Search

Background and Aim: The sinoatrial node is responsible for the intrinsic electrical activation that in mammals leads to coordinated rhythmic contractions of the heart, from where it is distributed through the atrial tissue to the atrioventricular node. This study aimed to conduct a histological and morphometric study of the components and cells in cardiac nodes altered by myocardial infarction (MI) and compare them with normal tissues in humans and pigs. Materials and Methods: We analyzed 10 human hearts and 10 pig hearts that died from MI and compared them with 10 healthy control hearts from each species. Histological sections of 5 μm thickness were obtained using a microtome and stained with hematoxylin–eosin and Masson’s trichrome. The identification and assessment of the percentage of connective tissue and cellularity in the cardiac nodes were performed. Results: We observed a decreased size of cardiac nodes in humans and pigs, as well as an increased percentage of fibrosis inside the nodes, and changes in the size of the nodal cells and surrounding cardiomyocytes (decrease or hypertrophy) were observed. Cartilaginous metaplasia was also found in the cardiac skeleton of all pig samples. Conclusion: In the present study, a significant increase in collagen fibers and a decrease in cellularity were found in cardiac nodes in samples from humans and pigs with MI. These findings would explain the presence of arrhythmias, which often lead to death.

Sex steroid hormones (SSH) are extremely versatile molecules with a myriad of physiological functions. Next to their well-known role in sexual development and reproduction, SSH play active roles in practically every tissue in the human body, including the oral cavity. It has long been demonstrated that periodontal tissues express SSH receptors and therefore are responsive to the presence of SSH. Interestingly, SSH not only interact with the periodontal tissues but also with other tissues in the oral cavity such as dental enamel, pulp, cementum, oral mucosa, and salivary glands. Questions concerning the possible physiological functions of these receptors and their role in maintenance of oral health, remain unanswered. The purpose of this scoping review was to gather and summarize all the available evidence on the role of SSH in physiological processes in the oral cavity in humans. Two comprehensive literature searches were performed. References were screened and selected based on title, abstract and full text according to our inclusion criteria. Both searches yielded 18,992 results of which 73 were included. Results were divided into four categories: (1) Periodontium; (2) Dental structure; (3) Mucosa; and (4) Salivary glands. The interaction of these tissues with progestagens, androgens and estrogens are summarized. Sex steroid hormones are an overlooked yet fundamental factor in oral homeostasis. They play important roles in the development and function of the periodontium, dental structure, mucosa and salivary glands. Dentists and healthcare providers should consider these hormonal factors when assessing and treating oral health conditions.

Recently, an African ancestry-specific Parkinson disease (PD) risk signal was identified at the gene encoding glucocerebrosidase (GBA1). This variant ( rs3115534 -G) is carried by ~50% of West African PD cases and imparts a dose-dependent increase in risk for disease. The risk variant has varied frequencies across African ancestry groups but is almost absent in European and Asian ancestry populations. GBA1 is a gene of high clinical and therapeutic interest. Damaging biallelic protein-coding variants cause Gaucher disease and monoallelic variants confer risk for PD and dementia with Lewy bodies, likely by reducing the function of glucocerebrosidase. Interestingly, the African ancestry-specific GBA1 risk variant is a noncoding variant, suggesting a different mechanism of action. Using full-length RNA transcript sequencing, we identified partial intron 8 expression in risk variant carriers (G) but not in nonvariant carriers (T). Antibodies targeting the N terminus of glucocerebrosidase showed that this intron-retained isoform is likely not protein coding and subsequent proteomics did not identify a shorter protein isoform, suggesting that the disease mechanism is RNA based. Clustered regularly interspaced short palindromic repeats editing of the reported index variant ( rs3115534 ) revealed that this is the sequence alteration responsible for driving the production of these transcripts containing intron 8. Follow-up analysis of this variant showed that it is in a key intronic branchpoint sequence and, therefore, has important implications in splicing and disease. In addition, when measuring glucocerebrosidase activity, we identified a dose-dependent reduction in risk variant carriers. Overall, we report the functional effect of a GBA1 noncoding risk variant, which acts by interfering with the splicing of functional GBA1 transcripts, resulting in reduced protein levels and reduced glucocerebrosidase activity. This understanding reveals a potential therapeutic target in an underserved and underrepresented population.

TGF-β, essential for development and immunity, is expressed as a latent complex (L-TGF-β) non-covalently associated with its prodomain and presented on immune cell surfaces by covalent association with GARP. Binding to integrin αvβ8 activates L-TGF-β1/GARP. The dogma is that mature TGF-β must physically dissociate from L-TGF-β1 for signaling to occur. Our previous studies discovered that αvβ8-mediated TGF-β autocrine signaling can occur without TGF-β1 release from its latent form. Here, we show that mice engineered to express TGF-β1 that cannot release from L-TGF-β1 survive without early lethal tissue inflammation, unlike those with TGF-β1 deficiency. Combining cryogenic electron microscopy with cell-based assays, we reveal a dynamic allosteric mechanism of autocrine TGF-β1 signaling without release where αvβ8 binding redistributes the intrinsic flexibility of L-TGF-β1 to expose TGF-β1 to its receptors. Dynamic allostery explains the TGF-β3 latency/activation mechanism and why TGF-β3 functions distinctly from TGF-β1, suggesting that it broadly applies to other flexible cell surface receptor/ligand systems.

Life-threatening thrombotic events and neurological symptoms are prevalent in COVID-19 and are persistent in patients with long COVID experiencing post-acute sequelae of SARS-CoV-2 infection1-4. Despite the clinical evidence1,5-7, the underlying mechanisms of coagulopathy in COVID-19 and its consequences in inflammation and neuropathology remain poorly understood and treatment options are insufficient. Fibrinogen, the central structural component of blood clots, is abundantly deposited in the lungs and brains of patients with COVID-19, correlates with disease severity and is a predictive biomarker for post-COVID-19 cognitive deficits1,5,8-10. Here we show that fibrin binds to the SARS-CoV-2 spike protein, forming proinflammatory blood clots that drive systemic thromboinflammation and neuropathology in COVID-19. Fibrin, acting through its inflammatory domain, is required for oxidative stress and macrophage activation in the lungs, whereas it suppresses natural killer cells, after SARS-CoV-2 infection. Fibrin promotes neuroinflammation and neuronal loss after infection, as well as innate immune activation in the brain and lungs independently of active infection. A monoclonal antibody targeting the inflammatory fibrin domain provides protection from microglial activation and neuronal injury, as well as from thromboinflammation in the lung after infection. Thus, fibrin drives inflammation and neuropathology in SARS-CoV-2 infection, and fibrin-targeting immunotherapy may represent a therapeutic intervention for patients with acute COVID-19 and long COVID.

Copper plays critical roles as a metal active site cofactor and metalloallosteric signal for enzymes involved in cell proliferation and metabolism, making it an attractive target for cancer therapy. In this study, we investigated the efficacy of polydopamine nanoparticles (PDA NPs), classically applied for metal removal from water, as a therapeutic strategy for depleting intracellular labile copper pools in triple-negative breast cancer models through the metal-chelating groups present on the PDA surface. By using the activity-based sensing probe FCP-1, we could track the PDA-induced labile copper depletion while leaving total copper levels unchanged and link it to the selective MDA-MB-231 cell death. Further mechanistic investigations revealed that PDA NPs increased reactive oxygen species (ROS) levels, potentially through the inactivation of superoxide dismutase 1 (SOD1), a copper-dependent antioxidant enzyme. Additionally, PDA NPs were found to interact with the mitochondrial membrane, resulting in an increase in the mitochondrial membrane potential, which may contribute to enhanced ROS production. We employed an in vivo tumor model to validate the therapeutic efficacy of PDA NPs. Remarkably, in the absence of any additional treatment, the presence of PDA NPs alone led to a significant reduction in tumor volume by a factor of 1.66 after 22 days of tumor growth. Our findings highlight the potential of PDA NPs as a promising therapeutic approach for selectively targeting cancer by modulating copper levels and inducing oxidative stress, leading to tumor growth inhibition as shown in these triple-negative breast cancer models.

Oculomotor deficits are common in hereditary ataxia, but disproportionally neglected in clinical ataxia scales and as outcome measures for interventional trials. Quantitative assessment of oculomotor function has become increasingly available and thus applicable in multicenter trials and offers the opportunity to capture severity and progression of oculomotor impairment in a sensitive and reliable manner. In this consensus paper of the Ataxia Global Initiative Working Group On Digital Oculomotor Biomarkers, based on a systematic literature review, we propose harmonized methodology and measurement parameters for the quantitative assessment of oculomotor function in natural-history studies and clinical trials in hereditary ataxia. MEDLINE was searched for articles reporting on oculomotor/vestibular properties in ataxia patients and a study-tailored quality-assessment was performed. One-hundred-and-seventeen articles reporting on subjects with genetically confirmed (n=1134) or suspected hereditary ataxia (n=198), and degenerative ataxias with sporadic presentation (n=480) were included and subject to data extraction. Based on robust discrimination from controls, correlation with disease-severity, sensitivity to change, and feasibility in international multicenter settings as prerequisite for clinical trials, we prioritize a core-set of five eye-movement types: (i) pursuit eye movements, (ii) saccadic eye movements, (iii) fixation, (iv) eccentric gaze holding, and (v) rotational vestibulo-ocular reflex. We provide detailed guidelines for their acquisition, and recommendations on the quantitative parameters to extract. Limitations include low study quality, heterogeneity in patient populations, and lack of longitudinal studies. Standardization of quantitative oculomotor assessments will facilitate their implementation, interpretation, and validation in clinical trials, and ultimately advance our understanding of the evolution of oculomotor network dysfunction in hereditary ataxias.

