be induced during gestation by viral infection of the dam, or inherited as a recessive defect. Genetically affected calves are typically aborted late in gestation, display skull deformities and exhibit a marked reduction of osteoclasts. Although mutations in several genes are associated with osteopetrosis in humans and mice, the genetic basis of the cattle disorder was previously unknown. Results: We have conducted a whole-genome association analysis to identify the mutation responsible for inherited osteopetrosis in Red Angus cattle. Analysis of \u3e54,000 SNP genotypes for each of seven affected calves and nine control animals localized the defective gene to the telomeric end of bovine chromosome 4 (BTA4). Homozygosity analysis refined the interval to a 3.4-Mb region containing the SLC4A2 gene, encoding an anion exchanger protein necessary for proper osteoclast function. Examination of SLC4A2 from normal and affected animals revealed a ~2.8-kb deletion mutation in affected calves that encompasses exon 2 and nearly half of exon 3, predicted to prevent normal protein function. Analysis of RNA from a proven heterozygous individual confirmed the presence of transcripts lacking exons 2 and 3, in addition to normal transcripts. Genotyping of additional animals demonstrated complete concordance of the homozygous deletion genotype with the osteopetrosis phenotype. Histological examination of affected tissues revealed scarce, morphologically abnormal osteoclasts displaying evidence of apoptosis. Conclusions: These results indicate that a deletion mutation within bovine SLC4A2 is associated with osteopetrosis in Red Angus cattle. Loss of SLC4A2 function appears to induce premature cell death, and likely results in cytoplasmic alkalinization of osteoclasts which, in turn, may disrupt acidification of resorption lacunae

Automated Essay Scoring in Australian Schools: Key Issues and Recommendations

This summary outlines critical issues associated with the use of Automated Essay Scoring (AES) technology in the Australian education system. The key insights presented in this paper emerged from a collaborative, multi-stakeholder workshop held in July 2022 that explored an automated essay-scoring trial and generated future possibilities aligned with participant interests and expertise. Drawing on the workshop and our expert understanding of the wider landscape, we propose recommendations that can be adopted by various stakeholders, schools, and educational systems. There are compelling reasons for Australian schools and education departments to investigate the use of AES. AES could potentially alleviate aspects of teachers’ workload at a time when teacher attrition is historically high and teacher recruitment historically low. At the same time, AES also has the potential to de-professionalise and deskill of teachers. Educationalists are acutely aware that quality feedback can help students improve their learning across multiple subjects and domains, however parents and many are reluctant to hand that responsibility over to AES. In 2018, concerns among teachers, teachers’ unions, principles, and parents became apparent when the federal Department of Education, Skills and Employment attempted to implement a form of AES in The National Assessment Program – Literacy and Numeracy (NAPLAN). These concerns primarily registered around three issues: − de-professionalisation of teachers, − inequitable infrastructure in Australian schools, and − lack of transparency from examination authorities as to how marking decisions are made. The use of AES in NAPLAN ultimately proved to be politically unpopular, leading to its suspension. However, the growing implementation of AES in schools across the globe means that the use of this technology is likely to re-emerge as a controversial issue in Australia. Without political leadership in this area, it is ultimately up to educational institutions and agencies, policymakers, and school communities to assess the benefits and pitfalls of AES and navigate the way forward. Our recommendations will assist the emergence of good governance in this area. To begin, it is crucially important to identify whether AES will be used in high-stakes or low-stakes tests. High-stakes tests are defined as those with consequential outcomes for students or educators, such as the determination of progression of students or rankings of school institutions. If AES is to be used in Australian schools, the following issues must be considered: − the capacity of stakeholders, including principals, teachers, and parents, to understand how AES systems work − the infrastructure required to support the use of AES − the potential impacts of AES on assessment and workload practices which requires adequate professional development resources − competing interests and values between schools, departments, and institutions associated with using AES − how the use of AES relates to and integrates with broader policy frameworks. The investigation of these issues requires information sharing, dialogue, and negotiation among diverse stakeholders, including teachers, parents, students, leaders, and policymakers. In addition to this engagement, schools and other educational institutions must also discuss the implementation of AES tools with AES system developers and commercial vendors, so as to better understand the functions and limitations of the AES tool, as well as its implications for professional and assessment practices. Only then can decision-makers evaluate whether a specific AES system is worth the investment of funds and resources, including teacher workload, in both the medium and longer term. Although it appears as yet another drag on teacher time, the participatory and collaborative development of AES guidance, policy, and regulation is crucial. It ensures that pluralistic views and shared values are reflected in any innovations or reforms across the education sector. To ensure a collaborative foundation, the introduction of AES must be informed by stakeholder expertise across multiple locations and decision-making levels, including classrooms, schools, organisations, and state, territory, and national jurisdictions. For Australia, we recommend multi-scalar policy development informed by educators, policymakers, and representatives from educational technology companies engaging in cooperative learning and action