An Extended Helical Conformation in Domain 3a of Munc18-1 Provides a Template for SNARE (Soluble N-Ethylmaleimide-sensitive Factor Attachment Protein Receptor) Complex Assembly

Munc18-1, a SEC1/Munc18 protein and key regulatory protein in synaptic transmission, can either promote or inhibit SNARE complex assembly. Although the binary inhibitory interaction between Munc18-1 and closed syntaxin 1 is well described, the mechanism of how Munc18-1 stimulates membrane fusion remains elusive. Using a reconstituted assay that resolves vesicle docking, priming, clamping, and fusion during synaptic exocytosis, we show that helix 12 in domain 3a of Munc18-1 stimulates SNAREpin assembly and membrane fusion. A single point mutation (L348R) within helix 12 selectively abolishes VAMP2 binding and the stimulatory function of Munc18-1 in membrane fusion. In contrast, targeting a natural switch site (P335A) at the start of helix 12, which can result in an extended α-helical conformation, further accelerates lipid-mixing. Together with structural modeling, the data suggest that helix 12 provides a folding template for VAMP2, accelerating SNAREpin assembly and membrane fusion. Analogous SEC1/Munc18-SNARE interactions at other transport steps may provide a general mechanism to drive lipid bilayer merger. At the neuronal synapse, Munc18-1 may convert docked synaptic vesicles into a readily releasable pool.     

Transmedulla Neurons in the Sky Compass Network of the Honeybee (Apis mellifera) Are a Possible Site of Circadian Input

Honeybees are known for their ability to use the sun’s azimuth and the sky’s polarization pattern for spatial orientation. Sky compass orientation in bees has been extensively studied at the behavioral level but our knowledge about the underlying neuronal systems and mechanisms is very limited. Electrophysiological studies in other insect species suggest that neurons of the sky compass system integrate information about the polarization pattern of the sky, its chromatic gradient, and the azimuth of the sun. In order to obtain a stable directional signal throughout the day, circadian changes between the sky polarization pattern and the solar azimuth must be compensated. Likewise, the system must be modulated in a context specific way to compensate for changes in intensity, polarization and chromatic properties of light caused by clouds, vegetation and landscape. The goal of this study was to identify neurons of the sky compass pathway in the honeybee brain and to find potential sites of circadian and neuromodulatory input into this pathway. To this end we first traced the sky compass pathway from the polarization-sensitive dorsal rim area of the compound eye via the medulla and the anterior optic tubercle to the lateral complex using dye injections. Neurons forming this pathway strongly resembled neurons of the sky compass pathway in other insect species. Next we combined tracer injections with immunocytochemistry against the circadian neuropeptide pigment dispersing factor and the neuromodulators serotonin, and γ-aminobutyric acid. We identified neurons, connecting the dorsal rim area of the medulla to the anterior optic tubercle, as a possible site of neuromodulation and interaction with the circadian system. These neurons have conspicuous spines in close proximity to pigment dispersing factor-, serotonin-, and GABA-immunoreactive neurons. Our data therefore show for the first time a potential interaction site between the sky compass pathway and the circadian clock.     

The neural adhesion molecule L1CAM confers chemoresistance in human glioblastomas

Glioblastoma multiforme (GBM) represents the most common and malignant brain tumor. GBM tissues exhibit elevated expression of the transforming growth factor-beta1 (TGF-β1) and the adhesion molecule L1CAM. This study investigated the mechanism of L1CAM regulation in GBM cells and its role in the mediation of chemoresistance. L1CAM expression levels varied in GBM cells being highest in A172 cells and low in T98G cells. Inhibition of TGF-β1 signaling in A172 cells reduced L1CAM expression and vice versa stimulation with exogenous TGF-β1 led to upregulation of L1CAM in T98G cells. Additionally, TGF-β1 and L1CAM expression increased during differentiation of glioma stem-like cells. L1CAM expressing GBM cells and differentiated glioma stem-like cells showed a reduced apoptotic response after treatment with the chemotherapeutic drug temozolomide. Accordingly, siRNA-mediated knock-down of L1CAM in A172 cells and differentiated glioma stem-like cells increased chemosensitivity, whereas overexpression of L1CAM in T98G cells and glioma spheroids diminished the apoptotic response. Elevated L1CAM expression caused a diminished expression of caspase-8 in GBM and differentiated glioma stem-like cells. These data show that TGF-β1 dependent upregulation of L1CAM expression in GBM cells leads to the downregulation of caspase-8 and apoptosis resistance pointing to L1CAM as potential target for improved therapy of GBM patients.     

