Mutation of the ALS-/FTD-Associated RNA-Binding Protein FUS Affects Axonal Development.

Aberrant condensation and localization of the RNA-binding protein (RBP) fused in sarcoma (FUS) occur in variants of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Changes in RBP function are commonly associated with changes in axonal cytoskeletal organization and branching in neurodevelopmental disorders. Here, we asked whether branching defects also occur in vivo in a model of FUS-associated disease. We use two reported Xenopus models of ALS/FTD (of either sex), the ALS-associated mutant FUS(P525L) and a mimic of hypomethylated FUS, FUS(16R). Both mutants strongly reduced axonal complexity in vivo. We also observed an axon looping defect for FUS(P525L) in the target area, which presumably arises due to errors in stop cue signaling. To assess whether the loss of axon complexity also had a cue-independent component, we assessed axonal cytoskeletal integrity in vitro. Using a novel combination of fluorescence and atomic force microscopy, we found that mutant FUS reduced actin density in the growth cone, altering its mechanical properties. Therefore, FUS mutants may induce defects during early axonal development.

Expert opinion on the use of contraception in people with multiple sclerosis.

BACKGROUND: Current guidance on the selection of appropriate contraception for people with multiple sclerosis (PwMS) is lacking. OBJECTIVE: To address this gap, an expert-led consensus program developed recommendations to support clinicians in discussing family planning and contraception with women and men with multiple sclerosis (MS). METHODS: A multidisciplinary steering committee (SC) of 13 international clinical experts led the program, supported by an extended faculty of 32 experts representing 18 countries. A modified Delphi methodology was used for decision-making and consensus-building. The SC drafted 15 clinical questions focused on patient-centered care, selection of contraception, and timing of stopping/starting contraception and disease-modifying therapies (DMTs). Statements addressing each question were drafted based on evaluation of published evidence and the experts' clinical experience. Consensus was reached if ⩾75% of respondents agreed (scoring 7-9 on a 9-point scale) with each recommendation. RESULTS: Consensus was reached on 24 of 25 proposed recommendations, including how and when to discuss contraception, types and safety of contraceptives, and how to evaluate the most appropriate contraceptive options for specific patient groups, including those with significant disability or being treated with DMTs. CONCLUSION: These expert recommendations provide the first practical, relevant, and comprehensive guidance for clinicians on the selection of contraception in PwMS.

Fast, multicolour optical sectioning over extended fields of view with patterned illumination and machine learning.

Structured illumination can reject out-of-focus signal from a sample, enabling high-speed and high-contrast imaging over large areas with widefield detection optics. However, this optical sectioning technique is currently limited by image reconstruction artefacts and poor performance at low signal-to-noise ratios. We combine multicolour interferometric pattern generation with machine learning to achieve high-contrast, real-time reconstruction of image data that is robust to background noise and sample motion. We validate the method in silico and demonstrate imaging of diverse specimens, from fixed and live biological samples to synthetic biosystems, reconstructing data live at 11 Hz across a 44 × 44µm2 field of view, and demonstrate image acquisition speeds exceeding 154 Hz.

Molecular Mechanisms of Cationic Fusogenic Liposome Interactions with Bacterial Envelopes.

Although fusogenic liposomes offer a promising approach for the delivery of antibiotic payloads across the cell envelope of Gram-negative bacteria, there is still a limited understanding of the individual nanocarrier interactions with the bacterial target. Using super-resolution microscopy, we characterize the interaction dynamics of positively charged fusogenic liposomes with Gram-negative (Escherichia coli) and Gram-positive (Bacillus subtilis) bacteria. The liposomes merge with the outer membrane (OM) of Gram-negative bacteria, while attachment or lipid internalization is observed in Gram-positive cells. Employing total internal reflection fluorescence microscopy, we demonstrated liposome fusion with model supported lipid bilayers. For whole E. coli cells, however, we observed heterogeneous membrane integrations, primarily involving liposome attachment and hemifusion events. With increasing lipopolysaccharide length, the likelihood of full-fusion events was reduced. The integration of artificial lipids into the OM of Gram-negative cells led to membrane destabilization, resulting in decreased bacterial vitality, membrane detachment, and improved codelivery of vancomycin─an effective antibiotic against Gram-positive cells. These findings provide significant insights into the interactions of individual nanocarriers with bacterial envelopes at the single-cell level, uncovering effects that would be missed in bulk measurements. This highlights the importance of conducting single-particle and single-cell investigations to assess the performance of next-generation drug delivery platforms.

