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Abstract The GRAMD genes are involved in maintaining cholesterol homeostasis, apoptosis, cancer and production traits in livestock. A lipid-binding GRAM domain is implicated in lipid transport and metabolism. The functions of GRAMD proteins remain incompletely understood. The aim of the present study was therefore to investigate the associations between six GRAMD genes in cattle using data from the international genomic evaluation of the Interbull InterGenomics Centre and to evaluate genotype-phenotype associations in human, cattle, pig, mouse and, chicken. Genotyping of 55,013 bulls was performed using DNA microarrays and 11 SNPs were mapped to the five GRAMD genes. A phenome-wide association study (PheWAS) tested associations between the 11 SNPs and 36 traits. The integrated analysis of SNP effects, rankings, and clustering patterns revealed their potential for improving cattle productivity, health, and robustness, and established a baseline for the targeted improvement of cattle traits. This study lays the groundwork for functional experiments aimed at uncovering the mechanism of action of GRAMD genes and to evaluate the potential of using GRAMD sequence variants for selection programs in dairy cattle. The study presents an example of how the combination of GWAS and the PheWAS offers a promising toolbox for the systematic functional annotation of vertebrate genomes.

Abstract Various signaling pathways are essential for both the innate immune response and the maintenance of cell homeostasis, requiring coordinated interactions among them. In this study, a mutation in the caspase-1 recognition site within MyD88 abolished inflammasome-dependent negative regulation, causing phenotypic changes in mice with some similarities to human NEMO-deficiencies. The MyD88D162E mutation reduced MyD88 protein levels and colon inflammation in DSS-induced colitis mice but did not affect cytokine expression in bone marrow-derived macrophages (BMDMs). However, compared to MyD88wt counterparts, MyD88D162E BMDMs had increased oxidative stress and dysfunctional mitochondria, along with reduced prosurvival Bcl-xL and BTK expression, rendering cells more prone to apoptosis, exacerbated by ibrutinib treatment. NF-κB activation by lipopolysaccharide mitigated this sensitive phenotype. These findings underscore the importance of MyD88wt signaling for NF-κB activation, protecting against macrophage premature apoptosis at resting state. Targeting MyD88 quantity rather than just its signaling could be a promising strategy for MyD88-driven lymphoma treatment.

Background: The paediatric use of ophthalmic chloramphenicol in New Zealand (NZ) is relatively high; however, little more is known about its utilisation, including whether this is equitable. This study aimed to describe chloramphenicol utilisation in NZ children aged five years and under, by patient ethnicity, socioeconomic deprivation, and urban/non-urban domicile. Methods: This analysis included every publicly subsidised chloramphenicol dispensing received from birth to five years of age, for every child born in NZ in 2013. Cumulative proportion of first exposure, dispensing rate per person-year, and seasonality of dispensing were quantified. These were calculated following stratification by ethnicity, socioeconomic deprivation quintile, and urban/non-urban health district. For cumulative proportion of first exposure, odds ratios (OR) were calculated and multivariate logistic regression was performed. For dispensing rate, incidence rate ratios (IRR) were calculated and zero-inflated Poisson regression was performed. Results: Almost one-quarter of NZ children received their first dispensing within the first year of life. By five years of age, 55.2% of children had received their first dispensing. By five years of age, children of Pacific ethnicity, those in the highest deprivation quintile, and in those non-urban health districts had lower odds of receiving chloramphenicol (adjusted OR 0.90, 0.79, and 0.81, respectively, all p < 0.001). In contrast, children of Māori ethnicity had higher odds (adjusted OR 1.99, p < 0.001). Māori and Pacific ethnicity, and residence in non-urban health districts, were associated with fewer dispensings (adjusted IRR 0.88, 0.75 and 0.87, all p < 0.001). In contrast, deprivation quintile was not significantly associated with dispensing rate. Conclusion: Chloramphenicol utilisation is prevalent among NZ children, and utilisation may be lower among children of Pacific ethnicity and those in non-urban areas

Objectives To compare the treatment effectiveness of secukinumab in radiographic (r) versus non-radiographic (nr) axial spondyloarthritis (axSpA) patients treated in routine care across Europe.Methods Prospectively collected data on secukinumab-treated axSpA patients with known radiographic status were pooled from nine countries.Remission rates based on patient-reported outcomes (PROs; Numeric Rating Scale (0–10), for example, pain ≤2/Bath Ankylosing Spondylitis Disease Activity Index (BASDAI) ≤2 and Ankylosing Spondylitis Disease Activity Score (ASDAS) inactive disease (ID)

Background. Enhanced B-cell presentation of donor alloantigen relative to presentation of HLA-mismatched reference alloantigen is associated with acute cellular rejection (ACR), when expressed as a ratio called the antigen presenting index (API) in an exploratory cohort of liver and intestine transplant (LT and IT) recipients. Methods. To test clinical performance, we measured the API using the previously described 6-h assay in 84 LT and 54 IT recipients with median age 3.3 y (0.05–23.96). Recipients experiencing ACR within 60 d after testing were termed rejectors. Results. We first confirmed that B-cell uptake and presentation of alloantigen induced and thus reflected the alloresponse of T-helper cells, which were incubated without and with cytochalasin and primaquine to inhibit antigen uptake and presentation, respectively. Transplant recipients included 76 males and 62 females. Rejectors were tested at median 3.6 d before diagnosis. The API was higher among rejectors compared with nonrejectors (2.2 ± 0.2 versus 0.6 ± 0.04, P value = 1.7E-09). In logistic regression and receiver-operating-characteristic analysis, API ≥1.1 achieved sensitivity, specificity, and positive and negative predictive values for predicting ACR in 99 training set samples. Corresponding metrics ranged from 80% to 88% in 32 independent posttransplant samples, and 73% to 100% in 20 independent pretransplant samples. In time-to-event analysis, API ≥1.1 predicted higher incidence of late donor-specific anti-HLA antibodies after API measurements in LT recipients (P = 0.011) and graft loss in IT recipients (P = 0.008), compared with recipients with API

BackgroundArtificial intelligence (AI)–powered technologies are being increasingly used in almost all fields, including medicine. However, to successfully implement medical AI applications, ensuring trust and acceptance toward such technologies is crucial for their successful spread and timely adoption worldwide. Although AI applications in medicine provide advantages to the current health care system, there are also various associated challenges regarding, for instance, data privacy, accountability, and equity and fairness, which could hinder medical AI application implementation. ObjectiveThe aim of this study was to identify factors related to trust in and acceptance of novel AI-powered medical technologies and to assess the relevance of those factors among relevant stakeholders. MethodsThis study used a mixed methods design. First, a rapid review of the existing literature was conducted, aiming to identify various factors related to trust in and acceptance of novel AI applications in medicine. Next, an electronic survey including the rapid review–derived factors was disseminated among key stakeholder groups. Participants (N=22) were asked to assess on a 5-point Likert scale (1=irrelevant to 5=relevant) to what extent they thought the various factors (N=19) were relevant to trust in and acceptance of novel AI applications in medicine. ResultsThe rapid review (N=32 papers) yielded 110 factors related to trust and 77 factors related to acceptance toward AI technology in medicine. Closely related factors were assigned to 1 of the 19 overarching umbrella factors, which were further grouped into 4 categories: human-related (ie, the type of institution AI professionals originate from), technology-related (ie, the explainability and transparency of AI application processes and outcomes), ethical and legal (ie, data use transparency), and additional factors (ie, AI applications being environment friendly). The categorized 19 umbrella factors were presented as survey statements, which were evaluated by relevant stakeholders. Survey participants (N=22) represented researchers (n=18, 82%), technology providers (n=5, 23%), hospital staff (n=3, 14%), and policy makers (n=3, 14%). Of the 19 factors, 16 (84%) human-related, technology-related, ethical and legal, and additional factors were considered to be of high relevance to trust in and acceptance of novel AI applications in medicine. The patient’s gender, age, and education level were found to be of low relevance (3/19, 16%). ConclusionsThe results of this study could help the implementers of medical AI applications to understand what drives trust and acceptance toward AI-powered technologies among key stakeholders in medicine. Consequently, this would allow the implementers to identify strategies that facilitate trust in and acceptance of medical AI applications among key stakeholders and potential users.

