Ligand effects on gas adsorption in nanoporous phthalocyanine crystals.

X-ray diffraction is used to study the sorption of CO and NO in two phthalocyanine nanoporous crystals (PNCs) with 4,4' bipyridine or 4,4' bipyrimidine trans coordinated to open Co2+ sites, demonstrating how the trans coordinated ligands influence the gas sorption properties and structures of the PNCs.

Impact of essential genes on the success of genome editing experiments generating 3313 new genetically engineered mouse lines.

The International Mouse Phenotyping Consortium (IMPC) systematically produces and phenotypes mouse lines with presumptive null mutations to provide insight into gene function. The IMPC now uses the programmable RNA-guided nuclease Cas9 for its increased capacity and flexibility to efficiently generate null alleles in the C57BL/6N strain. In addition to being a valuable novel and accessible research resource, the production of 3313 knockout mouse lines using comparable protocols provides a rich dataset to analyze experimental and biological variables affecting in vivo gene engineering with Cas9. Mouse line production has two critical steps - generation of founders with the desired allele and germline transmission (GLT) of that allele from founders to offspring. A systematic evaluation of the variables impacting success rates identified gene essentiality as the primary factor influencing successful production of null alleles. Collectively, our findings provide best practice recommendations for using Cas9 to generate alleles in mouse essential genes, many of which are orthologs of genes linked to human disease.

Lawesson's Reagent: Providing a New Approach to the Forgotten 6-Thioverdazyl Radical.

A new method for the preparation of the underrepresented 1,5-dimethyl-6-thioverdazyl radicals has been developed employing Lawesson's reagent (LR). The synthetic route involves the direct thionation of the carbonyl group of the corresponding dialkylbishydrazone followed by cyclization to give the tetrazinanthione verdazyl precursor on a gram scale. Subsequent oxidation yields the 6-thioverdazyl radical. It was determined that thionation of substrates containing electron-withdrawing groups in the ortho- or para-positions was high yielding. In contrast, for the parent phenyl group or substrates bearing weakly electron-donating substituents, thionation efficiency was significantly reduced. This could be overcome by utilizing partial in situ cyclization, which occurs during work up, to generate the tetrazinanthione directly via a one-pot synthesis. Density functional theory suggests that the LR fragment interacts with the carbonyl prior to cycloaddition and subsequent to cycloreversion, leading to the thiocarbonyl. The electronic nature of the radical is characterized with electron paramagnetic resonance as well as the first report of 6-thioverdazyl redox properties.

Interventions for cold homes: a rapid review of the health impacts.

BACKGROUND: Cold homes are associated with an increased risk of adverse health outcomes for older people. To mitigate this risk, homes need to be heated to an appropriate temperature. This review aims to identify interventions designed to improve heating and temperatures within homes and summarize its impact on health, health service utilization and cost effectiveness. METHODS: A rapid review was conducted. Studies assessing the effects of structural, financial, or behavioural interventions designed to improve home temperatures of residents aged 18+ years were eligible. Searches were carried out in four databases. A search for grey literature, and backward and forward citation searching were performed. Data were summarized in a narrative synthesis and mapped using EPPI-Reviewer and EPPI-Mapper software. RESULTS: Eighteen studies reported across 19 publications were included. Structural interventions were associated with better mental health and quality of life, a reduction in health service utilization, and improvements in satisfaction with internal home temperature, social interactions and financial difficulties. The impact on physical health outcomes varied by age, gender and long-term conditions. Evidence about the impact of behavioural interventions was inconsistent. CONCLUSION: Structural improvements to increase home temperatures may offer the potential to improve some aspects of health. However, the impact on physical health, including which groups are most likely to benefit, is unclear. Key gaps include the lack of evidence about the impact of financial interventions, and the impact of all types of interventions, on quality of life, mortality and costs.

Characteristics of older unpaid carers in England: a study of social patterning from the English Longitudinal Study of Ageing.

