3D printing technologies have been considered an important technology due to the ease manufacturing of objects, freedom of design, waste minimization, and fast prototyping. In chemistry, this ...technology potentializes the fabrication of conductive electrodes in large scale for sensing applications. Herein, we reported the modification of a 3D printed graphene electrode with Prussian blue. The modified electrode (3DGrE/PB) was characterized by microscopy (SEM and AFM) and spectroscopic techniques, and its electrochemical properties were compared to the traditional electrodes: glassy carbon, gold, and platinum. The 3DGrE/PB was used in the sensing of hydrogen peroxide in real-world samples of milk and mouthwash, and the results obtained according to the technique of batch-injection analysis were satisfactory for the concentration range typically found in such samples. Thus, 3DGrE/PB can be used as a new platform for sensing of molecular targets.
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IJS, KILJ, NUK, PNG, UL, UM
We investigated whether morphine and its pro-drug codeine are substrates of the highly genetically polymorphic organic cation transporter OCT1 and whether OCT1 polymorphisms may affect morphine and ...codeine pharmacokinetics in humans.
Morphine showed low transporter-independent membrane permeability (0.5×10−6cm/s). Morphine uptake was increased up to 4-fold in HEK293 cells overexpressing human OCT1. The increase was concentration-dependent and followed Michaelis-Menten kinetics (KM=3.4μM, VMAX=27pmol/min/mg protein). OCT1-mediated morphine uptake was abolished by common loss-of-function polymorphisms in the OCT1 gene and was strongly inhibited by drug-drug interactions with irinotecan, verapamil and ondansetron. Morphine uptake in primary human hepatocytes was strongly reduced by MPP+, an inhibitor of organic cation transporters, and morphine was not a substrate of OCT3, the other organic cation transporter expressed in human hepatocytes. In concordance with the in vitro data, morphine plasma concentrations in healthy volunteers were significantly dependent on OCT1 polymorphisms. After codeine administration, the mean AUC of morphine was 56% higher in carriers of loss-of-function OCT1 polymorphisms compared to non-carriers (P=0.005). The difference remained significant after adjustment for CYP2D6 genotype (P=0.03). Codeine itself had high transporter-independent membrane permeability (8.2×10−6cm/s). Codeine uptake in HEK293 cells was not affected by OCT1 overexpression and OCT1 polymorphisms did not affect codeine AUCs.
In conclusion, OCT1 plays an important role in the hepatocellular uptake of morphine. Carriers of loss-of-function OCT1 polymorphisms may be at higher risk of adverse effects after codeine administration, especially if they are also ultra-rapid CYP2D6 metabolizers.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
The successive addition of H atoms to CO in the solid phase has been hitherto regarded as the primary route to form methanol in dark molecular clouds. However, recent Monte Carlo simulations of ...interstellar ices alternatively suggested the radical-molecule H-atom abstraction reaction CH3O + H2CO → CH3OH + HCO, in addition to CH3O + H → CH3OH, as a very promising and possibly dominating (70%–90%) final step to form CH3OH in those environments. Here, we compare the contributions of these two steps leading to methanol by experimentally investigating hydrogenation reactions on H2CO and D2CO ices, which ensures comparable starting points between the two scenarios. The experiments are performed under ultrahigh vacuum conditions and astronomically relevant temperatures, with H:H2CO (or D2CO) flux ratios of 10:1 and 30:1. The radical-molecule route in the partially deuterated scenario, CHD2O + D2CO → CHD2OD + DCO, is significantly hampered by the isotope effect in the D-abstraction process, and can thus be used as an artifice to probe the efficiency of this step. We observe a significantly smaller yield of D2CO + H products in comparison to H2CO + H, implying that the CH3O-induced abstraction route must play an important role in the formation of methanol in interstellar ices. Reflection-absorption infrared spectroscopy and temperature-programmed desorption-quadrupole mass spectrometry analyses are used to quantify the species in the ice. Both analytical techniques indicate constant contributions of ∼80% for the abstraction route in the 10–16 K interval, which agrees well with the Monte Carlo calculations. Additional H2CO + D experiments confirm these conclusions.
