Bone marrow disease in rhabdomyosarcoma visualized by 2-[18F]fluorodeoxyglucose positron emission tomography/computed tomography.

Bone marrow metastases-noted in 6% of patients with rhabdomyosarcoma-have been linked to very poor outcomes. Bilateral bone marrow sampling from iliac crests has been the gold standard for bone marrow examination in rhabdomyosarcoma, but sampling errors due to patchy bone marrow involvement may limit its sensitivity. Here, we report the case of a 6-year-old boy with embryonal rhabdomyosarcoma of the skull base and multiple 2-[18F]fluoro-2-deoxy-D-glucose (2-[18F]FDG)-avid bone marrow metastases visualized by positron emission tomography and computed tomography (2-[18F]FDG PET/CT). His bone marrow aspirates were tumor-free. This case illustrates the diagnostic value of 2-[18F]FDG PET/CT in the detection of bone marrow metastases in rhabdomyosarcoma patients, which may re-shape the definition of bone marrow disease and, ultimately, alter disease staging and risk stratification.

Engineering Material Properties of Transcription Factor Condensates to Control Gene Expression in Mammalian Cells and Mice.

Phase separation of biomolecules into condensates is a key mechanism in the spatiotemporal organization of biochemical processes in cells. However, the impact of the material properties of biomolecular condensates on important processes, such as the control of gene expression, remains largely elusive. Here, the material properties of optogenetically induced transcription factor condensates are systematically tuned, and probed for their impact on the activation of target promoters. It is demonstrated that transcription factors in rather liquid condensates correlate with increased gene expression levels, whereas stiffer transcription factor condensates correlate with the opposite effect, reduced activation of gene expression. The broad nature of these findings is demonstrated in mammalian cells and mice, as well as by using different synthetic and natural transcription factors. These effects are observed for both transgenic and cell-endogenous promoters. The findings provide a novel materials-based layer in the control of gene expression, which opens novel opportunities in optogenetic engineering and synthetic biology.

Maternal Sleep Health, Social Support, and Distress: A Mixed-Methods Analysis of Mothers of Infants and Young Children in Rural US.

OBJECTIVES: The purpose of this study was to explore sleep health in rural maternal populations through a social-ecological framework and identify risk and protective factors for this population. METHODS: 39 individuals who are mothers of infants or children under the age of 5 years completed an online survey, 35 of which completed a subsequent semi-structured interview. Recruitment was limited to one rural community and was in partnership with community healthcare providers. Results were integrated using a convergent, parallel mixed-methods design. RESULTS: Poor sleep health and high prevalence of insomnia symptoms in rural mothers were evident and associated with social support and maternal distress. Qualitative content from interviews indicated that well-established precipitating and perpetuating factors for insomnia may contribute to poor maternal sleep health. Results also revealed a gap in knowledge and language surrounding sleep health among rural mothers. CONCLUSIONS: Sleep health is challenged during the transition to motherhood and rural mothers have less access to specialized perinatal and behavioral health care than their urban counterparts. In this sample, poor sleep was attributable to distress in addition to nocturnal infant and child sleep patterns which has implications for psychoeducation and promotion of sleep health in mothers. Sleep is a modifiable health indicator that is associated with several other maternal health outcomes and should be considered an element of a comprehensive maternal health for prevention and intervention across individual, interpersonal, and societal domains of the social-ecological model of sleep health.

Synergistic Cytotoxicity of a Toxin Targeting the Epidermal Growth Factor Receptor and the Glycosylated Triterpenoid SO1861 in Prostate Cancer.