BACKGROUND: Pegunigalsidase alfa is a PEGylated α-galactosidase A enzyme replacement therapy. BALANCE (NCT02795676) assessed non-inferiority of pegunigalsidase alfa versus agalsidase beta in adults with Fabry disease with an annualised estimated glomerular filtration rate (eGFR) slope more negative than -2 mL/min/1.73 m2/year who had received agalsidase beta for ≥1 year. METHODS: Patients were randomly assigned 2:1 to receive 1 mg/kg pegunigalsidase alfa or agalsidase beta every 2 weeks for 2 years. The primary efficacy analysis assessed non-inferiority based on median annualised eGFR slope differences between treatment arms. RESULTS: Seventy-seven patients received either pegunigalsidase alfa (n=52) or agalsidase beta (n=25). At baseline, mean (range) age was 44 (18-60) years, 47 (61%) patients were male, median eGFR was 74.5 mL/min/1.73 m2 and median (range) eGFR slope was -7.3 (-30.5, 6.3) mL/min/1.73 m2/year. At 2 years, the difference between median eGFR slopes was -0.36 mL/min/1.73 m2/year, meeting the prespecified non-inferiority margin. Minimal changes were observed in lyso-Gb3 concentrations in both treatment arms at 2 years. Proportions of patients experiencing treatment-related adverse events and mild or moderate infusion-related reactions were similar in both groups, yet exposure-adjusted rates were 3.6-fold and 7.8-fold higher, respectively, with agalsidase beta than pegunigalsidase alfa. At the end of the study, neutralising antibodies were detected in 7 out of 47 (15%) pegunigalsidase alfa-treated patients and 6 out of 23 (26%) agalsidase beta-treated patients. There were no deaths. CONCLUSIONS: Based on rate of eGFR decline over 2 years, pegunigalsidase alfa was non-inferior to agalsidase beta. Pegunigalsidase alfa had lower rates of treatment-emergent adverse events and mild or moderate infusion-related reactions. TRIAL REGISTRATION NUMBER: NCT02795676.

EXECUTIVE SUMMARY: Microbes are all pervasive in their distribution and influence on the functioning and well-being of humans, life in general and the planet. Microbially-based technologies contribute hugely to the supply of important goods and services we depend upon, such as the provision of food, medicines and clean water. They also offer mechanisms and strategies to mitigate and solve a wide range of problems and crises facing humanity at all levels, including those encapsulated in the sustainable development goals (SDGs) formulated by the United Nations. For example, microbial technologies can contribute in multiple ways to decarbonisation and hence confronting global warming, provide sanitation and clean water to the billions of people lacking them, improve soil fertility and hence food production and develop vaccines and other medicines to reduce and in some cases eliminate deadly infections. They are the foundation of biotechnology, an increasingly important and growing business sector and source of employment, and the centre of the bioeconomy, Green Deal, etc. But, because microbes are largely invisible, they are not familiar to most people, so opportunities they offer to effectively prevent and solve problems are often missed by decision-makers, with the negative consequences this entrains. To correct this lack of vital knowledge, the International Microbiology Literacy Initiative-the IMiLI-is recruiting from the global microbiology community and making freely available, teaching resources for a curriculum in societally relevant microbiology that can be used at all levels of learning. Its goal is the development of a society that is literate in relevant microbiology and, as a consequence, able to take full advantage of the potential of microbes and minimise the consequences of their negative activities. In addition to teaching about microbes, almost every lesson discusses the influence they have on sustainability and the SDGs and their ability to solve pressing problems of societal inequalities. The curriculum thus teaches about sustainability, societal needs and global citizenship. The lessons also reveal the impacts microbes and their activities have on our daily lives at the personal, family, community, national and global levels and their relevance for decisions at all levels. And, because effective, evidence-based decisions require not only relevant information but also critical and systems thinking, the resources also teach about these key generic aspects of deliberation. The IMiLI teaching resources are learner-centric, not academic microbiology-centric and deal with the microbiology of everyday issues. These span topics as diverse as owning and caring for a companion animal, the vast range of everyday foods that are produced via microbial processes, impressive geological formations created by microbes, childhood illnesses and how they are managed and how to reduce waste and pollution. They also leverage the exceptional excitement of exploration and discovery that typifies much progress in microbiology to capture the interest, inspire and motivate educators and learners alike. The IMiLI is establishing Regional Centres to translate the teaching resources into regional languages and adapt them to regional cultures, and to promote their use and assist educators employing them. Two of these are now operational. The Regional Centres constitute the interface between resource creators and educators-learners. As such, they will collect and analyse feedback from the end-users and transmit this to the resource creators so that teaching materials can be improved and refined, and new resources added in response to demand: educators and learners will thereby be directly involved in evolution of the teaching resources. The interactions between educators-learners and resource creators mediated by the Regional Centres will establish dynamic and synergistic relationships-a global societally relevant microbiology education ecosystem-in which creators also become learners, teaching resources are optimised and all players/stakeholders are empowered and their motivation increased. The IMiLI concept thus embraces the principle of teaching societally relevant microbiology embedded in the wider context of societal, biosphere and planetary needs, inequalities, the range of crises that confront us and the need for improved decisioning, which should ultimately lead to better citizenship and a humanity that is more sustainable and resilient. ABSTRACT: The biosphere of planet Earth is a microbial world: a vast reactor of countless microbially driven chemical transformations and energy transfers that push and pull many planetary geochemical processes, including the cycling of the elements of life, mitigate or amplify climate change (e.g., Nature Reviews Microbiology, 2019, 17, 569) and impact the well-being and activities of all organisms, including humans. Microbes are both our ancestors and creators of the planetary chemistry that allowed us to evolve (e.g., Lifes engines: How microbes made earth habitable, 2023). To understand how the biosphere functions, how humans can influence its development and live more sustainably with the other organisms sharing it, we need to understand the microbes. In a recent editorial (Environmental Microbiology, 2019, 21, 1513), we advocated for improved microbiology literacy in society. Our concept of microbiology literacy is not based on knowledge of the academic subject of microbiology, with its multitude of component topics, plus the growing number of additional topics from other disciplines that become vitally important elements of current microbiology. Rather it is focused on microbial activities that impact us-individuals/communities/nations/the human world-and the biosphere and that are key to reaching informed decisions on a multitude of issues that regularly confront us, ranging from personal issues to crises of global importance. In other words, it is knowledge and understanding essential for adulthood and the transition to it, knowledge and understanding that must be acquired early in life in school. The 2019 Editorial marked the launch of the International Microbiology Literacy Initiative, the IMiLI. HERE, WE PRESENT: our concept of how microbiology literacy may be achieved and the rationale underpinning it; the type of teaching resources being created to realise the concept and the framing of microbial activities treated in these resources in the context of sustainability, societal needs and responsibilities and decision-making; and the key role of Regional Centres that will translate the teaching resources into local languages, adapt them according to local cultural needs, interface with regional educators and develop and serve as hubs of microbiology literacy education networks. The topics featuring in teaching resources are learner-centric and have been selected for their inherent relevance, interest and ability to excite and engage. Importantly, the resources coherently integrate and emphasise the overarching issues of sustainability, stewardship and critical thinking and the pervasive interdependencies of processes. More broadly, the concept emphasises how the multifarious applications of microbial activities can be leveraged to promote human/animal, plant, environmental and planetary health, improve social equity, alleviate humanitarian deficits and causes of conflicts among peoples and increase understanding between peoples (Microbial Biotechnology, 2023, 16(6), 1091-1111). Importantly, although the primary target of the freely available (CC BY-NC 4.0) IMiLI teaching resources is schoolchildren and their educators, they and the teaching philosophy are intended for all ages, abilities and cultural spectra of learners worldwide: in university education, lifelong learning, curiosity-driven, web-based knowledge acquisition and public outreach. The IMiLI teaching resources aim to promote development of a global microbiology education ecosystem that democratises microbiology knowledge.

Monosomy 7 and del(7q) are among the most common and poorly understood genetic alterations in myelodysplastic neoplasms and acute myeloid leukemia. Chromosome band 7q22 is a minimally deleted segment in myeloid malignancies with a del(7q). However, the rarity of second hit mutations supports the idea that del(7q22) represents a contiguous gene syndrome. We generated mice harboring a 1.5 Mb germline deletion of chromosome band 5G2 syntenic to human 7q22 that removes Cux1 and 27 additional genes. Hematopoiesis is perturbed in 5G2+/del mice but they do not spontaneously develop hematologic disease. Whereas alkylator exposure modestly accelerated tumor development, the 5G2 deletion did not cooperate with KrasG12D, NrasG12D, or the MOL4070LTR retrovirus in leukemogenesis. 5G2+/del mice are a novel platform for interrogating the role of hemopoietic stem cell attrition/stress, cooperating mutations, genotoxins, and inflammation in myeloid malignancies characterized by monosomy 7/del(7q).

Characterizing bedside oculomotor deficits is a critical factor in defining the clinical presentation of hereditary ataxias. Quantitative assessments are increasingly available and have significant advantages, including comparability over time, reduced examiner dependency, and sensitivity to subtle changes. To delineate the potential of quantitative oculomotor assessments as digital-motor outcome measures for clinical trials in ataxia, we searched MEDLINE for articles reporting on quantitative eye movement recordings in genetically confirmed or suspected hereditary ataxias, asking which paradigms are most promising for capturing disease progression and treatment response. Eighty-nine manuscripts identified reported on 1541 patients, including spinocerebellar ataxias (SCA2, n = 421), SCA3 (n = 268), SCA6 (n = 117), other SCAs (n = 97), Friedreich ataxia (FRDA, n = 178), Niemann-Pick disease type C (NPC, n = 57), and ataxia-telangiectasia (n = 85) as largest cohorts. Whereas most studies reported discriminatory power of oculomotor assessments in diagnostics, few explored their value for monitoring genotype-specific disease progression (n = 2; SCA2) or treatment response (n = 8; SCA2, FRDA, NPC, ataxia-telangiectasia, episodic-ataxia 4). Oculomotor parameters correlated with disease severity measures including clinical scores (n = 18 studies (SARA: n = 9)), chronological measures (e.g., age, disease duration, time-to-symptom onset; n = 17), genetic stratification (n = 9), and imaging measures of atrophy (n = 5). Recurrent correlations across many ataxias (SCA2/3/17, FRDA, NPC) suggest saccadic eye movements as potentially generic quantitative oculomotor outcome. Recommendation of other paradigms was limited by the scarcity of cross-validating correlations, except saccadic intrusions (FRDA), pursuit eye movements (SCA17), and quantitative head-impulse testing (SCA3/6). This work aids in understanding the current knowledge of quantitative oculomotor parameters in hereditary ataxias, and identifies gaps for validation as potential trial outcome measures in specific ataxia genotypes.

