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Abstract To assess if SARS-CoV-2 (COVID-19) systemic disease can be determined by available nucleoprotein assays, we compared the performance of three commercial SARS-CoV-2 nucleoprotein (N) assays in plasma. A total of 272 plasma samples collected in the period November–December 2021 were analyzed by the methods Simoa SARS CoV‐2 N Protein Advantage Kit [Quanterix Simoa], Solsten SARS-CoV-2 Antigen enzyme immunosorbent assay (ELISA) [Solsten ELISA], and Elecsys SARS‐CoV‐2 Antigen electrochemiluminescence immunoassay [Elecsys ECLIA]. Additionally, a dilution series of inactivated virus culture was analyzed by the three assays. The SARS CoV-2 PCR-status was not known for the patients. Linear correlation in the pairwise correlation between assays as well as linearity of dilution series of inactivated virus culture was estimated by Spearman score. Sensitivity and specificity were estimated by pairwise comparison. The three assays showed poor agreement on patient samples with regards to concentration. Performance on virus culture was excellent but with different level of detection (LOD). Positive vs negative results show comparable sensitivity and specificity of Quanterix Simoa and Solsten ELISA, with a higher LOD in Elecsys ECLIA and thus lower sensitivity and high specificity. N by all tested assays can be used as a marker for systemic COVID-19 disease.

Abstract. Introduction:. Fibromyalgia (FM) is a chronic fluctuating, nociplastic pain condition. Naltrexone is a µ-opioid-receptor antagonist; preliminary studies have indicated a pain-relieving effect of low-dose naltrexone (LDN) in patients with FM. The impetus for studying LDN is the assumption of analgesic efficacy and thus reduction of adverse effects seen from conventional pharmacotherapy. Objectives:. First, to examine if LDN is associated with analgesic efficacy compared with control in the treatment of patients with FM. Second, to ascertain the analgesic efficacy of LDN in an experimental pain model in patients with FM evaluating the competence of the descending inhibitory pathways compared with controls. Third, to examine the pharmacokinetics of LDN. Methods:. The study used a randomized, double-blind, placebo-controlled, crossover design and had a 3-phase setup. The first phase included baseline assessment and a treatment period (days −3 to 21), the second phase a washout period (days 22–32), and the third phase a baseline assessment followed by a treatment period (days 33–56). Treatment was with either LDN 4.5 mg or an inactive placebo given orally once daily. The primary outcomes were Fibromyalgia Impact Questionnaire revised (FIQR) scores and summed pain intensity ratings (SPIR). Results:. Fifty-eight patients with FM were randomized. The median difference (IQR) for FIQR scores between LDN and placebo treatment was −1.65 (18.55; effect size = 0.15; P = 0.3). The median difference for SPIR scores was −0.33 (6.33; effect size = 0.13; P = 0.4). Conclusion:. Outcome data did not indicate any clinically relevant analgesic efficacy of the LDN treatment in patients with FM.

Abstract This study aimed to develop a highly sensitive SARS-CoV-2 nucleocapsid antigen assay using the single molecule array (Simoa) technology and compare it with real time RT-PCR as used in routine clinical practice with the ambition to achieve a comparative technical and clinical sensitivity. Samples were available from 148 SARS-CoV-2 real time RT-PCR positive and 73 SARS-CoV-2 real time RT-PCR negative oropharyngeal swabs. For determination of technical sensitivity SARS-CoV-2 virus culture material was used. The samples were treated with lysis buffer and analyzed using both an in-house and a pre-commercial SARS-CoV-2 nucleocapsid antigen assay on Simoa. Both nucleocapsid antigen assays have a technical sensitivity corresponding to around 100 SARS-CoV-2 RNA molecules/mL. Using a cut-off at 0.1 pg/mL the pre-commercial SARS-CoV-2 nucleocapsid antigen assay had a sensitivity of 96% (95% CI 91.4–98.5%) and specificity of 100% (95% CI 95.1–100%). In comparison the in-house nucleocapsid antigen assay had sensitivity of 95% (95% CI 89.3–98.1%) and a specificity of 100% (95% CI 95.1–100%) using a cut-off at 0.01 pg/mL. The two SARS-CoV-2 nucleocapsid antigen assays correlated with r = 0.91 (P

