Femorotibial angle scan-rescan reproducibility: A high-precision calculation on a large cohort.

PURPOSE: Femorotibial angle (FTA) is a convenient measure of coronal knee alignment that can be extracted from a short knee radiograph, avoiding the additional radiation exposure and specialist equipment required for full-leg radiographs. While intra- and inter-reader reproducibility from the same image has been reported, the full scan-rescan reproducibility across images, as calculated in this study, has not. METHODS: In this study, 4589 FTA measurement pairs from 2586 subjects acquired a year apart were used to estimate FTA scan-rescan reproducibility using data from the Osteoarthritis Initiative. Subjects with radiographic progression of osteoarthritis or other conditions that may cause a change in coronal knee alignment were excluded. Measurement pairs were analysed using paired-samples  t $t$ tests to detect differences and compared to symptomatic changes in Western Ontario and McMaster Universities Arthritis Index scores for joint pain, stiffness and physical function to detect correlations. RESULTS: The 95% limit of agreement and the paired-samples correlation were calculated with high precision to be [-1.76°, +1.78°] and 0.938, considerably worse than the corresponding figures for intra- and inter-reader reproducibility, without relation to symptomatic or radiographic changes in knee condition. This error will weakly attenuate R 2 ${R}^{2}$ and r $r$ values from their true values in correlative studies involving FTA. The realistic maximum value for R 2 ${R}^{2}$ is 87% and for Pearson's r $r$ is 93%. CONCLUSION: The scan-rescan reproducibility in FTA is almost double the intra- and inter-reader reliability from a single scan. At almost ±2° accuracy, FTA is inappropriate for surgical use, but it is sufficiently reproducible to produce good correlations in studies predicting disease incidence and progression. LEVEL OF EVIDENCE: Level II, retrospective study.

Wireless implantable bioelectronics with a direct electron transfer lactate enzyme for detection of surgical site infection in orthopaedics.

Periprosthetic infection is one of the most devastating complications following orthopaedic surgery. Rapid detection of an infection can change the treatment pathway and improve outcomes for the patient. In here, we propose a miniaturized lactate biosensor developed on a flexible substrate and integrated on a small-form bone implant to detect infection. The methods for lactate biosensor fabrication and integration on a bone implant are fully described within this study. The system performance was comprehensively electrochemically characterised, including with L-lactate solutions prepared in phosphate-buffered saline and culture medium, and interferents such as acetaminophen and ascorbic acid. A proof-of-concept demonstration was then conducted with ex vivo ovine femoral heads incubated with and without exposure to Staphylococcus epidermidis. The sensitivity, current density and limit-of-detection levels achieved by the biosensor were 1.25 µA mM-1, 1.51 µA.M-1.mm-2 and 66 µM, respectively. The system was insensitive to acetaminophen, while sensitivity to ascorbic acid was half that of the sensitivity to L-lactate. In the ex vivo bone model, S. epidermidis infection was detected within 5 h of implantation, while the control sample led to no change in the sensor readings. This pioneering work demonstrates a pathway to improving orthopaedic outcomes by enabling early infection diagnosis.

Operational patient care pathway: building pragmatic medical interoperability through health engagements.

The future operational demand for medical support in Western militaries will likely outstrip available resources, necessitating burden-sharing through medical interoperability with allies and partners. However, the current North Atlantic Treaty Organization (NATO) model of interoperability through standardisation, while achieving high levels of commonality and integration along the operational patient care pathway (OPCP), is high-cost and resource-intensive. We have termed this model assured interoperability. Assured interoperability, while applicable to well-established partnerships with high-resource nations, is unlikely to be feasible when working with resource-limited partners or, potentially, when in a sustained conflict with a near-peer adversary. In these circumstances, there will be a requirement to develop a far less resource-intensive model of medical interoperability with lower levels of commonality, assurance and standardisation than assured interoperability, but that provides a 'good enough' OPCP for the operational context. We have termed this pragmatic interoperability. By considering these two types of interoperability, the complete continuum of medical interoperability can be mapped with the full spectrum of partners demonstrating increasing levels of interoperability from pragmatic through to assured interoperability, integrateability and interchangeability, reducing the gap between demand and provision of medical support for operations, increasing operational resilience. This is a paper commissioned as a part of the Defence Engagement special issue of BMJ Military Health.