Treatment of advanced prostate cancer lacks specificity and curative intent. Therefore, the need for new targeted therapeutic approaches is high. In the present study, we generated the new targeted toxin EGF-PE24mutΔREDLK binding to the epidermal growth factor receptor (EGFR) on the surface of prostate cancer cells. It consists of the human epidermal growth factor (EGF) as binding domain and a de-immunized variant of Pseudomonas Exotoxin A (PE), called PE24mutΔREDLK, as toxin domain. The toxin domain contains a deletion of the C-terminal KDEL-like motif REDLK to prevent its transport from sorting endosomes via the KDEL receptor mediated pathway into the cytosol, where it can inhibit cellular protein biosynthesis and induce apoptosis. Indeed, REDLK deletion resulted in a strong decrease in cytotoxicity of the targeted toxin in prostate cancer cells compared to the parental targeted toxin EGF-PE24mut. However, addition of the plant glycosylated triterpenoid SO1861, which is known to mediate the release of biomolecules from endolysosomal compartments into the cytosol, resulted in an up to almost 7,000-fold enhanced synergistic cytotoxicity. Moreover, combination of PE24mutΔREDLK with SO1861 led to a cytotoxicity that was even 16- to 300-fold enhanced compared to that of EGF-PE24mut. Endolysosomal entrapment of the non-toxic targeted toxin EGF-PE24mutΔREDLK followed by activation through enhanced endosomal escape therefore represents a new promising approach for the future treatment of advanced prostate cancer with high efficacy and diminished side effects.

Impaired systemic nucleocapsid antigen clearance in severe COVID-19.

The circulating nucleocapsid (NCP) antigen of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is detectable in coronavirus disease-2019 (COVID-19) patients. To better understand the biology of disease severity, we investigated NCP clearance kinetics in hospitalized COVID-19 patients. Serum NCP was quantified using a commercial NCP-specific enzyme-linked immunoassay in hospitalized COVID-19 patients (n = 63) during their hospital stay. Results were correlated to COVID-19 disease severity, inflammation parameters, antibody response, and results of SARS-CoV-2 PCR from nasopharyngeal swabs. We demonstrate that NCP antigen levels in serum remained elevated in 21/45 (46.7%) samples from patients in intensive care units (ICU) after >8 days postdiagnosis. The proportion of ICU patients with detectable antigenemia declined only gradually from 84.6% to 25.0% over several weeks. This was in contrast to complete NCP clearance in all non-ICU patients after 8 days, and also in contrast to mucosal clearance of the virus as measured by PCR. Antigen clearance was associated with higher IgG against S1 but not NCP. Clearance of NCP antigenemia is delayed in >40% of severely ill COVID-19 patients. Thus, NCP antigenemia detected after 8 days post COVID-19 diagnosis is a characteristic of patients requiring intensive care. Prospective trials should further investigate NCP antigen clearance kinetics as a mechanistic biomarker.

Real-time monitoring of cell surface protein arrival with split luciferases.

Each cell in a multicellular organism permanently adjusts the concentration of its cell surface proteins. In particular, epithelial cells tightly control the number of carriers, transporters and cell adhesion proteins at their plasma membrane. However, sensitively measuring the cell surface concentration of a particular protein of interest in live cells and in real time represents a considerable challenge. Here, we introduce a novel approach based on split luciferases, which uses one luciferase fragment as a tag on the protein of interest and the second fragment as a supplement to the extracellular medium. Once the protein of interest arrives at the cell surface, the luciferase fragments complement and generate luminescence. We compared the performance of split Gaussia luciferase and split Nanoluciferase by using a system to synchronize biosynthetic trafficking with conditional aggregation domains. The best results were achieved with split Nanoluciferase, for which luminescence increased more than 6000-fold upon recombination. Furthermore, we showed that our approach can separately detect and quantify the arrival of membrane proteins at the apical and basolateral plasma membrane in single polarized epithelial cells by detecting the luminescence signals with a microscope, thus opening novel avenues for characterizing the variations in trafficking in individual epithelial cells.

Soy Extract, Rich in Hydroxylated Isoflavones, Exhibits Antidiabetic Properties In Vitro and in Drosophila melanogaster In Vivo.