ABSTRACT Zoledronic acid is a bisphosphonate commonly used to treat bone diseases such as osteoporosis and cancer‐induced bone disease. Patients exhibit a variable sensitivity to zoledronic acid; the underlying explanation for this remains unclear. The objective of this study was to obtain more knowledge in this regard. We hypothesized that osteoclasts generated from different individuals would show a variable sensitivity to zoledronic acid in vitro. Osteoclasts were generated using monocytes from 46 healthy female blood donors (40 to 66 years). Matured osteoclasts were reseeded onto bone slices precoated with different concentrations of zoledronic acid. IC50 values were determined based on total eroded bone surface after 3 days of resorption. The IC50 for inhibition of osteoclastic bone resorption varied from 0.06 to 12.57μM zoledronic acid; thus, a more than 200‐fold difference in sensitivity to zoledronic acid among osteoclasts from different individuals was observed. Multiple linear regression analyses showed that the determined IC50 correlated with smoking status, and the average number of nuclei per osteoclast in vitro. Further analyses showed that: (i) increasing protein levels of mature cathepsin K in osteoclast cultures rendered the osteoclasts less sensitive to zoledronic acid; (ii) surprisingly, neither the gene nor the protein expression of farnesyl diphosphate synthase was found to correlate with the IC50; and (iii) trench‐forming osteoclasts were found to be more sensitive to zoledronic acid than pit‐forming osteoclasts within the same cell culture. Thus, we conclude that there indeed is a high degree of variation in the potency of zoledronic acid on osteoclasts when generated from different individuals. We propose that our findings can explain some of the varying clinical efficacy of zoledronic acid therapy observed in patients, and may therefore be of clinical importance, which should be investigated in a clinical trial combining in vitro and in vivo investigations. © 2020 The Authors. JBMR Plus published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research.

Background: Ischemic stroke causes increased blood–brain barrier permeability and release of markers of axonal damage and inflammation. To investigate diagnostic and prognostic roles of neurofilament light chain (NF-L), we assessed levels of NF-L, S100B, interleukin-6 (IL-6), E-selectin, vascular endothelial growth factor-A (VEGF-A), vascular cell adhesion molecule-1 (VCAM-1), and intercellular adhesion molecule-1 (ICAM-1) in patients with acute ischemic stroke or transient ischemic attack (TIA) and healthy controls.Methods: We studied neurofilament (NF) expression in 2 cases of human postmortem ischemic stroke, representing infarcts aged 3- to >7-days. In a prospective study, we measured plasma NF-L and inflammatory markers

Abstract Background Whole genome duplication plays a central role in plant evolution. There are two main classes of polyploid formation: autopolyploids which arise within one species by doubling of similar homologous genomes; in contrast, allopolyploidy (hybrid polyploidy) arise via hybridization and subsequent doubling of nonhomologous (homoeologous) genomes. The distinction between polyploid origins can be made using gene phylogenies, if alleles from each genome can be correctly retrieved. We examined whether two closely related tetraploid Mediterranean shrubs (Medicago arborea and M. strasseri) have an allopolyploid origin – a question that has remained unsolved despite substantial previous research. We sequenced and analyzed ten low-copy nuclear genes from these and related species, phasing all alleles. To test the efficacy of allele phasing on the ability to recover the evolutionary origin of polyploids, we compared these results to analyses using unphased sequences. Results In eight of the gene trees the alleles inferred from the tetraploids formed two clades, in a non-sister relationship. Each of these clades was more closely related to alleles sampled from other species of Medicago, a pattern typical of allopolyploids. However, we also observed that alleles from one of the remaining genes formed two clades that were sister to one another, as is expected for autopolyploids. Trees inferred from unphased sequences were very different, with the tetraploids often placed in poorly supported and different positions compared to results obtained using phased alleles. Conclusions The complex phylogenetic history of M. arborea and M. strasseri is explained predominantly by shared allotetraploidy. We also observed that an increase in woodiness is correlated with polyploidy in this group of species and present a new possibility that woodiness could be a transgressive phenotype. Correctly phased homoeologues are likely to be critical for inferring the hybrid origin of allopolyploid species, when most genes retain more than one homoeologue. Ignoring homoeologous variation by merging the homoeologues can obscure the signal of hybrid polyploid origins and produce inaccurate results.