Microelectrode Arrays Measure Blocking of Voltage-Gated Calcium Ion Channels on Supported Lipid Bilayers Derived from Primary Neurons.

Drug studies targeting neuronal ion channels are crucial to understand neuronal function and develop therapies for neurological diseases. The traditional method to study neuronal ion-channel activities heavily relies on the whole-cell patch clamp as the industry standard. However, this technique is both technically challenging and labour-intensive, while involving the complexity of keeping cells alive with low throughput. Therefore, the shortcomings are limiting the efficiency of ion-channel-related neuroscience research and drug testing. Here, this work reports a new system of integrating neuron membranes with organic microelectrode arrays (OMEAs) for ion-channel-related drug studies. This work demonstrates that the supported lipid bilayers (SLBs) derived from both neuron-like (neuroblastoma) cells and primary neurons are integrated with OMEAs for the first time. The increased expression of voltage-gated calcium (CaV) ion channels on differentiated SH-SY5Y SLBs  compared to non-differentiated ones is sensed electrically. Also, dose-response of the CaV ion-channel blocking effect on primary cortical neuronal SLBs from rats is monitored. The dose range causing ion channel blocking is comparable to literature. This system overcomes the major challenges from traditional methods (e.g., patch clamp) and showcases an easy-to-test, rapid, ultra-sensitive, cell-free, and high-throughput platform to monitor dose-dependent ion-channel blocking effects on native neuronal membranes.

The carbonyl-lock mechanism underlying non-aromatic fluorescence in biological matter.

Challenging the basis of our chemical intuition, recent experimental evidence reveals the presence of a new type of intrinsic fluorescence in biomolecules that exists even in the absence of aromatic or electronically conjugated chemical compounds. The origin of this phenomenon has remained elusive so far. In the present study, we identify a mechanism underlying this new type of fluorescence in different biological aggregates. By employing non-adiabatic ab initio molecular dynamics simulations combined with a data-driven approach, we characterize the typical ultrafast non-radiative relaxation pathways active in non-fluorescent peptides. We show that the key vibrational mode for the non-radiative decay towards the ground state is the carbonyl elongation. Non-aromatic fluorescence appears to emerge from blocking this mode with strong local interactions such as hydrogen bonds. While we cannot rule out the existence of alternative non-aromatic fluorescence mechanisms in other systems, we demonstrate that this carbonyl-lock mechanism for trapping the excited state leads to the fluorescence yield increase observed experimentally, and set the stage for design principles to realize novel non-invasive biocompatible probes with applications in bioimaging, sensing, and biophotonics.

High Aspect Ratio Nanoscale Pores through BCP-Based Metal Oxide Masks and Advanced Dry Etching.

The reliable and regular modification of the surface properties of substrates plays a crucial role in material research and the development of functional surfaces. A key aspect of this is the development of the surface pores and topographies. These can confer specific advantages such as high surface area as well as specific functions such as hydrophobic properties. Here, we introduce a combination of nanoscale self-assembled block-copolymer-based metal oxide masks with optimized deep reactive ion etching (DRIE) of silicon to permit the fabrication of porous topographies with aspect ratios of up to 50. Following the evaluation of our procedure and involved parameters using various techniques, such as AFM or SEM, the suitability of our features for applications relying on high light absorption as well as efficient thermal management is explored and discussed in further detail.

A safety mechanism enables tissue-specific resistance to protein aggregation during aging in C. elegans.

