Variations in pigment and carbohydrate content of gallbladder bile affect accurate quantitation of total protein when using the fluorescamine method

Background: Despite solute dilution and reduced total lipid concentrations, an unexplained increase in protein concentration has been reported to occur in the gallbladder bile of cholesterol gallstone patients. Methods: Solutes in gallbladder bile from gallstone-free controls and from four study groups were measured using standard methods. Total proteins were measured using amino acid analysis and a conventional fluorescamine method. Results: Bile salts and pigment content were greater in gallstone-free controls than in all other study groups, including morbidly obese gallstone-free subjects. Total biliary protein concentration, as determined by amino acid analysis in the gallstone-free control group was higher than in non-obese gallstone patients with multiple stones and in morbidly obese gallstone-free subjects. Total biliary proteins as measured with fluorescamine, however, did not show intergroup differences. A major problem of the conventional fluorescamine assay is shown to be an artefact arising from the high pigment content of the more concentrated samples. Conclusions: Very dilute gallbladder bile samples are often found in the presence of gallstone disease. This also occurs in morbidly obese subjects, even in the absence of gallstones. Although the contribution of protein secretion/absorption by the gallbladder can also be relevant, especially in the presence of morbid obesity, the protein concentration in gallbladder bile, when accurately measured, generally parallels the concentrations of non-absorbed biliary solutes, reflecting the efficiency of fluid absorption. Measurement of biliary proteins by the conventional fluorescamine method is unreliable in clinical studies in which intergroup differences in pigment content are commonly present

Human gallbladder mucosal function : effects on intraluminal fluid and lipid composition in health and disease.

Abstract: Gallbladder mucosal absorption of fluid during fasting is a well-known process. Indirect in vivo and recent in vitro evidence for physiologically relevant gallbladder absorption of cholesterol and phospholipids from bile has been observed in humans. The present study explored and compared by indirect means the relative efficiences of human gallbladder mucosal absorption of fluid and lipids in health and disease. Biliary lipids and pigment content were measured in fasting gallbladder bile samples obtained from gallstone-free controls and from four study groups: multiple and solitary cholesterol gallstone patients, and morbidly obese subjects with and without gallstones. Bile salts and pigment content were significantly greater in gallstone-free controls than in all other disease study groups, This was interpreted as evidence of more effective gallbladder mucosal fluid absorption in nonobese gallstone-free controls compared to that in all other groups, Correlation plot analyses of biliary lipids showed lower concentrations of phospholipids than expected from the index bile salt concentrations, The same was found for cholesterol concentrations but only in supersaturated samples, These findings were much more pronounced in gallstone free-controls and were accordingly interpreted as evidence of more efficient gallbladder absorption of both phospholipids and cholesterol in controls compared with that found in each of the disease study groups, Moreover, impaired gallbladder mucosal function, while invariably associated with cholesterol gallstone disease, was not found to be a necessary consequence of the physical presence of stones. It is concluded that efficient gallbladder mucosal absorption of both fluid and apolar lipids from bile is a normal physiological process that is often seriously impaired in the presence of either cholesterol gallstone disease or at least one of its precursor forms

COVID-19 in pregnancy—characteristics and outcomes of pregnant women admitted to hospital because of SARS-CoV-2 infection in the Nordic countries

cited By 0Introduction Population-based studies about the consequences of SARS-CoV-2 infection (COVID-19) in pregnancy are few and have limited generalizability to the Nordic population and healthcare systems. Material and methods This study examines pregnant women with COVID-19 in the five Nordic countries. Pregnant women were included if they were admitted to hospital between 1 March and 30 June 2020 and had a positive SARS-CoV-2 PCR test Results In the study areas, 214 pregnant women with a positive test were admitted to hospital, of which 56 women required hospital care due to COVID-19. The risk of admission due to COVID-19 was 0.4/1000 deliveries in Denmark, Finland and Norway, and 3.8/1000 deliveries in the Swedish regions. Women hospitalized because of COVID-19 were more frequently obese (p < 0.001) and had a migrant background (p < 0.001) compared with the total population of women who delivered in 2018. Twelve women (21.4%) needed intensive care. Among the 56 women admitted due to COVID-19, 48 women delivered 51 infants. Preterm delivery (n = 12, 25%, p < 0.001) and cesarean delivery (n = 21, 43.8%, p < 0.001) were more frequent in women with COVID-19 compared with women who delivered in 2018. No maternal deaths, stillbirths or neonatal deaths were reported. Conclusions The risk of admission due to COVID-19 disease in pregnancy was low in the Nordic countries. A fifth of the women required intensive care and we observed higher rates of preterm and cesarean deliveries. National public health policies appear to have had an impact on the risk of admission due to severe COVID-19 disease in pregnancy. Nordic collaboration is important in collecting robust data and assessing rare outcomes.Peer reviewe