Tumor necrosis factor receptor 1 and 2 (TNFR1 and TNFR2) have been found in brain parenchyma of stroke patients, and plasma levels are increased in the acute phase of stroke. We evaluated associations between TNFR1 and TNFR2 plasma levels and stroke severity, infarct size, and functional outcome. Furthermore, we examined cellular expression of TNFR1 and TNFR2 on leukocyte subpopulations to explore the origin of the increased receptor levels. Blood samples were taken from 33 acute ischemic stroke patients and 10 healthy controls. TNFR1 and TNFR2 plasma concentrations were measured and correlated against the Scandinavian Stroke Scale at admission, infarct volume, and the modified Rankin Scale score three months after stroke onset. Classical, intermediate, and non-classical monocytes as well as neutrophils were purified, and cellular expression of TNFR1 and TNFR2 was examined using flow cytometry. TNFR1 and TNFR2 plasma levels were both increased after ischemic stroke, but we found no correlation with patient outcome measurements. Compared to healthy controls, ischemic stroke patients had decreased non-classical monocyte and neutrophil populations expressing TNFR1 and increased neutrophils expressing TNFR2, and decreased non-classical populations co-expressing both TNFR1 and TNFR2. This study supports the hypothesis of an acute immunological response orchestrated by the peripheral immune system following an ischemic stroke. However, the origin of the increased TNFR1 and TNFR2 plasma levels could not be clearly linked to peripheral monocytes or neutrophils. Future studies are needed and will help clarify the potential role as treatment target.

It is well established that multinucleation is central for osteoclastic bone resorption. However, our knowledge on the mechanisms regulating how many nuclei an osteoclast will have is limited. The objective of this study was to investigate donor-related variations in the fusion potential of in vitro-generated osteoclasts. Therefore, CD14+ monocytes were isolated from 49 healthy female donors. Donor demographics were compared to the in vivo bone biomarker levels and their monocytes’ ability to differentiate into osteoclasts, showing that: (1) C-terminal telopeptide of type I collagen (CTX) and procollagen type I N-terminal propeptide (PINP) levels increase with age, (2) the number of nuclei per osteoclast in vitro increases with age, and (3) there is a positive correlation between the number of nuclei per osteoclast in vitro and CTX levels in vivo. Furthermore, the expression levels of the gene encoding dendritic cell-specific transmembrane protein (DCSTAMP) of osteoclasts in vitro correlated positively with the number of nuclei per osteoclast, CTX levels in vivo, and donor age. Our results furthermore suggest that these changes in gene expression may be mediated through age-related changes in DNA methylation levels. We conclude that both intrinsic factors and age-induced increase in fusion potential of osteoclasts could be contributing factors for the enhanced bone resorption in vivo, possibly caused by increased expression levels of DCSTAMP.

According to current recommendations, blood samples should be taken in the morning after 15 minutes' resting time. Some components exhibit diurnal variation and in response to pressures to expand opening hours and reduce waiting time, the aims of this study were to investigate the impact of resting time prior to blood sampling and diurnal variation on biochemical components, including albumin, thyrotropin (TSH), total calcium and sodium in plasma.All patients referred to an outpatient clinic for blood sampling were included in the period Nov 2011 until June 2014 (opening hours: 7am-3pm). Each patient's arrival time and time of blood sampling were registered. The impact of resting time and the time of day for all components was analysed using simple linear regression. The "maximum allowable bias" was used as quality indicator for the change in reference interval.Significant diurnal variation was found for albumin (n = 15,544; p

Next-generation sequencing technology has increased the capacity to generate molecular data for plant biological research, including phylogenetics, and can potentially contribute to resolving complex phylogenetic problems. The evolutionary history of Medicago L. (Leguminosae: Trifoliae) remains unresolved due to incongruence between published phylogenies. Identification of the processes causing this genealogical incongruence is essential for the inference of a correct species phylogeny of the genus and requires that more molecular data, preferably from low-copy nuclear genes, are obtained across different species. Here we report the development of 50 novel LCN markers in Medicago and assess the phylogenetic properties of each marker. We used the genomic resources available for Medicago truncatula Gaertn., hybridisation-based gene enrichment (sequence capture) techniques and Next-Generation Sequencing to generate sequences. This alternative proves to be a cost-effective approach to amplicon sequencing in phylogenetic studies at the genus or tribe level and allows for an increase in number and size of targeted loci. Substitution rate estimates for each of the 50 loci are provided, and an overview of the variation in substitution rates among a large number of low-copy nuclear genes in plants is presented for the first time. Aligned sequences of major species lineages of Medicago and its sister genus are made available and can be used in further probe development for sequence-capture of the same markers.

