Aquaporin-0 (AQP0) is the main water channel in the mammalian lens and is involved in accommodation and maintaining lens transparency. AQP0 binds the Ca2+-sensing protein calmodulin (CaM) and this interaction is believed to gate its water permeability by closing the water-conducting pore. Here we express recombinant and functional human AQP0 in P. pastoris and investigate how phosphorylation affects the interaction with CaM in vitro as well as the CaM-dependent water permeability of AQP0 in proteoliposomes. Using microscale thermophoresis (MST) and surface plasmon resonance (SPR) technology we show that the introduction of the single phospho-mimicking mutations S229D and S235D in AQP0 reduces CaM-binding. In contrast, CaM interacts with S231D with similar affinity as wild type, but in a different manner. Permeability studies of wild type AQP0 showed that the water conductance was significantly reduced by CaM in a Ca2+-dependent manner, whereas AQP0 S229D, S231D and S235D were all locked in an open state, insensitive to CaM. We propose a model in which phosphorylation of AQP0 control CaM-mediated gating in two different ways (1) phosphorylation of S229 or S235 abolishes binding (the pore remains open) and (2) phosphorylation of S231 results in CaM-binding without causing pore closure, the functional role of which remains to be elucidated. Our results suggest that site-dependent phosphorylation of AQP0 dynamically controls its CaM-mediated gating. Since the level of phosphorylation increases towards the lens inner cortex, AQP0 may become insensitive to CaM-dependent gating along this axis.
The mucus layer in the small intestinal is generally regarded as a barrier to drug absorption. However, the mucus layer is a complex system, and presently, only a few studies have been conducted to elucidate its physicochemical properties. The current study hypothesizes that the mucus layer contains solubility-enhancing surfactants and thus might aid the oral absorption of poorly water-soluble drugs. Mucus was sampled from sections of the small intestine of fasted rats to analyze the rheological properties and determine the mucus pH and concentrations of proteins and endogenous surfactants, i.e., bile salts, polar lipids, and neutral lipids. The mucus layer in the two proximal sections of the small intestine exhibited different rheological properties such as higher zero-shear viscosity and lower loss tangent and higher protein concentrations compared to all subsequent sections of the small intestine. The pH of the mucus layer was stable at ~ 6.5 throughout most of the small intestine, but increased to 7.5 in the ileum. The bile salt concentrations increased from the duodenum (16.0 ± 2.2 mM) until the mid jejunum (55.1 ± 9.5 mM), whereas the concentrations of polar lipids and neutral lipids decreased from the duodenum (17.4 ± 2.2 mM and 37.8 ± 1.6 mM, respectively) until the ileum (4.8 ± 0.4 mM and 10.7 ± 1.1 mM, respectively). In conclusion, the mucus layer of the rat small intestine contains endogenous surfactants at levels that might benefit solubilization and absorption of orally administered poorly water-soluble drugs.
KEA1 and KEA2 are K /H antiporters in the chloroplast inner envelope that adjust stromal pH in light to dark transitions. We previously determined that stromal pH is higher in the kea1kea2 mutant cells. Given that cytosol and chloroplast volume are about equal in mesophyll cells, we wanted to test if this in turn could cause ionic imbalances in the cytosol as well. In this study we indeed find that the cytosol is more acidic in the kea1kea2 mutant. As cytosol pH is tightly controlled by the activity of the plasma membrane proton ATPase, we tested the activity of this enzyme. Acidic pH is expected to phosphorylate and activate the proton ATPase to restore pH. We could however not detect differences in plasma membrane H -ATPase hydrolytic activity between Col-0 and the kea1kea2 mutant. No differences in the amount of plasma membrane H -ATPase enzyme, regulatory 14-3-3 proteins and phosphorylation of H -ATPase Thr932 between Col-0 and the kea1kea2 mutant were detected, indicating that cytosol acidification did not induce transcriptional regulation or post-translational phosphorylation of plasma membrane H -ATPase in the kea1kea2 mutant. Curiously however, plasma membrane in the double mutant was depolarized, indicating a reduced electrogenic transport of protons by the ATPase, or an increased conductance to other ions. We determined that reconstituted membrane vesicles isolated from kea1kea2 mutants have slightly lower ATP dependent proton transport activity and appear especially more permeant to potassium, but also show increased proton conductance.
