Miracle-dz/UniFAR-FTUs huggingface dataset:
PET imaging studies to investigate functional expression of mGluR2 using [11C]mG2P001

Metabotropic glutamate receptor 2 (mGluR2) has been extensively studied for the treatment of various neurological and psychiatric disorders. Understanding of the mGluR2 function is pivotal in supporting the drug discovery targeting mGluR2. Herein, the positive allosteric modulation of mGluR2 was investigated via the in vivo positron emission tomography (PET) imaging using 2-((4-(2-[11C]methoxy-4-(trifluoromethyl)phenyl)piperidin-1-yl)methyl)-1-methyl-1H-imidazo[4,5-b]pyridine ([11C]mG2P001). Distinct from the orthosteric compounds, pretreatment with the unlabeled mG2P001, a potent mGluR2 positive allosteric modulator (PAM), resulted in a significant increase instead of decrease of the [11C]mG2P001 accumulation in rat brain detected by PET imaging. Subsequent in vitro studies with [3H]mG2P001 revealed the cooperative binding mechanism of mG2P001 with glutamate and its pharmacological effect that contributed to the enhanced binding of [3H]mG2P001 in transfected CHO cells expressing mGluR2. The in vivo PET imaging and quantitative analysis of [11C]mG2P001 in non-human primates (NHPs) further validated the characteristics of [11C]mG2P001 as an imaging ligand for mGluR2. Self-blocking studies in primates enhanced accumulation of [11C]mG2P001. Altogether, these studies show that [11C]mG2P001 is a sensitive biomarker for mGluR2 expression and the binding is affected by the tissue glutamate concentration.
Goldilocks and RNA: where Mg2+ concentration is just right

Magnesium, the most abundant divalent cation in cells, catalyzes RNA cleavage but also promotes RNA folding. Because folding can protect RNA from cleavage, we predicted a “Goldilocks peak”, which is a local maximum in RNA lifetime at the Mg2+ concentration required for folding. Here we use simulation and experiment to discover an innate yet sophisticated mechanism of control of RNA lifetime. By simulation we characterized the RNA Goldilocks peak and its dependence on cleavage parameters and extent of folding. Supporting experiments with yeast tRNAPhe and Tetrahymena ribozyme P4-P6 domains show that structured RNA can inhabit a Goldilocks peak in vitro. The Goldilocks peaks are tunable by differences in cleavage rate constants, Mg2+ binding cooperativity, and Mg2+ affinity. Broad ranges of those folding and cleavage parameters produce Goldilocks peaks of different intensities. Goldilocks behavior allows ultrafine control of RNA chemical lifetime, whereas non-folding RNAs do not display a Goldilocks peak. In sum, the effects of Mg2+ on RNA persistence are expected to be pleomorphic, both protecting and degrading RNA. In evolutionary context, Goldilocks behavior may have shaped RNA in an early Earth environment containing Mg2+ and other metals.
Epigenetic Mediation of AKT1 rs1130233’s Effect on Delta-9-Tetrahydrocannabinol-Induced Medial Temporal Function during Fear Processing

High doses of delta-9-tetrahydrocannabinol (THC), the main psychoactive component of cannabis, have been shown to have anxiogenic effects. Also, THC effects have been shown to be modulated by genotype, including the single nucleotide polymorphism (SNP) rs1130233 at the protein kinase AKT1 gene, a key component of the dopamine signaling cascade. As such, it is likely that epigenetic methylation around this SNP may affect AKT gene expression, which may in turn impact on the acute effects of THC on brain function. We investigated the genetic (AKT1 rs1130233) and epigenetic modulation of brain function during fear processing in a 2-session, double-blind, cross-over, randomized placebo-controlled THC administration, in 36 healthy males. Fear processing was assessed using an emotion (fear processing) paradigm, under functional magnetic resonance imaging (fMRI). Complete genetic and fMRI data was available for 34 participants. THC caused an increase in anxiety and transient psychotomimetic symptoms and para-hippocampal gyrus/ amygdala activation. Number of A alleles at the AKT1 rs1130233 SNP, and percentage methylation at the CpG11-12 site, were independently associated with a greater effect of THC on activation in a network of brain regions including left and right parahippocampal gyri, respectively. AKT1 rs1130233 moderation of the THC effect on left parahippocampal activation persisted after covarying for methylation percentage, and was partially mediated in sections of the left parahippocampal gyrus/ hippocampus by methylation percentage. These results may offer an example of how genetic and epigenetic variations influence the psychotomimetic and neurofunctional effects of THC.
The Ventral Midline Thalamus Mediates Successful Deliberation by Coordinating Prefrontal and Hippocampal Neural Activity