Background Vitamin K deficiency is highly prevalent in patients on dialysis and may contribute to their low bone mineral density (BMD) and increased risk of fracture. This study investigated the effect of menaquinone-7 (MK-7) supplementation on BMD in patients on chronic dialysis. Methods In a multicentre, double-blind, placebo-controlled intervention trial, 123 patients on chronic dialysis were randomised to a daily oral supplement of either MK-7 360 µg or placebo for 2 years. BMD of the distal radius (1/3, mid, ultradistal and total), femoral neck, lumbar spine (L1–L4) and whole body was assessed by dual-energy X-ray absorptiometry. Serum levels of vitamin K1 and MK-7 and plasma levels of total osteocalcin, dephosphorylated-uncarboxylated matrix Gla protein and protein induced by vitamin K absence II were measured to assess vitamin K status. Results After 2 years, an accelerated BMD loss of the 1/3 distal radius was found with MK-7 supplementation {mean difference of changes relative to placebo −0.023 g/cm2 [95% confidence interval (CI) −0.039 to −0.008]}, whereas the decrease in lumbar spine BMD seen in the placebo group was prevented [mean difference of changes between groups 0.050 g/cm2 (95% CI 0.015–0.085)]. No significant effects were observed at the remaining skeletal sites. Vitamin K status strongly improved in MK-7-supplemented participants. Conclusion Compared with placebo, an accelerated BMD loss of the 1/3 distal radius was found after 2 years of MK-7 supplementation, whereas a decline in lumbar spine BMD was prevented. As such, MK-7 supplementation might modify BMD site-specifically in patients on dialysis. In aggregate, our findings do not support MK-7 supplementation to preserve bone in patients on dialysis. [ABSTRACT FROM AUTHOR]

Abstract Diagnosing patients in the medical emergency department is complex and this is expected to increase in many countries due to an ageing population. In this study we investigate the feasibility of training machine learning algorithms to assist physicians handling the complex situation in the medical emergency departments. This is expected to reduce diagnostic errors and improve patient logistics and outcome. We included a total of 9,190 consecutive patient admissions diagnosed and treated in two hospitals in this cohort study. Patients had a biochemical workup including blood and urine analyses on clinical decision totaling 260 analyses. After adding nurse-registered data we trained 19 machine learning algorithms on a random 80% sample of the patients and validated the results on the remaining 20%. We trained algorithms for 19 different patient outcomes including the main outcomes death in 7 (Area under the Curve (AUC) 91.4%) and 30 days (AUC 91.3%) and safe-discharge(AUC 87.3%). The various algorithms obtained areas under the Receiver Operating Characteristics -curves in the range of 71.8–96.3% in the holdout cohort (68.3–98.2% in the training cohort). Performing this list of biochemical analyses at admission also reduced the number of subsequent venipunctures within 24 h from patient admittance by 22%. We have shown that it is possible to develop a list of machine-learning algorithms with high AUC for use in medical emergency departments. Moreover, the study showed that it is possible to reduce the number of venipunctures in this cohort.