ABSTRACT Aromatoleum aromaticum EbN1T anaerobically degrades phenol, p-cresol, and p-ethylphenol, each via a distinct peripheral pathway. The compound-specific regulation of each pathway is proposed to occur on the transcriptional level in the case of phenol supposedly mediated by the one-component system PheR. To confirm its predicted function, we generated an unmarked, in-frame deletion mutant (ΔpheR). This mutant did not express the ppsA1 gene, which encodes the A1 subunit of phenol-activating phenylphosphate synthase. The expression of ppsA1 was restored by in trans complementation of pheR into the ΔpheR background. The responsiveness to phenol was studied in vivo in benzoate-limited anaerobic cultures by adding, upon benzoate depletion, single defined pulses of phenol (from 100 µM down to 0.1 nM). Time-resolved, targeted transcript profiling by qRT-PCR revealed a response threshold for ppsA1 expression of 30‒50 nM phenol. Notably, ppsA1 expression could not be induced by p-cresol or p-ethylphenol. Conversely, lack of expression was also observed for the additional target genes cmh (p-cresol degradation) and acsA1 (p-ethylphenol degradation) applying phenol or p-ethylphenol as well as phenol or p-cresol as stimuli. Thus, the sensory proteins PheR, PcrS, and EtpR should be highly selective for phenol, p-cresol, and p-ethylphenol, respectively. The implicated incapability of cross-stimulus binding was corroborated by comparing the predicted 3D structural models of the proteins’ sensory domains. While the ligand-binding pockets share the conserved hydroxy group-anchoring histidine and tryptophane, their distal faces in PcrS and EtpR are, compared to PheR, enlarged to accommodate the bulkier methyl (p-cresol) and ethyl group (p-ethylphenol), respectively. IMPORTANCE Aromatic compounds are globally abundant organic molecules with a multitude of natural and anthropogenic sources, underpinning the relevance of their biodegradation. A. aromaticum EbN1T is a well-studied environmental betaproteobacterium specialized on the anaerobic degradation of aromatic compounds. The here studied responsiveness toward phenol in conjunction with the apparent high ligand selectivity (non-promiscuity) of its PheR sensor and those of the related p-cresol (PcrS) and p-ethylphenol (EtpR) sensors are in accord with the substrate-specificity and biochemical distinctiveness of the associated degradation pathways. Furthermore, the present findings advance our general understanding of the substrate-specific regulation of the strain’s remarkable degradation network and of the concentration thresholds below which phenolic compounds become essentially undetectable and as a consequence should escape substantial biodegradation. Furthermore, the findings may inspire biomimetic sensor designs for detecting and quantifying phenolic contaminants in wastewater or environments.

It is not well-understood how type 1 diabetes (T1DM) affects skeletal muscle histological phenotype, particularly capillarisation. This study aimed to analyze skeletal muscle myosin heavy chain (MyHC) fibre type changes and 3D capillary network characteristics in experimental T1DM mice. Female C57BL/6J-OlaHsd mice were categorized into streptozotocin (STZ)-induced diabetic (n = 12) and age-matched non-diabetic controls (n =12). The muscle fibre phenotype of the soleus, gluteus maximus, and gastrocnemius muscles were characterized based on the expression of MyHC isoforms, while capillaries of the gluteus maximus were assessed with immunofluorescence staining, confocal laser microscopy and 3D image analysis. STZ-induced diabetic mice exhibited elevated glucose levels, reduced body weight, and prolonged thermal latency, verifying the T1DM phenotype. In both T1DM and non-diabetic mice, the gluteus maximus and gastrocnemius muscles predominantly expressed fast-twitch type 2b fibers, with no significant differences noted. However, the soleus muscle in non-diabetic mice had a greater proportion of type 2a fibers and comparable type 1 fiber densities (26.2 ± 14.6% vs 21.9 ± 13.5%) relative to diabetic mice. T1DM mice showed reduced fiber diameters (P = 0.026), and the 3D capillary network analysis indicated a higher capillary length per muscle volume in the gluteus maximus of diabetic mice compared to controls (P < 0.05). Overall, T1DM induced significant changes in the skeletal muscle, including shifts in MyHC fibre types, decreased fibre diameters, and increased relative capillarisation, possibly due to muscle fibre atrophy. Our findings emphasize the superior detail provided by the 3D analytical method for characterizing skeletal muscle capillary architecture, highlighting caution in interpreting 2D data for capillary changes in T1DM.

Abstract Objectives To determine whether exempting people (with high health needs and living in areas of high deprivation) from a $5 prescription charge reduces hospital use. Design Two-group parallel prospective randomised controlled trial. Setting People living in the community in various regions of New Zealand. Participants One thousand sixty one people who lived in areas of high socioeconomic deprivation, and either took medicines for diabetes, took antipsychotic medicines, or had chronic obstructive pulmonary disease (COPD). Of the 1053 who completed the study, just under half (49%) were Māori. Interventions Participants were individually randomized (1–1 ratio) to either be exempted from the standard $5 charge per prescription item for one year (2020-2021) (n = 591) or usual care (n = 469). Those in the intervention group did not pay the standard NZ$5 charge, and pharmacies billed the study for these. Participants continued to pay any other costs for prescription medicines. Those in the control group continued to pay all prescription charges for the year although they may have received one-off assistance from other agencies. Main outcome measures The primary outcome was length of stay (hospital bed-days). Secondary outcomes presented in this paper included: all-cause hospitalisations, hospitalisations for diabetes/mental health problems/COPD, deaths, and emergency department visits. Results The trial was under-powered because the recruitment target was not met. There was no statistically significant reduction in the primary outcome, hospital bed-days (IRR = 0.68, CI: 0.54 to 1.05). Participants in the intervention group were significantly less likely to be hospitalised during the study year than those in the control group (OR = 0.70, CI: 0.54 to 0.90). There were statistically significant reductions in the number of hospital admissions for mental health problems (IRR = 0.39, CI: 0.17 to 0.92), the number of admissions for COPD (IRR = 0.37, CI: 0.16 to 0.85), and length of stay for COPD (IRR 0.20, CI: 0.07 to 0.60). Apart from all-cause mortality and diabetes length of stay, all measures were better for the intervention group than the control group. Conclusions Eliminating a small co-payment appears to have had a substantial effect on patients’ risk of being hospitalised. Given the small amount of revenue gathered from the charges, and the comparative large costs of hospitalisations, the results suggest that these charges are likely to increase the overall cost of healthcare, as well as exacerbate ethnic inequalities. Trial registration Australian New Zealand Clinical Trials Registry (ANZCTR): ACTRN12618001486213 registered on 04/09/2018.