When faced with difficult choices, the possible outcomes are considered through a process known as deliberation. In rats, deliberation can be reflected by pause-and-reorienting behaviors, better known as Vicarious Trial and Errors (VTEs). While VTEs are thought to require medial prefrontal cortex (mPFC) and dorsal hippocampal (dHPC) interactions, no work has demonstrated such a dual requirement. The nucleus reuniens (Re) of the ventral midline thalamus is anatomically connected with both the mPFC and dHPC, is required for HPC-dependent spatial memory tasks, and is critical for mPFC-dHPC neural synchronization. Currently, it is unclear if, or how, the Re is involved in deliberation. Therefore, by examining the role of the Re on VTE behaviors, we can better understand the anatomical and physiological mechanisms supporting deliberation. Here, we examined the impact of Re suppression on VTE behaviors and mPFC-dHPC theta synchrony during well-learned performance of a HPC-dependent delayed alternation (DA) task. Pharmacological suppression of the Re increased VTE behaviors that resulted in erroneous choices. These errors were best characterized as perseverative behaviors, where at the decision-point, some rats repeatedly turned in the direction that previously yielded no reward, while simultaneously deliberating. These ‘failed’ deliberations were associated with a reduction in mPFC-dHPC theta coherence at the choice-point. Importantly, the reduction in mPFC-dHPC theta synchrony observed during VTE behaviors was almost entirely driven by Re-suppression induced changes to the mPFC theta oscillation. Our findings suggest that the Re is important for successful deliberation, and mPFC-dHPC interactions, by coordinating local theta oscillations in the mPFC.
Programmable cleavage of linear double-stranded DNA by combined action of Argonaute CbAgo from Clostridium butyricum and nuclease deficient RecBC helicase from E. coli

Prokaryotic Argonautes (pAgos) use small nucleic acids as specificity guides to cleave single-stranded DNA at complementary sequences. DNA targeting function of pAgos creates attractive opportunities for DNA manipulations that require programmable DNA cleavage. Discovery of mesophilic Argonautes active at physiological temperature places pAgos closer to their possible application for genome editing as a simpler alternative to CRISPR/Cas nucleases. Currently, the use of mesophilic pAgos as programmable DNA endonucleases is hampered by their poor action on double-stranded DNA (dsDNA), mainly due to their inability to invade the DNA duplex. The present study demonstrates that efficient in vitro cleavage of double-stranded DNA by mesophilic Argonaute CbAgo from Clostridium butyricum can be activated via the DNA strand unwinding activity of nuclease deficient mutant of RecBC DNA helicase from Escherichia coli (referred to as RecBexo-C). Properties of CbAgo and characteristics of simultaneous cleavage of complementary DNA strands in concurrence with DNA strand unwinding by RecBexo-C were thoroughly explored using 0.3-25 kb DNA substrates. When combined with RecBexo-C helicase, CbAgo was capable of cleaving target sequences located 11-12.5 kb from the ends of linear dsDNA at 37ºC. Our study demonstrates that CbAgo with RecBexo-C can be programmed to generate dsDNA fragments flanked with custom-designed single-stranded overhangs suitable for ligation with compatible DNA fragments. At present, the combination of CbAgo and RecBexo-C represents the most efficient mesophilic DNA-guided DNA-cleaving programmable endonuclease for use in diagnostic and synthetic biology methods that require sequence-specific nicking/cleavage of dsDNA at any desired location.
TGFβ2 regulates human trabecular meshwork cell contractility via ERK and ROCK pathways with distinct signaling crosstalk dependent on the culture substrate

