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Abstract Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a debilitating chronic condition that is characterized by unresolved fatigue, post-exertion symptom exacerbation (PESE), cognitive dysfunction, orthostatic intolerance, and other symptoms. ME/CFS lacks established clinical biomarkers and requires further elucidation of disease mechanisms. A growing number of studies demonstrate signs of hematological and cardiovascular pathology in ME/CFS cohorts, including hyperactivated platelets, endothelial dysfunction, vascular dysregulation, and anomalous clotting processes. To build on these findings, and to identify potential biomarkers that can be related to pathophysiology, we measured differences in protein expression in platelet-poor plasma (PPP) samples from 15 ME/CFS study participants and 10 controls not previously infected with SARS-CoV-2, using DIA LC-MS/MS. We identified 24 proteins that are significantly increased in the ME/CFS group compared to the controls, and 21 proteins that are significantly downregulated. Proteins related to clotting processes – thrombospondin-1 (important in platelet activation), platelet factor 4, and protein S – were differentially expressed in the ME/CFS group, suggestive of a dysregulated coagulation system and abnormal endothelial function. Complement machinery was also significantly downregulated, including C9 which forms part of the membrane attack complex. Additionally, we identified a significant upregulation of lactotransferrin, protein S100-A9, and an immunoglobulin variant. The findings from this experiment further implicate the coagulation and immune system in ME/CFS, and bring to attention the pathology of or imposed on the endothelium. This study highlights potential systems and proteins that require further research with regards to their contribution to the pathogenesis of ME/CFS, symptom manifestation, and biomarker potential, and also gives insight into the hematological and cardiovascular risk for ME/CFS individuals affected by diabetes mellitus. Graphical abstract

Climate change is an indisputable threat to human health, especially for societies already confronted with rising social inequality, political and economic uncertainty, and a cascade of concurrent environmental challenges. Archaeological data about past climate and environment provide an important source of evidence about the potential challenges humans face and the long-term outcomes of alternative short-term adaptive strategies. Evidence from well-dated archaeological human skeletons and mummified remains speaks directly to patterns of human health over time through changing circumstances. Here, we describe variation in human epidemiological patterns in the context of past rapid climate change (RCC) events and other periods of past environmental change. Case studies confirm that human communities responded to environmental changes in diverse ways depending on historical, sociocultural, and biological contingencies. Certain factors, such as social inequality and disproportionate access to resources in large, complex societies may influence the probability of major sociopolitical disruptions and reorganizations-commonly known as "collapse." This survey of Holocene human-environmental relations demonstrates how flexibility, variation, and maintenance of Indigenous knowledge can be mitigating factors in the face of environmental challenges. Although contemporary climate change is more rapid and of greater magnitude than the RCC events and other environmental changes we discuss here, these lessons from the past provide clarity about potential priorities for equitable, sustainable development and the constraints of modernity we must address.

Objectives: To formulate the most current, evidence-based recommendations for the role of medication, physical medicine, and rehabilitation in the management of acute low back pain lasting

Background: Airway management is a core component of the treatment of cardiac arrest. Where a rescuer cannot establish a patent airway to provide oxygenation and ventilation using standard basic and advanced airway techniques, there may be a need to consider emergency front-of-neck airway access (eFONA, e.g., cricothyroidotomy), but there is limited evidence to inform this approach. Objectives: This scoping review aims to identify the evidence for the use of eFONA techniques in patients with cardiac arrest. Methods: In November 2023, we searched Medline, Embase, and Cochrane Central to identify studies on eFONA in adults. We included randomised controlled trials, non-randomised studies, and case series with at least five cases that described any use of eFONA. We extracted data, including study setting, population characteristics, intervention characteristics, and outcomes. Our analysis focused on four key areas: incidence of eFONA, eFONA success rates, clinical outcomes, and complications. Results: The search identified 21,565 papers, of which 18,934 remained after de-duplication. After screening, we included 69 studies (53 reported incidence, 40 reported success rate, 38 reported clinical outcomes; 36 studies reported complications). We identified only one randomised controlled trial. Across studies, there was a total of 4,457 eFONA attempts, with a median of 31 attempts (interquartile range 16–56.5) per study. There was marked heterogeneity across studies that precluded any pooling of data. There were no studies that included only patients in cardiac arrest. Conclusion: The available evidence for eFONA is extremely heterogeneous, with no studies specifically focusing on its use in adults with cardiac arrest.