In Europe, the Alpine ibex (Capra ibex) was on the brink of extinction in the 19th century. Therefore, different conservation measures were implemented, and several reintroductions were made in the Alpine arc, starting from the only surviving population in Gran Paradiso, Italy. An extreme historical bottleneck and additional reintroductions have strongly shaped the genetic make-up of recent populations, resulting in significant genetic drift and profound inbreeding across the species range. To support science-based conservation actions, molecular methods have been increasingly used. However, such analyses did not include populations in Slovenia. We analysed neutral loci (partial fragment of mitochondrial cytochrome b, mtDNA) and the adaptive major histocompatibility complex (MHC DRB exon 2) of the Alpine ibex from both Slovenian populations (Julian and Kamnik-Savinja Alps) to understand how past reintroductions and recent management have affected the genetic diversity of the species. Results showed that both populations are genetically severely depleted, carrying only one mtDNA haplotype and one functional allele for MHC DRB exon 2, Caib-DRB*01. This calls for further conservation actions, including the reintroduction of individuals with different genetic background. However, the Alpine ibex is currently considered a non-native species in Slovenia, which makes conservation actions extremely difficult and threatens the long-term survival of the species. Therefore, scientists and population managers are urging policy/decision makers to change the status of the species to the native one and consequently to allow reintroductions. These appeals are supported by previous archaeological data on the existence of bones assigned to Alpine ibex in the Julian Alps, and evidence of severe genetic depletion in current ibex populations confirmed in this study. PRVI VPOGLED V GENETSKO RAZNOLIKOST ALPSKEGA KOZOROGA (Capra ibex) V SLOVENIJI Izvleček: V Evropi je bil alpski kozorog (Capra ibex) v 19. stoletju na robu izumrtja. Izvajali so se različni ukrepi za njegovo ohranjanje. V alpskem loku je bilo izvedenih več ponovnih naselitev, najprej z edino ohranjeno populacijo v kraju Gran Paradiso v Italiji. Izredna okrnjenost vrste v preteklosti in dodatne ponovne naselitve so močno vplivale na genetsko sestavo populacije, kar je povzročilo znaten genetski zdrs in parjenje v sorodstvu na celotnem območju vrste. V podporo znanstveno utemeljenim ukrepom ohranjanja se vse pogosteje uporabljajo molekularne metode, vendar takšne analize niso vključevale populacij v Sloveniji. Da bi razumeli, kako je ponovno naseljevanje in nedavno upravljanje vplivalo na genetsko raznolikost vrste, smo analizirali nevtralne lokuse (delni fragment mitohondrijskega citokroma b, mtDNA) in adaptivni poglavitni histokompatibilnostni kompleks (MHC DRB ekson 2) alpskega kozoroga iz obeh slovenskih populacij (Julijske in Kamniško-Savinjske Alpe). Rezultati so pokazali, da sta obe populaciji genetsko zelo osiromašeni, saj nosita le en haplotip mtDNA in en funkcionalni alel za MHC DRB ekson 2, Caib-DRB*01. Zato so potrebni nadaljnji ukrepi za ohranjanje, vključno s ponovno naselitvijo živali z drugačnim genetskim ozadjem. Vendar alpski kozorog v Sloveniji trenutno velja za tujerodno vrsto, kar zelo otežuje ukrepe za njegovo ohranitev in ogroža dolgoročno preživetje vrste. Znanstveniki in upravljavci populacij zato pozivajo politike/odločevalce, naj spremenijo status vrste v avtohtono in posledično omogočijo ponovno naselitev. Ti pozivi so podprti s predhodnimi arheološkimi podatki o obstoju kosti alpskega kozoroga v Julijskih Alpah in z dokazi o izraziti genetski osiromašenosti sedanjih populacij kozoroga, potrjenimi v tej študiji. Ključne besede: Capra ibex; mitohondrijska DNA; MHC DRB exon2; ponovna naselitev; upravljanje

This issue’s cover illustration depicting the Alpine ibex against the backdrop of iconic Slovenia's Triglav mountain is inspired by the manuscript by Bužan et al. and serves as a poignant reminder of a lost legacy and biodiversity due to greed and overhunting. These magnificent creatures thrived entire European Alpine region before their extermination in the 19th century, with the only surviving wild population of Alpine ibex in the Gran Paradiso National Park (then a royal hunting reserve) in Italy. Reintroduction initiatives from this small population have since rebounded ibex to tens of thousands across the Alpine region. However, as the article on the Slovenian population of Alpine ibex by Bužan et al. in this issue clearly shows, while current numbers seem more promising, the genetic bottleneck they experienced from this near-extinction event has left lasting impacts on their genetic diversity. REŠEVANJE ALPSKEGA KOZOROGA: GENETSKI IN ZDRAVSTVENI PROBLEMI SLOVENSKIH KOZOROGOV S ŠIRŠIMI POSLEDICAMI ZA BIODIVERZITETO Ilustracija na naslovnici te številke, ki prikazuje alpskega kozoroga pred ikonično Triglavsko goro Slovenije, je navdihnjena po članku Bužan s sod., objavljenem v tej številki, in služi kot opomin izgubljeni dediščini in biodiverziteti zaradi pohlepa in prekomernega lova. Te veličastne živali so uspevale v celotni evropski Alpski regiji, preden so bile iztrebljene v 19. stoletju, z edino preživelo divjo populacijo v Narodnem parku Gran Paradiso (takrat kraljevi lovski rezervat) v Italiji. Ponovne naselitve iz te majhne populacije so od takrat od danes dvignile število kozorogov na deset tisoče v celotni Alpski regiji. Vendar, kot članek o slovenski populaciji alpskih kozorogov avtorice Bužan s sod. jasno kaže, čeprav se trenutna številka zdi obetavna, pa je genetsko ozko grlo, nastalo zaradi prekomernega lova, pustilo trajne posledice na njihovi genetski raznolikosti.

This paper discusses the challenges of producing CAR-T cells for cancer treatment and the potential for Artificial Intelligence (AI) for its improvement. CAR-T cell therapy was approved in 2018 as the first Advanced Therapy Medicinal Product (ATMP) for treating acute leukemia and lymphoma. ATMPs are cell- and gene-based therapies that show great promise for treating various cancers and hereditary diseases. While some new ATMPs have been approved, ongoing clinical trials are expected to lead to the approval of many more. However, the production of CAR-T cells presents a significant challenge due to the high costs associated with the manufacturing process, making the therapy very expensive (approx. $400,000). Furthermore, autologous CAR-T therapy is limited to a make-to-order approach, which makes scaling economical production difficult. First attempts are being made to automate this multi-step manufacturing process, which will not only directly reduce the high manufacturing costs but will also enable comprehensive data collection. AI technologies have the ability to analyze this data and convert it into knowledge and insights. In order to exploit these opportunities, this paper analyses the data potential in the automated CAR-T production process and creates a mapping to the capabilities of AI applications. The paper explores the possible use of AI in analyzing the data generated during the automated process and its capabilities to further improve the efficiency and cost-effectiveness of CAR-T cell production.

Amyloid self-assembly is linked to type 2 diabetes and Alzheimer’s disease. Here the authors designed constrained peptides which are nanomolar amyloid inhibitors of the key polypeptides IAPP and Aβ42 and act via supramolecular nanofiber co-assembly.

S. epidermidis is an important opportunistic pathogen causing chronic prosthetic joint infections associated with biofilm growth. Increased tolerance to antibiotic therapy often requires prolonged treatment or revision surgery. Phage therapy is currently used as compassionate use therapy and continues to be evaluated for its viability as adjunctive therapy to antibiotic treatment or as an alternative treatment for infections caused by S. epidermidis to prevent relapses. In the present study, we report the isolation and in vitro characterization of three novel lytic S. epidermidis phages. Their genome content analysis indicated the absence of antibiotic resistance genes and virulence factors. Detailed investigation of the phage preparation indicated the absence of any prophage-related contamination and demonstrated the importance of selecting appropriate hosts for phage development from the outset. The isolated phages infect a high proportion of clinically relevant S. epidermidis strains and several other coagulase-negative species growing both in planktonic culture and as a biofilm. Clinical strains differing in their biofilm phenotype and antibiotic resistance profile were selected to further identify possible mechanisms behind increased tolerance to isolated phages.

Immune therapy for cancer patients is a new and promising area that in the future may complement traditional chemotherapy. The cell expansion phase is a critical part of the process chain to produce a large number of high-quality, genetically modified immune cells from an initial sample from the patient. Smart sensors augment the ability of the control and monitoring system of the process to react in real-time to key control parameter variations, adapt to different patient profiles, and optimize the process. The aim of the current work is to develop and calibrate smart sensors for their deployment in a real bioreactor platform, with adaptive control and monitoring for diverse patient/donor cell profiles. A set of contrasting smart sensors has been implemented and tested on automated cell expansion batch runs, which incorporate advanced data-driven machine learning and statistical techniques to detect variations and disturbances of the key system features. Furthermore, a ‘consensus’ approach is applied to the six smart sensor alerts as a confidence factor which helps the human operator identify significant events that require attention. Initial results show that the smart sensors can effectively model and track the data generated by the Aglaris FACER bioreactor, anticipate events within a 30 min time window, and mitigate perturbations in order to optimize the key performance indicators of cell quantity and quality. In quantitative terms for event detection, the consensus for sensors across batch runs demonstrated good stability: the AI-based smart sensors (Fuzzy and Weighted Aggregation) gave 88% and 86% consensus, respectively, whereas the statistically based (Stability Detector and Bollinger) gave 25% and 42% consensus, respectively, the average consensus for all six being 65%. The different results reflect the different theoretical approaches. Finally, the consensus of batch runs across sensors gave even higher stability, ranging from 57% to 98% with an average consensus of 80%.