Background Early diagnosis of cerebral palsy (CP) is important to enable intervention at a time when neuroplasticity is at its highest. Current mean age at diagnosis is 13 months in Denmark. Recent research has documented that an early-diagnosis set-up can lower diagnostic age in high-risk infants. The aim of the current study is to lower diagnostic age of CP regardless of neonatal risk factors. Additionally, we want to investigate if an early intervention program added to standard care is superior to standard care alone. Methods The current multicentre study CP-EDIT (Early Diagnosis and Intervention Trial) with the GO-PLAY intervention included (Goal Oriented ParentaL supported home ActivitY program), aims at testing the feasibility of an early diagnosis set-up and the GO-PLAY early intervention. CP-EDIT is a prospective cohort study, consecutively assessing approximately 500 infants at risk of CP. We will systematically collect data at inclusion (age 3–11 months) and follow a subset of participants (n = 300) with CP or at high risk of CP until the age of two years. The GO-PLAY early intervention will be tested in 80 infants with CP or high risk of CP. Focus is on eight areas related to implementation and perspectives of the families: early cerebral magnetic resonance imaging (MRI), early genetic testing, implementation of the General Movements Assessment method, analysis of the GO-PLAY early intervention, parental perspective of early intervention and early diagnosis, early prediction of CP, and comparative analysis of the Hand Assessment for Infants, Hammersmith Infant Neurological Examination, MRI, and the General Movements method. Discussion Early screening for CP is increasingly possible and an interim diagnosis of “high risk of CP” is recommended but not currently used in clinical care in Denmark. Additionally, there is a need to accelerate identification in mild or ambiguous cases to facilitate appropriate therapy early. Most studies on early diagnosis focus on identifying CP in infants below five months corrected age. Little is known about early diagnosis in the 50% of all CP cases that are discernible later in infancy. The current study aims at improving care of patients with CP even before they have an established diagnosis. Trial registration ClinicalTrials.gov ID 22013292 (reg. date 31/MAR/2023) for the CP-EDIT cohort and ID 22041835 (reg. date 31/MAR/2023) for the GO-PLAY trial.
Tick-borne encephalitis (TBE) is a viral tick-borne infection occurring in many parts of Europe and Asia as described in this review. Increasing TBE case numbers have been reported over recent decades. In Denmark the infection is rare (1-14 annual cases). The rise in TBE in Denmark is mainly driven by microfoci outside of Bornholm, primarily North Zealand. Clinical illness has a bi-phasic presentation: "summer-flu" which may be followed by a neuroinfection. No specific treatment exists, and mortality is less-than 1%. A considerable percentage of patients may experience neurological sequelae. TBE is preventable through vaccination.
The KN Motif and AnKyrin Repeat Domain 1 (KANK1) is proposed as a tumour suppressor gene, as its expression is reduced or absent in several types of tumour tissue, and over-expressing the protein inhibited the proliferation of tumour cells in solid cancer models. We report a novel germline loss of heterozygosity mutation encompassing the KANK1 gene in a young patient diagnosed with myelodysplastic neoplasm (MDS) with no additional disease-related genomic aberrations. To study the potential role of KANK1 in haematopoiesis, we generated a new transgenic mouse model with a confirmed loss of KANK1 expression. KANK1 knockout mice did not develop any haematological abnormalities; however, the loss of its expression led to alteration in the colony forming and proliferative potential of bone marrow (BM) cells and a decrease in hematopoietic stem and progenitor cells (HSPCs) population frequency. A comprehensive marker expression analysis of lineage cell populations indicated a role for Kank1 in lymphoid cell development, and total protein analysis suggests the involvement of Kank1 in BM cells’ cytoskeleton formation and mobility.
Platform trials focus on the perpetual testing of many interventions in a disease or a setting. These trials have lasting organizational, administrative, data, analytic, and operational frameworks making them highly efficient. The use of adaptation often increases the probabilities of allocating participants to better interventions and obtaining conclusive results. The COVID-19 pandemic showed the potential of platform trials as a fast and valid way to improved treatments. This review gives an overview of key concepts and elements using the Intensive Care Platform Trial (INCEPT) as an example.