In the context of the growing prevalence of type 2 diabetes (T2DM), control of postprandial hyperglycemia is crucial for its prevention. Blood glucose levels are determined by various factors including carbohydrate hydrolyzing enzymes, the incretin system and glucose transporters. Furthermore, inflammatory markers are recognized predictors of diabetes outcome. Although there is some evidence that isoflavones may exhibit anti-diabetic properties, little is known about to what extent their corresponding hydroxylated metabolites may affect glucose metabolism. We evaluated the ability of a soy extract before (pre-) and after (post-) fermentation to counteract hyperglycemia in vitro and in Drosophila melanogaster in vivo. Fermentation with Aspergillus sp. JCM22299 led to an enrichment of hydroxy-isoflavones (HI), including 8-hydroxygenistein, 8-hydroxyglycitein and 8-hydroxydaidzein, accompanied by an enhanced free radical scavenging activity. This HI-rich extract demonstrated inhibitory activity towards α-glucosidase and a reduction of dipeptidyl peptidase-4 enzyme activity. Both the pre- and post-fermented extracts significantly inhibited the glucose transport via sodium-dependent glucose transporter 1. Furthermore, the soy extracts reduced c-reactive protein mRNA and secreted protein levels in interleukin-stimulated Hep B3 cells. Finally, supplementation of a high-starch D. melanogaster diet with post-fermented HI-rich extract decreased the triacylglyceride content of female fruit flies, confirming its anti-diabetic properties in an in vivo model.

Stabilization of membrane topologies by proteinaceous remorin scaffolds.

In plants, the topological organization of membranes has mainly been attributed to the cell wall and the cytoskeleton. Additionally, few proteins, such as plant-specific remorins have been shown to function as protein and lipid organizers. Root nodule symbiosis requires continuous membrane re-arrangements, with bacteria being finally released from infection threads into membrane-confined symbiosomes. We found that mutations in the symbiosis-specific SYMREM1 gene result in highly disorganized perimicrobial membranes. AlphaFold modelling and biochemical analyses reveal that SYMREM1 oligomerizes into antiparallel dimers and may form a higher-order membrane scaffolding structure. This was experimentally confirmed when expressing this and other remorins in wall-less protoplasts is sufficient where they significantly alter and stabilize de novo membrane topologies ranging from membrane blebs to long membrane tubes with a central actin filament. Reciprocally, mechanically induced membrane indentations were equally stabilized by SYMREM1. Taken together we describe a plant-specific mechanism that allows the stabilization of large-scale membrane conformations independent of the cell wall.

Minerals and Trace Elements in 990 Beverages and Their Contribution to Dietary Reference Values for German Consumers.

Beverages are an integral part of human nutrition, yet little is known about their contribution to daily intakes of minerals and trace elements in German consumers. Using inductively coupled plasma-mass spectrometry, we determined the concentration of five minerals and six trace elements in beverage samples (n = 990, assigned to different beverage groups) collected throughout Germany. For a calculation of their relative contribution to the mineral supply, available beverage consumption data was combined with our quantitative analysis to calculate the average contribution of beverage groups to meet the respective dietary reference values currently used in Germany, Austria and Switzerland (D-A-CH region). Based on their presence in beverages and their consumption, the top three minerals are phosphorous, calcium and magnesium, and they, therefore, may reasonably contribute to the reference values. Among the trace elements, beverages mostly contributed to the manganese supply, whereas at the same time, concentrations of iron, cobalt and copper were low across all tested groups. Our study provides an overview of the assumed mineral and trace element intake via beverages in Germany and may, thus, serve as a foundation for a mineral and trace element database of beverages that needs to be expanded in the future.

Socio-Ecological Context of Sleep: Gender Differences and Couples' Relationships as Exemplars.

PURPOSE OF REVIEW: We summarized recent findings on insufficient sleep and insomnia, two prominent sleep issues that impact public health. We demonstrate the socio-ecologial impact of sleep health with findings on gender and couples' relationships as exemplars. RECENT FINDINGS: Robust gender differences in sleep duration and insomnia are due to biological and socio-ecological factors. Gender differences in insufficient sleep vary by country of origin and age whereas gender differences in insomnia reflect minoritized identities (e.g., sexual, gender). Co-sleeping with a partner is associated with longer sleep and more awakenings. Gender differences and couples' sleep were affected by intersecting social and societal influences, which supports a socio-ecological approach to sleep. Recent and seminal contributions to sleep health highlight the importance of observing individual sleep outcomes in a socio-ecological context. Novel methodology, such as global measures of sleep health, can inform efforts to improve sleep and, ultimately, public health.

Shedding light on current trends in molecular optogenetics.

