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BACKGROUND: Yellow fever virus (YFV) infections are a major global disease concern with high mortality in humans, and as such it is critical to identify clinical correlates of disease severity. While nonstructural protein 1 (NS1) of the related dengue virus is implicated in contributing to vascular leak, little is known about the role of YFV NS1 in severe YF and mechanisms of vascular dysfunction in YFV infections. METHODS: Using serum samples from laboratory-confirmed YF patients with severe (n = 39) or non-severe (n = 18) disease in a well-defined hospital observational cohort in Brazil, plus samples from healthy uninfected controls (n = 11), we investigated factors associated with disease severity and endothelial dysfunction. FINDINGS: We found significantly increased levels of NS1, as well as syndecan-1, a marker of vascular leak, in serum from severe YF as compared to non-severe YF or control groups. We also showed that hyperpermeability of endothelial cell monolayers treated with serum from severe YF patients was significantly higher compared to non-severe YF and control groups, as measured by transendothelial electrical resistance (TEER). Further, we demonstrated that YFV NS1 induces shedding of syndecan-1 from the surface of human endothelial cells. Notably, YFV NS1 serum levels significantly correlated with syndecan-1 serum levels, TEER values, and signs of disease severity. Syndecan-1 levels also significantly correlated with clinical laboratory parameters of disease severity, viral load, hospitalization, and death. INTERPRETATION: This study provides further evidence for endothelial dysfunction as a mechanism of YF pathogenesis in humans and suggests serum quantification of YFV NS1 and syndecan-1 as valuable tools for disease diagnosis and/or prognosis. FUNDING: This work was supported by the US NIH and FAPESP.

In the modern "omics" era, measurement of the human exposome is a critical missing link between genetic drivers and disease outcomes. High-resolution mass spectrometry (HRMS), routinely used in proteomics and metabolomics, has emerged as a leading technology to broadly profile chemical exposure agents and related biomolecules for accurate mass measurement, high sensitivity, rapid data acquisition, and increased resolution of chemical space. Non-targeted approaches are increasingly accessible, supporting a shift from conventional hypothesis-driven, quantitation-centric targeted analyses toward data-driven, hypothesis-generating chemical exposome-wide profiling. However, HRMS-based exposomics encounters unique challenges. New analytical and computational infrastructures are needed to expand the analysis coverage through streamlined, scalable, and harmonized workflows and data pipelines that permit longitudinal chemical exposome tracking, retrospective validation, and multi-omics integration for meaningful health-oriented inferences. In this article, we survey the literature on state-of-the-art HRMS-based technologies, review current analytical workflows and informatic pipelines, and provide an up-to-date reference on exposomic approaches for chemists, toxicologists, epidemiologists, care providers, and stakeholders in health sciences and medicine. We propose efforts to benchmark fit-for-purpose platforms for expanding coverage of chemical space, including gas/liquid chromatography-HRMS (GC-HRMS and LC-HRMS), and discuss opportunities, challenges, and strategies to advance the burgeoning field of the exposome.

The first-generation Spike-alone-based COVID-19 vaccines have successfully contributed to reducing the risk of hospitalization, serious illness, and death caused by SARS-CoV-2 infections. However, waning immunity induced by these vaccines failed to prevent immune escape by many variants of concern (VOCs) that emerged from 2020 to 2024, resulting in a prolonged COVID-19 pandemic. We hypothesize that a next-generation Coronavirus (CoV) vaccine incorporating highly conserved non-Spike SARS-CoV-2 antigens would confer stronger and broader cross-protective immunity against multiple VOCs. In the present study, we identified ten non-Spike antigens that are highly conserved in 8.7 million SARS-CoV-2 strains, twenty-one VOCs, SARS-CoV, MERS-CoV, Common Cold CoVs, and animal CoVs. Seven of the 10 antigens were preferentially recognized by CD8+ and CD4+ T-cells from unvaccinated asymptomatic COVID-19 patients, irrespective of VOC infection. Three out of the seven conserved non-Spike T cell antigens belong to the early expressed Replication and Transcription Complex (RTC) region, when administered to the golden Syrian hamsters, in combination with Spike, as nucleoside-modified mRNA encapsulated in lipid nanoparticles (LNP) (i.e., combined mRNA/LNP-based pan-CoV vaccine): (i) Induced high frequencies of lung-resident antigen-specific CXCR5+CD4+ T follicular helper (TFH) cells, GzmB+CD4+ and GzmB+CD8+ cytotoxic T cells (TCYT), and CD69+IFN-γ+TNFα+CD4+ and CD69+IFN-γ+TNFα+CD8+ effector T cells (TEFF); and (ii) Reduced viral load and COVID-19-like symptoms caused by various VOCs, including the highly pathogenic B.1.617.2 Delta variant and the highly transmittable heavily Spike-mutated XBB1.5 Omicron sub-variant. The combined mRNA/LNP-based pan-CoV vaccine could be rapidly adapted for clinical use to confer broader cross-protective immunity against emerging highly mutated and pathogenic VOCs.

BACKGROUND: Cross-reactive SARS-CoV-2-specific memory CD4+ and CD8+ T cells are present in up to 50% of unexposed, pre-pandemic, healthy individuals (UPPHIs). However, the characteristics of cross-reactive memory CD4+ and CD8+ T cells associated with subsequent protection of asymptomatic coronavirus disease 2019 (COVID-19) patients (i.e., unvaccinated individuals who never develop any COVID-19 symptoms despite being infected with SARS-CoV-2) remains to be fully elucidated. METHODS: This study compares the antigen specificity, frequency, phenotype, and function of cross-reactive memory CD4+ and CD8+ T cells between common cold coronaviruses (CCCs) and SARS-CoV-2. T-cell responses against genome-wide conserved epitopes were studied early in the disease course in a cohort of 147 unvaccinated COVID-19 patients who were divided into six groups based on the severity of their symptoms. RESULTS: Compared to severely ill COVID-19 patients and patients with fatal COVID-19 outcomes, the asymptomatic COVID-19 patients displayed significantly: (i) higher rates of co-infection with the 229E alpha species of CCCs (α-CCC-229E); (ii) higher frequencies of cross-reactive functional CD134+CD137+CD4+ and CD134+CD137+CD8+ T cells that cross-recognized conserved epitopes from α-CCCs and SARS-CoV-2 structural, non-structural, and accessory proteins; and (iii) lower frequencies of CCCs/SARS-CoV-2 cross-reactive exhausted PD-1+TIM3+TIGIT+CTLA4+CD4+ and PD-1+TIM3+TIGIT+CTLA4+CD8+ T cells, detected both ex vivo and in vitro. CONCLUSIONS: These findings (i) support a crucial role of functional, poly-antigenic α-CCCs/SARS-CoV-2 cross-reactive memory CD4+ and CD8+ T cells, induced following previous CCCs seasonal exposures, in protection against subsequent severe COVID-19 disease and (ii) provide critical insights into developing broadly protective, multi-antigen, CD4+, and CD8+ T-cell-based, universal pan-Coronavirus vaccines capable of conferring cross-species protection.