Studies indicate that cocaine abuse in Denmark is rising. The drug can damage the midface's nasal tissues, cartilage, and bone. Diagnosing the cocaine-induced midline destructive lesions condition is difficult as patients may not admit to drug use. Thus, this review finds that physicians should ask about cocaine abuse in younger patients who present with midline destructions of unknown origin. Mild symptoms are reversible with total abstinence, which is why it is important to involve addiction services early. Besides drug abstinence, comprehensive treatment involves assistance from GPs, physiatrists, rhinologists, and plastic surgeons.
Polygenic risk scores (PRS) identify at-risk individuals for many common diseases. A discussion of strengths and limitations is carried out in this review. PRS complement traditional genetic testing and have shown utility in establishing a proper diagnosis and guiding primary and secondary prevention. Some individuals with high PRS have risks similar to those with monogenic predisposition. Limitations include potential misinterpretations, problems with application across ancestries, and limited usefulness in low-heritability traits. Despite its shortcomings PRS are predicted to play major roles in the future of personal medicine and genetic testing.
Background Globally, too many children are not sufficient physically active and have unhealthy food habits with the highest prevalence seen in families with low socio-economic-status (SES). Previous research has stated that schools are considered a key setting for promoting children physical activity (PA), food literacy (FL) and to improve their social health. School gardens is exactly such an intervention where teaching is relocated to an outdoor setting. Relocation of teaching, which is a central ingredient of school gardens, has shown to be positively related to increased PA in both boys and girls during the school day. School gardens aim towards practical experience and education in food and climate. PA is not I focus, but the school garden learning environment provides pupils opportunities to be active. Therefore, the FoodACT study aims to investigate how a school gardening intervention impact pupils school motivation, CL, FL and PA with a special attention on children with low SES. Methods The FoodACT study is designed in three sub-studies. An efficacy study, quantifying the effects of school gardens using a pre-post quasi-experimental design measuring the pupils’ FL, CL and SM. A within-subject-design association study, quantifying the influence of the school garden activities on pupils PA at two sessional periods during the intervention. An association study, investigating the contextual characteristics of the school garden activities using the systematic observations instrument, The Physical Activity Research and Assessment tool for Garden Observation (PARAGON), which can capture pupils’ movements and motions categorised in ‘overall PA-level’, ‘garden-related tasks’, ‘garden related motions’, ‘social associations’, and ‘interaction’. The observations is supplemented by pupil focus-group interviews about their experience of the school garden intervention. Results Study design, recruitment experience and pilot-study data on PA and garden activity observations will be presented. Funding The FoodACT study is funded by the Novo Nordisk Foundation (Grant no: 077522).
Two compounds, Na3[Eu(DPA)3]·14H2O and [Eu(DPA)(HDPA)(H2O)2]·4H2O, were created and the structure determined using single crystal X‐ray diffraction. The single crystal luminescent properties were compared and related to the Eu3+ coordination geometry. The formation of single crystals from solutions of Eu(CF3SO3)3 and H2DPA was found change with the pH value of the H2DPA solution. Mixtures of Na3[Eu(DPA)3]·14H2O and [Eu(DPA)(HDPA)(H2O)2]·4H2O were observed with a pH ratio between the two structures. While visual inspection showed that all samples contained both Na3[Eu(DPA)3]·14H2O and [Eu(DPA)(HDPA)(H2O)2]·4H2O, the PXRD and luminescence data did not immediately reveal that the samples were pure. Having discovered that the samples were indeed mixtures, quantification was attempted by Rietveld refinement of the PXRD data, and the luminescence spectra were compared to those from single crystals. As the data was not found to reveal that the samples were mixtures, even though we knew that this was the case, we must urge caution when inferring structure‐property relationships from powder samples. In this case we were able to isolate monophasic systems and do a comparative study, but this requires that the samples are identified as mixtures.