PurposeThe focus of SPINE20 is to develop evidence-based policy recommendations for the G20 countries to work with governments to reduce the burden of spine disease, and disability.MethodsOn September 17-18, 2021, SPINE20 held its annual meeting in Rome, Italy. Prior to the meeting, the SPINE20 created six proposed recommendations. These recommendations were uploaded to the SPINE20 website 10 days before the meeting and opened to the public for comments. The recommendations were discussed at the meeting allowing the participants to object and provide comments.ResultsIn total, 27 societies endorsed the following recommendations. SPINE20 calls upon the G20 countries: (1) to expand telehealth for the access to spine care, especially in light of the current situation with COVID-19. (2) To adopt value-based interprofessional spine care as an approach to improve patient outcomes and reduce disability. (3) To facilitate access and invest in the development of a competent rehabilitation workforce to reduce the burden of disability related to spine disorders. (4) To adopt a strategy to promote daily physical activity and exercises among the elderly population to maintain an active and independent life with a healthy spine, particularly after COVID-19 pandemic. (5) To engage in capacity building with emerging countries and underserved communities for the benefit of spine patients. (6) To promote strategies to transfer evidence-based advances into patient benefit through effective implementation processes.ConclusionsSPINE20's initiatives will make governments and decision makers aware of efforts to reduce needless suffering from disabling spine pain through education that can be instituted across the globe.

Glioblastoma Multiforme (GBM) is the most invasive and prevalent Central Nervous System (CNS) malignancy. It is characterised by diffuse infiltrative growth and metabolic dysregulation that impairs the extent of surgical resection (EoR), contributing to its poor prognosis. 5-Aminolevulinic acid (5-ALA) fluorescence-guided surgical resection (FGR) takes advantage of the preferential generation of 5-ALA-derived fluorescence signal in glioma cells, thereby improving visualisation and enhancing the EoR. However, despite 5-ALA FGR is a widely used technique in the surgical management of malignant gliomas, the infiltrative tumour margins usually show only vague or no visible fluorescence and thus a significant amount of residual tumour tissue may hence remain in the resection cavity, subsequently driving tumour recurrence. To investigate the molecular mechanisms that govern the preferential accumulation of 5-ALA in glioma cells, we investigated the precise subcellular localisation of 5-ALA signal using Correlative Light and Electron Microscopy (CLEM) and colocalisation analyses in U118MG glioma cells. Our results revealed strong 5-ALA signal localisation in the autophagy compartment – specifically autolysosomes and lysosomes. Flow cytometry was employed to investigate whether autophagy enhancement through spermidine treatment (SPD) or nutrient deprivation/caloric restriction (CR) would enhance 5-ALA fluorescence signal generation. Indeed, SPD, CR and a combination of SPD/CR treatment significantly increased 5-ALA signal intensity, with a most robust increase in signal intensity observed in the combination treatment of SPD/CR. When using 3-D glioma spheroids to assess the effect of 5-ALA on cellular ultrastructure, we demonstrate that 5-ALA exposure leads to cytoplasmic disruption, vacuolarisation and large-scale mitophagy induction. These findings not only suggest a critical role for the autophagy compartment in 5-ALA engagement and signal generation but also point towards a novel and practically feasible approach to enhance 5-ALA fluorescence signal intensity. The findings may highlight that indeed autophagy control may serve as a promising avenue to promote an improved resection and GBM prognosis.

Introduction: Helicopter Emergency Medical Services (HEMS) is integrated into modern emergency medical services because of its suggested mortality benefit in certain patient populations, it is an expensive resource and appropriate use/feasibility in low- to middle income countries (LMIC) is highly debated. To maximise benefit, correct patient selection in HEMS is paramount. To achieve this, current practices first need to be described. The study aims to describe a population of patients utilising HEMS in South Africa, in terms of flight data, patient demographics, provisional diagnosis, as well as clinical characteristics and interventions. Methods: A retrospective flight- and patient-chart review were conducted, extracting clinical and mission data of a single aeromedical operator in South Africa, over a 12-month period (July 2017 – June 2018) in Gauteng, Free State, Mpumalanga and North-West provinces. Results: A total of 916 cases were included (203 primary cases, 713 interfacility transport (IFT) cases). Most patients transported were male (n=548, 59.8%) and suffered blunt trauma (n=379, 41.4%). Medical pathology (n=247, 27%) and neonatal transfers (n=184, 20.1%) follows. Flights occurred mainly in daylight hours (n=729, 79.6%) with median mission times of 1-hour 53 minutes (primary missions), and 3 hours 10 minutes (IFT missions). Median on-scene times were 26 minutes (primary missions) and 55 minutes (IFT missions). Almost half were transported with an endotracheal tube (n=428, 46.7%), with a large number receiving no respiratory support (n=414, 45.2%). No patients received fibrinolysis, defibrillation, cardioversion or cardiac pacing. Intravenous fluid therapy (n=867, 94.7%) was almost universal, with common administration of sedation (n=430, 46.9%) and analgesia (n=329, 35.9%). Conclusion: Apart from the lack of universal call-out criteria and response to the high burden of trauma, HEMS seem to fulfil an important critical care transport role. It seems that cardiac pathologies are under-represented in this study and might have an important implication for crew training requirements.