BACKGROUND: The coronavirus disease 2019 (COVID-19) pandemic has created one of the largest global health crises in almost a century. Although the current rate of Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections has decreased significantly, the long-term outlook of COVID-19 remains a serious cause of morbidity and mortality worldwide, with the mortality rate still substantially surpassing even that recorded for influenza viruses. The continued emergence of SARS-CoV-2 variants of concern (VOCs), including multiple heavily mutated Omicron sub-variants, has prolonged the COVID-19 pandemic and underscores the urgent need for a next-generation vaccine that will protect from multiple SARS-CoV-2 VOCs. METHODS: We designed a multi-epitope-based coronavirus vaccine that incorporated B, CD4+, and CD8+ T- cell epitopes conserved among all known SARS-CoV-2 VOCs and selectively recognized by CD8+ and CD4+ T-cells from asymptomatic COVID-19 patients irrespective of VOC infection. The safety, immunogenicity, and cross-protective immunity of this pan-variant SARS-CoV-2 vaccine were studied against six VOCs using an innovative triple transgenic h-ACE-2-HLA-A2/DR mouse model. RESULTS: The pan-variant SARS-CoV-2 vaccine (i) is safe , (ii) induces high frequencies of lung-resident functional CD8+ and CD4+ TEM and TRM cells , and (iii) provides robust protection against morbidity and virus replication. COVID-19-related lung pathology and death were caused by six SARS-CoV-2 VOCs: Alpha (B.1.1.7), Beta (B.1.351), Gamma or P1 (B.1.1.28.1), Delta (lineage B.1.617.2), and Omicron (B.1.1.529). CONCLUSION: A multi-epitope pan-variant SARS-CoV-2 vaccine bearing conserved human B- and T- cell epitopes from structural and non-structural SARS-CoV-2 antigens induced cross-protective immunity that facilitated virus clearance, and reduced morbidity, COVID-19-related lung pathology, and death caused by multiple SARS-CoV-2 VOCs.

The use of different approaches for design and results presentation of studies for the clinical performance evaluation of continuous glucose monitoring (CGM) systems has long been recognized as a major challenge in comparing their results. However, a comprehensive characterization of the variability in study designs is currently unavailable. This article presents a scoping review of clinical CGM performance evaluations published between 2002 and 2022. Specifically, this review quantifies the prevalence of numerous options associated with various aspects of study design, including subject population, comparator (reference) method selection, testing procedures, and statistical accuracy evaluation. We found that there is a large variability in nearly all of those aspects and, in particular, in the characteristics of the comparator measurements. Furthermore, these characteristics as well as other crucial aspects of study design are often not reported in sufficient detail to allow an informed interpretation of study results. We therefore provide recommendations for reporting the general study design, CGM system use, comparator measurement approach, testing procedures, and data analysis/statistical performance evaluation. Additionally, this review aims to serve as a foundation for the development of a standardized CGM performance evaluation procedure, thereby supporting the goals and objectives of the Working Group on CGM established by the Scientific Division of the International Federation of Clinical Chemistry and Laboratory Medicine.

The Hippo pathway is known for its crucial involvement in development, regeneration, organ size control, and cancer. While energy stress is known to activate the Hippo pathway and inhibit its effector YAP, the precise role of the Hippo pathway in energy stress response remains unclear. Here, we report a YAP-independent function of the Hippo pathway in facilitating autophagy and cell survival in response to energy stress, a process mediated by its upstream components MAP4K2 and STRIPAK. Mechanistically, energy stress disrupts the MAP4K2-STRIPAK association, leading to the activation of MAP4K2. Subsequently, MAP4K2 phosphorylates ATG8-family member LC3, thereby facilitating autophagic flux. MAP4K2 is highly expressed in head and neck cancer, and its mediated autophagy is required for head and neck tumor growth in mice. Altogether, our study unveils a noncanonical role of the Hippo pathway in energy stress response, shedding light on this key growth-related pathway in tissue homeostasis and cancer.

The adult human breast is comprised of an intricate network of epithelial ducts and lobules that are embedded in connective and adipose tissue1-3. Although most previous studies have focused on the breast epithelial system4-6, many of the non-epithelial cell types remain understudied. Here we constructed the comprehensive Human Breast Cell Atlas (HBCA) at single-cell and spatial resolution. Our single-cell transcriptomics study profiled 714,331 cells from 126 women, and 117,346 nuclei from 20 women, identifying 12 major cell types and 58 biological cell states. These data reveal abundant perivascular, endothelial and immune cell populations, and highly diverse luminal epithelial cell states. Spatial mapping using four different technologies revealed an unexpectedly rich ecosystem of tissue-resident immune cells, as well as distinct molecular differences between ductal and lobular regions. Collectively, these data provide a reference of the adult normal breast tissue for studying mammary biology and diseases such as breast cancer.

BackgroundThere is a paucity of targeted therapies for patients with pseudomyxoma peritonei (PMP) secondary to low-grade appendiceal mucinous neoplasms (LAMNs). Dysregulated metabolism has emerged as a hallmark of cancer, and the relationship of metabolomics and cancer is an area of active scientific exploration. We sought to characterize phenotypic differences found in peritoneal metastases (PM) derived from LAMN versus adenocarcinoma.MethodsTumors were washed with phosphate-buffered saline (PBS), microdissected, then dissociated in ice-cold methanol dried and reconstituted in pyridine. Samples were derivatized in tert-butyldimethylsilyl (TBDMS) and subjected to gas chromatography-coupled mass spectrometry. Metabolites were assessed based on a standard library. RNA sequencing was performed, with pathway and network analyses on differentially expressed genes.ResultsEight peritoneal tumor samples were obtained and analyzed: LAMNs (4), and moderate to poorly differentiated adenocarcinoma (colon [1], appendix [3]). Decreases in pyroglutamate, fumarate, and cysteine in PM from LAMNs were found compared with adenocarcinoma. Analyses showed the differential gene expression was dominated by the prevalence of metabolic pathways, particularly lipid metabolism. The gene retinol saturase (RETSAT), downregulated by LAMN, was involved in the multiple metabolic pathways that involve lipids. Using network mapping, we found IL1B signaling to be a potential top-level modulation candidate.ConclusionsDistinct metabolic signatures may exist for PM from LAMN versus adenocarcinoma. A multitude of genes are differentially regulated, many of which are involved in metabolic pathways. Additional research is needed to identify the significance and applicability of targeting metabolic pathways in the potential development of novel therapeutics for these challenging tumors.

Pooling of samples can increase throughput and reduce costs for large-scale SARS-CoV-2 testing when incidence is low. In a cross-sectional study of serial SARS-CoV-2 sampling of staff and residents at three nursing homes, laboratory labor constraints limited the feasibility of pooling prior to the maximal incidence that favored cost savings. IMPORTANCE This study highlights the pragmatic considerations surrounding SARS-CoV-2 sample pooling beyond accuracy and costs. We performed a cost analysis to determine the percent positivity at which pooling would reduce costs versus single testing. We found that the need for a stable amount of daily work hours staffed by a highly trained workforce was a major limitation in pooling as test positivity increased. For the COVID-19 pandemic and future pandemic threats, laboratories should carefully consider the thresholds at which sample pooling is beneficial, with a particular focus on the impact on laboratory staff.