During aging, proteostasis capacity declines and distinct proteins become unstable and can accumulate as protein aggregates inside and outside of cells. Both in disease and during aging, proteins selectively aggregate in certain tissues and not others. Yet, tissue-specific regulation of cytoplasmic protein aggregation remains poorly understood. Surprisingly, we found that the inhibition of 3 core protein quality control systems, namely chaperones, the proteasome, and macroautophagy, leads to lower levels of age-dependent protein aggregation in Caenorhabditis elegans pharyngeal muscles, but higher levels in body-wall muscles. We describe a novel safety mechanism that selectively targets newly synthesized proteins to suppress their aggregation and associated proteotoxicity. The safety mechanism relies on macroautophagy-independent lysosomal degradation and involves several previously uncharacterized components of the intracellular pathogen response (IPR). We propose that this protective mechanism engages an anti-aggregation machinery targeting aggregating proteins for lysosomal degradation.

Cardioprotective effect of the quercetin on cardiovascular remodeling and atherosclerosis in rodents fed a high-fat diet: A systematic review.

Cardiovascular diseases (CVD) are the leading cause of death globally, estimated at 17.9 million premature deaths. Several risk factors contribute to the development of CVD, including unhealthy diet rich in saturated fat. Quercetin (Q) is a important natural flavonoid with cardioprotective effect. However, it is crucial to understand and clarify which dosages and intervention times quercetin promotes better cardioprotective effects when exposed to a High-Fat Diet (HFD). We aim was to carry out a review to identify and compare experimental studies that investigated the quercetin effect on cardiac parameters in rodents fed a HFD. This literature search was performed through the specialized databases PubMed, Embase, Web of Science and Lilacs in May 2022. The following information was collected and assessed: Species of animals, dietary fat content, intervention protocol (quercetin), and main results of alterations associated with cardiac change. A total of 116 articles were selected from the database and 30 articles were included in this study. The administration form of quercetin was used in the diet supplemented in 73.4% (n = 22) of the studies. The dosage ranged between 10 and 100 mg/kg, 0.01%-0.36%, and 4-8 g/kg diet. The treatment time ranged between 14 and 63 days in 48.4% studies and most of the selected studies observed changes in the: Serum concentrations of lipids (60%, n = 18) mainly decrease in TC and TG, left ventricle (LV) (16.13%, n = 5) includes attenuation of the cardiac hypertrophy; inhibition of atherosclerotic progression (32%, n = 10) with decrease in lesions and plaque formation; improvement in the expression of gene and protein associated with cardiac functionality and oxidative stress (51.6%; n = 16). Quercetin supplementation at different concentrations/doses promotes important cardioprotective effects in experimental models exposed to a HFD. The supplemented diet was shown to be the better administration option. The methodological variation presented in the articles selected in this review proves that the most appropriate intervention protocol, as well as the most effective route of administration, promotes these effects.

Diversity of dynamic voltage patterns in neuronal dendrites revealed by nanopipette electrophysiology.

Dendrites and dendritic spines are the essential cellular compartments in neuronal communication, conveying information through transient voltage signals. Our understanding of these compartmentalized voltage dynamics in fine, distal neuronal dendrites remains poor due to the difficulties inherent to accessing and stably recording from such small, nanoscale cellular compartments for a sustained time. To overcome these challenges, we use nanopipettes that permit long and stable recordings directly from fine neuronal dendrites. We reveal a diversity of voltage dynamics present locally in dendrites, such as spontaneous voltage transients, bursting events and oscillating periods of silence and firing activity, all of which we characterized using segmentation analysis. Remarkably, we find that neuronal dendrites can display spontaneous hyperpolarisation events, and sustain transient hyperpolarised states. The voltage patterns were activity-dependent, with a stronger dependency on synaptic activity than on action potentials. Long-time recordings of fine dendritic protrusions show complex voltage dynamics that may represent a previously unexplored contribution to dendritic computations.

Endometriosis features and dienogest tolerability in women with depression: a case-control study.