Abstract We report a diagnostically challenging case of a SMARCA4‐deficient undifferentiated tumour to emphasize its potential to mimic other malignant tumours on histology, especially in small biopsies and where rhabdoid morphology is lacking. A 48‐year‐old man, who was known for chronic obstructive pulmonary disease and polysubstance use, presented with dyspnoea and an anterior mediastinal mass that had grown rapidly over a seven‐month period. The rapid growth and location in the anterior mediastinum raised clinical suspicion for lymphoma or a germ cell tumour. Microscopic examination of a transthoracic, ultrasound‐guided, core needle biopsy revealed relatively uniform, malignant epithelioid cells with clear cytoplasm, but lacking any rhabdoid features. Tumour necrosis was prominent. The immunohistochemistry panel was negative for lymphoma markers, but positive for SALL4 (a marker typically associated with germ cell tumours), CD34, EMA, and HepPar1, while expression of SMARCA4 and claudin‐4 was entirely lost. Only focal cytokeratin expression was demonstrated. SMARCB1 (INI1) expression was retained. The diagnosis of SMARCA4‐DUT was made based on these findings. Unfortunately, the tumour was already at an advanced stage at diagnosis (stage IVA) and the patient had a poor performance status. He was treated with palliative radiotherapy with no significant improvement in performance status and passed away 3 months after diagnosis. The case highlights the importance of considering SMARCA4‐DUT in the differential diagnosis of an undifferentiated, rapidly growing thoracic tumour and the potential for misdiagnosis on a small tissue sample, particularly as rhabdoid morphology may be absent.

Abstract Background Post-acute sequelae of COVID-19 (PASC), also now known as long COVID, has become a major global health and economic burden. Previously, we provided evidence that there is a significant insoluble fibrin amyloid microclot load in the circulation of individuals with long COVID, and that these microclots entrap a substantial number of inflammatory molecules, including those that might prevent clot breakdown. Scientifically, the most challenging aspect of this debilitating condition is that traditional pathology tests such as a serum CRP (C-reactive protein) may not show any significant abnormal inflammatory markers, albeit these tests measure only the soluble inflammatory molecules. Elevated, or abnormal soluble biomarkers such as IL-6, D-Dimer or fibrinogen indicate an increased risk for thrombosis or a host immune response in COVID-19. The absence of biomarkers in standard pathology tests, result in a significant amount of confusion for patients and clinicians, as patients are extremely sick or even bed-ridden but with no regular identifiable reason for their disease. Biomarkers that are currently available cannot detect the molecules present in the microclots we identified and are therefore unable to confirm their presence or the mechanisms that drive their formation. Methods Here we analysed the protein content of double-digested microclots of 99 long COVID patients and 29 healthy controls. The patients suffering from long COVID reported their symptoms through a questionnaire completed by themselves or their attending physician. Results Our long COVID cohort’s symptoms were found to be in line with global findings, where the most prevalent symptoms were constant fatigue (74%,) cognitive impairment (71%) and depression and anxiety (30%). Our most noteworthy findings were a reduced level of plasma Kallikrein compared to our controls, an increased level of platelet factor 4 (PF4) von Willebrand factor (VWF), and a marginally increased level of α-2 antiplasmin (α-2-AP). We also found a significant presence of antibodies entrapped inside these microclots. Conclusion Our results confirm the presence of pro-inflammatory molecules that may also contribute to a failed fibrinolysis phenomenon, which could possibly explain why individuals with long COVID suffer from chronic fatigue, dyspnoea, or cognitive impairment. In addition, significant platelet hyperactivation was noted. Hyperactivation will result in the granular content of platelets being shed into the circulation, including PF4. Overall, our results provide further evidence of both a failed fibrinolytic system in long COVID/PASC and the entrapment of many proteins whose presence might otherwise go unrecorded. These findings might have significant implications for individuals with pre-existing comorbidities, including cardiovascular disease and type 2 diabetes.