BackgroundVisual impairment could worsen sleep/wake disorders and cognitive decline.ObjectiveTo examine interrelations among self-reported visual impairment, sleep, and cognitive decline in the Hispanic Community Health Study/Study of Latinos (HCHS/SOL) Miami-site.MethodHCHS/SOL Miami-site participants ages 45-74 years (n = 665) at Visit-1, who returned for cognitive test 7-years later (SOL-INCA). Participants completed the National Eye Institute Visual Functioning Questionnaire (NEI-VFQ), validated sleep questionnaires and test for obstructive sleep apnea (OSA) at Visit-1. We obtained verbal episodic learning and memory, verbal fluency, processing speed, and executive functioning at Visit-1 and at SOL-INCA. Processing speed/executive functioning were added to SOL-INCA. We examined global cognition and change using a regression-based reliable change index, adjusting for the time lapse between Visit-1 and SOL-INCA. We used regression models to test whether 1) persons with OSA, self-reported sleep duration, insomnia, and sleepiness have an increased risk for visual impairment, 2a) visual impairment is associated with worse cognitive function and/or decline, and 2b) sleep disorders attenuate these associations.ResultSleepiness (β= 0.04; p

BackgroundWhile others have reported severe acute respiratory syndrome-related coronavirus 2(SARS-CoV-2) seroprevalence studies in health care workers (HCWs), we leverage the use of a highly sensitive coronavirus antigen microarray to identify a group of seropositive health care workers who were missed by daily symptom screening that was instituted prior to any epidemiologically significant local outbreak. Given that most health care facilities rely on daily symptom screening as the primary method to identify SARS-CoV-2 among health care workers, here, we aim to determine how demographic, occupational, and clinical variables influence SARS-CoV-2 seropositivity among health care workers.MethodsWe designed a cross-sectional survey of HCWs for SARS-CoV-2 seropositivity conducted from May 15th to June 30th 2020 at a 418-bed academic hospital in Orange County, California. From an eligible population of 5,349 HCWs, study participants were recruited in two ways: an open cohort, and a targeted cohort. The open cohort was open to anyone, whereas the targeted cohort that recruited HCWs previously screened for COVID-19 or work in high-risk units. A total of 1,557 HCWs completed the survey and provided specimens, including 1,044 in the open cohort and 513 in the targeted cohort. Demographic, occupational, and clinical variables were surveyed electronically. SARS-CoV-2 seropositivity was assessed using a coronavirus antigen microarray (CoVAM), which measures antibodies against eleven viral antigens to identify prior infection with 98% specificity and 93% sensitivity.ResultsAmong tested HCWs (n = 1,557), SARS-CoV-2 seropositivity was 10.8%, and risk factors included male gender (OR 1.48, 95% CI 1.05-2.06), exposure to COVID-19 outside of work (2.29, 1.14-4.29), working in food or environmental services (4.85, 1.51-14.85), and working in COVID-19 units (ICU: 2.28, 1.29-3.96; ward: 1.59, 1.01-2.48). Amongst 1,103 HCWs not previously screened, seropositivity was 8.0%, and additional risk factors included younger age (1.57, 1.00-2.45) and working in administration (2.69, 1.10-7.10).ConclusionSARS-CoV-2 seropositivity is significantly higher than reported case counts even among HCWs who are meticulously screened. Seropositive HCWs missed by screening were more likely to be younger, work outside direct patient care, or have exposure outside of work.

Limited data are available on breakthrough COVID-19 in patients with hematologic malignancy (HM) after anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccination. Adult patients with HM, ≥1 dose of anti-SARS-CoV-2 vaccine, and breakthrough COVID-19 between January 2021 and March 2022 were analyzed. A total of 1548 cases were included, mainly lymphoid malignancies (1181 cases, 76%). After viral sequencing in 753 cases (49%), the Omicron variant was prevalent (517, 68.7%). Most of the patients received ≤2 vaccine doses before COVID-19 (1419, 91%), mostly mRNA-based (1377, 89%). Overall, 906 patients (59%) received COVID-19-specific treatment. After 30-day follow-up from COVID-19 diagnosis, 143 patients (9%) died. The mortality rate in patients with the Omicron variant was 7.9%, comparable to other variants, with a significantly lower 30-day mortality rate than in the prevaccine era (31%). In the univariable analysis, older age (P < .001), active HM (P < .001), and severe and critical COVID-19 (P = .007 and P < .001, respectively) were associated with mortality. Conversely, patients receiving monoclonal antibodies, even for severe or critical COVID-19, had a lower mortality rate (P < .001). In the multivariable model, older age, active disease, critical COVID-19, and 2-3 comorbidities were correlated with a higher mortality, whereas monoclonal antibody administration, alone (P < .001) or combined with antivirals (P = .009), was protective. Although mortality is significantly lower than in the prevaccination era, breakthrough COVID-19 in HM is still associated with considerable mortality. Death rate was lower in patients who received monoclonal antibodies, alone or in combination with antivirals.

ObjectiveThe COVID-19 pandemic prompted families to receive Early Head Start (EHS) home-based services virtually. This qualitative study evaluated parental perceptions of EHS tele-home visits.MethodsEHS parents who had transitioned to tele-home visits using any video-chat platform were recruited to participate in a Spanish or English virtual focus group that assessed their perceptions of tele-home visits. Using an iterative, consensus-seeking inductive content analysis approach, themes and subthemes were identified.ResultsThirty-five mothers of children newborn to 3-years-old, where the majority were Latino and Spanish-speaking, participated in four focus groups. Several patterns pertaining to technology, child engagement, child learning and development, and parent-home visitor relationship emerged in the qualitative analysis. Mothers revealed varying degrees of digital proficiency, device preference, and technology challenges. Mothers reported variability in child engagement and concerns with missed socialization opportunities for children as a results of tele-home visits, but also reported increased self-efficacy in supporting child development, positive relationships with their home visitor, and overall satisfaction with services.ConclusionParents revealed tele-home visits have the potential to be a viable service delivery method for EHS home-based programs. While parents perceived increased engagement and an uncompromised parent-home visitor relationship, they revealed areas of needed support that would optimize the use of tele-home visits.

BackgroundThe Pediatric Emergency Medicine (PEM) Point-of-care Ultrasound (POCUS) Network (P2Network) was established in 2014 to provide a platform for international collaboration among experts, including multicenter research. The objective of this study was to use expert consensus to identify and prioritize PEM POCUS topics, to inform future collaborative multicenter research.MethodsOnline surveys were administered in a two-stage, modified Delphi study. A steering committee of 16 PEM POCUS experts was identified within the P2Network, with representation from the United States, Canada, Italy, and Australia. We solicited the participation of international PEM POCUS experts through professional society mailing lists, research networks, social media, and "word of mouth." After each round, responses were refined by the steering committee before being reissued to participants to determine the ranking of all the research questions based on means and to identify the high-level consensus topics. The final stage was a modified Hanlon process of prioritization round (HPP), which emphasized relevance, impact, and feasibility.ResultsFifty-four eligible participants (16.6%) provided 191 items to Survey 1 (Round 1). These were refined and consolidated into 52 research questions by the steering committee. These were issued for rating in Survey 2 (Round 2), which had 45 participants. At the completion of Round 2, all questions were ranked with six research questions reaching high-level consensus. Thirty-one research questions with mean ratings above neutral were selected for the HPP round. Highly ranked topics included clinical applications of POCUS to evaluate and manage children with shock, cardiac arrest, thoracoabdominal trauma, suspected cardiac failure, atraumatic limp, and intussusception.ConclusionsThis consensus study has established a research agenda to inform future international multicenter PEM POCUS trials. This study has highlighted the ongoing need for high-quality evidence for PEM POCUS applications to guide clinical practice.

The tumor suppressor p53 is the most frequently mutated protein in human cancer. The majority of these mutations are missense mutations in the DNA binding domain of p53. Restoring p53 tumor suppressor function could have a major impact on the therapy for a wide range of cancers. Here we report a virtual screening approach that identified several small molecules with p53 reactivation activities. The UCI-LC0023 compound series was studied in detail and was shown to bind p53, induce a conformational change in mutant p53, restore the ability of p53 hotspot mutants to associate with chromatin, reestablish sequence-specific DNA binding of a p53 mutant in a reconstituted in vitro system, induce p53-dependent transcription programs, and prevent progression of tumors carrying mutant p53, but not p53null or p53WT alleles. Our study demonstrates feasibility of a computation-guided approach to identify small molecule corrector drugs for p53 hotspot mutations.