Molecular optogenetics is a highly dynamic research field. In the past two years, the field was characterized by the development of new allosteric switches as well as the forward integration of optogenetics research towards application. Further, two areas of research have significantly gathered momentum, the use of optogenetics to control liquid-liquid phase separation as well as the application of optogenetic tools in the extracellular space. Here, we review these areas and discuss future directions.

Epidermal Growth Factor Based Targeted Toxin for the Treatment of Bladder Cancer.

BACKGROUND/AIM: Reports on over-expression of the epidermal growth factor receptor (EGFR) in bladder cancer and its function in tumorigenesis have suggested to target this antigen. MATERIALS AND METHODS: We generated the targeted toxin EGF-PE40 consisting of the human epidermal growth factor (EGF) as the binding domain and PE40, a truncated version of Pseudomonas Exotoxin A, as the toxin domain. EGF-PE40 was tested on EGFR-expressing bladder cancer cells in view of binding via flow cytometry, and cytotoxicity via WST viability assay. Induction of apoptosis was examined by western blot. RESULTS: The targeted toxin specifically triggered cytotoxicity in the bladder cancer cells with 50% inhibitory concentration (IC50) values in the low nanomolar or picomolar range, and was about 1,250- to 1,500-fold more cytotoxic than the EGFR inhibitor erlotinib. Cytotoxicity of EGF-PE40 was based on the induction of apoptosis. CONCLUSION: EGF-PE40 represents a promising candidate for the future treatment of bladder cancer.

Liquid-liquid phase separation of light-inducible transcription factors increases transcription activation in mammalian cells and mice.

Light-inducible gene switches represent a key strategy for the precise manipulation of cellular events in fundamental and applied research. However, the performance of widely used gene switches is limited due to low tissue penetrance and possible phototoxicity of the light stimulus. To overcome these limitations, we engineer optogenetic synthetic transcription factors to undergo liquid-liquid phase separation in close spatial proximity to promoters. Phase separation of constitutive and optogenetic synthetic transcription factors was achieved by incorporation of intrinsically disordered regions. Supported by a quantitative mathematical model, we demonstrate that engineered transcription factor droplets form at target promoters and increase gene expression up to fivefold. This increase in performance was observed in multiple mammalian cells lines as well as in mice following in situ transfection. The results of this work suggest that the introduction of intrinsically disordered domains is a simple yet effective means to boost synthetic transcription factor activity.

Social-ecological considerations for the sleep health of rural mothers.

Using a social-ecological framework, we identify social determinants that interact to influence sleep health, identify gaps in the literature, and make recommendations for targeting sleep health in rural mothers. Rural mothers experience unique challenges and protective factors in maintaining adequate sleep health during the postpartum and early maternal years. Geographic isolation, barriers to comprehensive behavioral medicine services, and intra-rural ethno-racial disparities are discussed at the societal (e.g., public policy), social (e.g., community) and individual levels (e.g., stress) of the social-ecological model. Research on sleep health would benefit from attention to methodological considerations of factors affecting rural mothers such as including parity in population-level analyses or applying community-based participatory research principles. Future sleep health programs would benefit from using existing social support networks to disseminate sleep health information, integrating behavioral health services into clinical care frameworks, and tailoring culturally-appropriate Telehealth/mHealth programs to enhance the sleep health of rural mothers.

Pseudomonas Exotoxin A Based Toxins Targeting Epidermal Growth Factor Receptor for the Treatment of Prostate Cancer.

The epidermal growth factor receptor (EGFR) was found to be a valuable target on prostate cancer (PCa) cells. However, EGFR inhibitors mostly failed in clinical studies with patients suffering from PCa. We therefore tested the targeted toxins EGF-PE40 and EGF-PE24mut consisting of the natural ligand EGF as binding domain and PE40, the natural toxin domain of Pseudomonas Exotoxin A, or PE24mut, the de-immunized variant thereof, as toxin domains. Both targeted toxins were expressed in the periplasm of E.coli and evoked an inhibition of protein biosynthesis in EGFR-expressing PCa cells. Concentration- and time-dependent killing of PCa cells was found with IC50 values after 48 and 72 h in the low nanomolar or picomolar range based on the induction of apoptosis. EGF-PE24mut was found to be about 11- to 120-fold less toxic than EGF-PE40. Both targeted toxins were more than 600 to 140,000-fold more cytotoxic than the EGFR inhibitor erlotinib. Due to their high and specific cytotoxicity, the EGF-based targeted toxins EGF-PE40 and EGF-PE24mut represent promising candidates for the future treatment of PCa.