OBJECTIVE: Primary aim of this study was to investigate endometriosis characteristics of patients with psychiatric conditions or depression. The secondary aim was to study tolerability of dienogest in this context. METHODS: This observational case-control study included endometriosis data from patients visiting our clinic from 2015-2021. We collected information from patient charts and in phone interviews based on a structured survey. Patients with surgical confirmed endometriosis were included. RESULTS: 344 patients fulfilled the inclusion criteria: n = 255 no psychiatric disorder, n = 119 any psychiatric disorder and n = 70 depression. Patients with depression (EM-D, p=.018; p=.035) or psychiatric condition (EM-P, p=.020; p=.048) suffered more often from dyspareunia and dyschezia. EM-P patients had more often primary dysmenorrhoea with higher pain scores (p=.045). rASRM stage or localisation of lesions did not differ. EM-D and EM-P patients discontinued dienogest treatment more often related to worsening of mood (p= .001, p=.002). CONCLUSION: EM-D or EM-P had a higher prevalence of pain symptoms. This could not be attributed to differences in rASRM stage or location of endometriosis lesions. Strong primary dysmenorrhoea might predispose to develop chronic pain-based psychological symptoms. Therefore, early diagnosis and treatment are relevant. Gynaecologist should be aware of the potential impact of dienogest on mood.

Women with endometriosis and psychiatric disorders especially have more dyschezia and dyspareunia, independent from rASRM stage, depth of infiltration and localisation of endometriosis lesions. Dienogest has an impact on mood especially in already prone patients.Trial registration: trial registration number: NCT04816357. https://clinicaltrials.gov/ct2/show/NCT04816357Date of registration: 22.03.2021, date of enrolment of the first subject: 25.03.2021.

The Flavonoid Agathisflavone Directs Brain Microglia/Macrophages to a Neuroprotective Anti-Inflammatory and Antioxidant State via Regulation of NLRP3 Inflammasome.

Agathisflavone, purified from Cenostigma pyramidale (Tul.) has been shown to be neuroprotective in in vitro models of glutamate-induced excitotoxicity and inflammatory damage. However, the potential role of microglial regulation by agathisflavone in these neuroprotective effects is unclear. Here we investigated the effects of agathisflavone in microglia submitted to inflammatory stimulus in view of elucidating mechanisms of neuroprotection. Microglia isolated from cortices of newborn Wistar rats were exposed to Escherichia coli lipopolysaccharide (LPS, 1 µg/mL) and treated or not with agathisflavone (1 µM). Neuronal PC12 cells were exposed to a conditioned medium from microglia (MCM) treated or not with agathisflavone. We observed that LPS induced microglia to assume an activated inflammatory state (increased CD68, more rounded/amoeboid phenotype). However, most microglia exposed to LPS and agathisflavone, presented an anti-inflammatory profile (increased CD206 and branched-phenotype), associated with the reduction in NO, GSH mRNA for NRLP3 inflammasome, IL1-ß, IL-6, IL-18, TNF, CCL5, and CCL2. Molecular docking also showed that agathisflavone bound at the NLRP3 NACTH inhibitory domain. Moreover, in PC12 cell cultures exposed to the MCM previously treated with the flavonoid most cells preserved neurites and increased expression of ß-tubulin III. Thus, these data reinforce the anti-inflammatory activity and the neuroprotective effect of agathisflavone, effects associated with the control of NLRP3 inflammasome, standing out it as a promising molecule for the treatment or prevention of neurodegenerative diseases.

α-Synuclein fibril and synaptic vesicle interactions lead to vesicle destruction and increased lipid-associated fibril uptake into iPSC-derived neurons.

Monomeric alpha-synuclein (aSyn) is a well characterised protein that importantly binds to lipids. aSyn monomers assemble into amyloid fibrils which are localised to lipids and organelles in insoluble structures found in Parkinson's disease patient's brains. Previous work to address pathological aSyn-lipid interactions has focused on using synthetic lipid membranes, which lack the complexity of physiological lipid membranes. Here, we use physiological membranes in the form of synaptic vesicles (SV) isolated from rodent brain to demonstrate that lipid-associated aSyn fibrils are more easily taken up into iPSC-derived cortical i3Neurons. Lipid-associated aSyn fibril characterisation reveals that SV lipids are an integrated part of the fibrils and while their fibril morphology differs from aSyn fibrils alone, the core fibril structure remains the same, suggesting the lipids lead to the increase in fibril uptake. Furthermore, SV enhance the aggregation rate of aSyn, yet increasing the SV:aSyn ratio causes a reduction in aggregation propensity. We finally show that aSyn fibrils disintegrate SV, whereas aSyn monomers cause clustering of SV using small angle neutron scattering and high-resolution imaging. Disease burden on neurons may be impacted by an increased uptake of lipid-associated aSyn which could enhance stress and pathology, which in turn may have fatal consequences for neurons.