Active osseointegration in an ex vivo porcine bone model.

Achieving osseointegration is a fundamental requirement for many orthopaedic, oral, and craniofacial implants. Osseointegration typically takes three to 6 months, during which time implants are at risk of loosening. The aim of this study was to investigate whether osseointegration could be actively enhanced by delivering controllable electromechanical stimuli to the periprosthetic bone. First, the osteoconductivity of the implant surface was confirmed using an in vitro culture with murine preosteoblasts. The effects of active treatment on osseointegration were then investigated in a 21-day ex vivo model with freshly harvested cancellous bone cylinders (n = 24; Ø10 mm × 5 mm) from distal porcine femora, with comparisons to specimens treated by a distant ultrasound source and static controls. Cell viability, proliferation and distribution was evident throughout culture. Superior ongrowth of tissue onto the titanium discs during culture was observed in the actively stimulated specimens, with evidence of ten-times increased mineralisation after 7 and 14 days of culture (p < 0.05) and 2.5 times increased expression of osteopontin (p < 0.005), an adhesive protein, at 21 days. Moreover, histological analyses revealed increased bone remodelling at the implant-bone interface in the actively stimulated specimens compared to the passive controls. Active osseointegration is an exciting new approach for accelerating bone growth into and around implants.

Atrogin-1 promotes muscle homeostasis by regulating levels of endoplasmic reticulum chaperone BiP.

Skeletal muscle wasting results from numerous pathological conditions affecting both the musculoskeletal and nervous systems. A unifying feature of these pathologies is the upregulation of members of the E3 ubiquitin ligase family, resulting in increased proteolytic degradation of target proteins. Despite the critical role of E3 ubiquitin ligases in regulating muscle mass, the specific proteins they target for degradation and the mechanisms by which they regulate skeletal muscle homeostasis remain ill-defined. Here, using zebrafish loss-of-function models combined with in vivo cell biology and proteomic approaches, we reveal a role of atrogin-1 in regulating the levels of the endoplasmic reticulum chaperone BiP. Loss of atrogin-1 resulted in an accumulation of BiP, leading to impaired mitochondrial dynamics and a subsequent loss in muscle fiber integrity. We further implicated a disruption in atrogin-1-mediated BiP regulation in the pathogenesis of Duchenne muscular dystrophy. We revealed that BiP was not only upregulated in Duchenne muscular dystrophy, but its inhibition using pharmacological strategies, or by upregulating atrogin-1, significantly ameliorated pathology in a zebrafish model of Duchenne muscular dystrophy. Collectively, our data implicate atrogin-1 and BiP in the pathogenesis of Duchenne muscular dystrophy and highlight atrogin-1's essential role in maintaining muscle homeostasis.

Passive Biotelemetric Detection of Tibial Debonding in Wireless Battery-Free Smart Knee Implants.

Aseptic loosening is the dominant failure mechanism in contemporary knee replacement surgery, but diagnostic techniques are poorly sensitive to the early stages of loosening and poorly specific in delineating aseptic cases from infections. Smart implants have been proposed as a solution, but incorporating components for sensing, powering, processing, and communication increases device cost, size, and risk; hence, minimising onboard instrumentation is desirable. In this study, two wireless, battery-free smart implants were developed that used passive biotelemetry to measure fixation at the implant-cement interface of the tibial components. The sensing system comprised of a piezoelectric transducer and coil, with the transducer affixed to the superior surface of the tibial trays of both partial (PKR) and total knee replacement (TKR) systems. Fixation was measured via pulse-echo responses elicited via a three-coil inductive link. The instrumented systems could detect loss of fixation when the implants were partially debonded (+7.1% PKA, +32.6% TKA, both p < 0.001) and fully debonded in situ (+6.3% PKA, +32.5% TKA, both p < 0.001). Measurements were robust to variations in positioning of the external reader, soft tissue, and the femoral component. With low cost and small form factor, the smart implant concept could be adopted for clinical use, particularly for generating an understanding of uncertain aseptic loosening mechanisms.