Background: Low- to middle-income countries account for 70% of global cancer deaths. Evidence of the changing prevalence of head and neck cancer in Africa in terms of gender, race and epidemiology will inform future research and health planning. Aim: To synthesise epidemiological literature for head and neck cancer in Africa from 2010 to 2020. Method: A scoping review was completed. The Joanna Briggs Institute Population, context and concept framework confirmed the inclusion criteria. Studies from Africa that included participant demographics, the types, stages, signs and symptoms of head and neck cancer were selected. Five databases were used. Descriptive statistics was completed. Results: The Preferred Reporting Items for Systematic Reviews and MetaAnalysis guided the reporting of the findings. Of the 1891 articles downloaded, 66 were included in the final review. Nigeria produced the most studies and oral cancer at 74% was most prevalent. Substance abuse was the most prevalent cause. Diagnosis of head and neck cancers were in the late stage (stage IV) when signs and symptoms were severe. Males of lower socioeconomic status tended to have less health seeking behaviour. Conclusion: Countries from North Africa produce the most research outputs on head and neck cancers. Gender differences were noted and may be linked to lifestyle choices. A range of head and neck cancers (HNCs) are prevalent however late diagnosis and severe symptomatology impact treatment options. Contribution: Earlier diagnosis and intervention to prevent late-stage diagnosis is necessary. Awareness campaigns linked to evidence on causes, habits and lifestyle choices, signs and symptoms are needed.

Background It is generally understood that the profile of spine surgery varies by setting, based on factors such as the age profile of the population, the economic context and access to healthcare. Relatively little is known about the profile of spine surgery in South Africa, although one previous report from the public healthcare sector suggested a high burden of trauma and infection-related surgery. To our knowledge, there has been no formal investigation in the private sector. A holistic understanding of spine surgery within our setting may be beneficial for applications such as resource allocation and informing the training needs of future specialists. Objectives To provide insight into spinal surgery in the South African private healthcare sector by describing spine surgery characteristics within a large open medical scheme, including both patient and surgeon profiles. Methods This retrospective review included adult inpatient spine surgeries funded by the largest open medical scheme in South Africa between 2008 and 2017. An anonymized dataset extracted from the scheme records included patient demographics, ICD-10 codes, procedure codes and surgeon specialization. Descriptive patient and surgery data were presented. Furthermore, the association between patient and surgery factors and surgeon specialization was investigated using univariate and multivariate analyses. Results A total of 49,576 index spine surgeries were included. The largest proportion of surgeries involved members 40-59 years old (n=23,543, 48%), approximately half involved female members (n=25,293, 51%) and most were performed by neurosurgeons (n=35,439, 72%). At least 37,755 (76%) surgeries were for degenerative pathology, 2,100 (4%) for trauma and 242 (0.5%) for infection. Adjusted risk ratios (aRR) significantly associated with orthopaedic surgeon specialization included cervical spine region aRR = 0.49 (95% C.I. 0.39-0.61), trauma aRR = 1.50 (95% C.I. 1.20-1.88), deformity aRR = 1.77 (95% C.I. 1.33-2.35) and blood transfusion aRR = 1.46 (95% C.I. 1.12-1.91). Conclusion Spine surgery in South Africa’s largest open medical scheme was dominated by surgery for degenerative pathology in older adults and was performed largely by neurosurgeons. This constituted a stark contrast to a previous report from the public sector and highlighted a mismatch between exposure during public sector registrar training and private practice post-specialization. The findings support the need for private-public collaboration as well as the importance of spine fellowships for all specialists intending to practice spine surgery.

A 77-year-old woman with suspected lung carcinoma had multiple bladder masses and lymphadenopathy outside the normal urinary bladder drainage area. Fine needle aspiration and immunocytochemistry of the cervical lymph node complex and transurethral biopsy of the bladder masses confirmed metastatic small-cell neuroendocrine carcinoma. Contribution: Clinical correlation, imaging findings, tumour markers and immunohistochemistry are necessary for metastatic bladder tumour work-up.