High-throughput and rapid screening testing is highly desirable to effectively combat the rapidly evolving COVID-19 pandemic co-presents with influenza and seasonal common cold epidemics. Here, we present a general workflow for iterative development and validation of an antibody-based microarray assay for the detection of a respiratory viral panel: (a) antibody screening to quickly identify optimal reagents and assay conditions, (b) immunofluorescence assay design including signal amplification for low viral titers, (c) assay characterization with recombinant proteins, inactivated viral samples and clinical samples, and (d) multiplexing to detect a panel of common respiratory viruses. Using RT-PCR-confirmed SARS-CoV-2 positive and negative pharyngeal swab samples, we demonstrated that the antibody microarray assay exhibited a clinical sensitivity and specificity of 77.2% and 100%, respectively, which are comparable to existing FDA-authorized antigen tests. Moreover, the microarray assay is correlated with RT-PCR cycle threshold (Ct) values and is particularly effective in identifying high viral titers. The multiplexed assay can selectively detect SARS-CoV-2 and influenza virus, which can be used to discriminate these viral infections that share similar symptoms. Such protein microarray technology is amenable for scale-up and automation and can be broadly applied as a both diagnostic and research tool.

Aminotransferases (ATs) are pyridoxal 5'-phosphate-dependent enzymes that catalyze the transamination reactions between amino acid donor and keto acid acceptor substrates. Modern AT enzymes constitute ∼2% of all classified enzymatic activities, play central roles in nitrogen metabolism, and generate multitude of primary and secondary metabolites. ATs likely diverged into four distinct AT classes before the appearance of the last universal common ancestor and further expanded to a large and diverse enzyme family. Although the AT family underwent an extensive functional specialization, many AT enzymes retained considerable substrate promiscuity and multifunctionality because of their inherent mechanistic, structural, and functional constraints. This review summarizes the evolutionary history, diverse metabolic roles, reaction mechanisms, and structure-function relationships of the AT family enzymes, with a special emphasis on their substrate promiscuity and multifunctionality. Comprehensive characterization of AT substrate specificity is still needed to reveal their true metabolic functions in interconnecting various branches of the nitrogen metabolic network in different organisms.

Sjögrens disease is a complex autoimmune disease with twelve established susceptibility loci. This genome-wide association study (GWAS) identifies ten novel genome-wide significant (GWS) regions in Sjögrens cases of European ancestry: CD247, NAB1, PTTG1-MIR146A, PRDM1-ATG5, TNFAIP3, XKR6, MAPT-CRHR1, RPTOR-CHMP6-BAIAP6, TYK2, SYNGR1. Polygenic risk scores yield predictability (AUROC = 0.71) and relative risk of 12.08. Interrogation of bioinformatics databases refine the associations, define local regulatory networks of GWS SNPs from the 95% credible set, and expand the implicated gene list to >40. Many GWS SNPs are eQTLs for genes within topologically associated domains in immune cells and/or eQTLs in the main target tissue, salivary glands.

Multiple myeloma (MM) is a plasma cell malignancy that affects an increasing number of patients worldwide. Despite all the efforts to understand its pathogenesis and develop new treatment modalities, MM remains an incurable disease. Novel immunotherapies, such as CAR T cell therapy (CAR) and bispecific T cell engagers (BiTE), are intensively targeting different surface antigens, such as BMCA, SLAMF7 (CS1), GPRC5D, FCRH5 or CD38. However, stem cell transplantation is still indispensable in transplant-eligible patients. Studies suggest that the early use of immunotherapy may improve outcomes significantly. In this review, we summarize the currently available clinical literature on CAR and BiTE in MM. Furthermore, we will compare these two T cell-based immunotherapies and discuss potential therapeutic approaches to promote development of new clinical trials, using T cell-based immunotherapies, even as bridging therapies to a transplant.

The hippocampus is implicated in memory formation, and neurons in the hippocampus take part in replay sequences that have been proposed to reflect memory of explored space. By recording from large ensembles of hippocampal neurons as rats explored various tracks, we show that sustained replay appears after a single experience. Further, we found that with repeated experience in a novel environment, replay slows down, taking more time to traverse the same trajectory. This effect was dependent on experience, not passage of time, and was environment specific. By investigating the slow-gamma (25-50 Hz) hover-and-jump dynamics within replays, we show that replay slows by adding more hover locations, increasing the resolution of the behavioral trajectory. We provide evidence that inhibition and cortical engagement both increase as replay slows. Thus, replays can reflect single experiences and evolve with re-exposure, in a manner consistent with the encoding of greater detail into replay memories with experience.

While severe coronavirus 2019 (COVID-19) is associated with immune activation at the maternal-fetal interface, responses to asymptomatic/mild severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection during pregnancy remain unknown. Here, we assess immunological adaptations in blood and term decidua in response to asymptomatic/mild disease in pregnant women. We report attenuated antigen presentation and type I interferon (IFN) signaling pathways, loss of tissue-resident decidual macrophages, and upregulated cytokine/chemokine signaling in monocyte-derived decidual macrophages. Furthermore, we describe increased frequencies of activated tissue-resident T cells and decreased abundance of regulatory T cells with infection while frequencies of cytotoxic CD4/CD8 T cells are increased in the blood. In contrast to decidual macrophages, type I IFN signaling is higher in decidual T cells. Finally, infection leads to a narrowing of T cell receptor diversity in both blood and decidua. Collectively, these observations indicate that asymptomatic/mild COVID-19 during pregnancy results in remodeling of the immunological landscape of the maternal-fetal interface, with a potential for long-term adverse outcomes for the offspring.

BackgroundBody composition measures, specifically low weight or reduced muscle mass, are associated with mortality in COPD, but the effect of longitudinal body composition changes is undefined.Research questionIs the longitudinal loss of fat-free mass (FFM) associated with increased mortality, including in those with initially normal or elevated body composition metrics?Study design and methodsParticipants with complete data for at least one visit in the COPDGene study (n = 9,268) and the ECLIPSE study (n = 1,760) were included and monitored for 12 and 8 years, respectively. Pectoralis muscle area (PMA) was derived from thoracic CT scans and used as a proxy for FFM. A longitudinal mixed submodel for PMA and a Cox proportional hazards submodel for survival were fitted on a joint distribution, using a shared random intercept parameter and Markov chain Monte Carlo parameter estimation.ResultsBoth cohorts demonstrated a left-shifted distribution of baseline FFM, not reflected in BMI, and an increase in all-cause mortality risk associated with longitudinal loss of PMA. For each 1-cm2 PMA loss, mortality increased 3.1% (95% CI, 2.4%-3.7%; P < .001) in COPDGene, and 2.4% (95% CI, 0.9%-4.0%; P < .001) in ECLIPSE. Increased mortality risk was independent of enrollment values for BMI and disease severity [BODE (body mass, airflow obstruction, dyspnea, and exercise capacity) index quartiles] and was significant even in participants with initially greater than average PMA.InterpretationLongitudinal loss of PMA is associated with increased all-cause mortality, regardless of BMI or initial muscle mass. Consideration of novel screening tests and further research into mechanisms contributing to muscle decline may improve risk stratification and identify novel therapeutic targets in ever smokers.

The recent classification of colon cancer into molecular subtypes revealed that patients with the poorest prognosis harbor tumors with the lowest levels of Wnt signaling. This is contrary to the general understanding that overactive Wnt signaling promotes tumor progression from early initiation stages through to the later stages including invasion and metastasis. Here, we directly test this assumption by reducing the activity of ß-catenin-dependent Wnt signaling in colon cancer cell lines at either an upstream or downstream step in the pathway. We determine that Wnt-reduced cancer cells exhibit a more aggressive disease phenotype, including increased mobility in vitro and disruptive invasion into mucosa and smooth muscle in an orthotopic mouse model. RNA sequencing reveals that interference with Wnt signaling leads to an upregulation of gene programs that favor cell migration and invasion and a downregulation of inflammation signatures in the tumor microenvironment. We identify a set of upregulated genes common among the Wnt perturbations that are predictive of poor patient outcomes in early-invasive colon cancer. Our findings suggest that while targeting Wnt signaling may reduce tumor burden, an inadvertent side effect is the emergence of invasive cancer.ImplicationsDecreased Wnt signaling in colon tumors leads to a more aggressive disease phenotype due to an upregulation of gene programs favoring cell migration in the tumor and downregulation of inflammation programs in the tumor microenvironment; these impacts must be carefully considered in developing Wnt-targeting therapies.