Genome editing tools, particularly the Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) systems (e.g., CRISPR/Cas9), and their repurposing into epigenetic editing platforms, offer enormous potential as safe and customizable therapies for cancer. Specifically, various transcriptional abnormalities in human malignancies, such as silencing of tumor suppressors and ectopic re-expression of oncogenes, have been successfully targeted with virtually no off-target effects using CRISPR activation and repression systems. In these systems, the nuclease-deactivated Cas9 protein (dCas9) is fused to one or more domains inducing selective activation or repression of the targeted genes. Despite these advances, the efficient in vivo delivery of these molecules into the target cancer cells represents a critical barrier to accomplishing translation into a clinical therapy setting for cancer. Major obstacles include the large size of dCas9 fusion proteins, the necessity of multimodal delivery of protein and gRNAs, and the potential of these formulations to elicit detrimental immune responses.In this context, viral methods for delivering CRISPR face several limitations, such as the packaging capacity of the viral genome, the potential for integration of the nucleic acids into the host cells genome, and immunogenicity of viral proteins, posing serious safety concerns. The rapid development of mRNA vaccines in response to the COVID-19 pandemic has rekindled interest in mRNA-based approaches for CRISPR/dCas9 delivery. Simultaneously, due to their high loading capacity, scalability, customizable surface modification for cell targeting, and low immunogenicity, lipid nanoparticles (LNPs) have been widely explored as nonviral vectors. In this chapter, we first describe the design of optimized dCas9-effector mRNAs and gRNAs for epigenetic editing. We outline formulations of LNPs suitable for dCas9 mRNA delivery. Additionally, we provide a protocol for the co-encapsulation of the dCas9-effector mRNAs and gRNA into these LNPs, along with detailed methods for delivering these formulations to both cell lines (in vitro) and mouse models of breast cancer (in vivo).

Long-read sequencing technologies substantially overcome the limitations of short-reads but have not been considered as a feasible replacement for population-scale projects, being a combination of too expensive, not scalable enough or too error-prone. Here we develop an efficient and scalable wet lab and computational protocol, Napu, for Oxford Nanopore Technologies long-read sequencing that seeks to address those limitations. We applied our protocol to cell lines and brain tissue samples as part of a pilot project for the National Institutes of Health Center for Alzheimer's and Related Dementias. Using a single PromethION flow cell, we can detect single nucleotide polymorphisms with F1-score comparable to Illumina short-read sequencing. Small indel calling remains difficult within homopolymers and tandem repeats, but achieves good concordance to Illumina indel calls elsewhere. Further, we can discover structural variants with F1-score on par with state-of-the-art de novo assembly methods. Our protocol phases small and structural variants at megabase scales and produces highly accurate, haplotype-specific methylation calls.

Bacteria of the genus Brucella are facultative intracellular parasites that cause brucellosis, a severe animal and human disease. Recently, a group of taxonomists merged the brucellae with the primarily free-living, phylogenetically related Ochrobactrum spp. in the genus Brucella. This change, founded only on global genomic analysis and the fortuitous isolation of some opportunistic Ochrobactrum spp. from medically compromised patients, has been automatically included in culture collections and databases. We argue that clinical and environmental microbiologists should not accept this nomenclature, and we advise against its use because (i) it was presented without in-depth phylogenetic analyses and did not consider alternative taxonomic solutions; (ii) it was launched without the input of experts in brucellosis or Ochrobactrum; (iii) it applies a non-consensus genus concept that disregards taxonomically relevant differences in structure, physiology, population structure, core-pangenome assemblies, genome structure, genomic traits, clinical features, treatment, prevention, diagnosis, genus description rules, and, above all, pathogenicity; and (iv) placing these two bacterial groups in the same genus creates risks for veterinarians, medical doctors, clinical laboratories, health authorities, and legislators who deal with brucellosis, a disease that is particularly relevant in low- and middle-income countries. Based on all this information, we urge microbiologists, bacterial collections, genomic databases, journals, and public health boards to keep the Brucella and Ochrobactrum genera separate to avoid further bewilderment and harm.

By converting physical forces into electrical signals or triggering intracellular cascades, stretch-activated ion channels allow the cell to respond to osmotic and mechanical stress. Knowledge of the pathophysiological mechanisms underlying associations of stretch-activated ion channels with human disease is limited. Here, we describe 17 unrelated individuals with severe early-onset developmental and epileptic encephalopathy (DEE), intellectual disability, and severe motor and cortical visual impairment associated with progressive neurodegenerative brain changes carrying ten distinct heterozygous variants of TMEM63B, encoding for a highly conserved stretch-activated ion channel. The variants occurred de novo in 16/17 individuals for whom parental DNA was available and either missense, including the recurrent p.Val44Met in 7/17 individuals, or in-frame, all affecting conserved residues located in transmembrane regions of the protein. In 12 individuals, hematological abnormalities co-occurred, such as macrocytosis and hemolysis, requiring blood transfusions in some. We modeled six variants (p.Val44Met, p.Arg433His, p.Thr481Asn, p.Gly580Ser, p.Arg660Thr, and p.Phe697Leu), each affecting a distinct transmembrane domain of the channel, in transfected Neuro2a cells and demonstrated inward leak cation currents across the mutated channel even in isotonic conditions, while the response to hypo-osmotic challenge was impaired, as were the Ca2+ transients generated under hypo-osmotic stimulation. Ectopic expression of the p.Val44Met and p.Gly580Cys variants in Drosophila resulted in early death. TMEM63B-associated DEE represents a recognizable clinicopathological entity in which altered cation conductivity results in a severe neurological phenotype with progressive brain damage and early-onset epilepsy associated with hematological abnormalities in most individuals.

BackgroundDifficult facemask ventilation is perilous in children whose tracheas are difficult to intubate. We hypothesised that certain physical characteristics and anaesthetic factors are associated with difficult mask ventilation in paediatric patients who also had difficult tracheal intubation.MethodsWe queried a multicentre registry for children who experienced "difficult" or "impossible" facemask ventilation. Patient and case factors known before mask ventilation attempt were included for consideration in this regularised multivariable regression analysis. Incidence of complications, and frequency and efficacy of rescue placement of a supraglottic airway device were also tabulated. Changes in quality of mask ventilation after injection of a neuromuscular blocking agent were assessed.ResultsThe incidence of difficult mask ventilation was 9% (483 of 5453 patients). Infants and patients having increased weight, being less than 5th percentile in weight for age, or having Treacher-Collins syndrome, glossoptosis, or limited mouth opening were more likely to have difficult mask ventilation. Anaesthetic induction using facemask and opioids was associated with decreased risk of difficult mask ventilation. The incidence of complications was significantly higher in patients with "difficult" mask ventilation than in patients without. Rescue placement of a supraglottic airway improved ventilation in 71% (96 of 135) of cases. Administration of neuromuscular blocking agents was more frequently associated with improvement or no change in quality of ventilation than with worsening.ConclusionsCertain abnormalities on physical examination should increase suspicion of possible difficult facemask ventilation. Rescue use of a supraglottic airway device in children with difficult or impossible mask ventilation should be strongly considered.

PURPOSE: To communicate with linguistically diverse patients, medical students and physicians often use their non-English-language skills. However, there is no standard protocol to determine whether those skills are adequate before patient care. This causes many physicians, institutions, educators, and learners to forgo non-English-language proficiency assessment altogether. The purpose of this study is to report on the development, refinement, and interrater reliability of the Physician Oral Language Observation Matrix (POLOM), a rater-based tool assessing 6 language skill categories observed during clinical interactions: comprehension, fluency/fluidity, vocabulary, pronunciation, grammar, and communication. This study focused on the use of the POLOM in Spanish interactions. METHOD: The authors adapted an existing language observation tool for use in clinical settings, creating the preliminary POLOM. Next, they iteratively refined the tool from April to July 2021 using videorecorded medical student-standardized patient encounters from a U.S.-based medical Spanish program. In each refinement iteration, 4 bilingual raters (2 physicians and 2 linguists) independently rated 3 to 6 encounters and convened to discuss ratings with the goals of improving instrument instructions, descriptors, and subsequent rater agreement. Using the final POLOM, raters independently rated 50 videos in rotating interdisciplinary pairs. Generalizability theory was applied to estimate reliability via interrater agreement (dependability) coefficients (range 0-1) for each POLOM category and the total score. RESULTS: POLOM total score dependability equaled 0.927 (single rater) and 0.962 (averaged across 2 raters). The highest mean score was observed for the comprehension category (4.15; range 1-5) while the lowest was for communication (3.01; range 1-5). CONCLUSIONS: Raters achieved a high level of agreement on POLOM assessments of students medical oral Spanish proficiency. The POLOM is the first assessment tool that provides examinees and instructors with both a holistic and detailed review of clinician non-English oral language skills as contextualized for patient care.

People vary both in their embrace of their society's traditions, and in their perception of hazards as salient and necessitating a response. Over evolutionary time, traditions have offered avenues for addressing hazards, plausibly resulting in linkages between orientations toward tradition and orientations toward danger. Emerging research documents connections between traditionalism and threat responsivity, including pathogen-avoidance motivations. Additionally, because hazard-mitigating behaviors can conflict with competing priorities, associations between traditionalism and pathogen avoidance may hinge on contextually contingent tradeoffs. The COVID-19 pandemic provides a real-world test of the posited relationship between traditionalism and hazard avoidance. Across 27 societies (N = 7844), we find that, in a majority of countries, individuals' endorsement of tradition positively correlates with their adherence to costly COVID-19-avoidance behaviors; accounting for some of the conflicts that arise between public health precautions and other objectives further strengthens this evidence that traditionalism is associated with greater attention to hazards.