Transforming growth factor beta 2 (TGFβ2) is a major contributor to the pathologic changes occurring in human trabecular meshwork (HTM) cells in primary open-angle glaucoma. Receptor binding of TGFβ2 activates non-canonical extracellular-signal-regulated kinase (ERK) and Rho-associated kinase (ROCK) signaling pathways, both broadly affecting HTM cell behavior. However, exactly how these signaling pathways converge to regulate pathologic HTM cell contractility associated with glaucomatous dysfunction is unclear. Here, we investigated the molecular mechanism underlying TGFβ2-induced pathologic HTM cell contractility, and the crosstalk between ERK and ROCK signaling pathways. We compared soft biomimetic hydrogels composed of collagen type I, elastin-like polypeptide, and hyaluronic acid with conventional stiff glass coverslips. Results show that HTM cell morphology and filamentous (F)-actin organization was affected by the underlying culture substrate: TGFβ2 increased HTM cell contractility via ERK and ROCK signaling pathways by differentially regulating F-actin, α-smooth muscle actin, fibronectin, and phospho-myosin light chain in cells grown on hydrogels compared to glass. Importantly, we showed that ERK inhibition further increased TGFβ2-induced phospho-myosin light chain levels in HTM cells on hydrogels, but not on glass, which translated into hypercontractility of three-dimensional (3D) HTM cell-laden hydrogels. ROCK inhibition had precisely the opposite effects and potently relaxed the TGFβ2-induced hydrogels. This suggests that ERK signaling negatively regulates ROCK-mediated HTM cell contractility, and that impairment of this crosstalk balance contributes to the pathologic contraction associated with the glaucomatous stressor TGFβ2. These findings emphasize the critical importance of using 3D tissue-mimetic extracellular matrix substrates for investigating HTM cell physiology and glaucomatous pathophysiology in vitro.
Hypervariable-Locus Melting Typing (HLMT): a novel, fast and inexpensive sequencing-free approach to pathogen typing based on High Resolution Melting (HRM) analysis

Objectives Subspecies pathogen typing is a pivotal tool to detect the emergence of high-risk clones in hospital settings and to limit their spreading among patients. Unfortunately, the most used subspecies typing methods (i.e. Pulsed-field Gel Electrophoresis - PFGE, Multi-Locus Sequence Typing - MLST and Whole Genome Sequencing - WGS) are too expensive and time consuming to be suitable for real-time surveillance. Here we present Hypervariable-Locus Melting Typing (HLMT), a novel subspecies typing approach based on High Resolution Melting (HRM) analysis, which allows pathogen typing in a few hours and with ∼5 euros per sample. Methods HLMT types the strains by clustering them using melting temperatures (HLMT-clustering) and/or by assigning them to Melting Types (MTs) on the basis of a reference dataset (HLMT-assignment). We applied HLMT (clustering and typing) to 134 Klebsiella pneumoniae strains collected during outbreaks or surveillance programs in four hospitals. Then, we compared HLMT typing results to PFGE, MLST and WGS. Results HLMT-clustering distinguishes most of the K. pneumoniae high-risk clones with a sensitivity comparable to PFGE and MLST. It also drawed surveillance epidemiological curves comparable to those obtained by MLST, PFGE and WGS typing. Furthermore, the results obtained by HLMT-assignment were coherent to MLST for 96% of the typed strains with a Jaccard index of 0.912. Conclusions HLMT is a fast and scalable method for pathogen typing, suitable for real-time hospital microbiological surveillance. HLMT is also inexpensive and thus it is applicable to infection control programs in low-middle income countries.
Intrinsic Network Activity Reflects the Fluctuating Experience of Tonic Pain

Although we know sensation is continuous, research on long-lasting and continuously changing stimuli is scarce and the dynamic nature of ongoing cortical processing is largely neglected. In a longitudinal study with 152 fMRI sessions, participants were asked to continuously rate the intensity of applied tonic heat pain for 20 minutes. Using group independent component analysis and dual-regression, we extracted the subjects’ time courses of intrinsic network activity. The relationship between the dynamic fluctuation of network activity with the varying time courses of three pain processing entities was computed: pain intensity, the direction of pain intensity changes and temperature. We were able to dissociate the spatio-temporal patterns of objective (temperature) and subjective (pain intensity/changes of pain intensity) aspects of pain processing in the human brain. We found two somatosensory networks with distinct functions: one network which encodes the small fluctuations in temperature and consists mainly of bilateral SI. A second right-lateralised network that encodes the intensity of the subjective experience of pain consists of SI, SII, the PCC, and the thalamus. We revealed the somatosensory dynamics that build up towards a current subjective percept of pain. The timing suggests a cascade of subsequent processing steps towards the current pain percept.
Macrophage Circadian Rhythms are Differentially Affected Based on Stimuli