Abstract Background Traumatic brain injury (TBI) constitutes a global epidemic. Overall outcome is poor, with mortality ranging from 10 to 70% and significant long-term morbidity. Several experimental reports have claimed effect on traumatic edema, but no clinical trials have shown effect on edema or outcome. Antisecretory factor, an endogenous protein, is commercially available as Salovum®, which is classified as a medical food by the European Union and has shown effect in experimental trauma models and feasibility with signs of effect in 2 pilot case series. The aim of this study is to assess the effect of antisecretory factor in adult patients with severe traumatic brain injury as measured by 30-day mortality, treatment intensity level (TIL), and intracranial pressure (ICP). Methods/design This is a single-center, double-blind, randomized, placebo-controlled clinical phase 2 trial, investigating the clinical superiority of Salovum® given as a food supplement to adults with severe TBI (GCS

Objectives: The osteological paradox recognizes that the presence of lesions is not always directly related with increased mortality. When combined with the clinical, historical, and epidemiological literature on scurvy, survivorship analysis, a form of statistical analysis to assess the relationship between the presence of diseases in the archeological record and survival, helps determine the overall burden of the disease both in terms of morbidity and mortality. This article explores the relationship between scurvy and survivorship in 26 adults from Man Bac, a Neolithic site from northern Vietnam together with prepublished evidence of scurvy in the nonadult population (n = 44). Methods: Diagnosis of scurvy included differential diagnosis combined with the Snoddy, A. M. E., Buckley, H. R., Elliott, G. E., Standen, V. G., Arriaza, B. T., & Halcrow, S. E. (2018). Macroscopic features of scurvy in human skeletal remains: A literature synthesis and diagnostic guide. American Journal of Physical Anthropology, 167(4), 876–895. https://doi.org/10.1002/ajpa.23699 threshold criteria and the Brickley, M. B., & Morgan, B. (2023). Assessing diagnostic certainty for scurvy and rickets in human skeletal remains. American Journal of Biological Anthropology, 181, 637–645 diagnostic certainty approaches. Kaplan–Meier survival curves were produced to assess the relationship between the presence of probable scurvy and age‐at‐death. Results: The prevalence of probable scurvy in adults (35%) was considerably lower than reported for the nonadults (80%). Almost all lesions observed in the adults were in a mixed stage of healing. Kaplan–Meier analysis demonstrated no difference in survivorship between infants and children (<15 years) with or without probable scurvy, whereas a meaningful difference was observed for the adults and adolescents (15+ years). Conclusions: The findings demonstrate that scurvy considerably decreased survivorship to older age categories. The degree of lesion remodeling, however, indicates that scurvy was not necessarily the direct cause of death but contributed to an overall disease burden that was ultimately fatal. [ABSTRACT FROM AUTHOR]

Background: Spatial clumping of plants results in intense competition. Aims: We analysed the extent of clumping of Proteaceae seedlings across the Cape Floristic Region after fire. Methods: To demonstrate the extent of clumping in 23 species, we analysed the seedling and adult densities in 290 post-fire surveys each of 100 × 1 m2 plots using standard indices of clumping. Key results: We detected clumping of Cape Proteaceae seedlings, whereas parent plants were less clumped. The clumping is not due to limited safe sites for seedlings because the number of plots at a site with at least one seedling was positively related to the number of seedlings at the site. Sites with seedlings were enriched in nutrients relative to those without. Conclusions: The possible benefits of clumping are saturation of granivores and co-location of buried seeds with nutrient-rich patches of fire-derived ash and debris. A cause of clumping is seeds sticking together. Clumping, which is strongly intraspecific, has implications for trait evolution and the dominance of reseeders over resprouters. Strong clumping reduces the usefulness of the widely used seedling:parent index to determine whether fires were favourable or not. Implications: Clumping has implications for understanding seed dispersal distances, seedling versus adult traits, fire responses and demography. Seedlings of Cape Proteaceae tend to occur in clumps and these clumps thin out with time to produce adult stands that are less clumped. We analysed seedling and adult densities and discuss the implications for trait evolution and dominance of reseeders over resprouters of strongly intra-specific clumping. [ABSTRACT FROM AUTHOR]