Abstract
Methane is typically thought to be formed in the solid state on top of cold interstellar icy grain mantles via the successive atomic hydrogenation of a carbon atom. In the current work we ...investigate the role of molecular hydrogen in the CH
4
reaction network. We make use of an ultrahigh vacuum cryogenic setup combining an atomic carbon atom beam with atomic and/or molecular beams of hydrogen and deuterium on a water ice. These experiments lead to the formation of methane isotopologues detected in situ through reflection absorption infrared spectroscopy. Most notably, CH
4
is experimentally formed by combining C atoms with only H
2
on amorphous solid water, albeit more slowly than in experiments where H atoms are also present. Furthermore, CH
2
D
2
is detected in an experiment involving C atoms with H
2
and D
2
on H
2
O ice. CD
4
, however, is only formed when D atoms are present in the experiment. These findings have been rationalized by means of computational and theoretical chemical insights. This leads to the following conclusions: (a) the reaction C + H
2
→ CH
2
takes place, although it is not barrierless for all binding sites on water, (b) the reaction CH + H
2
→ CH
3
is barrierless, but has not yet been included in astrochemical models, (c) the reactions CH
2
+ H
2
→ CH
3
+ H and CH
3
+ H
2
→ CH
4
+ H can take place only via a tunneling mechanism, and (d) molecular hydrogen possibly plays a more important role in the solid-state formation of methane than assumed so far.
SARS-CoV-2, like other coronaviruses, builds a membrane-bound replication organelle (RO) to enable RNA replication
. The SARS-CoV-2 RO is composed of double membrane vesicles (DMVs) tethered to the ...endoplasmic reticulum (ER) by thin membrane connectors
, but the viral proteins and the host factors involved are currently unknown. Here we identify the viral non-structural proteins (NSPs) that generate the SARS-CoV-2 RO. NSP3 and NSP4 generate the DMVs while NSP6, through oligomerization and an amphipathic helix, zippers ER membranes and establishes the connectors. The NSP6ΔSGF mutant, which arose independently in the α, β, γ, η, ι, and λ variants of SARS-CoV-2, behaves as a gain-of-function mutant with a higher ER-zippering activity. We identified three main roles for NSP6: to act as a filter in RO-ER communication allowing lipid flow but restricting access of ER luminal proteins to the DMVs, to position and organize DMV clusters, and to mediate contact with lipid droplets (LDs) via the LD-tethering complex DFCP1-Rab18. NSP6 thus acts as an organizer of DMV clusters and can provide a selective track to refurbish them with LD-derived lipids. Importantly, both properly formed NSP6 connectors and LDs are required for SARS-CoV-2 replication. Our findings, uncovering the biological activity of NSP6 of SARS-CoV-2 and of other coronaviruses, have the potential to fuel the search for broad antiviral agents.