BACKGROUND: A growing number of older people provide unpaid care, but contemporary research evidence on this group is limited. AIM: This study aims to describe the characteristics of older people who provide unpaid care and how these vary by socioeconomic position. METHODS: Using recent information from the English Longitudinal Study of Ageing (ELSA wave 9, 2019), we analysed cross-sectional data on 1,282 unpaid carers aged ≥50. Data on sociodemographics, health, social wellbeing, care intensity and caregiver-recipient relationships were extracted. Total net non-pension wealth quintiles were used as a relative measure of socioeconomic position. Differences between the poorest and richest wealth quintiles were examined through logistic regression. FINDINGS: Most older carers in ELSA were female and looking after another older person. Poor mental and physical health and social isolation were common, and socially patterned. Compared with carers in the middle wealth group, the poorest group were more likely to be living with the person they cared for (odds ratio (OR) 1.56 [95% confidence interval (CI) 1.03-2.36]) and more likely to experience loneliness (OR 2.29 [95% CI 1.42-3.69]), dependency (i.e. the need for help with activities of daily living) (OR 1.62 [95% CI 1.05-2.51]), chronic pain (OR 1.81 [95% CI 1.23-2.67]), a higher number of diseases (OR 1.75 [95% CI 1.15-2.65]) and fair/poor self-rated health (OR 2.59 [95% CI 1.79-3.76]). The poorest carers were also less likely to have a high quality of life (OR 0.51 [95% CI 0.33-0.80]) or be in work (OR 0.33 [95% CI 0.19-0.59]). CONCLUSION: Our findings suggest that financially disadvantaged unpaid carers (and their households) may have the greatest needs for intervention and support. Focussing resources on this group has potential to address social inequalities.

Long-read sequencing for fast and robust identification of correct genome-edited alleles: PCR-based and Cas9 capture methods.

BACKGROUND: Recent developments in CRISPR/Cas9 genome-editing tools have facilitated the introduction of precise alleles, including genetic intervals spanning several kilobases, directly into the embryo. However, the introduction of donor templates, via homology directed repair, can be erroneous or incomplete and these techniques often produce mosaic founder animals. Thus, newly generated alleles must be verified at the sequence level across the targeted locus. Screening for the presence of the desired mutant allele using traditional sequencing methods can be challenging due to the size of the interval to be sequenced, together with the mosaic nature of founders. METHODOLOGY/PRINCIPAL FINDINGS: In order to help disentangle the genetic complexity of these animals, we tested the application of Oxford Nanopore Technologies long-read sequencing at the targeted locus and found that the achievable depth of sequencing is sufficient to offset the sequencing error rate associated with the technology used to validate targeted regions of interest. We have assembled an analysis workflow that facilitates interrogating the entire length of a targeted segment in a single read, to confirm that the intended mutant sequence is present in both heterozygous animals and mosaic founders. We used this workflow to compare the output of PCR-based and Cas9 capture-based targeted sequencing for validation of edited alleles. CONCLUSION: Targeted long-read sequencing supports in-depth characterisation of all experimental models that aim to produce knock-in or conditional alleles, including those that contain a mix of genome-edited alleles. PCR- or Cas9 capture-based modalities bring different advantages to the analysis.

Mutant Tbl1x male mice have a short life span and do not breed: unexpected findings.

Humans with the mutation Y509C in transducin beta like 1 X-linked (TBL1X HGNC ID HGNC:11585) have been reported to present with the combination of central congenital hypothyroidism and impaired hearing. TBL1X belongs to the WD40 repeat-containing protein family, is part of NCoR and SMRT corepressor complexes, and thereby involved in thyroid hormone signaling. In order to investigate the effects of the Y509C mutation in TBL1X on cellular thyroid hormone action, we aimed to generate a hemizygous male mouse cohort carrying the Tbl1x Y459C mutation which is equivalent to the human TBL1X Y509C mutation using CRISPR/Cas9 technology. Hemizygous male mice were small at birth and inactive. Their life span (median life span 93 days) was very short compared with heterozygous female mice (survived to >200 days with no welfare issues). About 52% of mice did not survive to weaning (133 mice). Of the remaining 118 mice, only 8 were hemizygous males who were unable to mate whereby it was impossible to generate homozygous female mice. In conclusion, the Tbl1x Y459C mutation in male mice has a marked negative effect on birth weight, survival, and fertility of male mice. The present findings are unexpected as they are in contrast to the mild phenotype in human males carrying the equivalent TBL1X Y509C mutation.

The small molecule raptinal can simultaneously induce apoptosis and inhibit PANX1 activity.