BACKGROUND: The benefits and risks of augmenting or switching antidepressants in older adults with treatment-resistant depression have not been extensively studied. METHODS: We conducted a two-step, open-label trial involving adults 60 years of age or older with treatment-resistant depression. In step 1, patients were randomly assigned in a 1:1:1 ratio to augmentation of existing antidepressant medication with aripiprazole, augmentation with bupropion, or a switch from existing antidepressant medication to bupropion. Patients who did not benefit from or were ineligible for step 1 were randomly assigned in step 2 in a 1:1 ratio to augmentation with lithium or a switch to nortriptyline. Each step lasted approximately 10 weeks. The primary outcome was the change from baseline in psychological well-being, assessed with the National Institutes of Health Toolbox Positive Affect and General Life Satisfaction subscales (population mean, 50; higher scores indicate greater well-being). A secondary outcome was remission of depression. RESULTS: In step 1, a total of 619 patients were enrolled; 211 were assigned to aripiprazole augmentation, 206 to bupropion augmentation, and 202 to a switch to bupropion. Well-being scores improved by 4.83 points, 4.33 points, and 2.04 points, respectively. The difference between the aripiprazole-augmentation group and the switch-to-bupropion group was 2.79 points (95% CI, 0.56 to 5.02; P = 0.014, with a prespecified threshold P value of 0.017); the between-group differences were not significant for aripiprazole augmentation versus bupropion augmentation or for bupropion augmentation versus a switch to bupropion. Remission occurred in 28.9% of patients in the aripiprazole-augmentation group, 28.2% in the bupropion-augmentation group, and 19.3% in the switch-to-bupropion group. The rate of falls was highest with bupropion augmentation. In step 2, a total of 248 patients were enrolled; 127 were assigned to lithium augmentation and 121 to a switch to nortriptyline. Well-being scores improved by 3.17 points and 2.18 points, respectively (difference, 0.99; 95% CI, -1.92 to 3.91). Remission occurred in 18.9% of patients in the lithium-augmentation group and 21.5% in the switch-to-nortriptyline group; rates of falling were similar in the two groups. CONCLUSIONS: In older adults with treatment-resistant depression, augmentation of existing antidepressants with aripiprazole improved well-being significantly more over 10 weeks than a switch to bupropion and was associated with a numerically higher incidence of remission. Among patients in whom augmentation or a switch to bupropion failed, changes in well-being and the occurrence of remission with lithium augmentation or a switch to nortriptyline were similar. (Funded by the Patient-Centered Outcomes Research Institute; OPTIMUM ClinicalTrials.gov number, NCT02960763.).

SARS-CoV-2 (COVID-19) has caused over 80 million infections 973,000 deaths in the United States, and mutations are linked to increased transmissibility. This study aimed to determine the effect of SARS-CoV-2 variants on respiratory features, mortality, and to determine the effect of vaccination status. A retrospective review of medical records (n = 55,406 unique patients) using the University of California Health COvid Research Data Set (UC CORDS) was performed to identify respiratory features, vaccination status, and mortality from 01/01/2020 to 04/26/2022. Variants were identified using the CDC data tracker. Increased odds of death were observed amongst unvaccinated individuals and fully vaccinated, partially vaccinated, or individuals who received any vaccination during multiple waves of the pandemic. Vaccination status was associated with survival and a decreased frequency of many respiratory features. More recent SARS-CoV-2 variants show a reduction in lower respiratory tract features with an increase in upper respiratory tract features. Being fully vaccinated results in fewer respiratory features and higher odds of survival, supporting vaccination in preventing morbidity and mortality from COVID-19.

BACKGROUND: End-stage renal disease is associated with a high risk of cardiovascular events. It is unknown, however, whether mild-to-moderate kidney dysfunction is causally related to coronary heart disease (CHD) and stroke. METHODS: Observational analyses were conducted using individual-level data from 4 population data sources (Emerging Risk Factors Collaboration, EPIC-CVD [European Prospective Investigation into Cancer and Nutrition-Cardiovascular Disease Study], Million Veteran Program, and UK Biobank), comprising 648 135 participants with no history of cardiovascular disease or diabetes at baseline, yielding 42 858 and 15 693 incident CHD and stroke events, respectively, during 6.8 million person-years of follow-up. Using a genetic risk score of 218 variants for estimated glomerular filtration rate (eGFR), we conducted Mendelian randomization analyses involving 413 718 participants (25 917 CHD and 8622 strokes) in EPIC-CVD, Million Veteran Program, and UK Biobank. RESULTS: There were U-shaped observational associations of creatinine-based eGFR with CHD and stroke, with higher risk in participants with eGFR values 105 mL·min-1·1.73 m-2, compared with those with eGFR between 60 and 105 mL·min-1·1.73 m-2. Mendelian randomization analyses for CHD showed an association among participants with eGFR 105 mL·min-1·1.73 m-2. Results were not materially different after adjustment for factors associated with the eGFR genetic risk score, such as lipoprotein(a), triglycerides, hemoglobin A1c, and blood pressure. Mendelian randomization results for stroke were nonsignificant but broadly similar to those for CHD. CONCLUSIONS: In people without manifest cardiovascular disease or diabetes, mild-to-moderate kidney dysfunction is causally related to risk of CHD, highlighting the potential value of preventive approaches that preserve and modulate kidney function.

PurposeAutosomal dominant cone rod dystrophy 7 (CORD7) was initially linked to the gene RIMS1 and reported in a 4-generation British family in 1998. The purpose of this study was to investigate the legitimacy of this association, and to correctly characterize the genetic cause of this condition.MethodsThe allele frequency of RIMS1 c.2459G>A, p.Arg820His, was investigated in the Genomes Aggregation Dataset (gnomAD) datasets and whole genome sequencing (WGS) was performed for 4 members of the CORD7 family with filtering of rare pathogenic variants in a virtual gene panel comprising all genes known to be associated with inherited retinal dystrophy (IRD). Cytogenetic analysis was performed to rule out interchromosomal translocation.ResultsRIMS1 p.Arg820His has a maximal carrier frequency of >1:5000 in Europeans. A previously well-characterized PROM1 variant: c.1118C>T, p.Arg373Cys, was detected in 9 affected members of the CORD7 family who underwent WGS or direct sequencing. One affected family member is now known to have macular dystrophy in the absence of RIMS1 p.Arg820His. Clinical analysis of affected family members and 27 individuals with retinopathy associated with the same - PROM1 - variant showed consistent phenotypes.ConclusionsThe case for pathogenicity of RIMS1 p.Arg820His is not strong based on its presence on 10 alleles in the gnomAD dataset and absence from additional CORD affected individuals. The finding of a known pathogenic variant in PROM1 correlates well with the phenotypic characteristics of the affected individuals, and is likely to account for the condition. Clear evidence of association between RIMS1 and a retinal dystrophy is yet to be described.

Objetivo: comparar los métodos de referencia de microdilución en caldo de la CLSI M27-A2 y EUCAST, identificando la utilidad y las principales diferencias de cada uno de ellos para los agentes antifúngicos anfotericina B (1), fluconazol (FCZ) e itraconazol (ITZ), contra aislamientos clínicos de Candidaspp. de pacientes con cáncer. Materiales y métodos: se estudiaron 136 aislamientos de C. albicans, 36 de C. tropicalis y 17 de Candidaspp. Se utilizó el índice Kappa ponderado para medir el grado de acuerdo entre los dos métodos. Resultados: se estableció que el grado de concordancia entre los dos métodos para el total de los aislamientos fue alto con AB (κ: 1) y FCZ (κ: 0.74) y bajo al utilizar ITZ (κ: 0.49). La concordancia fue variable y especie-específica: para ITZ y FCZ en C. albicans fue de 0,45 y 0,64; en C. tropicalis, de 0,48 y 0,91; y en Candidaspp. de 0,73 y 0,87, respectivamente. Discusión: este estudio sugiere que las pruebas de sensibilidad antifúngica para los dos métodos son equivalentes en lo esencial. Deben considerarse las diferencias y discrepancias asociadas a la especie implicada, el tipo de antifúngico utilizado y los tiempos de incubación, que puede producir variaciones al interpretar los resultados obtenidos de acuerdo con la metodología empleada.Objective: compare the broth microdilution testing reference standards CLSI M27-A2 and EUCAST, identifying the usefulness of each one of them and their main differences, against the antifungal agents amphotericin B (1), fluconazole (FCZ), and itraconazole (ITZ) using clinical isolates of Candidaspp. in cancer patients. Methods: isolates of C. albicans (n=136), C. tropicalis (n=36), and Candidaspp. (n=17) were tested by the two methods. The Kappa index was used to establish the degree of agreement between the methods. Results: the degree of agreement between the two methods was high for AB (κ: 1) and FCZ (κ: 0.74) and was low for ITZ (κ: 0.49). Agreement was variable and specific for the various species: for ITZ and FCZ in C. albicans, it was 0.45 and 0.64, respectively. In C. tropicalis, it was of 0.48 and 0.91, and in Candidaspp., it was 0.73 and 0.87 respectively. Discussion: this study suggests that antifungal susceptibility testing using both methods is equivalent. Attention should be focused on differences and discrepancies associated with the species tested, the type of antifungal agent, and the incubation times, which can cause variations at the moment of interpreting the results obtained.