Several mental illnesses, characterized by aberrant stress reactivity, often arise after early-life adversity (ELA). However, it is unclear how ELA affects stress-related brain circuit maturation, provoking these enduring vulnerabilities. We find that ELA increases functional excitatory synapses onto stress-sensitive hypothalamic corticotropin-releasing hormone (CRH)-expressing neurons, resulting from disrupted developmental synapse pruning by adjacent microglia. Microglial process dynamics and synaptic element engulfment were attenuated in ELA mice, associated with deficient signaling of the microglial phagocytic receptor MerTK. Accordingly, selective chronic chemogenetic activation of ELA microglia increased microglial process dynamics and reduced excitatory synapse density to control levels. Notably, selective early-life activation of ELA microglia normalized adult acute and chronic stress responses, including stress-induced hormone secretion and behavioral threat responses, as well as chronic adrenal hypertrophy of ELA mice. Thus, microglial actions during development are powerful contributors to mechanisms by which ELA sculpts the connectivity of stress-regulating neurons, promoting vulnerability to stress and stress-related mental illnesses.

Rab27A is a small GTPase, which mediates transport and docking of secretory vesicles at the plasma membrane via protein-protein interactions (PPIs) with effector proteins. Rab27A promotes the growth and invasion of multiple cancer types such as breast, lung and pancreatic, by enhancing secretion of chemokines, metalloproteases and exosomes. The significant role of Rab27A in multiple cancer types and the minor role in adults suggest that Rab27A may be a suitable target to disrupt cancer metastasis. Similar to many GTPases, the flat topology of the Rab27A-effector PPI interface and the high affinity for GTP make it a challenging target for inhibition by small molecules. Reported co-crystal structures show that several effectors of Rab27A interact with the Rab27A SF4 pocket (WF-binding pocket) via a conserved tryptophan-phenylalanine (WF) dipeptide motif. To obtain structural insight into the ligandability of this pocket, a novel construct was designed fusing Rab27A to part of an effector protein (fRab27A), allowing crystallisation of Rab27A in high throughput. The paradigm of KRas covalent inhibitor development highlights the challenge presented by GTPase proteins as targets. However, taking advantage of two cysteine residues, C123 and C188, that flank the WF pocket and are unique to Rab27A and Rab27B among the >60 Rab family proteins, we used the quantitative Irreversible Tethering (qIT) assay to identify the first covalent ligands for native Rab27A. The binding modes of two hits were elucidated by co-crystallisation with fRab27A, exemplifying a platform for identifying suitable lead fragments for future development of competitive inhibitors of the Rab27A-effector interaction interface, corroborating the use of covalent libraries to tackle challenging targets.

Efficient HIV-1 replication depends on balanced levels of host cell components including cellular splicing factors as the family of serine/arginine-rich splicing factors (SRSF, 1-10). Type I interferons (IFN-I) play a crucial role in the innate immunity against HIV-1 by inducing the expression of IFN-stimulated genes (ISGs) including potent host restriction factors. The less well known IFN-repressed genes (IRepGs) might additionally affect viral replication by downregulating host dependency factors that are essential for the viral life cycle; however, so far, the knowledge about IRepGs involved in HIV-1 infection is very limited. In this work, we could demonstrate that HIV-1 infection and the associated ISG induction correlated with low SRSF1 levels in intestinal lamina propria mononuclear cells (LPMCs) and peripheral blood mononuclear cells (PBMCs) during acute and chronic HIV-1 infection. In HIV-1-susceptible cell lines as well as primary monocyte-derived macrophages (MDMs), expression levels of SRSF1 were transiently repressed upon treatment with specific IFNα subtypes in vitro. Mechanically, 4sU labeling of newly transcribed mRNAs revealed that IFN-mediated SRSF1 repression is regulated on early RNA level. SRSF1 knockdown led to an increase in total viral RNA levels, but the relative proportion of the HIV-1 viral infectivity factor (Vif) coding transcripts, which is essential to counteract APOBEC3G-mediated host restriction, was significantly reduced. In the presence of high APOBEC3G levels, however, increased LTR activity upon SRSF1 knockdown facilitated the overall replication, despite decreased vif mRNA levels. In contrast, SRSF1 overexpression significantly impaired HIV-1 post-integration steps including LTR transcription, alternative splice site usage, and virus particle production. Since balanced SRSF1 levels are crucial for efficient viral replication, our data highlight the so far undescribed role of SRSF1 acting as an IFN-modulated cellular dependency factor decisively regulating HIV-1 post-integration steps.

BackgroundMexican immigrants in the United States face mental health challenges, disparities, and limited access to healthcare; however, mental health promotion efforts specifically targeting this population have been insufficient. The objective of this study was to develop and test a mental health promotion intervention based on protective mental health factors and coping strategies for Mexican immigrants recruited through a free, consulate-based program in Atlanta.Material and methodsWorking with the Ventanilla de Salud program, we conducted a longitudinal study in three phases: formative research and design, pre-intervention assessment and post-implementation evaluation. The intervention was designed based on the health promotion model and interviews with stakeholders. Qualitative information was collected by semi-structured interviews with participants before and after the intervention. Quantitative outcomes were knowledge about protective factors and coping mechanisms, and psychosocial distress. Differences were assessed using the Wilcoxon non-parametrical test. Intent-to-treat analysis was conducted with all participants who signed the informed consent (carrying last observation forward), and a complete case analysis was conducted with those who attended at least 70% of the sessions and completed the post- implementation evaluation.ResultsTwenty-five participants were enrolled in the intervention. Mean age was 38 years, and the majority were women. Only nine participants attended at least 70% of the sessions and completed the final evaluation. Men, those who did not complete high school, and workers in service or construction jobs were more likely to drop out. Knowledge about protective factors [pre- vs. post-intervention median (inter-quartile range) = 111 (100, 120) vs. 115 (100, 124)] and coping mechanisms [96 (85, 104) vs. 99 (90, 110)], as well as psychosocial distress [3 (2, 3) vs. 2 (2, 3)] improved after the intervention in both intent-to treat and complete case analyses (p < 0.05). Qualitative results also support improvements in targeted protective factors.DiscussionThe intervention was successful in improving psychological distress among Mexican immigrants. These results support the implementation of evidence-based mental health promotion interventions among Mexican immigrants via free and familiar programs. A limitation was the high attrition; future studies should explore approaches to improve retention in this population.