Atrial fibrillation (AF) is the most common cardiac arrhythmia. AF reduces the patients' quality of life and increases the risks of heart failure, ischaemic stroke, and death. The aetiology of AF is complex and involves multiple pathophysiological pathways. Comorbidities often coexist in patients with AF and contribute to the pathogenesis. The pathogenesis, the most common comorbidities, and possible individualized treatment options of AF are discussed in this review.
Lemierre's syndrome is rare and characterized by an oropharyngeal infection with thrombophlebitis of the internal jugular vein (IJV). Septic microemboli can spread to the lungs or abdomen. This case describes a patient who presented with a sore throat, unilateral swelling on the neck and diffuse abdominal pain. Imaging showed a thrombus in the right IJV. The patient developed septic shock and was treated with antibiotics, anticoagulants, and intensive care support. The most common causative organism is Fusobacterium necrophorum. Early diagnosis and treatment are crucial for reducing mortality.
The genomic landscape of colorectal cancer (CRC) is shaped by inactivating mutations in tumour suppressors such as APC, and oncogenic mutations such as mutant KRAS. Here we used genetically engineered mouse models, and multimodal mass spectrometry-based metabolomics to study the impact of common genetic drivers of CRC on the metabolic landscape of the intestine. We show that untargeted metabolic profiling can be applied to stratify intestinal tissues according to underlying genetic alterations, and use mass spectrometry imaging to identify tumour, stromal and normal adjacent tissues. By identifying ions that drive variation between normal and transformed tissues, we found dysregulation of the methionine cycle to be a hallmark of APC-deficient CRC. Loss of Apc in the mouse intestine was found to be sufficient to drive expression of one of its enzymes, adenosylhomocysteinase (AHCY), which was also found to be transcriptionally upregulated in human CRC. Targeting of AHCY function impaired growth of APC-deficient organoids in vitro, and prevented the characteristic hyperproliferative/crypt progenitor phenotype driven by acute deletion of Apc in vivo, even in the context of mutant Kras. Finally, pharmacological inhibition of AHCY reduced intestinal tumour burden in ApcMin/+ mice indicating its potential as a metabolic drug target in CRC.
DNA-stabilized silver nanoclusters (DNA-AgNCs) are easily tunable emitters with intriguing photophysical properties. Here, a DNA-AgNC with dual emission in the red and near-infrared (NIR) regions is presented. Mass spectrometry data showed that two DNA strands stabilize 18 silver atoms with a nanocluster charge of 12+. Besides determining the composition and charge of DNA2[Ag18]12+, steady-state and time-resolved methods were applied to characterize the picosecond red fluorescence and the relatively intense microsecond-lived NIR luminescence. During this process, the luminescence-to-fluorescence ratio was found to be excitation-intensity-dependent. This peculiar feature is very rare for molecular emitters and allows the use of DNA2[Ag18]12+ as a nanoscale excitation intensity probe. For this purpose, calibration curves were constructed using three different approaches based either on steady-state or time-resolved emission measurements. The results showed that processes like thermally activated delayed fluorescence (TADF) or photon upconversion through triplet-triplet annihilation (TTA) could be excluded for DNA2[Ag18]12+. We, therefore, speculate that the ratiometric excitation intensity response could be the result of optically activated delayed fluorescence.
DNA‐stabilized silver nanoclusters (DNA‐AgNCs) are easily tunable emitters with intriguing photophysical properties. Here, a DNA‐AgNC with dual emission in the red and near‐infrared (NIR) regions is presented. Mass spectrometry data showed that two DNA strands stabilize 18 silver atoms with a nanocluster charge of 12+. Besides determining the composition and charge of DNA2[Ag18]12+, steady‐state and time‐resolved methods were applied to characterize the picosecond red fluorescence and the relatively intense microsecond‐lived NIR luminescence. During this process, the luminescence‐to‐fluorescence ratio was found to be excitation‐intensity‐dependent. This peculiar feature is very rare for molecular emitters and allows the use of DNA2[Ag18]12+ as a nanoscale excitation intensity probe. For this purpose, calibration curves were constructed using three different approaches based either on steady‐state or time‐resolved emission measurements. The results showed that processes like thermally activated delayed fluorescence (TADF) or photon upconversion through triplet‐triplet annihilation (TTA) could be excluded for DNA2[Ag18]12+. We, therefore, speculate that the ratiometric excitation intensity response could be the result of optically activated delayed fluorescence.