Technological advances in both genome sequencing and prenatal imaging are increasing our ability to accurately recognize and diagnose Mendelian conditions prenatally. Phenotype-driven early genetic diagnosis of fetal genetic disease can help to strategize treatment options and clinical preventive measures during the perinatal period, to plan in utero therapies, and to inform parental decision-making. Fetal phenotypes of genetic diseases are often unique and at present are not well understood; more comprehensive knowledge about prenatal phenotypes and computational resources have an enormous potential to improve diagnostics and translational research. The Human Phenotype Ontology (HPO) has been widely used to support diagnostics and translational research in human genetics. To better support prenatal usage, the HPO consortium conducted a series of workshops with a group of domain experts in a variety of medical specialties, diagnostic techniques, as well as diseases and phenotypes related to prenatal medicine, including perinatal pathology, musculoskeletal anomalies, neurology, medical genetics, hydrops fetalis, craniofacial malformations, cardiology, neonatal-perinatal medicine, fetal medicine, placental pathology, prenatal imaging, and bioinformatics. We expanded the representation of prenatal phenotypes in HPO by adding 95 new phenotype terms under the Abnormality of prenatal development or birth (HP:0001197) grouping term, and revised definitions, synonyms, and disease annotations for most of the 152 terms that existed before the beginning of this effort. The expansion of prenatal phenotypes in HPO will support phenotype-driven prenatal exome and genome sequencing for precision genetic diagnostics of rare diseases to support prenatal care.

RASopathies are a group of genetic disorders that are caused by genes that affect the canonical Ras/mitogen-activated protein kinase (MAPK) signaling pathway. Despite tremendous progress in understanding the molecular consequences of these genetic anomalies, little movement has been made in translating these findings to the clinic. This year, the seventh International RASopathies Symposium focused on expanding the research knowledge that we have gained over the years to enhance new discoveries in the field, ones that we hope can lead to effective therapeutic treatments. Indeed, for the first time, research efforts are finally being translated to the clinic, with compassionate use of Ras/MAPK pathway inhibitors for the treatment of RASopathies. This biannual meeting, organized by the RASopathies Network, brought together basic scientists, clinicians, clinician scientists, patients, advocates, and their families, as well as representatives from pharmaceutical companies and the National Institutes of Health. A history of RASopathy gene discovery, identification of new disease genes, and the latest research, both at the bench and in the clinic, were discussed.

PurposeTo provide precise age-specific risk estimates of cancers other than female breast and ovarian cancers associated with pathogenic variants (PVs) in BRCA1 and BRCA2 for effective cancer risk management.MethodsWe used data from 3,184 BRCA1 and 2,157 BRCA2 families in the Consortium of Investigators of Modifiers of BRCA1/2 to estimate age-specific relative (RR) and absolute risks for 22 first primary cancer types adjusting for family ascertainment.ResultsBRCA1 PVs were associated with risks of male breast (RR = 4.30; 95% CI, 1.09 to 16.96), pancreatic (RR = 2.36; 95% CI, 1.51 to 3.68), and stomach (RR = 2.17; 95% CI, 1.25 to 3.77) cancers. Associations with colorectal and gallbladder cancers were also suggested. BRCA2 PVs were associated with risks of male breast (RR = 44.0; 95% CI, 21.3 to 90.9), stomach (RR = 3.69; 95% CI, 2.40 to 5.67), pancreatic (RR = 3.34; 95% CI, 2.21 to 5.06), and prostate (RR = 2.22; 95% CI, 1.63 to 3.03) cancers. The stomach cancer RR was higher for females than males (6.89 v 2.76; P = .04). The absolute risks to age 80 years ranged from 0.4% for male breast cancer to approximately 2.5% for pancreatic cancer for BRCA1 carriers and from approximately 2.5% for pancreatic cancer to 27% for prostate cancer for BRCA2 carriers.ConclusionIn addition to female breast and ovarian cancers, BRCA1 and BRCA2 PVs are associated with increased risks of male breast, pancreatic, stomach, and prostate (only BRCA2 PVs) cancers, but not with the risks of other previously suggested cancers. The estimated age-specific risks will refine cancer risk management in men and women with BRCA1/2 PVs.

ImportanceRadiotherapy combined with androgen deprivation therapy (ADT) is a standard of care for high-risk prostate cancer. However, the interplay between radiotherapy dose and the required minimum duration of ADT is uncertain.ObjectiveTo determine the specific ADT duration threshold that provides a distant metastasis-free survival (DMFS) benefit in patients with high-risk prostate cancer receiving external beam radiotherapy (EBRT) or EBRT with a brachytherapy boost (EBRT+BT).Design, settings, and participantsThis was a cohort study of 3 cohorts assembled from a multicenter retrospective study (2000-2013); a post hoc analysis of the Randomized Androgen Deprivation and Radiotherapy 03/04 (RADAR; 2003-2007) randomized clinical trial (RCT); and a cross-trial comparison of the RADAR vs the Deprivación Androgénica y Radio Terapía (Androgen Deprivation and Radiation Therapy; DART) 01/05 RCT (2005-2010). In all, the study analyzed 1827 patients treated with EBRT and 1108 patients treated with EBRT+BT from the retrospective cohort; 181 treated with EBRT and 203 with EBRT+BT from RADAR; and 91 patients treated with EBRT from DART. The study was conducted from October 15, 2020, to July 1, 2021, and the data analyses, from January 5 to June 15, 2021.ExposuresHigh-dose EBRT or EBRT+BT for an ADT duration determined by patient-physician choice (retrospective) or by randomization (RCTs).Main outcomes and measuresThe primary outcome was DMFS; secondary outcome was overall survival (OS). Natural cubic spline analysis identified minimum thresholds (months).ResultsThis cohort study of 3 studies totaling 3410 men (mean age [SD], 68 [62-74] years; race and ethnicity not collected) with high-risk prostate cancer found a significant interaction between the treatment type (EBRT vs EBRT+BT) and ADT duration (binned to

BackgroundThe diagnostic rate of Mendelian disorders in sequencing studies continues to increase, along with the pace of novel disease gene discovery. However, variant interpretation in novel genes not currently associated with disease is particularly challenging and strategies combining gene functional evidence with approaches that evaluate the phenotypic similarities between patients and model organisms have proven successful. A full spectrum of intolerance to loss-of-function variation has been previously described, providing evidence that gene essentiality should not be considered as a simple and fixed binary property.MethodsHere we further dissected this spectrum by assessing the embryonic stage at which homozygous loss-of-function results in lethality in mice from the International Mouse Phenotyping Consortium, classifying the set of lethal genes into one of three windows of lethality: early, mid, or late gestation lethal. We studied the correlation between these windows of lethality and various gene features including expression across development, paralogy and constraint metrics together with human disease phenotypes. We explored a gene similarity approach for novel gene discovery and investigated unsolved cases from the 100,000 Genomes Project.ResultsWe found that genes in the early gestation lethal category have distinct characteristics and are enriched for genes linked with recessive forms of inherited metabolic disease. We identified several genes sharing multiple features with known biallelic forms of inborn errors of the metabolism and found signs of enrichment of biallelic predicted pathogenic variants among early gestation lethal genes in patients recruited under this disease category. We highlight two novel gene candidates with phenotypic overlap between the patients and the mouse knockouts.ConclusionsInformation on the developmental period at which embryonic lethality occurs in the knockout mouse may be used for novel disease gene discovery that helps to prioritise variants in unsolved rare disease cases.

Alterations in the levels of serum sphingolipids and phospholipids have been reported in Gaucher disease and in Parkinson's disease, suggesting a potential role of these lipids as biomarkers. This project's objective is to detect novel associations and novel candidate biomarkers in the largest Spanish Gaucher and Parkinson diseases of the Iberian Peninsula. For that, 278 participants were included: 100 sporadic Parkinson's patients, 70 Gaucher patients, 15 GBA1-mutation-carrier Parkinson's patients and 93 controls. A serum lipidomics array including 10 phospholipid groups, 368 species, was performed using high-performance liquid chromatography-mass spectrometry. Lipid levels were compared between groups via multiple-regression analyses controlling for clinical and demographic parameters. Additionally, lipid levels were compared within the Gaucher and Parkinson's groups controlling for medication and/or disease severity. Results were controlled for robustness by filtering of non-detectable lipid values. There was an increase in the levels of phosphatidylcholine, with a simultaneous decrease in lyso-phosphatidylcholine, in the Gaucher, Parkinson's and GBA1-mutation-carrier Parkinson's patients vs. controls. Phosphatidylethanolamine, lyso- and plasmalogen-phosphatidylethanolamine were also increased in Gaucher and Parkinson's. Gaucher patients also showed an increase in lyso-phosphatidylserine and phosphatidylglycerol. While in the Gaucher and Parkinson's groups, velaglucerase alpha and dopamine agonists, respectively, showed positive associations with the lipid changes, miglustat treatment in Gaucher patients normalized the altered phosphatidylcholine/lyso-phosphatidylcholine ratio. In conclusion, Gaucher and Parkinson's patients showed changes in various serum phospholipid levels when compared with healthy controls, further supporting the role of such lipids in disease development and, possibly, as putative biomarkers. This hypothesis was reinforced by the normalizing effect of miglustat, and by controlling for data robustness, even though the limited number of participants, especially in the sub-distribution by treatment groups in GD requires validation in a larger number of patients.

Metabolic syndrome (MetS) is a complex condition encompassing a constellation of cardiometabolic abnormalities. Oxylipins are a superfamily of lipid mediators regulating many cardiometabolic functions. Plasma oxylipin signature could provide a new clinical tool to enhance the phenotyping of MetS pathophysiology. A high-throughput validated mass spectrometry method, allowing for the quantitative profiling of over 130 oxylipins, was applied to identify and validate the oxylipin signature of MetS in two independent nested case/control studies involving 476 participants. We identified an oxylipin signature of MetS (coined OxyScore), including 23 oxylipins and having high performances in classification and replicability (cross-validated AUCROC of 89%, 95% CI: 85-93% and 78%, 95% CI: 72-85% in the Discovery and Replication studies, respectively). Correlation analysis and comparison with a classification model incorporating the MetS criteria showed that the oxylipin signature brings consistent and complementary information to the clinical criteria. Being linked with the regulation of various biological processes, the candidate oxylipins provide an integrative phenotyping of MetS regarding the activation and/or negative feedback regulation of crucial molecular pathways. This may help identify patients at higher risk of cardiometabolic diseases. The oxylipin signature of patients with metabolic syndrome enhances MetS phenotyping and may ultimately help to better stratify the risk of cardiometabolic diseases.