Introduction: The purpose is to report on the fourth set of recommendations developed by SPINE20 to advocate for evidence-based spine care globally under the theme of “One Earth, One Family, One Future WITHOUT Spine DISABILITY”. Research question: Not applicable. Material and methods: Recommendations were developed and refined through two modified Delphi processes with international, multi-professional panels. Results: Seven recommendations were delivered to the G20 countries calling them to: - establish, prioritize and implement accessible National Spine Care Programs to improve spine care and health outcomes. - eliminate structural barriers to accessing timely rehabilitation for spinal disorders to reduce poverty. - implement cost-effective, evidence-based practice for digital transformation in spine care, to deliver self-management and prevention, evaluate practice and measure outcomes. - monitor and reduce safety lapses in primary care including missed diagnoses of serious spine pathologies and risk factors for spinal disability and chronicity. - develop, implement and evaluate standardization processes for spine care delivery systems tailored to individual and population health needs. - ensure accessible and affordable quality care to persons with spine disorders, injuries and related disabilities throughout the lifespan. - promote and facilitate healthy lifestyle choices (including physical activity, nutrition, smoking cessation) to improve spine wellness and health. Discussion and conclusion: SPINE20 proposes that focusing on the recommendations would facilitate equitable access to health systems, affordable spine care delivered by a competent healthcare workforce, and education of persons with spine disorders, which will contribute to reducing spine disability, associated poverty, and increase productivity of the G20 nations.

This article outlines the research and development of a blockchain assessment framework which enables the assessment of the technical suitability, high-level design, adoption approach, economic feasibility, and business value potential of a blockchain solution with a particular organization for a specific process. The framework is a comprehensive, high-level, and generic assessment approach that enables better decision-making regarding blockchain exploration. Blockchain is a novel technology with the potential to disrupt several industries through its possession of many desirable functional characteristics, including, but not limited to, immutability, transparency, decentralization, and secure. Cryptocurrencies and these desirable characteristics have created hype around blockchain, consequently leading to blockchain projects with minimal understanding of what the technology is capable of and beneficial for, resulting in excessively high failure rates. Attempts have been made by researchers to reduce these high failure rates by creating a better understanding of blockchain, as well as creating assessment approaches. However, these approaches tend to apply to specific narrow use cases, or the approach is not comprehensive and only considers one aspect of blockchain assessment. This emphasizes the need for a comprehensive and generic blockchain assessment approach to aid with better decision-making regarding blockchain within organizations. This article aims at addressing this need by creating a blockchain assessment framework to aids with deciding whether it is worthwhile investing more time, effort, and money into blockchain exploration. The context of the study is set in the introduction, this is then followed by a brief explanation of the blockchain technology. Thereafter, the blockchain assessment framework is presented, followed by a brief explanation of the demonstration and validation of the framework using a case study and expert analysis. The framework is most valuable during the initial stages of blockchain exploration and creates momentum for further blockchain exploration in an organization. The study concludes with the limitations and future research recommendations.

Viruses have evolved mechanisms to modulate cellular pathways to facilitate infection. One such pathway is the formation of stress granules (SG), which are ribonucleoprotein complexes that assemble during translation inhibition following cellular stress. Inhibition of SG assembly has been observed under numerous virus infections across species, suggesting a conserved fundamental viral strategy. However, the significance of SG modulation during virus infection is not fully understood. The 1A protein encoded by the model dicistrovirus, Cricket paralysis virus (CrPV), is a multifunctional protein that can bind to and degrade Ago-2 in an E3 ubiquitin ligase-dependent manner to block the antiviral RNA interference pathway and inhibit SG formation. Moreover, the R146 residue of 1A is necessary for SG inhibition and CrPV infection in both Drosophila S2 cells and adult flies. Here, we uncoupled CrPV-1A's functions and provide insight into its underlying mechanism for SG inhibition. CrPV-1A mediated inhibition of SGs requires the E3 ubiquitin-ligase binding domain and the R146 residue, but not the Ago-2 binding domain. Wild-type but not mutant CrPV-1A R146A localizes to the nuclear membrane which correlates with nuclear enrichment of poly(A)+ RNA. Transcriptome changes in CrPV-infected cells are dependent on the R146 residue. Finally, Nup358/RanBP2 is targeted and degraded in CrPV-infected cells in an R146-dependent manner and the depletion of Nup358 blocks SG formation. We propose that CrPV utilizes a multiprong strategy whereby the CrPV-1A protein interferes with a nuclear event that contributes to SG inhibition in order to promote infection.