Macrophages are white blood cells of the innate immune system that play disparate roles in homeostasis and immune responses. As a result, they have the capability to alter their phenotypes to pro-inflammatory (M1) or anti-inflammatory (M2) subtypes in response to their environment. 8-15% of the macrophage transcriptome has circadian oscillations, including genes closely related to their functioning. As circadian rhythms are also associated with cellular phenotypes, we hypothesized that the polarization of macrophages to opposing subtypes might differently affect their circadian rhythms. We tested this by tracking the circadian rhythms of the mouse model macrophage cell line, RAW 264.7, which was stably transfected with Bmal1:luc and Per2:luc reporters, representing a positive and a negative component of the core molecular clock. The strength of rhythmicity was assessed using three measures: the relative power of the circadian band in the power spectral density, the rhythmicity index computed as the height of the third peak of the correlogram, and the maximum value of the chi square periodorgram. The period and amplitudes of the de-trended, smoothed time-series were estimated both by fitting to a damped cosine curve and by identifying the peak and trough of each cycle. M1 polarization decreased amplitudes and rhythmicities of both Bmal1:luc and Per2:luc, but did not significantly affect periods, while M2 polarization increased periods and caused no substantial alterations to amplitudes or rhythmicity. As macrophage phenotypes are also altered in the presence of cancer cells, we then tested the circadian effects of conditioned media from two mouse breast cancer cell lines on macrophage circadian rhythms. Media from highly agressive 4T1 cells caused loss of rhythmicity, while media from less aggressive EMT6 cells yielded no changes. As macrophages are known to play roles in tumors, and oncogenic features are associated with circadian rhythms, we also tested whether conditioned media from macrophages can alter circadian rhythms of cancer cells. We found that conditioned media from RAW 264.7 cells resulted in lower circadian rhythmicities and periods, but higher circadian amplitudes in human osteosarcoma, U2OS-Per2:luc cells. Taken together, our study shows that different circadian characteristics exist based on macrophage phenotypes, and suggests further that there is an association between circadian rhythms and macrophage polarization state. Additionally, our data shows that macrophages treated with breast cancer-conditioned media have circadian phenotypes similar to those of the M1 subtype, and cancer cells treated with macrophage-conditioned media have circadian alterations, providing insight to another level of cross-talk between macrophages and cancer cells that merits further investigation.
Activation of Adipocyte mTORC1 Increases Milk Lipids in a Mouse Model of Lactation

Human milk is the recommended nutrient source for newborns. The mammary gland comprises multiple cell types including epithelial cells and adipocytes. The contributions of mammary adipocytes to breast milk composition and the intersections between mammary nutrient sensing and milk lipids are not fully understood. A major nutrient sensor in most tissues is the mechanistic target of rapamycin 1 (mTORC1). To assess the role of excess nutrient sensing on mammary gland structure, function, milk composition, and offspring weights, we used an Adiponectin-Cre driven Tsc1 knockout model of adipocyte mTORC1 hyperactivation. Our results show that the knockout dams have higher milk fat contributing to higher milk caloric density and heavier offspring weight during lactation. Additionally, milk of knockout dams displayed a lower percentage of saturated fatty acids, higher percentage of monounsaturated fatty acids, and a lower milk ω6: ω3 ratio driven by increases in docosahexaenoic acid (DHA). Mammary gland gene expression analyses identified changes in eicosanoid metabolism, adaptive immune function, and contractile gene expression. Together, these results suggest a novel role of adipocyte mTORC1 in mammary gland function and morphology, milk composition, and offspring growth.
Two faces of perceptual awareness during the attentional blink: Gradual and discrete.