Effects of a high-fat diet on the bone structure of Wistar rats: a systematic review.

CONTEXT: Saturated fats found in diets known as high-fat, cafeteria, or Western diets appear to have a negative effect on bone structure; however, few studies have focused on investigating this association, and the data available in the literature remain controversial. OBJECTIVE: The aim of the current review was to investigate the effects of a high-fat dietary intake on the bone structure of Wistar rats. DATA SOURCES: A search for articles was carried out in the Pubmed/MEDLINE, Web of Science, Embase, and Scopus databases. DATA EXTRACTION: In total, 447 articles were found in the initial search; 5 articles were included in the systematic review, after application of the exclusion criteria. DATA ANALYSIS: The review was guided by the PICOS strategy and based on the PRISMA protocol for animal reviews. CONCLUSION: High-fat diets appear to affect bone structure of Wistar rats. Diet composition and exposure time are the factors determining the strength of the effect.

ERnet: a tool for the semantic segmentation and quantitative analysis of endoplasmic reticulum topology.

The ability to quantify structural changes of the endoplasmic reticulum (ER) is crucial for understanding the structure and function of this organelle. However, the rapid movement and complex topology of ER networks make this challenging. Here, we construct a state-of-the-art semantic segmentation method that we call ERnet for the automatic classification of sheet and tubular ER domains inside individual cells. Data are skeletonized and represented by connectivity graphs, enabling precise and efficient quantification of network connectivity. ERnet generates metrics on topology and integrity of ER structures and quantifies structural change in response to genetic or metabolic manipulation. We validate ERnet using data obtained by various ER-imaging methods from different cell types as well as ground truth images of synthetic ER structures. ERnet can be deployed in an automatic high-throughput and unbiased fashion and identifies subtle changes in ER phenotypes that may inform on disease progression and response to therapy.

Feasibility of Coacervate-Like Nanostructure for Instant Drug Nanoformulation.

Despite the enormous advancements in nanomedicine research, a limited number of nanoformulations are available on the market, and few have been translated to clinics. An easily scalable, sustainable, and cost-effective manufacturing strategy and long-term stability for storage are crucial for successful translation. Here, we report a system and method to instantly formulate NF achieved with a nanoscale polyelectrolyte coacervate-like system, consisting of anionic pseudopeptide poly(l-lysine isophthalamide) derivatives, polyethylenimine, and doxorubicin (Dox) via simple "mix-and-go" addition of precursor solutions in seconds. The coacervate-like nanosystem shows enhanced intracellular delivery of Dox to patient-derived multidrug-resistant (MDR) cells in 3D tumor spheroids. The results demonstrate the feasibility of an instant drug formulation using a coacervate-like nanosystem. We envisage that this technique can be widely utilized in the nanomedicine field to bypass the special requirement of large-scale production and elongated shelf life of nanomaterials.

Endometriosis Features in Women With and Without Migraine.

Background: This study examines endometriosis (EM) features in women with EM and migraines (MG) (EM-MG) and women with EM alone (EM-O). The comorbidity of MG and EM is well known. However, knowledge about differences in symptoms, clinical manifestations, and severity of EM between EM-MG and EM-O is scarce. Materials and Methods: We conducted a cross-sectional observational study of premenopausal patients with biopsy-confirmed EM treated in our department from 2015 to 2021. All patients underwent surgical treatment for EM. Information about infiltration depth and localization of EM was available. We interviewed patients using a structured questionnaire that includes questions about clinical characteristics, symptoms, and treatment history. We reported categorical variables as frequencies and continuous variables as means with standard deviations. We compared subgroups (EM-MG vs. EM-O) using an independent sample t-test, the Wilcoxon-Mann-Whitney test, chi-square test, and Fisher's exact test. The significance level was 0.05. Results: We included 344 participants: 250 with EM-O and 94 with EM-MG. EM-MG had less severe revised American Society of Reproductive Medicine scores (p = 0.023), more deliveries (p = 0.009), more and higher scores of dysmenorrhea at menarche (p = 0.044; p = 0.036), prolonged heavy menstrual bleeding (p = 0.009), more and prolonged pain during menstrual bleeding (p = 0.011, p = 0.039), and more dyschezia (p < 0.001) compared with EM-O. Conclusion: Migraineurs experienced more intense EM symptoms at lower EM stages. This discrepancy strongly indicates pain sensitizations and a lower pain threshold in patients with EM-MG. Knowledge about EM features allows early diagnosis and treatment of women with potential EM-MG, both highly disabling conditions. Clinical Trials.gov (NCT04816357).