The transition from acute to persistent pain: the identification of distinct trajectories among women presenting to an emergency department.

Posttraumatic stress disorder (PTSD) symptoms and other negative psychosocial factors have been implicated in the transition from acute to persistent pain. Women (N = 375) who presented to an inner-city emergency department (ED) with complaints of acute pain were followed up for 3 months. They completed a comprehensive battery of questionnaires at an initial visit and provided ratings of pain intensity at the site of pain presented in the ED during 3 monthly phone calls. Latent class growth analyses were used to detect possible trajectories of change in pain intensity from the initial visit to 3 months later. A 3-trajectory solution was found, which identified 3 groups of participants. One group (early recovery; n = 93) had recovered to virtually no pain by the initial visit, whereas a second group (delayed recovery; n = 120) recovered to no pain only after 1 month. A third group (no recovery; n = 162) still reported elevated pain at 3 months after the ED visit. The no recovery group reported significantly greater PTSD symptoms, anger, sleep disturbance, and lower social support at the initial visit than both the early recovery and delayed recovery groups. Results suggest that women with high levels of PTSD symptoms, anger, sleep disturbance, and low social support who experience an acute pain episode serious enough to prompt an ED visit may maintain elevated pain at this pain site for at least 3 months. Such an array of factors may place women at an increased risk of developing persistent pain following acute pain.

Favorable outcomes of hematopoietic stem cell transplantation in children and adolescents with Diamond-Blackfan anemia.

Diamond-Blackfan anemia (DBA) is a congenital pure red cell aplasia associated with congenital abnormalities and cancer predisposition. Allogeneic hematopoietic stem cell transplantation (HSCT) can correct the hematological phenotype and is indicated in transfusion-dependent patients. In 70 children reported to the German DBA and French HSCT registries, HSCT was performed from 1985 to 2017. Median age at HSCT was 5.5 years (range, 0.9-17.3 years). Two-thirds of patients (64%) were transplanted from a matched sibling donor (MSD), and most procedures were performed after the year 1999 (73%). Primary engraftment was achieved in all patients. One patient developed secondary graft failure. Cumulative incidence of acute graft-versus-host disease (GVHD) was 24% for °II-IV (95% confidence interval [CI], 16% to 37%) and 7% for °III-IV (95% CI, 3% to 17%); cumulative incidence of chronic GVHD was 11% (95% CI, 5% to 22%). The probability of chronic GVHD-free survival (cGFS) was 87% (95% CI, 79% to 95%) and significantly improved over time (<2000: 68% [95% CI, 47% to 89%] vs ≥2000: 94% [95% CI, 87% to 100%], P < .01). cGFS was comparable following HSCT from a MSD and an unrelated donor (UD). Of note, no severe chronic GVHD or deaths were reported following MSD-HSCT after 1999. The difference of cGFS in children transplanted <10 years of age compared with older patients did not reach statistical significance (<10 years: 90% [95% CI, 81% to 99%] vs 10-18 years 78% [95% CI, 58% to 98%]). In summary, these data indicate that HSCT is efficient and safe in young DBA patients and should be considered if a MSD or matched UD is available. HSCT for transfusion dependency only must be critically discussed in older patients.

Supplementation with nitrate only modestly affects lipid and glucose metabolism in genetic and dietary-induced murine models of obesity.