Background: Work poses increased risk of injury not only for workers but also for the public, yet the broader impact of work-related injury is not quantified. This study, utilising population data from New Zealand, estimates the societal burden of work-related fatal injury (WRFI) by including bystanders and commuters. Methods: This observational study selected deaths due to unintentional injury, in persons aged 0–84 years using International Classification of Disease external cause codes, matched to coronial records, and reviewed for work-relatedness. Work-relatedness was determined by the decedent's circumstances at the time of the incident: working for pay, profit, in kind, or an unpaid capacity (worker); commuting to or from work (commuter); or a bystander to another's work activity (bystander). To estimate the burden of WRFI, frequencies, percentages, rates, and years-of-life lost (YLL) were estimated. Results: In total 7,707 coronial records were reviewed of which 1,884 were identified as work-related, contributing to 24% of the deaths and 23% of the YLL due to injury. Of these deaths close to half (49%) occurred amongst non-working bystanders and commuters. The overall burden of WRFI was widespread across age, sex, ethnic and deprivation sub-groups. Injury deaths due to machinery (97%) and due to being struck by another object (69%) were predominantly work-related. Interpretation: When utilising a more inclusive definition of work-relatedness the contribution of work to the societal burden of fatal injuries is substantial, conservatively estimated at one quarter of all injury deaths in New Zealand. Other estimates of WRFI likely exclude a similar number of fatalities occurring among commuters and bystanders. The findings, also relevant to other OECD nations, can guide where public health efforts can be used, alongside organisational actions, to reduce WRFI for all those impacted.

Background & Aims: Bile salt export pump (BSEP) deficiency frequently necessitates liver transplantation in childhood. In contrast to two predicted protein truncating mutations (PPTMs), homozygous p.D482G or p.E297G mutations are associated with relatively mild phenotypes, responsive to surgical interruption of the enterohepatic circulation (siEHC). The phenotype of patients with a compound heterozygous genotype of one p.D482G or p.E297G mutation and one PPTM has remained unclear. We aimed to assess their genotype-phenotype relationship. Methods: From the NAPPED database, we selected patients with homozygous p.D482G or p.E297G mutations (BSEP1/1; n = 31), with one p.D482G or p.E297G, and one PPTM (BSEP1/3; n = 30), and with two PPTMs (BSEP3/3; n = 77). We compared clinical presentation, native liver survival (NLS), and the effect of siEHC on NLS. Results: The groups had a similar median age at presentation (0.7-1.3 years). Overall NLS at age 10 years was 21% in BSEP1/3 vs. 75% in BSEP1/1 and 23% in BSEP3/3 (p

Aims In May 2020, a nationwide, web-based system for remote entry of patient-reported outcomes (PROs) in inflammatory rheumatic diseases was launched and implemented in routine care (DANBIO-from-home). After 1.5 years of use, we explored clinical characteristics of patients who did versus did not use the system, and the time to first entry of PROs.Methods All patients followed in DANBIO were informed about DANBIO-from-home by electronic invitations or when attending their clinic. Characteristics of patients who did/did not use DANBIO-from-home in the period after implementation were explored by multivariable logistic regression analyses including demographic and clinical variables (gender, age group, diagnosis, disease duration, use of biological disease-modifying agent (bDMARD), Health Assessment Questionnaire (HAQ), Patient Acceptable Symptom Scale (PASS)). Time from launch to first entry was presented as cumulative incidence curves by age group (80 years).Results Of 33 776 patients, 68% entered PROs using DANBIO-from-home at least once. Median (IQR) time to first entry was 27 (11–152) days. Factors associated with data entry in multivariate analyses (OR (95% CI)) were: female gender (1.19 (1.12 to 1.27)), bDMARD treatment (1.41 (1.33 to 1.50)), age 40–60 years (1.79 (1.63 to 1.97)), 61–80 years (1.87 (1.70 to 2.07), or age >80 years (0.57 (0.50 to 0.65)) (reference: age

Abstract Background Local breeds retained unique genetic variability important for adaptive potential especially in light of challenges related to climate change. Our first objective was to perform, for the first time, a genome-wide diversity characterization using Illumina GoatSNP50 BeadChip of autochthonous Drežnica goat breed from Slovenia, and five and one local breeds from neighboring Austria and Italy, respectively. For optimal conservation and breeding programs of endangered local breeds, it is important to detect past admixture events and strive for preservation of purebred representatives of each breed with low or without admixture. In the second objective, we hence investigated the effect of inclusion or exclusion of outliers from datasets on genetic diversity and population structure parameters. Results Distinct genetic origin of the Drežnica goat was demonstrated as having closest nodes to Austrian and Italian breeds. A phylogenetic study of these breeds with other goat breeds having SNP data available in the DRYAD repository positioned them in the alpine, European and global context. Swiss breeds clustered with cosmopolitan alpine breeds and were closer to French and Spanish breeds. On the other hand, the Drežnica goat, Austrian and Italian breeds were closer to Turkish breeds. Datasets where outliers were excluded affected estimates of genetic diversity parameters within the breed and increased the pairwise genetic distances between most of the breeds. Alpine breeds, including Drežnica, Austrian and Italian goats analyzed here, still exhibit relatively high levels of genetic variability, homogeneous genetic structure and strong geographical partitioning. Conclusions Genetic diversity analyses revealed that the Slovenian Drežnica goat has a distinct genetic identity and is closely related to the neighboring Austrian and Italian alpine breeds. These results expand our knowledge on phylogeny of goat breeds from easternmost part of the European Alps. The here employed outlier test and datasets optimization approaches provided an objective and statistically powerful tool for removal of admixed outliers. Importance of this test in selecting the representatives of each breed is warranted to obtain more objective diversity parameters and phylogenetic analysis. Such parameters are often the basis of breeding and management programs and are therefore important for preserving genetic variability and uniqueness of local rare breeds.

Background: People with language problems following stroke (aphasia) benefit from speech and language therapy. Optimising speech and language therapy for aphasia recovery is a research priority. Objectives: The objectives were to explore patterns and predictors of language and communication recovery, optimum speech and language therapy intervention provision, and whether or not effectiveness varies by participant subgroup or language domain. Design: This research comprised a systematic review, a meta-analysis and a network meta-analysis of individual participant data. Setting: Participant data were collected in research and clinical settings. Interventions: The intervention under investigation was speech and language therapy for aphasia after stroke. Main outcome measures: The main outcome measures were absolute changes in language scores from baseline on overall language ability, auditory comprehension, spoken language, reading comprehension, writing and functional communication. Data sources and participants: Electronic databases were systematically searched, including MEDLINE, EMBASE, Cumulative Index to Nursing and Allied Health Literature, Linguistic and Language Behavior Abstracts and SpeechBITE (searched from inception to 2015). The results were screened for eligibility, and published and unpublished data sets (randomised controlled trials, non-randomised controlled trials, cohort studies, case series, registries) with at least 10 individual participant data reporting aphasia duration and severity were identified. Existing collaborators and primary researchers named in identified records were invited to contribute electronic data sets. Individual participant data in the public domain were extracted. Review methods: Data on demographics, speech and language therapy interventions, outcomes and quality criteria were independently extracted by two reviewers, or available as individual participant data data sets. Meta-analysis and network meta-analysis were used to generate hypotheses. Results: We retrieved 5928 individual participant data from 174 data sets across 28 countries, comprising 75 electronic (3940 individual participant data), 47 randomised controlled trial (1778 individual participant data) and 91 speech and language therapy intervention (2746 individual participant data) data sets. The median participant age was 63 years (interquartile range 53–72 years). We identified 53 unavailable, but potentially eligible, randomised controlled trials (46 of these appeared to include speech and language therapy). Relevant individual participant data were filtered into each analysis. Statistically significant predictors of recovery included age (functional communication, individual participant data: 532, n = 14 randomised controlled trials) and sex (overall language ability, individual participant data: 482, n = 11 randomised controlled trials; functional communication, individual participant data: 532, n = 14 randomised controlled trials). Older age and being a longer time since aphasia onset predicted poorer recovery. A negative relationship between baseline severity score and change from baseline (p