Activation of Olfactory Receptors on Mouse Pulmonary Macrophages Promotes Monocyte Chemotactic Protein-1 Production

Background

Emerging evidence suggests that non-olfactory tissues and cells can express olfactory receptors (ORs), however, the exact function of ectopic OR expression remains unknown. We have previously shown in mouse models that a unique cooperation between interferon-γ (IFN-γ) and lipopolysaccharide (LPS) drives the activation of pulmonary macrophages and leads to the induction of pathogenic responses in the respiratory tract. Further, through gene array studies, we have shown that activation of macrophages by these molecules results in the selective expression of a number of ORs. In this study, we validated the expression of these ORs in mouse airway and pulmonary macrophages in response to IFN-γ and LPS (γ/LPS) stimulation, and further explored the effect of odorant stimulation on macrophage function.

Methodology/Principal Findings

OR expression in airway and pulmonary macrophages in response to IFN-γ, LPS or γ/LPS treatments was assessed by microarray and validated by q-PCR. OR expression (e.g. OR622) on macrophages was confirmed by visualization in immunofluoresence assays. Functional responses to odorants were assessed by quantifying inflammatory cytokine and chemokine expression using q-PCR and cell migration was assessed by a modified Boyden chamber migration assay. Our results demonstrate that eight ORs are expressed at basal levels in both airway and pulmonary macrophages, and that γ/LPS stimulation cooperatively increased this expression. Pulmonary macrophages exposed to the combined treatment of γ/LPS+octanal (an odorant) exhibited a 3-fold increase in MCP-1 protein production, compared to cells treated with γ/LPS alone. Supernatants from γ/LPS+octanal exposed macrophages also increased macrophage migration in vitro.

Conclusions/Significance

Eight different ORs are expressed at basal levels in pulmonary macrophages and expression is upregulated by the synergistic action of γ/LPS. Octanal stimulation further increased MCP-1 production and the motility of macrophages. Our results suggest that ORs may mediate macrophage function by regulating MCP-1 production and cell migration.     

Comparison of NOGA Endocardial Mapping and Cardiac Magnetic Resonance Imaging for Determining Infarct Size and Infarct Transmurality for Intramyocardial Injection Therapy Using Experimental Data

Objectives

We compared the accuracy of NOGA endocardial mapping for delineating transmural and non-transmural infarction to the results of cardiac magnetic resonance imaging (cMRI) with late gadolinium enhancement (LE) for guiding intramyocardial reparative substance delivery using data from experimental myocardial infarction studies.

Methods

Sixty domestic pigs underwent diagnostic NOGA endocardial mapping and cMRI-LE 60 days after induction of closed-chest reperfused myocardial infarction. The infarct size was determined by LE of cMRI and by delineation of the infarct core on the unipolar voltage polar map. The sizes of the transmural and non-transmural infarctions were calculated from the cMRI transmurality map using signal intensity (SI) cut-offs of>75% and>25% and from NOGA bipolar maps using bipolar voltage cut-off values of <0.8 mV and <1.9 mV. Linear regression analysis and Bland-Altman plots were used to determine correlations and systematic differences between the two images. The overlapping ratios of the transmural and non-transmural infarcted areas were calculated.

Results

Infarct size as determined by 2D NOGA unipolar voltage polar mapping correlated with the 3D cMRI-LE findings (r = 0.504, p<0.001) with a mean difference of 2.82% in the left ventricular (LV) surface between the two images. Polar maps of transmural cMRI and bipolar maps of NOGA showed significant association for determining of the extent of transmural infarction (r = 0.727, p<0.001, overlap ratio of 81.6±11.1%) and non-transmural infarction (r = 0.555, p<0.001, overlap ratio of 70.6±18.5%). NOGA overestimated the transmural scar size (6.81% of the LV surface) but slightly underestimated the size of the non-transmural infarction (−3.04% of the LV surface).

Conclusions

By combining unipolar and bipolar voltage maps, NOGA endocardial mapping is useful for accurate delineation of the targeted zone for intramyocardial therapy and is comparable to cMRI-LE. This may be useful in patients with contraindications for cMRI who require targeted intramyocardial regenerative therapy.     

Nonneutral admixture of immigrant genotypes in African Drosophila melanogaster populations from Zimbabwe

Drosophila melanogaster originated in Africa and colonized the rest of the world only recently (approximately 10,000 to 15,000 years ago). Using 151 microsatellite loci, we investigated patterns of gene flow between African D. melanogaster populations representing presumptive ancestral variation and recently colonized European populations. Although we detected almost no evidence for alleles of non-African ancestry in a rural D. melanogaster population from Zimbabwe, an urban population from Zimbabwe showed evidence for admixture. Interestingly, the degree of admixture differed among chromosomes. X chromosomes of both rural and urban populations showed almost no non-African ancestry, but the third chromosome in the urban population showed up to 70% of non-African alleles. When chromosomes were broken into contingent microsatellite blocks, even higher estimates of admixture and significant heterogeneity in admixture was observed among these blocks. The discrepancy between the X chromosome and the third chromosome is not consistent with a neutral admixture hypothesis. The higher number of European alleles on the third chromosome could be due to stronger selection against foreign alleles on the X chromosome or to more introgression of (beneficial) alleles on the third chromosome.     