Zip nucleic acids are potent hydrolysis probes for quantitative PCR

Zip nucleic acids (ZNAs) are oligonucleotides conjugated with cationic spermine units that increase affinity for their target. ZNAs were recently shown to enable specific and sensitive reactions when used as primers for polymerase chain reaction (PCR) and reverse-transcription. Here, we report their use as quantitative PCR hydrolysis probes. Ultraviolet duplex melting data demonstrate that attachment of cationic residues to the 3′ end of an oligonucleotide does not alter its ability to discriminate nucleotides nor the destabilization pattern relative to mismatch location in the oligonucleotide sequence. The stability increase provided by the cationic charges allows the use of short dual-labeled probes that significantly improve single-nucleotide polymorphism genotyping. Longer ZNA probes were shown to display reduced background fluorescence, therefore, generating greater sensitivity and signal level as compared to standard probes. ZNA probes thus provide broad flexibility in assay design and also represent an effective alternative to minor groove binder- and locked nucleic-acid-containing probes

Light-up properties of complexes between thiazole orange-small molecule conjugates and aptamers

The full understanding of dynamics of cellular processes hinges on the development of efficient and non-invasive labels for intracellular RNA species. Light-up aptamers binding fluorogenic ligands show promise as specific labels for RNA species containing those aptamers. Herein, we took advantage of existing, non-light-up aptamers against small molecules and demonstrated a new class of light-up probes in vitro. We synthesized two conjugates of thiazole orange dye to small molecules (GMP and AMP) and characterized in vitro their interactions with corresponding RNA aptamers. The conjugates preserved specific binding to aptamers while showing several 100-fold increase in fluorescence of the dye (the ‘light-up’ property). In the presence of free small molecules, conjugates can be displaced from aptamers serving also as fluorescent sensors. Our in vitro results provide the proof-of-concept that the small-molecule conjugates with light-up properties can serve as a general approach to label RNA sequences containing aptamers

Impact of direct cell co-cultures on human adipose-derived stromal cells and nucleus pulposus cells

Biologic and cellular treatment strategies aiming for curing intervertebral disc degeneration (IDD) have been proposed recently. Given the convenient availability and expansion potential, adipose-derived stromal cells (ADSCs) might be an ideal cell candidate. However, the interaction between ADSCs and nucleus pulposus (NP) cells still remains ambiguous, especially in direct co-cultures of the two types of cells. Nevertheless, NP markers in ADSCs after co-cultures were unidentified. Here, we addressed the interaction of human ADSCs and NP cells in a direct co-culture system for the first time. As a result, ADSCs could differentiate to the NP cell phenotype with a significant up-regulated expression of multiple genes and proteins in extracellular matrix (ECM) (SOX9, COL2A1, ACAN, and COL6A2), relative NP markers (FOXF1, PAX1, CA12, and HBB) and pertinent growth factors (CDMP-1, TGF-beta1, IGF-1, and CTGF). Moreover, the gene expression of COL2A1, ACAN, and COL6A2 of degenerate NP cells was also up-regulated. Collectively, these results suggest that direct co-cultures of ADSCs and NP cells may exert a reciprocal impact, that is, both stimulating ADSCs differentiation to the NP cell phenotype and inducing NP cells to regain functional phenotype. Accordingly, ADSCs might be a potential candidate in the development of cellular treatment strategies for IDD.published_or_final_versio

Snap-to-it probes: chelate-constrained nucleobase oligomers with enhanced binding specificity

We describe snap-to-it probes, a novel probe technology to enhance the hybridization specificity of natural and unnatural nucleic acid oligomers using a simple and readily introduced structural motif. Snap-to-it probes were prepared from peptide nucleic acid (PNA) oligomers by modifying each terminus with a coordinating ligand. The two coordinating ligands constrain the probe into a macrocyclic configuration through formation of an intramolecular chelate with a divalent transition metal ion. On hybridization with a DNA target, the intramolecular chelate in the snap-to-it probe dissociates, resulting in the probe ‘snapping-to’ and binding the target nucleic acid. Thermal transition analysis of snap-to-it probes with complementary and single-mismatch DNA targets revealed that the transition between free and target-bound probe conformations was a reversible equilibrium, and the intramolecular chelate provided a thermodynamic barrier to target binding that resulted in a significant increase in mismatch discrimination. A 4–6°C increase in specificity (ΔTm) was observed from snap-to-it probes bearing either terminal iminodiacetic acid ligands coordinated with Ni2+, or terminal dihistidine and nitrilotriacetic acid ligands coordinated with Cu2+. The difference in specificity of the PNA oligomer relative to DNA was more than doubled in snap-to-it probes. Snap-to-it probes labeled with a fluorophore-quencher pair exhibited target-dependent fluorescence enhancement upon binding with target DNA