ImportanceProstate-specific membrane antigen (PSMA) positron emission tomography/computed tomography (PET/CT) can detect low-volume, nonlocalized (ie, regional or metastatic) prostate cancer that was occult on conventional imaging. However, the long-term clinical implications of PSMA PET/CT upstaging remain unclear.ObjectivesTo evaluate the prognostic significance of a nomogram that models an individual's risk of nonlocalized upstaging on PSMA PET/CT and to compare its performance with existing risk-stratification tools.Design, setting, and participantsThis cohort study included patients diagnosed with high-risk or very high-risk prostate cancer (ie, prostate-specific antigen [PSA] level >20 ng/mL, Gleason score 8-10, and/or clinical stage T3-T4, without evidence of nodal or metastatic disease by conventional workup) from April 1995 to August 2018. This multinational study was conducted at 15 centers. Data were analyzed from December 2020 to March 2021.ExposuresCurative-intent radical prostatectomy (RP), external beam radiotherapy (EBRT), or EBRT plus brachytherapy (BT), with or without androgen deprivation therapy.Main outcomes and measuresPSMA upstage probability was calculated from a nomogram using the biopsy Gleason score, percentage positive systematic biopsy cores, clinical T category, and PSA level. Biochemical recurrence (BCR), distant metastasis (DM), prostate cancer-specific mortality (PCSM), and overall survival (OS) were analyzed using Fine-Gray and Cox regressions. Model performance was quantified with the concordance (C) index.ResultsOf 5275 patients, the median (IQR) age was 66 (60-72) years; 2883 (55%) were treated with RP, 1669 (32%) with EBRT, and 723 (14%) with EBRT plus BT; median (IQR) PSA level was 10.5 (5.9-23.2) ng/mL; 3987 (76%) had Gleason grade 8 to 10 disease; and 750 (14%) had stage T3 to T4 disease. Median (IQR) follow-up was 5.1 (3.1-7.9) years; 1221 (23%) were followed up for at least 8 years. Overall, 1895 (36%) had BCR, 851 (16%) developed DM, and 242 (5%) died of prostate cancer. PSMA upstage probability was significantly prognostic of all clinical end points, with 8-year C indices of 0.63 (95% CI, 0.61-0.65) for BCR, 0.69 (95% CI, 0.66-0.71) for DM, 0.71 (95% CI, 0.67-0.75) for PCSM, and 0.60 (95% CI, 0.57-0.62) for PCSM (P

Immunocompromised individuals, particularly autologous hematopoietic stem cell transplant (auHSCT) recipients, are at high risk for herpes zoster (HZ). We provide an in-depth description of humoral and cell-mediated immune (CMI) responses by age (protocol-defined) or underlying disease (post-hoc) as well as efficacy by underlying disease (post-hoc) of the adjuvanted recombinant zoster vaccine (RZV) in a randomized observer-blind phase III trial (ZOE-HSCT, NCT01610414). 1846 adult auHSCT recipients were randomized to receive a first dose of either RZV or placebo 50-70 days post-auHSCT, followed by the second dose at 1-2 months (M) later. In cohorts of 114-1721 participants, at 1 M post-second vaccine dose: Anti-gE antibody geometric mean concentrations (GMCs) and median gE-specific CD4[2+] T-cell frequencies (CD4 T cells expressing ≥2 of four assessed activation markers) were similar between 18-49 and ≥50-year-olds. Despite lower anti-gE antibody GMCs in non-Hodgkin B-cell lymphoma (NHBCL) patients, CD4[2+] T-cell frequencies were similar between NHBCL and other underlying diseases. The proportion of polyfunctional CD4 T cells increased over time, accounting for 79.6% of gE-specific CD4 T cells at 24 M post-dose two. Vaccine efficacy against HZ ranged between 42.5% and 82.5% across underlying diseases and was statistically significant in NHBCL and multiple myeloma patients. In conclusion, two RZV doses administered early post-auHSCT induced robust, persistent, and polyfunctional gE-specific immune responses. Efficacy against HZ was also high in NHBCL patients despite the lower humoral response.

BackgroundWith the advancement of imaging technology, focal therapy (FT) has been gaining acceptance for the treatment of select patients with localized prostate cancer (CaP). We aim to provide details of a formal physician consensus on the utilization of FT for patients with CaP who are discontinuing active surveillance (AS).MethodsA 3-stage Delphi consensus on CaP and FT was conducted. Consensus was defined as agreement by ≥80% of physicians. An in-person meeting was attended by 17 panelists to formulate the consensus statement.ResultsFifty-six respondents participated in this interdisciplinary consensus study (82% urologist, 16% radiologist, 2% radiation oncology). The participants confirmed that there is a role for FT in men discontinuing AS (48% strongly agree, 39% agree). The benefit of FT over radical therapy for men coming off AS is: less invasive (91%), has a greater likelihood to preserve erectile function (91%), has a greater likelihood to preserve urinary continence (91%), has fewer side effects (86%), and has early recovery post-treatment (80%). Patients will need to undergo mpMRI of the prostate and/or a saturation biopsy to determine if they are potential candidates for FT. Our limitations include respondent's biases and that the participants of this consensus may not represent the larger medical community.ConclusionsFT can be offered to men coming off AS between the age of 60 to 80 with grade group 2 localized cancer. This consensus from a multidisciplinary, multi-institutional, international expert panel provides a contemporary insight utilizing FT for CaP in select patients who are discontinuing AS.

BackgroundPrehospital electrocardiogram(s) (ECG) can improve early detection of acute coronary syndrome (ST-segment elevation myocardial infarction [STEMI], non-STEMI, and unstable angina) and inform prehospital activation of cardiac catheterization lab; thus, reducing total ischemic time and improving patient outcomes. Less is known, however, about the association of prehospital ECG ischemic findings and long term adverse clinical events. With this in mind, this study was designed to examine the: 1) frequency of prehospital ECGs for acute myocardial ischemia (ST-elevation, ST-depression, and/or T-wave inversion); and, 2) whether any of these specific ECG features are associated with adverse clinical events within 30 day of initial presentation to the emergency department (ED).MethodsWe included consecutive patients ≥ 21 years during a five-year period (2013-2017), who were transported by ambulance to the ED with non-traumatic chest pain and/or anginal equivalent(s) and had a prehospital 12‑lead ECG. Two cardiologists (LG, EC), blinded to clinical data, interpreted the 12‑lead ECGs applying current guideline based ischemia criteria. Adverse clinical events, return to ED, and rehospitalization evaluated at 30-days.ResultsWe identified 3646 patients (mean age, 59.7 years ±15.7; 45% female) with ECGs, of which N = 3587 had data on the three ischemic markers of interest. Of these, 1762 (49.1%) had ECG evidence of ischemia. In adjusted logistic regression models, those with T-wave inversion had a higher odds (OR = 1.59) of new onset heart failure, while ST-elevation was associated with lower odds (OR = 0.69). Patients with ST-depression had higher odds of new onset heart failure and death within 30 days (OR = 1.29, 1.49 respectively), but this association attenuated after controlling for other ECG features.ConclusionsST-depression and/or T-wave inversion are independent predictors of new onset heart failure, within 30 days of initial ED presentation. Our study in a large cohort of patients, suggests that using ECG ST-elevation alone may not capture patients with ischemia who may benefit from aggressive anti-ischemic therapies to reduce myocardial damage with resultant heart failure.

BackgroundExperimental studies and small clinical trials have suggested that treatment with intranasal oxytocin may reduce social impairment in persons with autism spectrum disorder. Oxytocin has been administered in clinical practice to many children with autism spectrum disorder.MethodsWe conducted a 24-week, placebo-controlled phase 2 trial of intranasal oxytocin therapy in children and adolescents 3 to 17 years of age with autism spectrum disorder. Participants were randomly assigned in a 1:1 ratio, with stratification according to age and verbal fluency, to receive oxytocin or placebo, administered intranasally, with a total target dose of 48 international units daily. The primary outcome was the least-squares mean change from baseline on the Aberrant Behavior Checklist modified Social Withdrawal subscale (ABC-mSW), which includes 13 items (scores range from 0 to 39, with higher scores indicating less social interaction). Secondary outcomes included two additional measures of social function and an abbreviated measure of IQ.ResultsOf the 355 children and adolescents who underwent screening, 290 were enrolled. A total of 146 participants were assigned to the oxytocin group and 144 to the placebo group; 139 and 138 participants, respectively, completed both the baseline and at least one postbaseline ABC-mSW assessments and were included in the modified intention-to-treat analyses. The least-squares mean change from baseline in the ABC-mSW score (primary outcome) was -3.7 in the oxytocin group and -3.5 in the placebo group (least-squares mean difference, -0.2; 95% confidence interval, -1.5 to 1.0; P = 0.61). Secondary outcomes generally did not differ between the trial groups. The incidence and severity of adverse events were similar in the two groups.ConclusionsThis placebo-controlled trial of intranasal oxytocin therapy in children and adolescents with autism spectrum disorder showed no significant between-group differences in the least-squares mean change from baseline on measures of social or cognitive functioning over a period of 24 weeks. (Funded by the National Institute of Child Health and Human Development; SOARS-B ClinicalTrials.gov number, NCT01944046.).