Integrative and comparative genomic analyses of mammalian macrophage responses to intracellular mycobacterial pathogens.

Mycobacterium tuberculosis, the causative agent of human tuberculosis (hTB), is a close evolutionary relative of Mycobacterium bovis, which causes bovine tuberculosis (bTB), one of the most damaging infectious diseases to livestock agriculture. Previous studies have shown that the pathogenesis of bTB disease is comparable to hTB disease, and that the bovine and human alveolar macrophage (bAM and hAM, respectively) transcriptomes are extensively reprogrammed in response to infection with these intracellular mycobacterial pathogens. In this study, a multi-omics integrative approach was applied with functional genomics and GWAS data sets across the two primary hosts (Bos taurus and Homo sapiens) and both pathogens (M. bovis and M. tuberculosis). Four different experimental infection groups were used: 1) bAM infected with M. bovis, 2) bAM infected with M. tuberculosis, 3) hAM infected with M. tuberculosis, and 4) human monocyte-derived macrophages (hMDM) infected with M. tuberculosis. RNA-seq data from these experiments 24 h post-infection (24 hpi) was analysed using three computational pipelines: 1) differentially expressed genes, 2) differential gene expression interaction networks, and 3) combined pathway analysis. The results were integrated with high-resolution bovine and human GWAS data sets to detect novel quantitative trait loci (QTLs) for resistance to mycobacterial infection and resilience to disease. This revealed common and unique response macrophage pathways for both pathogens and identified 32 genes (12 bovine and 20 human) significantly enriched for SNPs associated with disease resistance, the majority of which encode key components of the NF-κB signalling pathway and that also drive formation of the granuloma.

Preferential differential gene expression within the WC1.1+ γδ T cell compartment in cattle naturally infected with Mycobacterium bovis.

Bovine tuberculosis (bTB), caused by infection with Mycobacterium bovis, continues to cause significant issues for the global agriculture industry as well as for human health. An incomplete understanding of the host immune response contributes to the challenges of control and eradication of this zoonotic disease. In this study, high-throughput bulk RNA sequencing (RNA-seq) was used to characterise differential gene expression in γδ T cells - a subgroup of T cells that bridge innate and adaptive immunity and have known anti-mycobacterial response mechanisms. γδ T cell subsets are classified based on expression of a pathogen-recognition receptor known as Workshop Cluster 1 (WC1) and we hypothesised that bTB disease may alter the phenotype and function of specific γδ T cell subsets. Peripheral blood was collected from naturally M. bovis-infected (positive for single intradermal comparative tuberculin test (SICTT) and IFN-γ ELISA) and age- and sex-matched, non-infected control Holstein-Friesian cattle. γδ T subsets were isolated using fluorescence activated cell sorting (n = 10-12 per group) and high-quality RNA extracted from each purified lymphocyte subset (WC1.1+, WC1.2+, WC1- and γδ-) was used to generate transcriptomes using bulk RNA-seq (n = 6 per group, representing a total of 48 RNA-seq libraries). Relatively low numbers of differentially expressed genes (DEGs) were observed between most cell subsets; however, 189 genes were significantly differentially expressed in the M. bovis-infected compared to the control groups for the WC1.1+ γδ T cell compartment (absolute log2 FC ≥ 1.5 and FDR P adj. ≤ 0.1). The majority of these DEGs (168) were significantly increased in expression in cells from the bTB+ cattle and included genes encoding transcription factors (TBX21 and EOMES), chemokine receptors (CCR5 and CCR7), granzymes (GZMA, GZMM, and GZMH) and multiple killer cell immunoglobulin-like receptor (KIR) proteins indicating cytotoxic functions. Biological pathway overrepresentation analysis revealed enrichment of genes with multiple immune functions including cell activation, proliferation, chemotaxis, and cytotoxicity of lymphocytes. In conclusion, γδ T cells have important inflammatory and regulatory functions in cattle, and we provide evidence for preferential differential activation of the WC1.1+ specific subset in cattle naturally infected with M. bovis.