Background Exotic and ornamental fish are highly popular companion animals in industrialized human societies resulting in a significant transcontinental trade of fish, invertebrates or aquatic plants. A major issue is the diseases associated with these organisms as they have a major impact on health of the fish in both public and private household aquaria. A secondary issue is the trade with these products, which potentially may expand the distribution area and spread a range of diseases to new habitats. Results We here describe how Poecilia reticulata Peters, 1859 (guppy), produced in a private household aquarium, were invaded by cercariae of an exotic trematode released by imported snails. The fish presented with severe clinical signs (tremor, flashing, scraping of body against objects). A standard parasitological examination and morphometric identification showed the disease to be caused by scale pocket infections with a species within the genus Transversotrema patialense (Soparkar, 1924), an exotic digenean trematode. Molecular identification by PCR, sequencing and BLAST analysis indicated that the parasite was an undescribed variant and the isolate was therefore named T. patialense var. hafniensis. Conclusions GenBank sequences for species within the genus Transversotrema are sparse and fragmented and we here present the entire 2646 bp sequence of encoding ribosomal RNA (partial 18S, ITS1, 5.8 S, ITS2, partial 28S) and a 1107 bp sequence of mitochondrial DNA (CO1). These are recommended for future diagnostics use and separation of sister species within T. patialense sensu lato. The fish were not purchased from a pet shop but produced in the home aquarium. This indicated that an infection pressure existed in the aquarium, where the source of infection was found to be exotic intermediate host snails Melanoides tuberculata (Müller, 1774), which originally were imported and purchased from a pet shop. They were recovered from the facility and shown to release infective cercariae, which infected naïve guppies. We frame that trade with infected snails may not only challenge the health of fish in aquaria but also represent a risk for spread of exotic fish diseases to new geographic regions where climate changes may facilitate their establishment.
The retreat of glaciers worldwide is causing changes in species diversity, community composition and species interactions. However, the impact of glacier retreat on interaction diversity and ecological networks remains poorly understood. An integrative understanding of network dynamics is of major importance to supporting biodiversity and ecosystem functioning after glacier extinction. Here, we address how glacier retreat affects the frequency, complexity, and diversity of plant–insect interactions, both directly and indirectly through biodiversity change. We surveyed flower visitors and analyzed visitation networks along the foreland of the Mont Miné glacier (Valais, Switzerland). As glacier retreat impacts both plant and insect communities, we observe sharp changes in the diversity of plant–insect interactions and the structure of pollination networks. We find an increase in the frequency of interactions following glacier retreat, but an ultimate decrease with glacier extinction. After controlling for the effects of floral diversity, interaction frequency showed a regular ‘universal’ pattern. Accordingly, the complexity of pollination networks and interaction diversity tends to change at constant rates with glacier retreat. Our results indicate that glacier retreat decreases biodiversity and influences the stability of ecological networks. The good news is that increasing floral diversity counteracts these impacts by increasing interaction diversity and supporting complexity. Slowing down woody encroachment and enhancing floral diversity may therefore be key strategies for halting the erosion of ecological networks while increasing biodiversity and ecosystem functioning.
Photoswitches are molecular systems that are chemically transformed subsequent to interaction with light and they find potential application in many new technologies. The design and discovery of photoswitch candidates require intricate molecular engineering of a range of properties to optimize a candidate to a specific applications, a task which can be tackled efficiently using quantum chemical screening procedures. In this paper, we perform a large scale screening of approximately half a million bicyclic diene photoswitches in the context of molecular solar thermal energy storage using ab initio quantum chemical methods. We further device an efficient strategy for scoring the systems based on their predicted solar energy conversion efficiency and elucidate potential pitfalls of this approach. Our search through the chemical space of bicyclic dienes reveals systems with unprecedented solar energy conversion efficiencies and storage densities that show promising design guidelines for next generation molecular solar thermal energy storage systems.
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