Abstract Preeclampsia (PE) is characterized by de novo hypertension (HTN) and is often associated with intrauterine growth restriction (IUGR). Hallmarks of PE are placental ischemia, decreased nitric oxide (NO) bioavailability, oxidative stress (OS), and organ damage in the kidneys and brain. This study aims to characterize a new model of PE using pregnant IUGR rats from hypertensive placental ischemic dams. It is hypothesized that pregnant IUGR rats from hypertensive placental ischemic dams will have elevated blood pressure (BP), OS, and organ damage. In this study, pregnant rats are divided into two groups: normal pregnant (NP) and hypertensive placental ischemic dams (RUPP). Offspring from NP and RUPP dams were mated at 10 weeks of age to generate pregnant IUGR (IUGR Preg) and pregnant control (CON Preg) rats. BP and other markers of PE were evaluated during late gestation. Pregnant IUGR rats had elevated BP and systemic OS. The maternal body weight of pregnant IUGR rats and their pups' weights were decreased, while the brains were enlarged with elevated OS. In summary, pregnant IUGR rats, born from hypertensive placental ischemic dams, have HTN and increased systemic and brain OS, with larger brain sizes and smaller pups. Furthermore, this study shows that pregnant IUGR rats exhibit a preeclamptic‐like phenotype, suggesting a new epigenetic model of PE.

Study objectives: To develop a non-invasive and practical wearable method for long-term tracking of infants’ sleep. Methods: An infant wearable, NAPping PAnts (NAPPA), was constructed by combining a diaper cover and a movement sensor (triaxial accelerometer and gyroscope), allowing either real-time data streaming to mobile devices or offline feature computation stored in the sensor memory. A sleep state classifier (wake, N1/REM, N2/N3) was trained and tested for NAPPA recordings (N = 16649 epochs of 30 s), using hypnograms from co-registered polysomnography (PSG) as a training target in 33 infants (age 2 weeks to 18 months; Mean = 4). User experience was assessed from an additional group of 16 parents. Results: Overnight NAPPA recordings were successfully performed in all infants. The sleep state classifier showed good overall accuracy (78 %; Range 74–83 %) when using a combination of five features related to movement and respiration. Sleep depth trends were generated from the classifier outputs to visualise sleep state fluctuations, which closely aligned with PSG-derived hypnograms in all infants. Consistently positive parental feedback affirmed the effectiveness of the NAPPA-design. Conclusions: NAPPA offers a practical and feasible method for out-of-hospital assessment of infants’ sleep behaviour. It can directly support large-scale quantitative studies and development of new paradigms in scientific research and infant healthcare. Moreover, NAPPA provides accurate and informative computational measures for body positions, respiration rates, and activity levels, each with their respective clinical and behavioural value.

Abstract To improve prostate cancer (PCa) diagnosis, it is imperative to identify novel biomarkers and establish effective screening techniques. Here, we introduce electrochemical biosensing of β-2-Microglobulin (β2M) in urine as a potential diagnostic tool for PCa. The immunosensor is composed of a screen-printed graphene electrode coated with anti β2M antibodies. The sensor is capable of detecting the protein directly in urine without any sample pretreatment within 45 min including sample incubation and a lower limit of detection of 204 µg/L. The sensor demonstrated a significant difference in the β2M-creatinine ratio in urine between control and both local- and metastatic PCa (mPCa) (P = 0.0302 and P = 0.0078 respectively), and between local- and mPCa (P = 0.0302). This first example of electrochemical sensing of β2M for the diagnosis of PCa may set the stage for an affordable, on-site screening technique for PCa.

Rapid on-site electrochemical screening for prostate cancer (PCa) in the clinic holds the potential for early diagnosis of PCa patients. Currently, the accuracy of commonly used biomarkers is insufficient; however, several protein biomarker candidates show promise for diagnostic or prognostic patient screening. Among these candidates are Prostate specific antigen (PSA), Human glandular kallikrein 2 (hK2), Annexins 3 (Annex 3), Beta-2-microglobulin (β2M), Microseminoprotein-beta (MSMB), Serum amyloid A (SAA), and Engrailed-2 (EN2). This review provides an overview of the advancements in electrochemical biosensor development specifically targeting the detection of these seven biomarkers. The selection of these biomarkers was based on their suitability for protein-level detection using electrochemical sensors, as well as their presence in biological fluids. Moreover, we emphasize that relying solely on a single biomarker may not be sufficient to obtain precise diagnostic information. It is now widely accepted that panels of biomarkers measured in a multiplexed fashion are of greater clinical utility than single biomarkers. Therefore, we suggest that future research prioritize the utilization of multiplex electrochemical methods for measuring panels of protein biomarkers, offering a simple and efficient approach for regular PCa monitoring.