Extinction of high-power laser radiation under adverse weather conditions.

The propagation of laser radiation over long distances can be significantly affected by atmospheric extinction due to precipitation as well as aerosol particles and molecules. The knowledge of the contribution of precipitation is critical to the operation of a variety of laser-based systems. The study of high-power laser transmission around 1 µm is of particular interest because several atmospheric transmission windows are located in this region. To investigate the effect of adverse weather conditions on laser transmission, free-space laser transmission experiments are conducted on the DLR test range in Lampoldshausen, Germany. A high-power laser with a wavelength of 1.03 µm is used for the transmission measurements in combination with calibrated power monitors. Local weather conditions are continuously monitored by meteorological instruments during the experiments. Extinction coefficients are derived from transmission measurements showing that the extinction for snow is 7 times higher than for rain, and the extinction for drizzle/rain is 4 times higher than for rain at a given precipitation rate of 1 mm/h. For a mixture of rain and snow, the extinction is comparable to that of rain, indicating that the water content strongly influences the optical properties and thus the extinction of laser radiation in mixed precipitation. A good relationship is found between the measured extinction coefficient and visibility for drizzle and rain and a slightly larger scatter of the data for snow. Furthermore, measured extinction coefficients are compared to the extinction coefficients based on the experimental size distributions of precipitation particles and geometric optics. A reasonable agreement is obtained for rain, with no improvement taking the forward-scattering into the detector aperture into account, and a much better agreement is obtained for snow when the forward-scattering contribution is included.

Cell surface plasticity in response to shape change in the whole organism.

Plasma membrane rupture can result in catastrophic cell death. The skeletal muscle fiber plasma membrane, the sarcolemma, provides an extreme example of a membrane subject to mechanical stress since these cells specifically evolved to generate contraction and movement. A quantitative model correlating ultrastructural remodeling of surface architecture with tissue changes in vivo is required to understand how membrane domains contribute to the shape changes associated with tissue deformation in whole animals. We and others have shown that loss of caveolae, small invaginations of the plasma membrane particularly prevalent in the muscle sarcolemma, renders the plasma membrane more susceptible to rupture during stretch.1,2,3 While it is thought that caveolae are able to flatten and be absorbed into the bulk membrane to buffer local membrane expansion, a direct demonstration of this model in vivo has been unachievable since it would require measurement of caveolae at the nanoscale combined with detailed whole-animal morphometrics under conditions of perturbation. Here, we describe the development and application of the "active trapping model" where embryonic zebrafish are immobilized in a curved state that mimics natural body axis curvature during an escape response. The model is amenable to multiscale, multimodal imaging including high-resolution whole-animal three-dimensional quantitative electron microscopy. Using the active trapping model, we demonstrate the essential role of caveolae in maintaining sarcolemmal integrity and quantify the specific contribution of caveolar-derived membrane to surface expansion. We show that caveolae directly contribute to an increase in plasma membrane surface area under physiologically relevant membrane deformation conditions.

Toxic Shock Syndrome After Orthopaedic Surgery.