ObjectiveTo evaluate the barriers and facilitators to implementing Alberta Family Integrated Care (AB-FICare [2019 Benzies]), a model of care for integrating parents into level II neonatal intensive care units (NICUs) care teams, from the perspective of healthcare providers (HCP) and hospital administrators.DesignQualitative process evaluation substudy.SettingTen level II NICUs in six cities across Alberta, Canada.ParticipantsHCP and hospital administrators (n=32) who were involved in the cluster-randomised controlled trial of AB-FICare in level II NICUs.MethodsPost-implementation semi-structured interviews were conducted via phone or in-person. The Consolidated Framework for Implementation Research was used to develop interview guides, code transcripts and analyse data.ResultsKey facilitators to implementation of AB-FICare included (1) a receptive implementation climate, (2) compatibility of the intervention with individual and organisational practices, (3) available resources and access to knowledge and information for HCP and hospital administrators, (4) engagement of key stakeholders across the organisation, (5) engagement of and outcomes for intervention participants, and (6) reflecting and evaluating on implementation progress and patient and family outcomes. Barriers were (1) design quality and packaging of the intervention, (2) relative priority of AB-FICare in relation to other initiatives, and (3) learning climate within the organisation. Mixed influences on implementation depending on contextual factors were coded to eight constructs: intervention source, cost, peer pressure, external policy and incentives, staff needs and resources, structural characteristics, organisational incentives and rewards, and knowledge, beliefs and attitudes.ConclusionsThe characteristics of an organisation and the implementation process had largely positive influences, which can be leveraged for implementation of AB-FICare in the NICU. We recommend site-specific consultations to mitigate barriers and assess how swing factors might impact implementation given the local context, with the goal that strategies can be put in place to manage their influence on implementation.Trial registration numberNCT02879799.

Glucose-galactose malabsorption (GGM) is due to mutations in the gene coding for the intestinal sodium glucose cotransporter SGLT1 (SLC5A1). Here we identify the rare variant Gln457Arg (Q457R) in a large pedigree of patients in the Västerbotten County in Northern Sweden with the clinical phenotype of GGM. The functional effect of the Q457R mutation was determined in protein expressed in Xenopus laevis oocytes using biophysical and biochemical methods. The mutant failed to transport the specific SGLT1 sugar analog α-methyl-D-glucopyranoside (αMDG). Q457R SGLT1 was synthesized in amounts comparable to the wild-type (WT) transporter. SGLT1 charge measurements and freeze-fracture electron microscopy demonstrated that the mutant protein was inserted into the plasma membrane. Electrophysiological experiments, both steady-state and presteady-state, demonstrated that the mutant bound sugar with an affinity lower than the WT transporter. Together with our previous studies on Q457C and Q457E mutants, we established that the positive charge on Q457R prevented the translocation of sugar from the outward-facing to inward-facing conformation. This is contrary to other GGM cases where missense mutations caused defects in trafficking SGLT1 to the plasma membrane. Thirteen GGM patients are now added to the pedigree traced back to the late 17th century. The frequency of the Q457R variant in Västerbotten County genomes, 0.0067, is higher than in the general Swedish population, 0.0015, and higher than the general European population, 0.000067. This explains the high number of GGM cases in this region of Sweden.

The natural history of radiorecurrent high-risk prostate cancer (HRPCa) is not well-described. To better understand its clinical course, we evaluated rates of distant metastases (DM) and prostate cancer-specific mortality (PCSM) in a cohort of 978 men with radiorecurrent HRPCa who previously received either external beam radiation therapy (EBRT, n = 654, 67%) or EBRT + brachytherapy (EBRT + BT, n = 324, 33%) across 15 institutions from 1997 to 2015. In men who did not die, median follow-up after treatment was 8.9 yr and median follow-up after biochemical recurrence (BCR) was 3.7 yr. Local and systemic therapy salvage, respectively, were delivered to 21 and 390 men after EBRT, and eight and 103 men after EBRT + BT. Overall, 435 men developed DM, and 248 were detected within 1 yr of BCR. Measured from time of recurrence, 5-yr DM rates were 50% and 34% after EBRT and EBRT + BT, respectively. Measured from BCR, 5-yr PCSM rates were 27% and 29%, respectively. Interval to BCR was independently associated with DM (p

The gut epithelium serves to maximize the surface for nutrient and fluid uptake, but at the same time must provide a tight barrier to pathogens and remove damaged intestinal epithelial cells (IECs) without jeopardizing barrier integrity. How the epithelium coordinates these tasks remains a question of significant interest. We used imaging and an optical flow analysis pipeline to study the dynamicity of untransformed murine and human intestinal epithelia, cultured atop flexible hydrogel supports. Infection with the pathogen Salmonella Typhimurium (STm) within minutes elicited focal contractions with inward movements of up to ∼1,000 IECs. Genetics approaches and chimeric epithelial monolayers revealed contractions to be triggered by the NAIP/NLRC4 inflammasome, which sensed type-III secretion system and flagellar ligands upon bacterial invasion, converting the local tissue into a contraction epicenter. Execution of the response required swift sublytic Gasdermin D pore formation, ion fluxes, and the propagation of a myosin contraction pulse across the tissue. Importantly, focal contractions preceded, and could be uncoupled from, the death and expulsion of infected IECs. In both two-dimensional monolayers and three-dimensional enteroids, multiple infection-elicited contractions coalesced to produce shrinkage of the epithelium as a whole. Monolayers deficient for Caspase-1(-11) or Gasdermin D failed to elicit focal contractions but were still capable of infected IEC death and expulsion. Strikingly, these monolayers lost their integrity to a markedly higher extent than wild-type counterparts. We propose that prompt NAIP/NLRC4/Caspase-1/Gasdermin D/myosin-dependent contractions allow the epithelium to densify its cell packing in infected regions, thereby preventing tissue disintegration due to the subsequent IEC death and expulsion process.

Overexpressed tumor-associated antigens [for example, epidermal growth factor receptor (EGFR) and human epidermal growth factor receptor 2 (HER2)] are attractive targets for therapeutic T cells, but toxic "off-tumor" cross-reaction with normal tissues that express low levels of target antigen can occur with chimeric antigen receptor (CAR)-T cells. Inspired by natural ultrasensitive response circuits, we engineered a two-step positive-feedback circuit that allows human cytotoxic T cells to discriminate targets on the basis of a sigmoidal antigen-density threshold. In this circuit, a low-affinity synthetic Notch receptor for HER2 controls the expression of a high-affinity CAR for HER2. Increasing HER2 density thus has cooperative effects on T cells-it increases both CAR expression and activation-leading to a sigmoidal response. T cells with this circuit show sharp discrimination between target cells expressing normal amounts of HER2 and cancer cells expressing 100 times as much HER2, both in vitro and in vivo.

ObjectiveThe development of effective cancer treatments depends on the availability of cell lines that faithfully recapitulate the cancer in question. This study definitively re-assigns the histologic identities of two ovarian cancer cell lines, COV434 (originally described as a granulosa cell tumour) and TOV-112D (originally described as grade 3 endometrioid carcinoma), both of which were recently suggested to represent small cell carcinoma of the ovary, hypercalcemic type (SCCOHT), based on their shared gene expression profiles and sensitivity to EZH2 inhibitors.MethodsFor COV434 and TOV-112D, we re-reviewed the original pathology slides and obtained clinical follow-up on the patients, when available, and performed immunohistochemistry for SMARCA4, SMARCA2 and additional diagnostic markers on the original formalin-fixed, paraffin-embedded (FFPE) clinical material, when available. For COV434, we further performed whole exome sequencing and validated SMARCA4 mutations by Sanger sequencing. We studied the growth of the cell lines at baseline and upon re-expression of SMARCA4 in vitro for both cell lines and evaluated the serum calcium levels in vivo upon injection into immunodeficient mice for COV434 cells.ResultsThe available morphological, immunohistochemical, genetic, and clinical features indicate COV434 is derived from SCCOHT, and TOV-112D is a dedifferentiated carcinoma. Transplantation of COV434 into mice leads to increased serum calcium level. Re-expression of SMARCA4 in either COV434 and TOV-112D cells suppressed their growth dramatically.ConclusionsCOV434 represents a bona fide SCCOHT cell line. TOV-112D is a dedifferentiated ovarian carcinoma cell line.

BackgroundMost evidence for interpregnancy interval (IPI) and adverse birth outcomes come from studies that are prone to incomplete control for confounders that vary between women. Comparing pregnancies to the same women can address this issue.MethodsWe conducted an international longitudinal cohort study of 5,521,211 births to 3,849,193 women from Australia (1980-2016), Finland (1987-2017), Norway (1980-2016) and the United States (California) (1991-2012). IPI was calculated based on the time difference between two dates-the date of birth of the first pregnancy and the date of conception of the next (index) pregnancy. We estimated associations between IPI and preterm birth (PTB), spontaneous PTB, and small-for-gestational age births (SGA) using logistic regression (between-women analyses). We also used conditional logistic regression comparing IPIs and birth outcomes in the same women (within-women analyses). Random effects meta-analysis was used to calculate pooled adjusted odds ratios (aOR).ResultsCompared to an IPI of 18-23 months, there was insufficient evidence for an association between IPI 24 month IPIs.ConclusionsWe found consistently elevated odds of adverse birth outcomes following long IPIs. IPI shorter than 6 months were associated with elevated risk of spontaneous PTB, but there was insufficient evidence for increased risk of other adverse birth outcomes. Current recommendations of waiting at least 24 months to conceive after a previous pregnancy, may be unnecessarily long in high-income countries.

ObjectivesMore children are surviving critical illness but are at risk of residual or new health conditions. An evidence-informed and stakeholder-recommended core outcome set is lacking for pediatric critical care outcomes. Our objective was to create a multinational, multistakeholder-recommended pediatric critical care core outcome set for inclusion in clinical and research programs.DesignA two-round modified Delphi electronic survey was conducted with 333 invited research, clinical, and family/advocate stakeholders. Stakeholders completing the first round were invited to participate in the second. Outcomes scoring greater than 69% "critical" and less than 15% "not important" advanced to round 2 with write-in outcomes considered. The Steering Committee held a virtual consensus conference to determine the final components.SettingMultinational survey.PatientsStakeholder participants from six continents representing clinicians, researchers, and family/advocates.Measurements and main resultsOverall response rates were 75% and 82% for each round. Participants voted on seven Global Domains and 45 Specific Outcomes in round 1, and six Global Domains and 30 Specific Outcomes in round 2. Using overall (three stakeholder groups combined) results, consensus was defined as outcomes scoring greater than 90% "critical" and less than 15% "not important" and were included in the final PICU core outcome set: four Global Domains (Cognitive, Emotional, Physical, and Overall Health) and four Specific Outcomes (Child Health-Related Quality of Life, Pain, Survival, and Communication). Families (n = 21) suggested additional critically important outcomes that did not meet consensus, which were included in the PICU core outcome set-extended.ConclusionsThe PICU core outcome set and PICU core outcome set-extended are multistakeholder-recommended resources for clinical and research programs that seek to improve outcomes for children with critical illness and their families.