Background: To improve and compare outcomes in healthcare, it is necessary to standardise outcome measurements. There are no widely accepted standardised outcome measures reflecting quality of care for bladder cancer (BCa) patients. Objective: The aim of this study was to create a standardised set of outcomes for patients with muscle-invasive or metastatic BCa, using the value-based healthcare principles. Design, setting, and participants: A multidisciplinary working group of 25 healthcare professionals and patient representatives was assembled, to develop the set. Outcome measurements and statistical analysis: We used an online RAND-modified Delphi process to prioritise, discuss, and reach consensus regarding the outcomes, case-mix variables, and treatment factors. Results and limitations: Recognising the heterogeneity of patients with BCa, the working group defined the scope as patients with muscle-invasive and metastatic BCa. A total of 24 outcomes, including ten patient-reported outcomes, were included in the standard set of outcomes, covering survival, complication rates, recurrence of disease, readmissions after treatment, and quality of life (QoL). Fourteen case-mix variables were included. The EQ-5D and European Organisation for Research and Treatment of Cancer quality of life (EORTC-QLQ) questionnaires were recommended to measure QoL. Conclusions: We developed the first standardised set of patient-centred outcomes for muscle-invasive and metastatic BCa. The sue of this set enables institutions to monitor, compare, and improve the quality of BCa care, on an international level. Patient summary: Our group of healthcare professionals and patient representatives recommended a standardised set of patient-centred outcomes to be followed during the treatment of patients with muscle-invasive or metastatic bladder cancer, in order to monitor, compare, and improve the quality of care.

Ocular antibiotics are integral to the prevention and treatment of bacterial ocular infections. This study aimed to describe their utilisation across New Zealand according to patient and healthcare factors. Every subsidy-eligible community dispensing of ocular chloramphenicol, fusidic acid and ciprofloxacin in New Zealand, between 2010 and 2019, was included in this analysis. Number of dispensings/1000 population/year was quantified, stratified by patient age and urban/non-urban health districts. Dispensing rates by ethnicity were determined and were age adjusted. The proportion of dispensings by socioeconomic deprivation quintile was also determined. Chloramphenicol was the most commonly dispensed antibiotic; however, its utilisation decreased over time. Ciprofloxacin use was higher in children, while chloramphenicol use was higher in older patients. Ciprofloxacin usage was higher among Māori and Pasifika ethnicities, while fusidic acid use was lower. Chloramphenicol usage was higher among Pasifika. Antibiotic utilisation was higher in urban health districts, and in the most deprived quintile; both were most marked with ciprofloxacin. The utilisation of publicly funded ocular antibiotics across New Zealand varied between patient subgroups. These findings will help improve the prevention, management and outcomes of bacterial ocular infections, and support wider initiatives in antibiotic stewardship and medicine access equity.

All-optical switching is important for integrated-photonics and communication devices. Here the authors demonstrate all-optical switching of an Epsilon-Near-Zero plasmon resonance using indium tin oxide thin film.

Abstract Background Molecular tweezers (MTs) are broad-spectrum inhibitors of abnormal protein aggregation. A lead MT, called CLR01, has been demonstrated to inhibit the aggregation and toxicity of multiple amyloidogenic proteins in vitro and in vivo. Previously, we evaluated the effect of CLR01 in the 3 × Tg mouse model of Alzheimer’s disease, which overexpresses mutant human presenilin 1, amyloid β-protein precursor, and tau and found that subcutaneous administration of the compound for 1 month led to a robust reduction of amyloid plaques, neurofibrillary tangles, and microgliosis. CLR01 also has been demonstrated to inhibit tau aggregation in vitro and tau seeding in cell culture, yet because in Alzheimer’s disease (AD) and in the 3 × Tg model, tau hyperphosphorylation and aggregation are thought to be downstream of Aβ insults, the study in this model left open the question whether CLR01 affected tau in vivo directly or indirectly. Methods To determine if CLR01 could ameliorate tau pathology directly in vivo, we tested the compound similarly using the P301S-tau (line PS19) mouse model. Mice were administered 0.3 or 1.0 mg/kg per day CLR01 and tested for muscle strength and behavioral deficits, including anxiety- and disinhibition-like behavior. Their brains then were analyzed by immunohistochemical and biochemical assays for pathological forms of tau, neurodegeneration, and glial pathology. Results CLR01 treatment ameliorated muscle-strength deterioration, anxiety-, and disinhibition-like behavior. Improved phenotype was associated with decreased levels of pathologic tau forms, suggesting that CLR01 exerts a direct effect on tau in vivo. Limitations of the study included a relatively short treatment period of the mice at an age in which full pathology is not yet developed. In addition, high variability in this model lowered the statistical significance of the findings of some outcome measures. Conclusions The findings suggest that CLR01 is a particularly attractive candidate for the treatment of AD because it targets simultaneously the two major pathogenic proteins instigating and propagating the disease, amyloid β-protein (Aβ), and tau, respectively. In addition, our study suggests that CLR01 can be used for the treatment of other tauopathies in the absence of amyloid pathology.

CAR-T cell therapy is a promising treatment for acute leukemia and lymphoma. CAR-T cell therapies take a pioneering role in autologous gene therapy with three EMA-approved products. However, the chance of clinical success remains relatively low as the applicability of CAR-T cell therapy suffers from long, labor-intensive manufacturing and a lack of comprehensive insight into the bioprocess. This leads to high manufacturing costs and limited clinical success, preventing the widespread use of CAR-T cell therapies. New manufacturing approaches are needed to lower costs to improve manufacturing capacity and shorten provision times. Semi-automated devices such as the Miltenyi Prodigy® were developed to reduce hands-on production time. However, these devices are not equipped with the process analytical technology necessary to fully characterize and control the process. An automated AI-driven CAR-T cell manufacturing platform in smart manufacturing hospitals (SMH) is being developed to address these challenges. Automation will increase the cost-effectiveness and robustness of manufacturing. Using Artificial Intelligence (AI) to interpret the data collected on the platform will provide valuable process insights and drive decisions for process optimization. The smart integration of automated CAR-T cell manufacturing platforms into hospitals enables the independent manufacture of autologous CAR-T cell products. In this perspective, we will be discussing current challenges and opportunities of the patient-specific but highly automated, AI-enabled CAR-T cell manufacturing. A first automation concept will be shown, including a system architecture based on current Industry 4.0 approaches for AI integration.

Abstract Background Long-acting local anaesthetics (e.g. bupivacaine hydrochloride) or sustained-release formulations of bupivacaine (e.g. liposomal bupivacaine) may be neurotoxic when applied in the setting of diabetic neuropathy. The aim of the study was to assess neurotoxicity of bupivacaine and liposome bupivacaine in streptozotocin (STZ) - induced diabetic mice after sciatic nerve block. We used the reduction in fibre density and decreased myelination assessed by G-ratio (defined as axon diameter divided by large fibre diameter) as indicators of local anaesthetic neurotoxicity. Results Diabetic mice had higher plasma levels of glucose (P

An amendment to this paper has been published and can be accessed via the original article.