Toxic shock syndrome (TSS) is a rare, life-threatening, acute multisystem disorder caused by exotoxin-producing streptococcal or staphylococcal bacteria. It is often characterised by pyrexia, diffuse erythroderma, malaise, confusion, and hypotension which may progress to multiorgan dysfunction and coma. A high index of suspicion along with immediate diagnosis and multidisciplinary management is required to improve the outcome of the disease. A 62-year-old male presented to the hospital a week after an open reduction and internal fixation of a left wrist fracture. He was confused, febrile, and hypotensive with a generalised maculopapular rash on admission. Surgical wound sepsis was a top differential diagnosis; however, other possible sources were considered. Diagnostic imaging and echocardiography effectively ruled out other possible aetiologies. Despite fluids, vasopressor support, and appropriate antibiotics, he showed no significant clinical improvement. Admission blood cultures grew Staphylococcus aureus and after a multidisciplinary meeting, he was taken to the theatre for wound exploration, debridement, and removal of the metal plate. He was eventually weaned off vasopressor support and recovered well. A high index of suspicion is important in recognising TSS in postoperative orthopaedic patients as wounds may appear healthy-looking and the onset of symptoms may be delayed. Early recognition, timely intervention, and multidisciplinary management are vital to the care of these patients.

Optimising the zebrafish Cre/Lox toolbox. Codon improved iCre, new gateway tools, Cre protein and guidelines.

We recently introduced the Cre/Lox technology in our laboratory for both transient (mRNA injections) and stable/transgenic experiments. We experienced significant issues such as silencing, mosaicism, and partial recombination using both approaches. Reviewing the literature gave us the impression that these issues are common among the zebrafish community using the Cre/Lox system. While some researchers took advantage of these problems for specific applications, such as cell and lineage tracing using the Zebrabow construct, we tried here to improve the efficiency and reliability of this system by constituting and testing a new set of tools for zebrafish genetics. First, we implemented a codon-improved Cre version (iCre) designed for rodent studies to counteract some of the aforementioned problems. This eukaryotic-like iCre version was engineered to i) reduce silencing, ii) increase mRNA stability, iii) enhance translational efficiency, and iv) improve nuclear translocation. Second, we established a new set of tol2-kit compatible vectors to facilitate the generation of either iCre-mRNA or iCre-transgenes for transient and transgenic experiments, respectively. We then validated the use of this material and are providing tips for users. Interestingly, during the validation steps, we found that maternal iCRE-mRNA and/or protein deposition from female transgenics systematically led to complete/homogeneous conversion of all tested Lox-responder-transgenes, as opposed to some residual imperfect conversion when using males-drivers or mRNA injections. Considering that we did not find any evidence of Cre-protein soaking and injections in the literature as it is usually conducted with cells, we tested these approaches. While soaking of cell-permeant CRE-protein did not lead to any detectable Lox-conversion, 1ng-10 ng protein injections led to robust and homogeneous Lox-recombination, suggesting that the use of protein could be a robust option for exogenous delivery. This approach may be particularly useful to manipulate housekeeping genes involved in development, sex determination and reproduction which are difficult to investigate with traditional knockout approaches. All in all, we are providing here a new set of tools that should be useful in the field.

Electromechanical and biological evaluations of 0.94Bi0.5Na0.5TiO3-0.06BaTiO3 as a lead-free piezoceramic for implantable bioelectronics.

Smart implantable electronic medical devices are being developed to deliver healthcare that is more connected, personalised, and precise. Many of these implantables rely on piezoceramics for sensing, communication, energy autonomy, and biological stimulation, but the piezoceramics with the strongest piezoelectric coefficients are almost exclusively lead-based. In this article, we evaluate the electromechanical and biological characteristics of a lead-free alternative, 0.94Bi0.5Na0.5TiO3-0.06BaTiO3 (BNT-6BT), manufactured via two synthesis routes: the conventional solid-state method (PIC700) and tape casting (TC-BNT-6BT). The BNT-6BT materials exhibited soft piezoelectric properties, with d33 piezoelectric coefficients that were inferior to commonly used PZT (PIC700: 116 pC/N; TC-BNT-6BT: 121 pC/N; PZT-5A: 400 pC/N). The material may be viable as a lead-free substitute for soft PZT where moderate performance losses up to 10 dB are tolerable, such as pressure sensing and pulse-echo measurement. No short-term harmful biological effects of BNT-6BT were detected and the material was conducive to the proliferation of MC3T3-E1 murine preosteoblasts. BNT-6BT could therefore be a viable material for electroactive implants and implantable electronics without the need for hermetic sealing.