Discovery of new small molecules that can activate distinct programmed cell death pathway is of significant interest as a research tool and for the development of novel therapeutics for pathological conditions such as cancer and infectious diseases. The small molecule raptinal was discovered as a pro-apoptotic compound that can rapidly trigger apoptosis by promoting the release of cytochrome c from the mitochondria and subsequently activating the intrinsic apoptotic pathway. As raptinal is very effective at inducing apoptosis in a variety of different cell types in vitro and in vivo, it has been used in many studies investigating cell death as well as the clearance of dying cells. While examining raptinal as an apoptosis inducer, we unexpectedly identified that in addition to its pro-apoptotic activities, raptinal can also inhibit the activity of caspase-activated Pannexin 1 (PANX1), a ubiquitously expressed transmembrane channel that regulates many cell death-associated processes. By implementing numerous biochemical, cell biological and electrophysiological approaches, we discovered that raptinal can simultaneously induce apoptosis and inhibit PANX1 activity. Surprisingly, raptinal was found to inhibit cleavage-activated PANX1 via a mechanism distinct to other well-described PANX1 inhibitors such as carbenoxolone and trovafloxacin. Furthermore, raptinal also interfered with PANX1-regulated apoptotic processes including the release of the 'find-me' signal ATP, the formation of apoptotic cell-derived extracellular vesicles, as well as NLRP3 inflammasome activation. Taken together, these data identify raptinal as the first compound that can simultaneously induce apoptosis and inhibit PANX1 channels. This has broad implications for the use of raptinal in cell death studies as well as in the development new PANX1 inhibitors.

Characterisation and prion transmission study in mice with genetic reduction of sporadic Creutzfeldt-Jakob disease risk gene Stx6.

Sporadic Creutzfeldt-Jakob disease (sCJD), the most common human prion disease, is thought to occur when the cellular prion protein (PrPC) spontaneously misfolds and assembles into prion fibrils, culminating in fatal neurodegeneration. In a genome-wide association study of sCJD, we recently identified risk variants in and around the gene STX6, with evidence to suggest a causal increase of STX6 expression in disease-relevant brain regions. STX6 encodes syntaxin-6, a SNARE protein primarily involved in early endosome to trans-Golgi network retrograde transport. Here we developed and characterised a mouse model with genetic depletion of Stx6 and investigated a causal role of Stx6 expression in mouse prion disease through a classical prion transmission study, assessing the impact of homozygous and heterozygous syntaxin-6 knockout on disease incubation periods and prion-related neuropathology. Following inoculation with RML prions, incubation periods in Stx6-/- and Stx6+/- mice differed by 12 days relative to wildtype. Similarly, in Stx6-/- mice, disease incubation periods following inoculation with ME7 prions also differed by 12 days. Histopathological analysis revealed a modest increase in astrogliosis in ME7-inoculated Stx6-/- animals and a variable effect of Stx6 expression on microglia activation, however no differences in neuronal loss, spongiform change or PrP deposition were observed at endpoint. Importantly, Stx6-/- mice are viable and fertile with no gross impairments on a range of neurological, biochemical, histological and skeletal structure tests. Our results provide some support for a pathological role of Stx6 expression in prion disease, which warrants further investigation in the context of prion disease but also other neurodegenerative diseases considering syntaxin-6 appears to have pleiotropic risk effects in progressive supranuclear palsy and Alzheimer's disease.

Systematic review of the relationship between burn-out and spiritual health in doctors.

OBJECTIVE: To investigate the relationship between burn-out and spiritual health among medical doctors. DESIGN: Systematic literature review and narrative synthesis of cross-sectional studies. SETTING: Any setting, worldwide. DATA SOURCES: Five databases were searched from inception to March 2022, including Medline, Embase, PsycINFO, Scopus and Web of Science. ELIGIBILITY CRITERIA: Any study design that involved medical doctors (and other healthcare staff if assessed alongside medical doctors), that measured (in any way) both burn-out (or similar) and spiritual health (or similar) medical doctors. DATA EXTRACTION AND SYNTHESIS: All records were double screened. Data extraction was performed by one reviewer and a proportion (10%) checked by a second reviewer. Quality was assessed using the Appraisal of Cross-sectional Studies tool. Due to the heterogeneity of the included studies, a narrative review was undertaken without a meta-analysis. RESULTS: Searches yielded 1049 studies. 40 studies met eligibility criteria and were included in this review. Low reported levels of spirituality were associated with high burn-out scores and vice versa. Religion was not significantly associated with lower levels of burn-out. Few studies reported statistically significant findings, few used validated spiritual scores and most were vulnerable to sampling bias. CONCLUSIONS: Published research suggests that burn-out is linked to spiritual health in medical doctors but not to religion. Robust research is needed to confirm these findings and develop effective interventions. PROSPERO REGISTRATION NUMBER: CRD42020200145.