In a series of experiments, the nature of perceptual awareness during the attentional blink was investigated. Previous work has considered the attentional blink as a discrete, all-or-none phenomenon, indicative of general access to conscious awareness. Using continuous report measures in combination with mixture modeling, the outcomes showed that perceptual awareness during the attentional blink can be a gradual phenomenon. Awareness was not exclusively discrete, but also exhibited a gradual characteristic whenever the spatial extent of attention induced by the first target spanned more than a single location. Under these circumstances, mental representations of blinked targets were impoverished, but did approach the actual identities of the targets. Conversely, when the focus of attention covered only a single location, there was no evidence for any partial knowledge of blinked targets. These two different faces of awareness during the attentional blink challenge current theories of both awareness and temporal attention, which cannot explain the existence of gradual awareness of targets during the attentional blink. To account for the current outcomes, an adaptive gating model is proposed that casts awareness on a continuum between gradual and discrete, rather than as being of either single kind. (PsycInfo Database Record (c) 2021 APA, all rights reserved).
Less rain and rainy days - lessons from 45 years of rainfall data (1971 to 2015) in the Kathmandu Valley, Nepal

Understanding spatio-temporal variability in rainfall patterns is crucial for evaluating water balances needed for water resources planning and management. This paper investigates spatio-temporal variability in rainfall and assesses the frequency of daily rainfall observations from seven stations in the Kathmandu Valley, Nepal, from 1971–2015. Daily rainfall totals were classified into five classes, namely, A (light rain, daily rainfall < 10 mm in a day), B (between 10–50 mm), C (between 50–100 mm), D (between 100–150 mm) and E (> 150 mm). The relationship between daily rainfall and rainfall frequency of various rainfall rate classes were analysed. Kriging method was used for interpolation in interpreting seasonal and annual rainfall data and spatial maps were generated using QGIS. The Mann-Kendall (MK) test was performed to determine the temporal trends and Theil-Sen’s (TS) slope estimator was used in quantifying the magnitude of trends. Mountain stations showed a decreasing trend in rainfall for all seasons, ranging from − 8.4 mm/year at Sankhu to -21.8 mm/year at Thankot, whereas, a mixed pattern was found on the Valley floor. Mean annual rainfall in the Valley was 1610 mm. Both annual rainfall and the number of rainy days decreased in the Kathmandu Valley over the study period. The study indicated a significant reduction in rainfall after 2000. Since springs and shallow groundwater are the primary sources of water supply for residents in the Kathmandu Valley, it is apparent that decreasing rainfall will have (and is already having) an adverse impact on domestic, industrial, and agricultural water supplies, and the livelihoods of people.
Storage Behaviour of Ettinger, Fuerte and Hass Avocados Grown on Mexican Rootstock in Cyprus

SummaryThe storage behaviour of avocado cvs Ettinger, Fuerte and Hass was studied at 2.2°, 4.4°, 6.6° and 8.8°C for 0, 1, 2, 3, 4 and 5 weeks. Ettinger stored well for one week at 4.4°, 6.6° or 8.8°C, Fuerte for three weeks at 4.4°C and Hass for three weeks at 2.2° or 4.4°C. Chilling injury, internal darkening and the time required for ripening after removal to 20°C were the limiting factors determining the storage temperature and period for each cultivar. Chilling injury symptoms were visible during storage and not only after removal to 20°C. Internal darkening, observed at both chilling and non-chilling temperatures, increased with length of storage. Weight loss during storage or after removal did not cause any detrimental effect on quality. Ettinger was the most sensitive cultivar to chilling injury and Hass the least sensitive. The response to temperature of the three cultivars differed from that reported for same cultivars in other countries, depending on temperature and length of storage.
Application of numerical modelling to scaling-up of electrically induced extraction from an organic mixture using an ionic liquid

Liquid-liquid extraction provides an environmentally friendly process as an alternative to azeotropic distillation, pervaporation and reverse osmosis because these techniques require the use of large amounts of energy, may involve volatile organic compounds, and operation at high pressure. Ionic liquids (ILs) continue to gain wide recognition as potential environmentally friendly solvents due to their unique properties. However due to their current high cost, their use in industry is seriously limited without an efficient methodology for recovery and recycle. In this paper we describe an innovative methodology for a liquid-liquid extraction process based on an electrically induced emulsion of an ionic liquid as the extracting solvent dispersed in an organic mixture. This offers a most efficient exploitation of the solvent. On the other hand we present our own design of a pilot (semi-industrial) scale extractor based on this methodology and which demonstrates effective recovery of the ionic liquid. In order to achieve this goal we used a numerical modelling tool implemented using our own simulation software based on the finite element method. We also used our original previous experience with generating and investigating liquid-liquid electrosprays using phase Doppler anemometry. Finally we present recommendations for contactor geometry and for the preferred operating conditions for the extractor.