Effects of Short Term Bioturbation by Common Voles on Biogeochemical Soil Variables

Bioturbation contributes to soil formation and ecosystem functioning. With respect to the active transport of matter by voles, bioturbation may be considered as a very dynamic process among those shaping soil formation and biogeochemistry. The present study aimed at characterizing and quantifying the effects of bioturbation by voles on soil water relations and carbon and nitrogen stocks. Bioturbation effects were examined based on a field set up in a luvic arenosol comprising of eight 50 × 50 m enclosures with greatly different numbers of common vole (Microtus arvalis L., ca. 35–150 individuals ha^–1 mth^–1). Eleven key soil variables were analyzed: bulk density, infiltration rate, saturated hydraulic conductivity, water holding capacity, contents of soil organic carbon (SOC) and total nitrogen (N), CO2 emission potential, C/N ratio, the stable isotopic signatures of ¹³C and ¹⁵N, and pH. The highest vole densities were hypothesized to cause significant changes in some variables within 21 months. Results showed that land history had still a major influence, as eight key variables displayed an additional or sole influence of topography. However, the δ¹⁵N at depths of 10–20 and 20–30 cm decreased and increased with increasing vole numbers, respectively. Also the CO2 emission potential from soil collected at a depth of 15–30 cm decreased and the C/N ratio at 5–10 cm depth narrowed with increasing vole numbers. These variables indicated the first influence of voles on the respective mineralization processes in some soil layers. Tendencies of vole activity homogenizing SOC and N contents across layers were not significant. The results of the other seven key variables did not confirm significant effects of voles. Thus overall, we found mainly a first response of variables that are indicative for changes in biogeochemical dynamics but not yet of those representing changes in pools.     

Long-term efficacy of antipsychotic drugs in initially acutely ill adults with schizophrenia: systematic review and network meta-analysis

Most acute phase antipsychotic drug trials in schizophrenia last only a few weeks, but patients must usually take these drugs much longer. We examined the long-term efficacy of antipsychotic drugs in acutely ill patients using network meta-analysis. We searched the Cochrane Schizophrenia Group register up to March 6, 2022 for randomized, blinded trials of at least 6-month duration on all second-generation and 18 first-generation antipsychotics. The primary outcome was change in overall symptoms of schizophrenia; secondary outcomes were all-cause discontinuation; change in positive, negative and depressive symptoms; quality of life, social functioning, weight gain, antiparkinson medication use, akathisia, serum prolactin level, QTc prolongation, and sedation. Confidence in the results was assessed by the CINeMA (Confidence in Network Meta-Analysis) framework. We included 45 studies with 11,238 participants. In terms of overall symptoms, olanzapine was on average more efficacious than ziprasidone (standardized mean difference, SMD=0.37, 95% CI: 0.26-0.49), asenapine (SMD=0.33, 95% CI: 0.21-0.45), iloperidone (SMD=0.32, 95% CI: 0.15-0.49), paliperidone (SMD=0.28, 95% CI: 0.11-0.44), haloperidol (SMD=0.27, 95% CI: 0.14-0.39), quetiapine (SMD=0.25, 95% CI: 0.12-0.38), aripiprazole (SMD=0.16, 95% CI: 0.04-0.28) and risperidone (SMD=0.12, 95% CI: 0.03-0.21). The 95% CIs for olanzapine versus aripiprazole and risperidone included the possibility of trivial effects. The differences between olanzapine and lurasidone, amisulpride, perphenazine, clozapine and zotepine were either small or uncertain. These results were robust in sensitivity analyses and in line with other efficacy outcomes and all-cause discontinuation. Concerning weight gain, the impact of olanzapine was higher than all other antipsychotics, with a mean difference ranging from –4.58 kg (95% CI: –5.33 to –3.83) compared to ziprasidone to –2.30 kg (95% CI: –3.35 to –1.25) compared to amisulpride. Our data suggest that olanzapine is more efficacious than a number of other antipsychotic drugs in the longer term, but its efficacy must be weighed against its side effect profile.     

Human presence drives bobcat interactions among the U.S. carnivore guild

Biodiversity and Conservation - Mammalian carnivores are elusive and enigmatic species that often play keystone roles in ecosystems through direct and indirect effects. Growing evidence shows that...     

AI for in-line vehicle sequence controlling: development and evaluation of an adaptive machine learning artifact to predict sequence deviations in a mixed-model production line

Flexible Services and Manufacturing Journal - Customers in the manufacturing sector, especially in the automotive industry, have a high demand for individualized products at price levels comparable...