Perinatal and post-weaning exposure to a high-fat diet causes histomorphometric, neuroplastic, and histopathological changes in the rat ileum.

Exposure to a diet with a high saturated fat content can influence the characteristics of the gastrointestinal tract, causing losses in the absorption of nutrients and favoring the appearance of diseases. The objective was to assess the effects of a high-fat diet (HFD) in the perinatal (pregnancy and lactation) and post-weaning period on the histomorphometry, neuroplasticity, and histopathology of the ileum. Wistar rats were divided into four subgroups: Control/Control (CC, n = 10) rats fed a control diet (C) throughout the trial period; Control/HFD (CH, n = 9) rats fed diet C (perinatal) and HFD after weaning; HFD/Control (HC, n = 10) rats fed HFD (perinatal) and diet C (post-weaning); HFD/HFD (HH, n = 9) rats fed HFD throughout the experimental period. There was atrophy of the Ileum wall with a reduction in the muscular tunic, submucosa, and mucosa thickness in the HH group of 37%, 28%, and 46%, respectively (p < 0.0001). The depth of the crypts decreased by 29% (p < 0.0001) and height increased by 5% (p < 0.0013). Villus height decreased by 41% and 18% in HH and HC groups (p < 0.0001) and width decreased by 11% in the HH (p < 0.0001). The height of the enterocytes decreased by 18% in the HH (p < 0.0001). There was a decrease in the area of the myenteric and submucosal plexus ganglia in the HH and HC groups (p < 0.0001). The number, occupation, and granules of Paneth cells increased in the HH and HC groups (p < 0.0001). Intraepithelial lymphocytes (IELs) increased in all groups exposed to the HFD. Goblet cells decreased in groups CH and HH (p < 0.0001). The evidence from this study suggests that the HFD had altered the histomorphometry, neuroplasticity, and histopathology of the ileum of the rats.

Decreased Water Mobility Contributes To Increased α-Synuclein Aggregation.

The solvation shell is essential for the folding and function of proteins, but how it contributes to protein misfolding and aggregation has still to be elucidated. We show that the mobility of solvation shell H2 O molecules influences the aggregation rate of the amyloid protein α-synuclein (αSyn), a protein associated with Parkinson's disease. When the mobility of H2 O within the solvation shell is reduced by the presence of NaCl, αSyn aggregation rate increases. Conversely, in the presence CsI the mobility of the solvation shell is increased and αSyn aggregation is reduced. Changing the solvent from H2 O to D2 O leads to increased aggregation rates, indicating a solvent driven effect. We show the increased aggregation rate is not directly due to a change in the structural conformations of αSyn, it is also influenced by a reduction in both the H2 O mobility and αSyn mobility. We propose that reduced mobility of αSyn contributes to increased aggregation by promoting intermolecular interactions.

Decreased Water Mobility Contributes To Increased α-Synuclein Aggregation.

The solvation shell is essential for the folding and function of proteins, but how it contributes to protein misfolding and aggregation has still to be elucidated. We show that the mobility of solvation shell H2O molecules influences the aggregation rate of the amyloid protein α-synuclein (αSyn), a protein associated with Parkinson's disease. When the mobility of H2O within the solvation shell is reduced by the presence of NaCl, αSyn aggregation rate increases. Conversely, in the presence CsI the mobility of the solvation shell is increased and αSyn aggregation is reduced. Changing the solvent from H2O to D2O leads to increased aggregation rates, indicating a solvent driven effect. We show the increased aggregation rate is not directly due to a change in the structural conformations of αSyn, it is also influenced by a reduction in both the H2O mobility and αSyn mobility. We propose that reduced mobility of αSyn contributes to increased aggregation by promoting intermolecular interactions.