In obesity, accumulation of lipid droplets in skeletal muscle fibers and a shift towards fast muscle fiber types can both contribute to insulin resistance. However, it is not yet clear how intramyocellular lipid accumulation and fiber type changes are associated. Therefore, we investigated to what extent the lipids accumulated in a fiber type-specific manner in the functionally similar fast-, intermediate- and slow-twitch gastrocnemius, plantaris, and soleus muscles, respectively, in high-fat diet-induced obese 54-week-old female C57BL/6JOlaHsd mice (n = 9) compared to control standard-diet-treated lean mice (n = 9). A high-fat diet was administered for 26 weeks. Fiber-type specific intramyocellular lipid content analysis and muscle fiber typing were performed using histochemical analysis of lipids with Sudan Black and immunohistochemical analysis of myosin heavy chains on serial sections of skeletal muscles. Compared to the lean mice, the lipid accumulation was most prominent in types 2a and 2x/d fibers (p < 0.05) of fast-twitch gastrocnemius and intermediate plantaris muscles in the obese mice, while in slow-twitch soleus muscle, there was no significant lipid accumulation in the obese animals. Furthermore, the slow-twitch soleus muscle of the obese mice with no significant change in muscle fiber diameters exhibited the most pronounced shift towards fast-type myosin heavy chain isoform expression (p < 0.05). In contrast, the fast-twitch and intermediate-twitch gastrocnemius and plantaris muscles, respectively, in which the muscle fiber diameters increased (p < 0.05), were more resistant toward myosin heavy chain expression changes. In conclusion, we demonstrated both muscle- and fiber-type specificity in intramyocellular lipid accumulation in obese mice, suggesting that in obesity, similar muscle fiber types in different muscles accumulate lipids differentially.

Objectives The aims of this study were to describe the following: (1) the time to change of therapy in patients with type 2 diabetes who had initiated metformin monotherapy as first-line treatment and (2) the sequence in which subsequent therapeutic regimens were introduced.Design Cohort study.Setting National study based on linked data from the New Zealand Ministry of Health’s National Collections of health and pharmaceutical dispensing data.Participants People with type 2 diabetes mellitus who initiated metformin monotherapy between 1 January 2006 and 30 September 2014 (n=93 874).Primary outcome measures Cumulative incidence curves were plotted to show the time taken to move from one regimen to another, while sunburst plots were used to illustrate the sequence in which regimens were introduced.Results About 10% and 35% of cohort members had moved to a second regimen 1 year and 5 years, respectively, after initiating metformin monotherapy; the majority received a regimen recommended by New Zealand treatment guidelines (mostly metformin and a sulphonylurea). Of those who started a recommended second regimen, 37% and 67% had moved to a third regimen after 1 and 5 years, respectively; the corresponding proportions for those who started an ‘other’ (not listed as recommended) second regimen were 53% and 75%. Most of those who received a third regimen after a recommended second regimen were dispensed an ‘other’ third regimen. Of those who moved to a third regimen from an ‘other’ second regimen, similar proportions received recommended and ‘other’ third regimens.Conclusions Real-world type 2 diabetes treatment patterns in New Zealand are complex and not always consistent with guidelines.

Introduction Prescription charges prevent many people from accessing the medicines they need to maintain or improve their health. In New Zealand, where most people pay $5 per prescription item, Māori and Pacific peoples, those living in most deprived areas and those with chronic health conditions are the most likely to report that cost prevents them from accessing medicines.Methods and analysis This randomised controlled trial (RCT) will evaluate the effect of removing prescription charges on health outcomes and healthcare utilisation patterns of people with low income and high health needs. We will enrol 2000 participants: half will be allocated to the intervention group and we will pay for their prescription charges for 12 months. The other half will receive usual care. The primary outcome will be hospital bed-days. Secondary outcomes will be: all-cause and diabetes/mental health-specific hospitalisations, prescription medicines dispensed (number and type), deaths, emergency department visits and quality of life as measured by the 5-level EQ-5D version. Costs associated with these outcomes will be compared in an economic substudy. A qualitative substudy will also help understand the impact of free prescriptions on participant well-being using in-depth interviews.Discussion Being unable to afford prescription medicines is only one of many factors that influence adherence to medicines, but removing prescription charges is relatively simple and in New Zealand would be cheap compared with other policy changes. This RCT will help identify the extent of the impact of a simple intervention to improve access to medicines on health outcomes and health service utilisation.Ethics and dissemination This study was approved by the Central Health and Disability Ethics Committee (NZ) in July 2019 (19/CEN/33). Findings will be reported in peer-reviewed publications, as well as in professional newsletters, mainstream media and through public meetings.Trial registration number ACTRN12618001486213p.

Abstract Background Improving New Zealand’s poor workplace safety record has become a high priority following high profile workplace fatal incidents in 2010 and 2014. Existing routine official data are unable to reliably inform occupational safety policy and action in New Zealand. This case review study will examine work-related fatal injury (WRFI) to: i) determine their burden, rates and distribution; ii) identify high-risk groups, causes and circumstances; iii) analyse secular trends, and iv) examine the impact of historic occupational safety legislative reform. Design and methods A comprehensive New Zealand WRFI dataset from 1975 to 2014 will be established using existing data for 1975–1994 combined with new data for 1995–2014 extracted from reviewed coronial case files. Data collection involves: 1) identifying likely cases of WRFI from national mortality records using selected injury external cause codes; 2) linking these to coronial case files, which will be retrieved and reviewed to determine work-relatedness; and 3) coding work-related cases. Annual WRFI frequencies and rates will be calculated and disaggregated by age, sex, employment status, occupation and industry to identify high-risk groups and compared across the time series. The circumstances of the WRFIs will be analysed in-depth. The impact of New Zealand’s Health and Safety in Employment 1992 Act, which resulted in deregulation of the previous legislative frameworks for occupational health and safety during a period of rapid labour market restructuring, will be examined by comparing rates before and after implementation of the Act. Discussion The resulting evidence will serve as the basis for policy development and practical interventions to reduce WRFI, targeting groups of high-risk workers, and for bench-marking of workplace safety performance in New Zealand.

Little is known regarding the pharmacological properties of extended-release local anesthetics in the setting of diabetic peripheral neuropathy. We investigated and compared the duration of sciatic nerve block following administration of clinically relevant concentrations of liposomal bupivacaine (LB) and bupivacaine hydrochloride (BH) in diabetic mice with peripheral neuropathy. In this prospective, randomized, and double-blind study, twenty-four female C57BL/6J-OlaHsd mice were assigned to a streptozotocin-induced type 1 diabetes group and a control group without diabetes. The presence of peripheral neuropathy was established by assessing the duration of thermal latency of the plantar and tail-flick tests, following which both groups were subdivided into two subgroups in which 35 mg/kg of 1.31% LB and 7 mg/kg of 0.25% BH were respectively administered for sciatic nerve block. The average sensory block duration with BH was 106 min and 117.1 min in the control and diabetic groups, respectively. With LB, the average sensory block duration was 118 min in the control mice, while in mice with diabetic peripheral neuropathy, the average block duration was significantly longer and above the 270 min limit set in our study. Accordingly, sensory block duration was longer with LB compared to BH, and diabetic peripheral neuropathy significantly increased sciatic nerve block duration with LB.

Comprehensive analysis of tumor infiltrating myeloid cells in the tumor microenvironment of penile squamous cell carcinoma (PSCC) is lacking. In this retrospective study, for the first time, PSCC resection specimens (N = 103) were annotated into the following compartments: intratumoral tumor (IT Tumor), intratumoral stroma (IT Stroma), peritumoral tumor (PT Tumor) and peritumoral stroma (PT Stroma) compartments. We then quantified CD14+, CD68+ and CD163+ myeloid cells within these compartments using an image analysis software and assessed their association with various clinical parameters, including high-risk human papillomavirus (hrHPV) status. In the total cohort, hrHPV status, grade of differentiation, age and tumor size were associated with myeloid cell densities. hrHPV+ tumors had higher infiltration rates of CD14+, CD68+ and CD163+ myeloid cells in the IT tumor compartment (p < 0.001, for all) compared to hrHPV− tumors. Furthermore, when examining the association between compartment-specific infiltration and differentiation grade, increased myeloid cell densities in the IT tumor compartment were associated with a more advanced histological grade (p < 0.001, for all). This association remained significant when the hrHPV− cohort (N = 60) was analyzed (CD14+ p = 0.001; CD68+ p < 0.001; CD163+ p = 0.004). Subgroup analysis in the hrHPV+ group (N = 43) showed that high infiltration rates of CD68+ and CD163+ cells in the PT tumor compartment were associated with lymph node (LN) metastasis (p = 0.031 and p = 0.026, respectively). Regarding the association between myeloid cell densities and disease-specific survival, the risk of death was found to decrease slightly as the number of myeloid cells in the IT tumor compartment increased (CD14+ p = 0.04; CD68+ p = 0.05; CD163+ p = 0.02). However, after adjusting for hrHPV, no independent association between myeloid densities and disease-specific survival were found. Altogether, these findings demonstrate the importance of assessing myeloid cell densities within the spatial context of the tumor. Further studies are needed to unravel the specific phenotype of myeloid cells residing in the different compartments, their effect on clinical parameters and the impact of hrHPV on the recruitment of myeloid cell populations in PSCC.