Variations across Europe in hospitalization and management of pregnant women with SARS-CoV-2 during the initial phase of the pandemic : Multi-national population-based cohort study using the International Network of Obstetric Survey Systems (INOSS)

Funding Information: The national studies reported the following funding sources: The BOSS project was funded by the Belgian Federal Public Service of Health. The NOSS collaboration was supported by the Nordic Federation of Societies of Obstetrics and Gynecology (grant no. 6505, 2020). NOSS‐Denmark was supported by grants from The Region of Southern Denmark and Region Zealand's shared fund for joint health research projects (Reg. no. A767), and EasyTrial provided the data collection software. NOSS‐Finland received grants from the Finnish Medical Society, and from Helsinki University. UKOSS received funding from the National Institute for Health Research HS&DR Programme (11/46/12). The national studies in Italy and the Netherlands did not have specific funding. The multi‐national study received partial funding support from the European Medicines Agency (EMA) under the Framework service contract nr EMA/2018/28/PE. The content of this paper expresses the opinions of the authors and may not be understood or quoted as being made on behalf of or reflecting the position of the EMA or any of its committees or working parties. The research leading to these results was conducted as part of the activities of the EU PE&PV (Pharmacoepidemiology and Pharmacovigilance) Research Network, which is a public academic partnership coordinated by Utrecht University, the Netherlands. The CONSIGN project was scientifically coordinated by the University Medical Center, Utrecht. Funding Information: OB declares support from the European Medicines agency (EMA). HE declares grants from the Nordic Federation of Societies of Obstetrics and Gynecology (NFOG) and Norwegian Research Council (grant no 320181). AA declares a grant from the Region of Southern Denmark and Region Zealand's shared fund for joint health research projects. OA declares grants from the Finnish Medical Association and NFOG. MK declares grants from the National Institute for Health and Care Research, Medical Research Council, Healthcare Quality Improvement Partnership and Wellbeing of Women during the course of the study. MS leads a department that conducts studies on COVID‐19 vaccines for the European Medicines Agency, Pfizer, AstraZeneca and Janssen. All support was according to the ENCePP code of conduct. None of the other authors (NV, RR, SD, EJ, EO, MAS, TS, RV, AV, KB) has anything to disclose. Publisher Copyright: © 2023 The Authors. Acta Obstetricia et Gynecologica Scandinavica published by John Wiley & Sons Ltd on behalf of Nordic Federation of Societies of Obstetrics and Gynecology (NFOG).Introduction: The majority of data on COVID-19 in pregnancy are not from sound population-based active surveillance systems. Material and methods: We conducted a multi-national study of population-based national or regional prospective cohorts using standardized definitions within the International Network of Obstetric Survey systems (INOSS). From a source population of women giving birth between March 1 and August 31, 2020, we included pregnant women admitted to hospital with a positive SARS-CoV-2 PCR test ≤7 days prior to or during admission and up to 2 days after birth. The admissions were further categorized as COVID-19-related or non-COVID-19-related. The primary outcome of interest was incidence of COVID-19-related hospital admission. Secondary outcomes included severe maternal disease (ICU admission and mechanical ventilation) and COVID-19-directed medical treatment. Results: In a source population of 816 628 maternities, a total of 2338 pregnant women were admitted with SARS-CoV-2; among them 940 (40%) were COVID-19-related admissions. The pooled incidence estimate for COVID-19-related admission was 0.59 (95% confidence interval 0.27–1.02) per 1000 maternities, with notable heterogeneity across countries (I2 = 97.3%, P = 0.00). In the COVID-19 admission group, between 8% and 17% of the women were admitted to intensive care, and 5%–13% needed mechanical ventilation. Thromboprophylaxis was the most frequent treatment given during COVID-19-related admission (range 14%–55%). Among 908 infants born to women in the COVID-19-related admission group, 5 (0.6%) stillbirths were reported. Conclusions: During the initial months of the pandemic, we found substantial variations in incidence of COVID-19-related admissions in nine European countries. Few pregnant women received COVID-19-directed medical treatment. Several barriers to rapid surveillance were identified. Investment in robust surveillance should be prioritized to prepare for future pandemics.Peer reviewe