BackgroundParents of infants in neonatal intensive care units (NICUs) are often unintentionally marginalized in pursuit of optimal clinical care. Family Integrated Care (FICare) was developed to support families as part of their infants' care team in level III NICUs. We adapted the model for level II NICUs in Alberta, Canada, and evaluated whether the new Alberta FICare™ model decreased hospital length of stay (LOS) in preterm infants without concomitant increases in readmissions and emergency department visits.MethodsIn this pragmatic cluster randomized controlled trial conducted between December 15, 2015 and July 28, 2018, 10 level II NICUs were randomized to provide Alberta FICare™ (n = 5) or standard care (n = 5). Alberta FICare™ is a psychoeducational intervention with 3 components: Relational Communication, Parent Education, and Parent Support. We enrolled mothers and their singleton or twin infants born between 32 0/7 and 34 6/7 weeks gestation. The primary outcome was infant hospital LOS. We used a linear regression model to conduct weighted site-level analysis comparing adjusted mean LOS between groups, accounting for site geographic area (urban/regional) and infant risk factors. Secondary outcomes included proportions of infants with readmissions and emergency department visits to 2 months corrected age, type of feeding at discharge, and maternal psychosocial distress and parenting self-efficacy at discharge.ResultsWe enrolled 654 mothers and 765 infants (543 singletons/111 twin cases). Intention to treat analysis included 353 infants/308 mothers in the Alberta FICare™ group and 365 infants/306 mothers in the standard care group. The unadjusted difference between groups in infant hospital LOS (1.96 days) was not statistically significant. Accounting for site geographic area and infant risk factors, infant hospital LOS was 2.55 days shorter (95% CI, - 4.44 to - 0.66) in the Alberta FICare™ group than standard care group, P = .02. Secondary outcomes were not significantly different between groups.ConclusionsAlberta FICare™ is effective in reducing preterm infant LOS in level II NICUs, without concomitant increases in readmissions or emergency department visits. A small number of sites in a single jurisdiction and select group infants limit generalizability of findings.Trial registrationClinicalTrials.gov Identifier NCT02879799 , retrospectively registered August 26, 2016.

BackgroundCurrent South American populations trace their origins mainly to three continental ancestries, i.e. European, Amerindian and African. Individual variation in relative proportions of each of these ancestries may be confounded with socio-economic factors due to population stratification. Therefore, ancestry is a potential confounder variable that should be considered in epidemiologic studies and in public health plans. However, there are few studies that have assessed the ancestry of the current admixed Chilean population. This is partly due to the high cost of genome-scale technologies commonly used to estimate ancestry. In this study we have designed a small panel of SNPs to accurately assess ancestry in the largest sampling to date of the Chilean mestizo population (n = 3349) from eight cities. Our panel is also able to distinguish between the two main Amerindian components of Chileans: Aymara from the north and Mapuche from the south.ResultsA panel of 150 ancestry-informative markers (AIMs) of SNP type was selected to maximize ancestry informativeness and genome coverage. Of these, 147 were successfully genotyped by KASPar assays in 2843 samples, with an average missing rate of 0.012, and a 0.95 concordance with microarray data. The ancestries estimated with the panel of AIMs had relative high correlations (0.88 for European, 0.91 for Amerindian, 0.70 for Aymara, and 0.68 for Mapuche components) with those obtained with AXIOM LAT1 array. The country's average ancestry was 0.53 ± 0.14 European, 0.04 ± 0.04 African, and 0.42 ± 0.14 Amerindian, disaggregated into 0.18 ± 0.15 Aymara and 0.25 ± 0.13 Mapuche. However, Mapuche ancestry was highest in the south (40.03%) and Aymara in the north (35.61%) as expected from the historical location of these ethnic groups. We make our results available through an online app and demonstrate how it can be used to adjust for ancestry when testing association between incidence of a disease and nongenetic risk factors.ConclusionsWe have conducted the most extensive sampling, across many different cities, of current Chilean population. Ancestry varied significantly by latitude and human development. The panel of AIMs is available to the community for estimating ancestry at low cost in Chileans and other populations with similar ancestry.

BackgroundThe BRCA1 c.3331_3334delCAAG founder mutation has been reported in hereditary breast and ovarian cancer families from multiple Hispanic groups. We aimed to evaluate BRCA1 c.3331_3334delCAAG haplotype diversity in cases of European, African, and Latin American ancestry.MethodsBC mutation carrier cases from Colombia (n = 32), Spain (n = 13), Portugal (n = 2), Chile (n = 10), Africa (n = 1), and Brazil (n = 2) were genotyped with the genome-wide single nucleotide polymorphism (SNP) arrays to evaluate haplotype diversity around BRCA1 c.3331_3334delCAAG. Additional Portuguese (n = 13) and Brazilian (n = 18) BC mutation carriers were genotyped for 15 informative SNPs surrounding BRCA1. Data were phased using SHAPEIT2, and identical by descent regions were determined using BEAGLE and GERMLINE. DMLE+ was used to date the mutation in Colombia and Iberia.ResultsThe haplotype reconstruction revealed a shared 264.4-kb region among carriers from all six countries. The estimated mutation age was ~ 100 generations in Iberia and that it was introduced to South America early during the European colonization period.ConclusionsOur results suggest that this mutation originated in Iberia and later introduced to Colombia and South America at the time of Spanish colonization during the early 1500s. We also found that the Colombian mutation carriers had higher European ancestry, at the BRCA1 gene harboring chromosome 17, than controls, which further supported the European origin of the mutation. Understanding founder mutations in diverse populations has implications in implementing cost-effective, ancestry-informed screening.

BackgroundHigh consumption of red and processed meat is commonly associated with increased cancer risk, particularly colorectal cancer. Antibodies against the red meat-derived carbohydrate N-glycolylneuraminic acid (Neu5Gc) exacerbate cancer in "human-like" mice. Human anti-Neu5Gc IgG and red meat are both independently proposed to increase cancer risk, yet how diet affects these antibodies is largely unknown.MethodsWe used world global data to demonstrate that colorectal cancer incidence and mortality are associated with increased national meat consumption. In a well-defined large cohort, we used glycomics to measure daily Neu5Gc intake from red meat and dairy, and investigated serum as well as affinity-purified anti-Neu5Gc antibodies. Based on 24-h dietary records, daily Neu5Gc intake was calculated for 19,621 subjects aged ≥ 18 years of the NutriNet-Santé study. Serum and affinity-purified anti-Neu5Gc antibodies were evaluated by ELISA and glycan microarrays in representative 120 individuals, each with at least eighteen 24-h dietary records (aged 45-60, Q1-Q4; aged > 60, Q1 and Q4; 10 men/women per quartile).ResultsWe found that high-Neu5Gc diet, gender, and age affect the specificity, levels, and repertoires of anti-Neu5Gc IgG immune responses, but not their affinity. Men consumed more Neu5Gc than women, mostly from red meat (p = 0.0015), and exhibited higher overall serum anti-Neu5Gc IgG levels by ELISA (3.94 ng/μl versus 2.22 ng/μl, respectively; p = 0.039). Detailed glycan microarray analysis against 56 different glycans revealed high Neu5Gc-specificity with increased anti-Neu5Gc IgG and altered repertoires, associated with higher consumption of Neu5Gc from red meat and cow dairy. Affinity purification of serum anti-Neu5Gc antibodies revealed increased levels and biased array repertoire patterns, without an increase in antibody affinity, in individuals consuming higher Neu5Gc levels. Furthermore, in a high-meat diet, antibody diversity patterns on glycan microarrays shifted towards Neu5Gcα3-linked glycans, increasing the α3/α6-glycans ratio score.ConclusionsWe found a clear link between the levels and repertoire of serum anti-Neu5Gc IgG and Neu5Gc intake from red meat and dairy. These precise rational methodologies allowed to develop a Gcemic index to simplify the assessment of Neu5Gc in foods that could potentially be adapted for dietary recommendations to reduce cancer risk.

The severe acute respiratory syndrome coronavirus 2 (coronavirus disease 2019 [COVID-19]) pandemic has challenged medical systems and clinicians globally to unforeseen levels. Rapid spread of COVID-19 has forced clinicians to care for patients with a highly contagious disease without evidence-based guidelines. Using a virtual modified nominal group technique, the Pediatric Difficult Intubation Collaborative (PeDI-C), which currently includes 35 hospitals from 6 countries, generated consensus guidelines on airway management in pediatric anesthesia based on expert opinion and early data about the disease. PeDI-C identified overarching goals during care, including minimizing aerosolized respiratory secretions, minimizing the number of clinicians in contact with a patient, and recognizing that undiagnosed asymptomatic patients may shed the virus and infect health care workers. Recommendations include administering anxiolytic medications, intravenous anesthetic inductions, tracheal intubation using video laryngoscopes and cuffed tracheal tubes, use of in-line suction catheters, and modifying workflow to recover patients from anesthesia in the operating room. Importantly, PeDI-C recommends that anesthesiologists consider using appropriate personal protective equipment when performing aerosol-generating medical procedures in asymptomatic children, in addition to known or suspected children with COVID-19. Airway procedures should be done in negative pressure rooms when available. Adequate time should be allowed for operating room cleaning and air filtration between surgical cases. Research using rigorous study designs is urgently needed to inform safe practices during the COVID-19 pandemic. Until further information is available, PeDI-C advises that clinicians consider these guidelines to enhance the safety of health care workers during airway management when performing aerosol-generating medical procedures. These guidelines have been endorsed by the Society for Pediatric Anesthesia and the Canadian Pediatric Anesthesia Society.