Defence Engagement (Health): a historical perspective.

Defence Engagement (DE) (Health) themes are reviewed in this paper through two historical perspectives set in very different contexts. The first-person narrative in Guerrilla Surgeon by Lindsay Rogers outlines the experience of a medical officer building capacity with Tito's Partisans behind enemy lines in Second World War Yugoslavia. This is contrasted with a more academic evaluation of strategic and medical benefits of DE (Health) delivered by the US military during the Vietnam War in Robert Wilensky's Military Medicine to Win Hearts and Minds: Aid to Civilians in the Vietnam War Both texts infer that clear objectives, supported by effective strategic communication, are required for the impact of DE (Health) to be fully realised. Wilensky, in particular, noted that the US military medical effort in Vietnam had no measurable impact on health or political goals in the conflict. Rogers' experience on a more individual level speaks to the promise of DE (Health) delivery contrasted against the lack of regional objectives and cites the loss of British influence when Soviet propaganda was more cohesive and coordinated, resulting in the shift in Partisan loyalty despite British efforts in supplying the bulk of military and medical material. While neither author offers a definitive guide on DE (Health), they offer clear examples of themes that should be considered and demonstrate the importance of evaluating activity and maintaining the historical record to provide an evidence base for future work. This is an article commissioned for the Defence Engagement special issue of BMJ Military Health.

Defence context for the UK's Defence Engagement (Health).

Defence Engagement (DE) has been a core UK Defence task since 2015. DE (Health) is the use of military medical capabilities to achieve DE effects within the health sector to achieve security and defence objectives. DE (Health) practitioners must understand the underlying defence context that shapes these objectives. The strategic context is becoming more uncertain with the return of great power competition layered on enduring threats from non-state actors and transnational challenges. The UK response has been to develop the Integrated Review, outlining four national security and international policy objectives. UK Defence has responded by developing the integrated operating concept, differentiating military activity between operating and warfighting. Engage is one of the three functions of operate activity, which is complementary to the other operate functions of protect and constrain. DE (Health) can play a unique role in engagement, given its ability to develop new partnerships through health-related activity. DE (Health) may be an enabler for other engagements or to enable the protect and constrain functions. This will be dependent on delivering improvement in health outcomes. Therefore, the DE (Health) practitioner must be conversant with both the contemporary defence and global health contexts to deliver effective DE (Health) activities. This is an article commissioned for the DE special issue of BMJ Military Health.

Predicting hip-knee-ankle and femorotibial angles from knee radiographs with deep learning.

BACKGROUND: Knee alignment affects the development and surgical treatment of knee osteoarthritis. Automating femorotibial angle (FTA) and hip-knee-ankle angle (HKA) measurement from radiographs could improve reliability and save time. Further, if HKA could be predicted from knee-only radiographs then radiation exposure could be reduced and the need for specialist equipment and personnel avoided. The aim of this research was to assess if deep learning methods could predict FTA and HKA angle from posteroanterior (PA) knee radiographs. METHODS: Convolutional neural networks with densely connected final layers were trained to analyse PA knee radiographs from the Osteoarthritis Initiative (OAI) database. The FTA dataset with 6149 radiographs and HKA dataset with 2351 radiographs were split into training, validation, and test datasets in a 70:15:15 ratio. Separate models were developed for the prediction of FTA and HKA and their accuracy was quantified using mean squared error as loss function. Heat maps were used to identify the anatomical features within each image that most contributed to the predicted angles. RESULTS: High accuracy was achieved for both FTA (mean absolute error 0.8°) and HKA (mean absolute error 1.7°). Heat maps for both models were concentrated on the knee anatomy and could prove a valuable tool for assessing prediction reliability in clinical application. CONCLUSION: Deep learning techniques enable fast, reliable and accurate predictions of both FTA and HKA from plain knee radiographs and could lead to cost savings for healthcare providers and reduced radiation exposure for patients.