Aims In Danish patients with inflammatory rheumatic diseases to explore self-protection strategies and health behaviour including adherence to disease-modifying antirheumatic treatment (DMARD) during the initial phase of the COVID-19 pandemic and again after the reopening of the society started. Furthermore, to identify characteristics of patients with high levels of anxiety and self-isolation.Methods Patients in routine care followed prospectively in the nationwide DANBIO registry were invited to answer an online questionnaire regarding disease activity and COVID-19 infection, behaviour in March and June 2020. Responses were linked to patient data in DANBIO. Characteristics potentially associated with anxiety, self-isolation and medication adherence (gender/age/diagnosis/education/work status/comorbidity/DMARD/smoking/EQ-5D/disease activity) were explored with multivariable logistic regression analyses.Results We included 12 789 patients (8168 rheumatoid arthritis/2068 psoriatic arthritis/1758 axial spondyloarthritis/795 other) of whom 65% were women and 36% treated with biological DMARD. Self-reported COVID-19 prevalence was 0.3%. Patients reported that they were worried to get COVID-19 infection (March/June: 70%/45%) and self-isolated more than others of the same age (48%/38%). The fraction of patients who changed medication due to fear of COVID-19 were 4.1%/0.6%. Female gender, comorbidities, not working, lower education, biological treatment and poor European Quality of life, 5 dimensions were associated with both anxiety and self-isolation.Conclusion In >12 000 patients with inflammatory arthritis, we found widespread anxiety and self-isolation, but high medication adherence, in the initial phase of the COVID-19 pandemic. This persisted during the gradual opening of society during the following months. Attention to patients’ anxiety and self-isolation is important during this and potential future epidemics.

Background Cardiovascular comorbidity is common among patients with chronic obstructive pulmonary disease (COPD) and there is concern that long-acting bronchodilators (long-acting muscarinic antagonists (LAMAs) and long-acting beta2 agonists (LABAs)) may further increase the risk of acute coronary events. Information about the impact of treatment intensification on acute coronary syndrome (ACS) risk in real-world settings is limited. We undertook a nationwide nested case–control study to estimate the risk of ACS in users of both a LAMA and a LABA relative to users of a LAMA.Methods We used routinely collected national health and pharmaceutical dispensing data to establish a cohort of patients aged >45 years who initiated long-acting bronchodilator therapy for COPD between 1 February 2006 and 30 December 2013. Fatal and non-fatal ACS events during follow-up were identified using hospital discharge and mortality records. For each case we used risk set sampling to randomly select up to 10 controls, matched by date of birth, sex, date of cohort entry (first LAMA and/or LABA dispensing), and COPD severity.Results From the cohort (n=83 417), we identified 5399 ACS cases during 281 292 person-years of follow-up. Compared with current use of LAMA therapy, current use of LAMA and LABA dual therapy was associated with a higher risk of ACS (OR 1.28 (95% CI 1.13 to 1.44)). The OR in an analysis restricted to fatal cases was 1.46 (95% CI 1.12 to 1.91).Conclusion In real-world clinical practice, use of two versus one long-acting bronchodilator by people with COPD is associated with a higher risk of ACS.

AimTo describe the patterns of discontinuation and reinitiation in new users of metformin monotherapy in New Zealand, overall and according to person- and healthcare-related factors.Materials and methodsWe created a cohort (n = 85,066) of all patients in New Zealand with type 2 diabetes mellitus who initiated metformin monotherapy between 1 January 2006 and 30 September 2014 from the national data collections, and followed them until the earlier of their death or 31 December 2015. Discontinuation was defined as a gap in possession of metformin monotherapy of ≥90 days. We explored patterns of discontinuation and reinitiation using competing risks methods.ResultsAfter 1 year of follow-up, 28% of cohort members had discontinued metformin monotherapy at least once; the corresponding figures after 2 and 5 years were 37% and 46%. The proportions who reinitiated metformin monotherapy within 1, 2, and 5 years of their first discontinuation were 23%, 49%, and 73%. Discontinuation after the first reinitiation was common (48% after 1 year). Discontinuation and reinitiation varied by age, ethnicity, and other person- and healthcare-related factors.DiscussionOur findings highlight the dynamic nature of metformin monotherapy use, show that substantial periods of non-use are common, and identify priority populations for interventions to facilitate adherence.

Eukaryotic cells release different types of extracellular vesicles (EVs) including exosomes, ectosomes, and microvesicles. Exosomes are nanovesicles, 30–200 nm in diameter, that carry cell- and cell-state-specific cargo of proteins, lipids, and nucleic acids, including mRNA and miRNA. Recent studies have shown that central nervous system (CNS)-derived exosomes may carry amyloidogenic proteins and facilitate their cell-to-cell transfer, thus playing a critical role in the progression of neurodegenerative diseases, such as tauopathies and synucleinopathies. CNS-derived exosomes also have been shown to cross the blood-brain-barrier into the bloodstream and therefore have drawn substantial attention as a source of biomarkers for various neurodegenerative diseases as they can be isolated via a minimally invasive blood draw and report on the biochemical status of the CNS. However, although isolating specific brain-cell-derived exosomes from the blood is theoretically simple and the approach has great promise, practical details are of crucial importance and may compromise the reproducibility and utility of this approach, especially when different laboratories use different protocols. In this review we discuss the role of exosomes in neurodegenerative diseases, the usefulness of CNS-derived blood exosomes as a source of biomarkers for these diseases, and practical challenges associated with the methodology of CNS-derived blood exosomes and subsequent biomarker analysis.

ObjectiveTo describe prescription medicine dispensing before and during pregnancy in New Zealand, 2005-2015.MethodsMembers of the New Zealand Pregnancy Cohort were linked with their dispensing records in a national database of prescription products dispensed from community pharmacies. We identified the proportion of pregnancies during which at least one prescription medicine was dispensed, the number of different medicines used and the most commonly dispensed medicine groups both during pregnancy and in the 270 days before conception. Dispensing during pregnancy was assessed by several maternal characteristics.Results874,884 pregnancies were included. Over the study timeframe, the proportion of pregnancies exposed to a non-supplement prescription medicine increased from 38.5% to 67.2%. The mean number of different non-supplement medicines dispensed during pregnancy increased from 2.5 to 3.2. Dispensing during pregnancy was weakly associated with body mass index, smoking status and ethnicity. Pregnancy exposure was highest for Antibacterials (26.0%), Analgesics (16.7%) and Antinausea & Vertigo Agents (11.0%).ConclusionsFrom 2005-2015, both the proportion of exposed pregnancies and the number of different medicines dispensed to pregnant women in New Zealand increased.

BackgroundMaintaining adherence to statins reduces the risk of an initial cardiovascular disease (CVD) event in high-risk individuals (primary prevention) and additional CVD events following the first event (secondary prevention). The effectiveness of statin therapy is limited by the level of adherence maintained by the patient. We undertook a nationwide study to compare adherence and discontinuation in primary and secondary prevention patients.MethodsDispensing data from New Zealand community pharmacies were used to identify patients who received their first statin dispensing between 2006 and 2011. The Medication Possession Ratio (MPR) and proportion who discontinued statin medication was calculated for the year following first statin dispensing for patients with a minimum of two dispensings. Adherence was defined as an MPR ≥ 0.8. Previous CVD was identified using hospital discharge records. Multivariable logistic regression was used to control for demographic and statin characteristics.ResultsBetween 2006 and 2011 289,666 new statin users were identified with 238,855 (82.5%) receiving the statin for primary prevention compared to 50,811 (17.5%) who received it for secondary prevention. The secondary prevention group was 1.55 (95% CI 1.51-1.59) times as likely to be adherent and 0.67 (95% CI 0.65-0.69) times as likely to discontinue statin treatment than the primary prevention group. An early gap in statin coverage increased the odds of discontinuing statin treatment.ConclusionAdherence to statin medication is higher in secondary prevention than primary prevention. Within each group, a range of demographic and treatment factors further influences adherence.