Parkinson’s disease (PD), the fastest-growing neurological disorder globally, has a complex etiology. A previous study by our group identified the p.G849D variant in neurexin 2 (NRXN2), encoding the synaptic protein, NRXN2α, as a possible causal variant of PD. Therefore, we aimed to perform functional studies using proteomics in an attempt to understand the biological pathways affected by the variant. We hypothesized that this may reveal insight into the pathobiology of PD. Wild-type and mutant NRXN2α plasmids were transfected into SH-SY5Y cells. Thereafter, total protein was extracted and prepared for mass spectrometry using a Thermo Scientific Fusion mass spectrometer equipped with a Nanospray Flex ionization source. The data were then interrogated against the UniProt H. sapiens database and afterward, pathway and enrichment analyses were performed using in silico tools. Overexpression of the wild-type protein led to the enrichment of proteins involved in neurodegenerative diseases, while overexpression of the mutant protein led to the decline of proteins involved in ribosomal functioning. Thus, we concluded that the wild-type NRXN2α may be involved in pathways related to the development of neurodegenerative disorders, and that biological processes related to the ribosome, transcription, and tRNA, specifically at the synapse, could be an important mechanism in PD. Future studies targeting translation at the synapse in PD could therefore provide further information on the pathobiology of the disease.

Abstract Background Severe acute respiratory syndrome coronavirus 2 (SARS-Cov-2)-induced infection, the cause of coronavirus disease 2019 (COVID-19), is characterized by acute clinical pathologies, including various coagulopathies that may be accompanied by hypercoagulation and platelet hyperactivation. Recently, a new COVID-19 phenotype has been noted in patients after they have ostensibly recovered from acute COVID-19 symptoms. This new syndrome is commonly termed Long COVID/Post-Acute Sequelae of COVID-19 (PASC). Here we refer to it as Long COVID/PASC. Lingering symptoms persist for as much as 6 months (or longer) after acute infection, where COVID-19 survivors complain of recurring fatigue or muscle weakness, being out of breath, sleep difficulties, and anxiety or depression. Given that blood clots can block microcapillaries and thereby inhibit oxygen exchange, we here investigate if the lingering symptoms that individuals with Long COVID/PASC manifest might be due to the presence of persistent circulating plasma microclots that are resistant to fibrinolysis. Methods We use techniques including proteomics and fluorescence microscopy to study plasma samples from healthy individuals, individuals with Type 2 Diabetes Mellitus (T2DM), with acute COVID-19, and those with Long COVID/PASC symptoms. Results We show that plasma samples from Long COVID/PASC still contain large anomalous (amyloid) deposits (microclots). We also show that these microclots in both acute COVID-19 and Long COVID/PASC plasma samples are resistant to fibrinolysis (compared to plasma from controls and T2DM), even after trypsinisation. After a second trypsinization, the persistent pellet deposits (microclots) were solubilized. We detected various inflammatory molecules that are substantially increased in both the supernatant and trapped in the solubilized pellet deposits of acute COVID-19 and Long COVID/PASC, versus the equivalent volume of fully digested fluid of the control samples and T2DM. Of particular interest was a substantial increase in α(2)-antiplasmin (α2AP), various fibrinogen chains, as well as Serum Amyloid A (SAA) that were trapped in the solubilized fibrinolytic-resistant pellet deposits. Conclusions Clotting pathologies in both acute COVID-19 infection and in Long COVID/PASC might benefit from following a regime of continued anticlotting therapy to support the fibrinolytic system function.

Abstract Three-dimensional models are considered a powerful tool for improving the concordance between in vitro and in vivo phenotypes. However, the duration of spheroid culture may influence the degree of correlation between these counterparts. When using immortalised cell lines as model systems, the assumption for consistency and reproducibility is often made without adequate characterization or validation. It is therefore essential to define the biology of each spheroid model by investigating proteomic dynamics, which may be altered relative to culture duration. As an example, we assessed the influence of culture duration on the relative proteome abundance of HepG2 cells cultured as spheroids, which are routinely used to model aspects of the liver. Quantitative proteomic profiling of whole cell lysates labelled with tandem-mass tags was conducted using liquid chromatography-tandem mass spectrometry (LC–MS/MS). In excess of 4800 proteins were confidently identified, which were shared across three consecutive time points over 28 days. The HepG2 spheroid proteome was divergent from the monolayer proteome after 14 days in culture and continued to change over the successive culture time points. Proteins representing the recognised core hepatic proteome, cell junction, extracellular matrix, and cell adhesion proteins were found to be continually modulated.