Subunit vaccines offer advantages over more traditional inactivated or attenuated whole-cell-derived vaccines in safety, stability, and standard manufacturing. To achieve an effective protein-based subunit vaccine, the protein antigen often needs to adopt a native-like conformation. This is particularly important for pathogen-surface antigens that are membrane-bound proteins. Cell-free methods have been successfully used to produce correctly folded functional membrane protein through the co-translation of nanolipoprotein particles (NLPs), commonly known as nanodiscs. This strategy can be used to produce subunit vaccines consisting of membrane proteins in a lipid-bound environment. However, cell-free protein production is often limited to small scale (

BackgroundObesity is a persistent public health concern and a risk factor for many chronic diseases including at least 13 different cancers. Adult Black females have the highest prevalence of obesity (57%) compared to other racial/gender groups in the U.S. Although behavioral weight loss (BWL) interventions have demonstrated effectiveness, Black females tend to lose less weight than White counterparts. The higher prevalence of chronic psychological stress reported by Black females may contribute to their disproportionate prevalence of obesity and observed suboptimal weight loss. This study will examine the effectiveness of a 12-month culturally-targeted, stress management-enhanced BWL intervention on weight loss and stress reduction among Black females in a fully-powered randomized, controlled trial.MethodsAdult Black females with obesity (n = 340) will be randomized to either a culturally targeted stress management-enhanced BWL intervention (BWL-Stress) or the same BWL intervention alone (BWL-alone). The primary outcome is weight change at month 6. Secondary outcomes will include changes in stress measures (e.g., perceived stress, cortisol), energy intake, and physical activity at month 6. We will also assess process measures (e.g., treatment adherence, treatment burden). Each outcome will also be evaluated at month 12 to assess longer-term effects of the intervention.DiscussionThis novel approach for enhancing an evidence-based BWL program with culturally-targeted stress management strategies for Black females addresses an understudied barrier to effective weight management among a population at high risk for obesity and obesity-related chronic diseases. This study will potentially elucidate psychological or behavioral mechanisms linking our novel intervention to study outcomes. If the intervention is proven to be effective, this study will have significant clinical and public health implications for weight management among Black females.Trial registrationThis study was registered on ClinicalTrials.gov , identifier NCT04335799t , on April 6, 2020.

In this study, peripheral blood mononuclear cells from young and old patients with COVID-19 were examined phenotypically, transcriptionally and functionally to reveal age-, time- and severity-specific adaptations. Gene signatures within memory B cells and plasmablasts correlated with reduced frequency of antigen-specific B cells and neutralizing antibodies in older patients with severe COVID-19. Moreover, these patients exhibited exacerbated T cell lymphopenia, which correlated with lower plasma interleukin-2, and diminished antigen-specific T cell responses. Single-cell RNA sequencing revealed augmented signatures of activation, exhaustion, cytotoxicity and type I interferon signaling in memory T and natural killer cells with age. Although cytokine storm was evident in both age groups, older individuals exhibited elevated levels of myeloid cell recruiting factors. Furthermore, we observed redistribution of monocyte and dendritic cell subsets and emergence of a suppressive phenotype with severe disease, which was reversed only in young patients over time. This analysis provides new insights into the impact of aging on COVID-19.

BackgroundEarly evaluations of healthcare professional (HCP) COVID-19 risk occurred during insufficient personal protective equipment and disproportionate testing, contributing to perceptions of high patient-care related HCP risk. We evaluated HCP COVID-19 seropositivity after accounting for community factors and coworker outbreaks.MethodsPrior to universal masking, we conducted a single-center retrospective cohort plus cross-sectional study. All HCP (1) seen by Occupational Health for COVID-like symptoms (regardless of test result) or assigned to (2) dedicated COVID-19 units, (3) units with a COVID-19 HCP outbreak, or (4) control units from 01/01/2020 to 04/15/2020 were offered serologic testing by an FDA-authorized assay plus a research assay against 67 respiratory viruses, including 11 SARS-CoV-2 antigens. Multivariable models assessed the association of demographics, job role, comorbidities, care of a COVID-19 patient, and geocoded socioeconomic status with positive serology.ResultsOf 654 participants, 87 (13.3%) were seropositive; among these 60.8% (N = 52) had never cared for a COVID-19 patient. Being male (OR 1.79, CI 1.05-3.04, p = 0.03), working in a unit with a HCP-outbreak unit (OR 2.21, CI 1.28-3.81, p

AbstractRapid Access Ice Drill is a new drilling technology capable of quickly accessing the glacial bed of Antarctic ice sheets, retrieving ice core and rock core samples, and providing boreholes for downhole logging of physical properties. Scientific goals include searching for old ice near the glacial bed and sampling subglacial bedrock. During field trials near McMurdo Station on a piedmont glacier at Minna Bluff in the 2019–20 austral summer, we successfully completed a ‘top-to-bottom’ operational sequence in three boreholes by (1) augering through firn, (2) creating a borehole packer seal in non-porous ice, (3) establishing fluid circulation, (4) quickly drilling a borehole in ice at penetration rates up to 1.2 m min−1, (5) acquiring a short ice core at depth, (6) penetrating the glacial bed at a depth of ~677 m, (7) recovering a 3.2 m core of ice, basal till and subglacial bedrock, (8) optically logging the borehole on wireline, (9) testing hydrofracture potential by overpressuring the borehole fluid and (10) operating in an environmentally benign yet rapid field mode. Minna Bluff testing, therefore, demonstrates the effectiveness of this integrated system to drill rapidly through thick ice and penetrate across the glacial bed to take cores of bedrock.

AbstractRapid Access Ice Drill is a new drilling technology capable of quickly accessing the glacial bed of Antarctic ice sheets, retrieving ice core and rock core samples, and providing boreholes for downhole logging of physical properties. Scientific goals include searching for old ice near the glacial bed and sampling subglacial bedrock. During field trials near McMurdo Station on a piedmont glacier at Minna Bluff in the 2019–20 austral summer, we successfully completed a ‘top-to-bottom’ operational sequence in three boreholes by (1) augering through firn, (2) creating a borehole packer seal in non-porous ice, (3) establishing fluid circulation, (4) quickly drilling a borehole in ice at penetration rates up to 1.2 m min−1, (5) acquiring a short ice core at depth, (6) penetrating the glacial bed at a depth of ~677 m, (7) recovering a 3.2 m core of ice, basal till and subglacial bedrock, (8) optically logging the borehole on wireline, (9) testing hydrofracture potential by overpressuring the borehole fluid and (10) operating in an environmentally benign yet rapid field mode. Minna Bluff testing, therefore, demonstrates the effectiveness of this integrated system to drill rapidly through thick ice and penetrate across the glacial bed to take cores of bedrock.

Chlamydia trachomatis is a sexually transmitted bacterium that infects over 130 million individuals worldwide annually. To implement a vaccine, we developed a cell-free co-translational system to express the Chlamydia muridarum major outer membrane protein (MOMP). This approach uses a nanolipoprotein particles (tNLP) made from ApoA1 protein, amphiphilic telodendrimer and lipids that self-assemble to form 10-25 nm discs. These tNLP provide a protein-encapsulated lipid support to solubilize and fold membrane proteins. The cell-free system co-translated MOMP and ApoA1 in the presence of telodendrimer mixed with lipids. The MOMP-tNLP complex was amenable to CpG and FSL-1 adjuvant addition. To investigate the ability of MOMP-tNLP+CpG+FSL-1 to induce protection against an intranasal (i.n.) C. muridarum challenge, female mice were vaccinated intramuscularly (i.m.) or i.n. and i.m. simultaneously 4 weeks apart. Following vaccination with MOMP-tNLP+CpG+FSL-1, mice mounted significant humoral and cell-mediated immune responses. Following the i.n. challenge, mice vaccinated with MOMP-tNLP+CpG+FSL-1 i.n. + i.m. group were protected as determined by the percentage change in body weight and by the number of C. muridarum inclusion forming units (IFU) recovered from the lungs. To our knowledge, this is the first time a MOMP-based vaccine formulated in tNLP has been shown to protect against C. muridarum.