Surface hoar crystals are common on the surface of mountain snow covers. Once buried, layers of large plate-shaped surface hoar crystals are prone to releasing dangerous snow-slab avalanches. Since snow microstructure influences the optical properties of snow, remote sensors could potentially detect the formation of surface hoar and other snow types associated with avalanche release. The spectral reflectance of 377 snow samples was measured with a field spectrometer between 750 and 2500 nm, including 161 samples of surface hoar. Morphological snow shapes associated with critical avalanche layers (surface hoar, near-surface faceted particles and depth hoar) had lower average reflectance factors than non-critical snow shapes at infrared wavelengths. Needle-shaped surface hoar was more reflective than plate-shaped surface hoar, but there were no significant differences between different sizes of surface hoar. Normalized difference indices calculated with reflectance from two wavelength bands is presented as a potential method to classify critical snow surfaces remotely, although melt-freeze crusts near the surface complicated the classification. Accordingly, further studying on the effect of melt-freeze crusts and quantification of the bidirectional reflective properties of critical snow types is needed.

Birt-Hogg-Dubé syndrome is associated with an increased risk for renal cell carcinoma. Surveillance is recommended, but the optimal imaging method and screening interval remain to be defined. The main aim of our study was to evaluate the outcomes of RCC surveillance to get insight in the safety of annual US in these patients. Surveillance data and medical records of 199 patients with Birt-Hogg-Dubé syndrome were collected retrospectively using medical files and a questionnaire. These patients were diagnosed in two Dutch hospitals and data were collected until June 2014. A first screening for renal cell carcinoma was performed in 172/199 patients (86%). Follow-up data were available from 121 patients. The mean follow-up period per patient was 4.2 years. Of the patients known to be under surveillance, 83% was screened at least annually and 94% at least every two years. Thirty-eight renal cell carcinomas had occurred in 23 patients. The mean age at diagnosis of the first tumour was 51. Eighteen tumours were visualized by ultrasound. Nine small tumours (7-27 mm) were visible on MRI or CT and not detected using ultrasound. Our data indicate that compliance to renal screening is relatively high. Furthermore, ultrasound might be a sensitive, cheap and widely available alternative for MRI or part of the MRIs for detecting clinically relevant renal tumours in BHD patients,but the limitations should be considered carefully. Data from larger cohorts are necessary to confirm these observations.

The objective of this study was to compare the effects of >Your< Iron Syrup, a novel oral liquid iron-containing food supplement, with the commonly prescribed iron sulphate (Fe-sulphate) in a mouse model of diet-induced iron deficiency. Standard inbred BALB/cOlaHsd mice were fed low-iron diet for 11 weeks to induce significant decrease in blood haemoglobin and haematocrit and were then supplemented by gavage with either >Your< Iron Syrup or Fe-sulphate for two weeks. In >Your< Iron Syrup group, several markers of iron deficiency, such as serum iron concentration, transferrin saturation and ferritin level were significantly improved in both female and male mice. Fe-sulphate induced similar responses, except that it did not significantly increase iron serum in females and serum ferritin in both sexes. Fe-sulphate significantly increased liver-iron content which >Your< Iron Syrup did not. Transcription of Hamp and selected inflammatory genes in the liver was comparable between the two supplementation groups and with the Control diet group. Some sex-specific effects were noted, which were more pronounced and less variable in males. In conclusion, >Your< Iron Syrup was efficient, comparable and in some parameters superior to Fe-sulphate in improving iron-related parameters without inducing a response of selected liver inflammation markers in a mouse model of diet-induced iron deficiency.

A Correction to this paper has been published: https://doi.org/10.1038/s41467-021-22020-7

The use of existing mouse models in cancer research is of utmost importance as they aim to explore the casual link between candidate cancer genes and carcinogenesis as well as to provide models to develop and test new therapies. However, faster progress in translating mouse cancer model research into the clinic has been hampered due to the limitations of these models to better reflect the complexities of human tumors. Traditionally, immunocompetent and immunodeficient mice with syngeneic and xenografted tumors transplanted subcutaneously or orthotopically have been used. These models are still being widely employed for many different types of studies, in part due to their widespread availability and low cost. Other types of mouse models used in cancer research comprise transgenic mice in which oncogenes can be constitutively or conditionally expressed and tumor-suppressor genes silenced using conventional methods, such as retroviral infection, microinjection of DNA constructs, and the so-called “gene-targeted transgene” approach. These traditional transgenic models have been very important in studies of carcinogenesis and tumor pathogenesis, as well as in studies evaluating the development of resistance to therapy. Recently, the clustered regularly interspaced short palindromic repeats (CRISPR)-based genome editing approach has revolutionized the field of mouse cancer models and has had a profound and rapid impact on the development of more effective systems to study human cancers. The CRISPR/Cas9-based transgenic models have the capacity to engineer a wide spectrum of mutations found in human cancers and provide solutions to problems that were previously unsolvable. Recently, humanized mouse xenograft models that accept patient-derived xenografts and CD34+ cells were developed to better mimic tumor heterogeneity, the tumor microenvironment, and cross-talk between the tumor and stromal/immune cells. These features make them extremely valuable models for the evaluation of investigational cancer therapies, specifically new immunotherapies. Taken together, improvements in both the CRISPR/Cas9 system producing more valid mouse models and in the humanized mouse xenograft models resembling complex interactions between the tumor and its environment might represent one of the successful pathways to precise individualized cancer therapy, leading to improved cancer patient survival and quality of life.

Cytochrome P450 lanosterol 14α-demethylase (CYP51) and its products, meiosis-activating sterols (MASs), were hypothesized by previous in vitro studies to have an important role in regulating meiosis and reproduction. To test this in vivo, we generated a conditional male germ cell-specific knockout of the gene Cyp51 in the mouse. High excision efficiency of Cyp51 allele in germ cells resulted in 85–89% downregulation of Cyp51 mRNA and protein levels in germ cells. Quantitative metabolic profiling revealed significantly higher levels of CYP51 substrates lanosterol and 24,25-dihydrolanosterol and substantially diminished levels of MAS, the immediate products of CYP51. However, germ cell-specific ablation of Cyp51, leading to lack of MAS, did not affect testicular morphology, daily sperm production, or reproductive performance in males. It is plausible that due to the similar structures of cholesterol intermediates, previously proposed biological function of MAS in meiosis progression can be replaced by some other yet-unidentified functionally redundant lipid molecule(s). Our results using the germ cell-specific knockout model provide first in vivo evidence that the de novo synthesis of MAS and cholesterol in male germ cells is most likely not essential for spermatogenesis and reproduction and that MASs, originating from germ cells, do not cell-autonomously regulate spermatogenesis and fertility.

Mammalian spermatogenesis is a complex developmental program in which a diploid progenitor germ cell transforms into highly specialized spermatozoa. One intriguing aspect of sperm production is the dynamic change in membrane lipid composition that occurs throughout spermatogenesis. Cholesterol content, as well as its intermediates, differs vastly between the male reproductive system and nongonadal tissues. Accumulation of cholesterol precursors such as testis meiosis-activating sterol and desmosterol is observed in testes and spermatozoa from several mammalian species. Moreover, cholesterogenic genes, especially meiosis-activating sterol-producing enzyme cytochrome P450 lanosterol 14α-demethylase, display stage-specific expression patterns during spermatogenesis. Discrepancies in gene expression patterns suggest a complex temporal and cell-type specific regulation of sterol compounds during spermatogenesis, which also involves dynamic interactions between germ and Sertoli cells. The functional importance of sterol compounds in sperm production is further supported by the modulation of sterol composition in spermatozoal membranes during epididymal transit and in the female reproductive tract, which is a prerequisite for successful fertilization. However, the exact role of sterols in male reproduction is unknown. This review discusses sterol dynamics in sperm maturation and describes recent methodological advances that will help to illuminate the complexity of sperm formation and function.