Bioreactor analyses of tissue ingrowth, ongrowth and remodelling around implants: An alternative to live animal testing.

Introduction: Preclinical assessment of bone remodelling onto, into or around novel implant technologies is underpinned by a large live animal testing burden. The aim of this study was to explore whether a lab-based bioreactor model could provide similar insight. Method: Twelve ex vivo trabecular bone cylinders were extracted from porcine femora and were implanted with additively manufactured stochastic porous titanium implants. Half were cultured dynamically, in a bioreactor with continuous fluid flow and daily cyclic loading, and half in static well plates. Tissue ongrowth, ingrowth and remodelling around the implants were evaluated with imaging and mechanical testing. Results: For both culture conditions, scanning electron microscopy (SEM) revealed bone ongrowth; widefield, backscatter SEM, micro computed tomography scanning, and histology revealed mineralisation inside the implant pores; and histology revealed woven bone formation and bone resorption around the implant. The imaging evidence of this tissue ongrowth, ingrowth and remodelling around the implant was greater for the dynamically cultured samples, and the mechanical testing revealed that the dynamically cultured samples had approximately three times greater push-through fixation strength (p < 0.05). Discussion: Ex vivo bone models enable the analysis of tissue remodelling onto, into and around porous implants in the lab. While static culture conditions exhibited some characteristics of bony adaptation to implantation, simulating physiological conditions with a bioreactor led to an accelerated response.

Caveolae sense oxidative stress through membrane lipid peroxidation and cytosolic release of CAVIN1 to regulate NRF2.

Caveolae have been linked to many biological functions, but their precise roles are unclear. Using quantitative whole-cell proteomics of genome-edited cells, we show that the oxidative stress response is the major pathway dysregulated in cells lacking the key caveola structural protein, CAVIN1. CAVIN1 deletion compromised sensitivity to oxidative stress in cultured cells and in animals. Wound-induced accumulation of reactive oxygen species and apoptosis were suppressed in Cavin1-null zebrafish, negatively affecting regeneration. Oxidative stress triggered lipid peroxidation and induced caveolar disassembly. The resulting release of CAVIN1 from caveolae allowed direct interaction between CAVIN1 and NRF2, a key regulator of the antioxidant response, facilitating NRF2 degradation. CAVIN1-null cells with impaired negative regulation of NRF2 showed resistance to lipid-peroxidation-induced ferroptosis. Thus, caveolae, via lipid peroxidation and CAVIN1 release, maintain cellular susceptibility to oxidative-stress-induced cell death, demonstrating a crucial role for this organelle in cellular homeostasis and wound response.

Integrative genomics analysis highlights functionally relevant genes for equine behaviour.

Behavioural plasticity enables horses entering an exercise training programme to adapt with reduced stress. We characterised SNPs associated with behaviour in yearling Thoroughbred horses using genomics analyses for two phenotypes: (1) handler-assessed coping with early training events [coping] (n = 96); and (2) variation in salivary cortisol concentration at the first backing event [cortisol] (n = 34). Using RNA-seq derived gene expression data for amygdala and hippocampus tissues from n = 2 Thoroughbred stallions, we refined the SNPs to those with functional relevance to behaviour by cross-referencing to the 500 most highly expressed genes in each tissue. The SNPs of high significance (q < 0.01) were in proximity to genes (coping - GABARAP, NDM, OAZ1, RPS15A, SPARCL1, VAMP2; cortisol - CEBPA, COA3, DUSP1, HNRNPH1, RACK1) with biological functions in social behaviour, autism spectrum disorder, suicide, stress-induced anxiety and depression, Alzheimer's disease, neurodevelopmental disorders, neuroinflammatory disease, fear-induced behaviours and alcohol and cocaine addiction. The strongest association (q = 0.0002) was with NDN, a gene previously associated with temperament in cattle. This approach highlights functionally relevant genes in the behavioural adaptation of Thoroughbred horses that will contribute to the development of genetic markers to improve racehorse welfare.