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
Glioblastoma (GB) is the most aggressive primary malignant brain tumor and is associated with short survival. O-GlcNAcylation is an intracellular glycosylation that regulates protein function, ...enzymatic activity, protein stability, and subcellular localization. Aberrant O-GlcNAcylation is related to the tumorigenesis of different tumors, and mounting evidence supports O-GlcNAc transferase (OGT) as a potential therapeutic target. Here, we used two human GB cell lines alongside primary human astrocytes as a non-tumoral control to investigate the role of O-GlcNAcylation in cell proliferation, cell cycle, autophagy, and cell death. We observed that hyper O-GlcNAcylation promoted increased cellular proliferation, independent of alterations in the cell cycle, through the activation of autophagy. On the other hand, hypo O-GlcNAcylation inhibited autophagy, promoted cell death by apoptosis, and reduced cell proliferation. In addition, the decrease in O-GlcNAcylation sensitized GB cells to the chemotherapeutic temozolomide (TMZ) without affecting human astrocytes. Combined, these results indicated a role for O-GlcNAcylation in governing cell proliferation, autophagy, cell death, and TMZ response, thereby indicating possible therapeutic implications for treating GB. These findings pave the way for further research and the development of novel treatment approaches which may contribute to improved outcomes and increased survival rates for patients facing this challenging disease.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Neurodegenerative diseases such as Alzheimer's disease (AD) have long been acknowledged as mere disorders of the central nervous system (CNS). However, in recent years the gut with its autonomous ...nervous system and the multitude of microbial commensals has come into focus. Changes in gut properties have been described in patients and animal disease models such as altered enzyme secretion or architecture of the enteric nervous system. The underlying cellular mechanisms have so far only been poorly investigated. An important organelle for integrating potentially toxic signals such as the AD characteristic A-beta peptide is the primary cilium. This microtubule-based signaling organelle regulates numerous cellular processes. Even though the role of primary cilia in a variety of developmental and disease processes has recently been recognized, the contribution of defective ciliary signaling to neurodegenerative diseases such as AD, however, has not been investigated in detail so far. The AD mouse model 5xFAD was used to analyze possible changes in gut functionality by organ bath measurement of peristalsis movement. Subsequently, we cultured primary enteric neurons from mutant mice and wild type littermate controls and assessed for cellular pathomechanisms. Neurite mass was quantified within transwell culturing experiments. Using a combination of different markers for the primary cilium, cilia number and length were determined using fluorescence microscopy. 5xFAD mice showed altered gut anatomy, motility, and neurite mass of enteric neurons. Moreover, primary cilia could be demonstrated on the surface of enteric neurons and exhibited an elongated phenotype in 5xFAD mice. In parallel, we observed reduced
expression, a key signaling molecule that regulates Wnt signaling, which is regulated in part via ciliary associated mechanisms. Both results could be recapitulated via in vitro treatments of enteric neurons from wild type mice with A-beta. So far, only a few reports on the probable role of primary cilia in AD can be found. Here, we reveal for the first time an architectural altered phenotype of primary cilia in the enteric nervous system of AD model mice, elicited potentially by neurotoxic A-beta. Potential changes on the sub-organelle level-also in CNS-derived neurons-require further investigations.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Tropical forests are being exposed to increasing levels of habitat loss and fragmentation, threatening the maintenance of global biodiversity. However, the effect that land-use change may have on the ...spatial dissimilarities in taxonomic and functional composition of remaining assemblages (i.e., taxonomic/functional β-diversity) remains poorly understood. We examined a large vegetation database from an old and severely fragmented Atlantic forest landscape to test two alternative hypotheses: (1) tree assemblages experience a taxonomic and functional homogenization (reduced β-diversity) between forest fragments and along forest edges, or alternatively, (2) these edge-affected forests show increased taxonomic and functional differentiation (increased β-diversity) when compared to forest interior (reference) stands. Taxonomic and functional β-diversity were examined via novel abundance-based metrics and considering functional traits related to plant dispersion, recruitment, and growth. Overall, taxonomic β-diversity among fragments was significantly higher than among edge and reference plots. Edge plots also showed higher β-diversity than reference plots, but only when considering dominant species. In functional terms, β-diversity among reference plots was also lower than among forest fragments and among edge plots. These patterns support the landscape-divergence hypothesis, which postulates that variable human disturbances among forest fragments and along forest edges can lead to contrasting trajectories of vegetation changes, thus increasing the compositional and functional differentiation of tree communities in these emerging environments. Our results also show that such differentiation can preserve landscape-wide biodiversity, thus overriding negative effects of habitat fragmentation on local (α) diversity. Therefore, our findings demonstrate that forest fragments and forest edges can be more valuable for maintaining species diversity and ecosystem function in fragmented tropical landscapes than previously thought.
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BFBNIB, FZAB, GIS, IJS, INZLJ, KILJ, NLZOH, NMLJ, NUK, OILJ, PNG, SAZU, SBCE, SBMB, UL, UM, UPUK, ZRSKP
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