Tandem High-Pressure Crystallography-Optical Spectroscopy Unpacks Noncovalent Interactions of Piezochromic Fluorescent Molecular Rotors.

To develop luminescent molecular materials with predictable and stimuli-responsive emission, it is necessary to correlate changes in their geometries, packing structures, and noncovalent interactions with the associated changes in their optical properties. Here, we demonstrate that high-pressure single-crystal X-ray diffraction can be combined with high-pressure UV-visible absorption and fluorescence emission spectroscopies to elucidate how subtle changes in structure influence optical outputs. A piezochromic aggregation-induced emitter, sym-heptaphenylcycloheptatriene (Ph7C7H), displays bathochromic shifts in its absorption and emission spectra at high pressure. Parallel X-ray measurements identify the pressure-induced changes in specific phenyl-phenyl interactions responsible for the piezochromism. Pairs of phenyl rings from neighboring molecules approach the geometry of a stable benzene dimer, while conformational changes alter intramolecular phenyl-phenyl interactions correlated with a relaxed excited state. This tandem crystallographic and spectroscopic analysis provides insights into how subtle structural changes relate to the photophysical properties of Ph7C7H and could be applied to a library of similar compounds to provide general structure-property relationships in fluorescent organic molecules with rotor-like geometries.

Pressure-induced postsynthetic cluster anion substitution in a MIL-53 topology scandium metal-organic framework.

Postsynthetic modification of metal-organic frameworks (MOFs) has proven to be a hugely powerful tool to tune physical properties and introduce functionality, by exploiting reactive sites on both the MOF linkers and their inorganic secondary building units (SBUs), and so has facilitated a wide range of applications. Studies into the reactivity of MOF SBUs have focussed solely on removal of neutral coordinating solvents, or direct exchange of linkers such as carboxylates, despite the prevalence of ancillary charge-balancing oxide and hydroxide ligands found in many SBUs. Herein, we show that the µ2-OH ligands in the MIL-53 topology Sc MOF, GUF-1, are labile, and can be substituted for µ2-OCH3 units through reaction with pore-bound methanol molecules in a very rare example of pressure-induced postsynthetic modification. Using comprehensive solid-state NMR spectroscopic analysis, we show an order of magnitude increase in this cluster anion substitution process after exposing bulk samples suspended in methanol to a pressure of 0.8 GPa in a large volume press. Additionally, single crystals compressed in diamond anvil cells with methanol as the pressure-transmitting medium have enabled full structural characterisation of the process across a range of pressures, leading to a quantitative single-crystal to single-crystal conversion at 4.98 GPa. This unexpected SBU reactivity - in this case chemisorption of methanol - has implications across a range of MOF chemistry, from activation of small molecules for heterogeneous catalysis to chemical stability, and we expect cluster anion substitution to be developed into a highly convenient novel method for modifying the internal pore surface and chemistry of a range of porous materials.

Plastic pollution on the world's coral reefs.

Coral reefs are losing the capacity to sustain their biological functions1. In addition to other well-known stressors, such as climatic change and overfishing1, plastic pollution is an emerging threat to coral reefs, spreading throughout reef food webs2, and increasing disease transmission and structural damage to reef organisms3. Although recognized as a global concern4, the distribution and quantity of plastics trapped in the world's coral reefs remains uncertain3. Here we survey 84 shallow and deep coral ecosystems at 25 locations across the Pacific, Atlantic and Indian ocean basins for anthropogenic macrodebris (pollution by human-generated objects larger than 5 centimetres, including plastics), performing 1,231 transects. Our results show anthropogenic debris in 77 out of the 84 reefs surveyed, including in some of Earth's most remote and near-pristine reefs, such as in uninhabited central Pacific atolls. Macroplastics represent 88% of the anthropogenic debris, and, like other debris types, peak in deeper reefs (mesophotic zones at 30-150 metres depth), with fishing activities as the main source of plastics in most areas. These findings contrast with the global pattern observed in other nearshore marine ecosystems, where macroplastic densities decrease with depth and are dominated by consumer items5. As the world moves towards a global treaty to tackle plastic pollution6, understanding its distribution and drivers provides key information to help to design the strategies needed to address this ubiquitous threat.