The WNT signaling system plays an important but paradoxical role in the regulation of pluripotency. In the cow, IWR-1, which inhibits canonical WNT activation and has WNT-independent actions, promotes the derivation of primed pluripotent embryonic stem cells from the blastocyst. Here, we describe a series of experiments to determine whether derivation of embryonic stem cells could be generated by replacing IWR-1 with other inhibitors of WNT signaling. Results confirm the importance of inhibition of canonical WNT signaling for the establishment of pluripotent embryonic stem cells in cattle and indicate that the actions of IWR-1 can be mimicked by the WNT secretion inhibitor IWP2 but not by the tankyrase inhibitor XAV939 or WNT inhibitory protein dickkopf 1. The role of Janus kinase-mediated signaling pathways for the maintenance of pluripotency of embryonic stem cells was also evaluated. Maintenance of pluripotency of embryonic stem cells lines was blocked by a broad inhibitor of Janus kinase, even though the cells did not express phosphorylated signal transducer and activator of transcription 3 (pSTAT3). Further studies with blastocysts indicated that IWR-1 blocks the activation of pSTAT3. A likely explanation is that IWR-1 blocks differentiation of embryonic stem cells into a pSTAT3+ lineage. In conclusion, results presented here indicate the importance of inhibition of WNT signaling for the derivation of pluripotent bovine embryonic stem cells, the role of Janus kinase signaling for maintenance of pluripotency, and the participation of IWR-1 in the inhibition of activation of STAT3.

BACKGROUND & AIMS: The prevalence and significance of digestive manifestations in coronavirus disease 2019 (COVID-19) remain uncertain. We aimed to assess the prevalence, spectrum, severity, and significance of digestive manifestations in patients hospitalized with COVID-19. METHODS: Consecutive patients hospitalized with COVID-19 were identified across a geographically diverse alliance of medical centers in North America. Data pertaining to baseline characteristics, symptomatology, laboratory assessment, imaging, and endoscopic findings from the time of symptom onset until discharge or death were abstracted manually from electronic health records to characterize the prevalence, spectrum, and severity of digestive manifestations. Regression analyses were performed to evaluate the association between digestive manifestations and severe outcomes related to COVID-19. RESULTS: A total of 1992 patients across 36 centers met eligibility criteria and were included. Overall, 53% of patients experienced at least 1 gastrointestinal symptom at any time during their illness, most commonly diarrhea (34%), nausea (27%), vomiting (16%), and abdominal pain (11%). In 74% of cases, gastrointestinal symptoms were judged to be mild. In total, 35% of patients developed an abnormal alanine aminotransferase or total bilirubin level; these were increased to less than 5 times the upper limit of normal in 77% of cases. After adjusting for potential confounders, the presence of gastrointestinal symptoms at any time (odds ratio, 0.93; 95% CI, 0.76-1.15) or liver test abnormalities on admission (odds ratio, 1.31; 95% CI, 0.80-2.12) were not associated independently with mechanical ventilation or death. CONCLUSIONS: Among patients hospitalized with COVID-19, gastrointestinal symptoms and liver test abnormalities were common, but the majority were mild and their presence was not associated with a more severe clinical course.

ObjectivesTo investigate if the nicotine metabolite ratio (NMR, the ratio of nicotine metabolites 3'-hydroxycotinine/cotinine) is a reliable phenotypic biomarker for nicotine clearance across races, and as a function of differences in the rate of nicotine, cotinine and 3'-hydroxycotinine glucuronidation and UGT genotypes.MethodsParticipants [Caucasians (Whites), African Americans (Blacks) and Asian-Americans (Asians)] received an oral solution of deuterium-labeled nicotine and its metabolite cotinine. Plasma and saliva concentrations of nicotine and cotinine were used to determine oral clearances. Rates of glucuronidation were assessed from urine glucuronide/parent ratios, and UGT2B10 and UGT2B17 genotypes from DNA.ResultsAmong the 227 participants, 96 (42%) were White, 67 (30%) Asian and 64 (28%) Black. Compared to the other two races, Whites had higher nicotine and cotinine total oral clearance, Blacks had lower nicotine and cotinine glucuronidation rates and Asians had lower 3'-hydroxycotinine glucuronidation rates. A strong positive correlation (correlations coefficients 0.77-0.84; P

ObjectivesTo investigate if the nicotine metabolite ratio (NMR, the ratio of nicotine metabolites 3'-hydroxycotinine/cotinine) is a reliable phenotypic biomarker for nicotine clearance across races, and as a function of differences in the rate of nicotine, cotinine and 3'-hydroxycotinine glucuronidation and UGT genotypes.MethodsParticipants [Caucasians (Whites), African Americans (Blacks) and Asian-Americans (Asians)] received an oral solution of deuterium-labeled nicotine and its metabolite cotinine. Plasma and saliva concentrations of nicotine and cotinine were used to determine oral clearances. Rates of glucuronidation were assessed from urine glucuronide/parent ratios, and UGT2B10 and UGT2B17 genotypes from DNA.ResultsAmong the 227 participants, 96 (42%) were White, 67 (30%) Asian and 64 (28%) Black. Compared to the other two races, Whites had higher nicotine and cotinine total oral clearance, Blacks had lower nicotine and cotinine glucuronidation rates and Asians had lower 3'-hydroxycotinine glucuronidation rates. A strong positive correlation (correlations coefficients 0.77-0.84; P

Over the last two decades, there have been three deadly human outbreaks of coronaviruses (CoVs) caused by SARS-CoV, MERS-CoV, and SARS-CoV-2, which has caused the current COVID-19 global pandemic. All three deadly CoVs originated from bats and transmitted to humans via various intermediate animal reservoirs. It remains highly possible that other global COVID pandemics will emerge in the coming years caused by yet another spillover of a bat-derived SARS-like coronavirus (SL-CoV) into humans. Determining the Ag and the human B cells, CD4+ and CD8+ T cell epitope landscapes that are conserved among human and animal coronaviruses should inform in the development of future pan-coronavirus vaccines. In the current study, using several immunoinformatics and sequence alignment approaches, we identified several human B cell and CD4+ and CD8+ T cell epitopes that are highly conserved in 1) greater than 81,000 SARS-CoV-2 genome sequences identified in 190 countries on six continents; 2) six circulating CoVs that caused previous human outbreaks of the common cold; 3) nine SL-CoVs isolated from bats; 4) nine SL-CoV isolated from pangolins; 5) three SL-CoVs isolated from civet cats; and 6) four MERS strains isolated from camels. Furthermore, the identified epitopes: 1) recalled B cells and CD4+ and CD8+ T cells from both COVID-19 patients and healthy individuals who were never exposed to SARS-CoV-2, and 2) induced strong B cell and T cell responses in humanized HLA-DR1/HLA-A*02:01 double-transgenic mice. The findings pave the way to develop a preemptive multiepitope pan-coronavirus vaccine to protect against past, current, and future outbreaks.

Effective methods for predicting COVID-19 disease trajectories are urgently needed. Here, enzyme-linked immunosorbent assay (ELISA) and coronavirus antigen microarray (COVAM) analysis mapped antibody epitopes in the plasma of COVID-19 patients (n = 86) experiencing a wide range of disease states. The experiments identified antibodies to a 21-residue epitope from nucleocapsid (termed Ep9) associated with severe disease, including admission to the intensive care unit (ICU), requirement for ventilators, or death. Importantly, anti-Ep9 antibodies can be detected within 6 days post-symptom onset and sometimes within 1 day. Furthermore, anti-Ep9 antibodies correlate with various comorbidities and hallmarks of immune hyperactivity. We introduce a simple-to-calculate, disease risk factor score to quantitate each patient's comorbidities and age. For patients with anti-Ep9 antibodies, scores above 3.0 predict more severe disease outcomes with a 13.42 likelihood ratio (96.7% specificity). The results lay the groundwork for a new type of COVID-19 prognostic to allow early identification and triage of high-risk patients. Such information could guide more effective therapeutic intervention.IMPORTANCE The COVID-19 pandemic has resulted in over two million deaths worldwide. Despite efforts to fight the virus, the disease continues to overwhelm hospitals with severely ill patients. Diagnosis of COVID-19 is readily accomplished through a multitude of reliable testing platforms; however, prognostic prediction remains elusive. To this end, we identified a short epitope from the SARS-CoV-2 nucleocapsid protein and also a disease risk factor score based upon comorbidities and age. The presence of antibodies specifically binding to this epitope plus a score cutoff can predict severe COVID-19 outcomes with 96.7% specificity.