To gain a better understanding of how nitrate may affect carbohydrate and lipid metabolism, female wild-type mice were fed a high-fat, high-fructose diet supplemented with either 0, 400, or 800 mg nitrate/kg diet for 28 days. Additionally, obese female db/db mice were fed a 5% fat diet supplemented with the same levels and source of nitrate. Nitrate decreased the sodium-dependent uptake of glucose by ileal mucosa in wild-type mice. Moreover, nitrate significantly decreased triglyceride content and mRNA expression levels of Pparγ in liver and Glut4 in skeletal muscle. Oral glucose tolerance as well as plasma cholesterol, triglyceride, insulin, leptin, glucose and the activity of ALT did not significantly differ between experimental groups but was higher in db/db mice than in wild-type mice. Nitrate changed liver fatty acid composition and mRNA levels of Fads only slightly. Further hepatic genes encoding proteins involved in lipid and carbohydrate metabolism were not significantly different between the three groups. Biomarkers of inflammation and autophagy in the liver were not affected by the different dietary treatments. Overall, the present data suggest that short-term dietary supplementation with inorganic nitrate has only modest effects on carbohydrate and lipid metabolism in genetic and dietary-induced mouse models of obesity.

High Dietary Kuding Tea Extract Supplementation Induces Hepatic Xenobiotic-Metabolizing Enzymes-A 6-Week Feeding Study in Mice.

Kuding tea (KT) is a traditional Chinese beverage rich in plant bioactives that may exhibit various health benefits. However, little is known about the safety of KT extract (KTE) when consumed long term at high doses as a dietary supplement. Therefore, in this study, we investigated aspects of the safety of KTE. Male C57BL/6 mice were fed a high-fat, high-fructose, Western-type diet (control) supplemented with either 12.88% γ-cyclodextrin (γCD), 7.12% KTE (comprising 0.15% ursolic acid, UA) encapsulated in 12.88% γCD (KTE-γCD), or 0.15% UA over a 6-week experimental period. The dietary treatments did not affect food intake, body weight or body composition. However, treatment with KTE-γCD, but not γCD and UA, increased liver weight and hepatic fat accumulation, which was accompanied by increased hepatic PPARγ and CD36 mRNA levels. KTE-γCD treatment elevated plasma cholesterol and CYP7A1 mRNA and protein levels compared to those in control mice. KTE-γCD substantially increased the mRNA and protein levels of hepatic CYP3A and GSTA1, which are central to the detoxification of drugs and xenobiotics. Furthermore, we observed a moderate elevation in hepatic CYP3A (5-fold change) and GSTA1 (1.7-fold change) mRNA levels in UA-fed mice. In vitro data collected in HepG2 cells indicated a dose-dependent increase in hepatic cytotoxicity in response to KTE treatment, which may have been partly mediated by UA. Overall, the present data may contribute to the safety assessment of KTE and suggest that KTE encapsulated in γCD affects liver fat storage and the hepatic phase I and phase II responses in mice.

Adherence to a Mediterranean-like Diet as a Protective Factor Against COPD: A Nested Case-Control Study.

A diet rich in nutrients has been suggested to have protective effects against the development of chronic obstructive pulmonary disease (COPD). Since the traditional Mediterranean diet is high in nutrients, including antioxidants, vitamins, and minerals, it is of interest to study as a protective factor against COPD. Our aim was therefore to study its associations with development of COPD using population-based prospective data from the Västerbotten Intervention Programme (VIP) cohort. Data on diet from 370 individuals, who later visited the Department of Medicine at the University Hospital, Umeå, Sweden, with a diagnosis of COPD, were compared to 1432 controls. Adherence to a Mediterranean diet was assessed by a modified version of the Mediterranean diet score (MDS). Cases were diagnosed with COPD 11.1 years (mean) (standard deviation [SD] 4.5 years) after first stating their dietary habits in the VIP at a mean age of 55.5 years (SD 6.6 years). Higher MDS was associated with a higher level of education and not living alone. After adjustment for co-habiting and education level, individuals with an intermediate MDS and those with the highest MDS had a lower odds of developing COPD (odds ratio [OR] 0.73, 95% confidence interval [CI] 0.56-0.95; OR 0.56, 95% CI 0.37-0.86, respectively). These results remained also after adjustment for smoking intensity, i.e., numbers of cigarettes smoked per day (OR 0.73, 95% CI 0.53-0.99; OR 0.59, 95% CI 0.35-0.97), respectively). To conclude, adherence to a Mediterranean-like diet seems to be inversely associated with the development of COPD.