High-pressure single-crystal diffraction at the Australian Synchrotron.

A new high-pressure single-crystal diffraction setup has been designed and implemented at the Australian Synchrotron for collecting molecular and protein crystal structures. The setup incorporates a modified micro-Merrill-Bassett cell and holder designed specifically to fit onto the horizontal air-bearing goniometer, allowing high-pressure diffraction measurements to be collected with little to no modification of the beamline setup compared with ambient data collections. Compression data for the amino acid, L-threonine, and the protein, hen egg-white lysozyme, were collected, showcasing the capabilities of the setup.

Investigating the Role of Heparanase in Breast Cancer Development Utilising the MMTV-PyMT Murine Model of Mammary Carcinoma.

Breast cancer is the second most common human malignancy and is a major global health burden. Heparanase (HPSE) has been widely implicated in enhancing the development and progression of solid tumours, including breast cancer. In this study, the well-established spontaneous mammary tumour-developing MMTV-PyMT murine model was utilised to examine the role of HPSE in breast cancer establishment, progression, and metastasis. The use of HPSE-deficient MMTV-PyMT (MMTV-PyMTxHPSE-/-) mice addressed the lack of genetic ablation models to investigate the role of HPSE in mammary tumours. It was demonstrated that even though HPSE regulated mammary tumour angiogenesis, mammary tumour progression and metastasis were HPSE-independent. Furthermore, there was no evidence of compensatory action by matrix metalloproteinases (MMPs) in response to the lack of HPSE expression in the mammary tumours. These findings suggest that HPSE may not play a significant role in the mammary tumour development of MMTV-PyMT animals. Collectively, these observations may have implications in the clinical setting of breast cancer and therapy using HPSE inhibitors.

Genotyping Genome-Edited Founders and Subsequent Generation.

Targeted nucleases allow the production of many types of genetic mutations directly in the early embryo. However, the outcome of their activity is a repair event of unpredictable nature, and the founder animals that are produced are generally of a mosaic nature. Here, we present the molecular assays and genotyping strategies that will support the screening of the first generation for potential founders and the validation of positive animals in the subsequent generation, according to the type of mutation generated.

Which interventions are effective at decreasing or increasing emergency department attendances or hospital admissions from long-term care facilities? A systematic review.

OBJECTIVE: UK long-term care facility residents account for 185 000 emergency hospital admissions each year. Avoidance of unnecessary hospital transfers benefits residents, reduces demand on the healthcare systems but is difficult to implement. We synthesised evidence on interventions that influence unplanned hospital admissions or attendances by long-term care facility residents. METHODS: This is a systematic review of randomised controlled trials. PubMed, MEDLINE, EMBASE, ISI Web of Science, CINAHL and the Cochrane Library were searched from 2012 to 2022, building on a review published in 2013. We included randomised controlled trials that evaluated interventions that influence (decrease or increase) acute hospital admissions or attendances of long-term care facility residents. Risk of bias and evidence quality were assessed using Cochrane Risk Of Bias-2 and Grading of Recommendations Assessment, Development and Evaluation. RESULTS: Forty-three randomised studies were included in this review. A narrative synthesis was conducted and the weight of evidence described with vote counting. Advance care planning and goals of care setting appear to be effective at reducing hospitalisations from long-term care facilities. Other effective interventions, in order of increasing risk of bias, were: nurse practitioner/specialist input, palliative care intervention, influenza vaccination and enhancing access to intravenous therapies in long-term care facilities. CONCLUSIONS: Factors that affect hospitalisation and emergency department attendances of long-term care facility residents are complex. This review supports the already established use of advance care planning and influenza vaccination to reduce unscheduled hospital attendances. It is likely that more than one intervention will be needed to impact on healthcare usage across the long-term care facility population. The findings of this review are useful to identify effective interventions that can be combined, as well as highlighting interventions that either need evaluation or are not effective at decreasing healthcare usage. PROSPERO REGISTRATION NUMBER: CRD42020169604.

Pressure and guest-mediated pore shape modification in a small pore MOF to 1200 bar.

Guest-mediated pore-shape modification of the metal-organic framework, Sc2BDC3 upon adsorption of n-pentane and isopentane is examined from 50-1200 bar. Rotation of the BDC linker responsible for the change in pore shape occurs at much lower pressures than previously reported, with distinct adsorption behaviour observed between pentane isomers.