Effective methods for predicting COVID-19 disease trajectories are urgently needed. Here, enzyme-linked immunosorbent assay (ELISA) and coronavirus antigen microarray (COVAM) analysis mapped antibody epitopes in the plasma of COVID-19 patients (n = 86) experiencing a wide range of disease states. The experiments identified antibodies to a 21-residue epitope from nucleocapsid (termed Ep9) associated with severe disease, including admission to the intensive care unit (ICU), requirement for ventilators, or death. Importantly, anti-Ep9 antibodies can be detected within 6 days post-symptom onset and sometimes within 1 day. Furthermore, anti-Ep9 antibodies correlate with various comorbidities and hallmarks of immune hyperactivity. We introduce a simple-to-calculate, disease risk factor score to quantitate each patient's comorbidities and age. For patients with anti-Ep9 antibodies, scores above 3.0 predict more severe disease outcomes with a 13.42 likelihood ratio (96.7% specificity). The results lay the groundwork for a new type of COVID-19 prognostic to allow early identification and triage of high-risk patients. Such information could guide more effective therapeutic intervention.IMPORTANCE The COVID-19 pandemic has resulted in over two million deaths worldwide. Despite efforts to fight the virus, the disease continues to overwhelm hospitals with severely ill patients. Diagnosis of COVID-19 is readily accomplished through a multitude of reliable testing platforms; however, prognostic prediction remains elusive. To this end, we identified a short epitope from the SARS-CoV-2 nucleocapsid protein and also a disease risk factor score based upon comorbidities and age. The presence of antibodies specifically binding to this epitope plus a score cutoff can predict severe COVID-19 outcomes with 96.7% specificity.

BackgroundThe estimated 2.2 million people who inject drugs (PWID) in the USA experience significant gaps in preventive healthcare and a high burden of infectious, psychiatric, and other chronic diseases. Many PWID rely on emergency medical services, which are costly and not designed to deliver preventive services, manage chronic conditions, or address social needs.ObjectiveThe objective of this study was to explore barriers and facilitators to primary care utilization from the perspectives of PWID in New England, a region highly affected by the overdose crisis.DesignParticipants completed semi-structured qualitative interviews exploring substance use and healthcare utilization patterns.ParticipantsWe recruited 78 PWID through community-based organizations (e.g., syringe service programs) in 16 urban and non-urban communities throughout Massachusetts and Rhode Island.ApproachThematic analysis identified barriers and facilitators to primary care utilization at the individual, interpersonal, and systemic levels.Key resultsAmong 78 PWID, 48 described recent primary care experiences; 33 had positive experiences and 15 described negative experiences involving discrimination or mistrust. Individual-level barriers to primary care utilization included perceived lack of need and competing priorities (e.g., avoiding opioid withdrawal, securing shelter beds). Interpersonal-level barriers included stigma and perceived low quality of care for PWID. Systemic-level barriers included difficulty navigating healthcare systems, inadequate transportation, long wait times, and frequent provider turnover. Participants with positive primary care experiences explained how appointment reminders, flexible hours, addiction medicine-trained providers, case management services, and transportation support facilitated primary care utilization and satisfaction.ConclusionsFindings regarding the multilevel barriers and facilitators to accessing primary care among PWID identify potential targets for programmatic interventions to improve primary care utilization in this population. Based on these findings, we make recommendations for improving the engagement of PWID in primary care as a means to advance individual and public health outcomes.

Bovine embryonic stem cells (bESCs) extend the lifespan of the transient pluripotent bovine inner cell mass in vitro. After years of research, derivation of stable bESCs was only recently reported. Although successful, bESC culture relies on complex culture conditions that require a custom-made base medium and mouse embryonic fibroblasts (MEF) feeders, limiting the widespread use of bESCs. We report here simplified bESC culture conditions based on replacing custom base medium with a commercially available alternative and eliminating the need for MEF feeders by using a chemically-defined substrate. bESC lines were cultured and derived using a base medium consisting of N2B27 supplements and 1% BSA (NBFR-bESCs). Newly derived bESC lines were easy to establish, simple to propagate and stable after long-term culture. These cells expressed pluripotency markers and actively proliferated for more than 35 passages while maintaining normal karyotype and the ability to differentiate into derivatives of all three germ lineages in embryoid bodies and teratomas. In addition, NBFR-bESCs grew for multiple passages in a feeder-free culture system based on vitronectin and Activin A medium supplementation while maintaining pluripotency. Simplified conditions will facilitate the use of bESCs for gene editing applications and pluripotency and lineage commitment studies.

Chlamydia trachomatis is a sexually transmitted bacterium that infects over 130 million individuals worldwide annually. To implement a vaccine, we developed a cell-free co-translational system to express the Chlamydia muridarum major outer membrane protein (MOMP). This approach uses a nanolipoprotein particles (tNLP) made from ApoA1 protein, amphiphilic telodendrimer and lipids that self-assemble to form 10-25 nm discs. These tNLP provide a protein-encapsulated lipid support to solubilize and fold membrane proteins. The cell-free system co-translated MOMP and ApoA1 in the presence of telodendrimer mixed with lipids. The MOMP-tNLP complex was amenable to CpG and FSL-1 adjuvant addition. To investigate the ability of MOMP-tNLP+CpG+FSL-1 to induce protection against an intranasal (i.n.) C. muridarum challenge, female mice were vaccinated intramuscularly (i.m.) or i.n. and i.m. simultaneously 4 weeks apart. Following vaccination with MOMP-tNLP+CpG+FSL-1, mice mounted significant humoral and cell-mediated immune responses. Following the i.n. challenge, mice vaccinated with MOMP-tNLP+CpG+FSL-1 i.n. + i.m. group were protected as determined by the percentage change in body weight and by the number of C. muridarum inclusion forming units (IFU) recovered from the lungs. To our knowledge, this is the first time a MOMP-based vaccine formulated in tNLP has been shown to protect against C. muridarum.

We analyzed data from two ongoing COVID-19 longitudinal serological surveys in Orange County, CA., between April 2020 and March 2021. A total of 8476 finger stick blood specimens were collected before and after a vaccination campaign. IgG levels were determined using a multiplex antigen microarray containing antigens from SARS-CoV-2, SARS, MERS, Common CoV, and Influenza. Twenty-six percent of specimens from unvaccinated Orange County residents in December 2020 were SARS-CoV-2 seropositive; out of 852 seropositive individuals 77 had symptoms and 9 sought medical care. The antibody response was predominantly against nucleocapsid (NP), full length, and S2 domain of spike. Anti-receptor binding domain (RBD) reactivity was low and not cross-reactive against SARS S1 or SARS RBD. A vaccination campaign at the University of California Irvine Medical Center (UCIMC) started on December, 2020 and 6724 healthcare workers were vaccinated within 3 weeks. Seroprevalence increased from 13% pre-vaccination to 79% post-vaccination in January, 93% in February, and 99% in March. mRNA vaccination induced higher antibody levels than natural exposure, especially against the RBD domain and cross-reactivity against SARS RBD and S1 was observed. Nucleocapsid protein antibodies can be used to distinguish vaccinees to classify pre-exposure to SARS-CoV-2 Previously infected individuals developed higher antibody titers to the vaccine than non pre-exposed individuals. Hospitalized patients in intensive care with severe disease reach significantly higher antibody levels than mild cases, but lower antibody levels compared to the vaccine. These results indicate that mRNA vaccination rapidly induces a much stronger and broader antibody response than SARS-CoV-2 infection.