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Animals with small nervous systems have a limited number of sensory neurons that must encode information from a changing environment. This problem is particularly exacerbated in nematodes that populate a wide variety of distinct ecological niches but only have a few sensory neurons available to encode multiple modalities. How does sensory diversity prevail within this neuronal constraint? To identify the genetic basis for patterning different nervous systems, we demonstrate that sensory neurons in the Pristionchus pacificus respond to various salt sensory cues in a manner that is partially distinct from that of the distantly related nematode C. elegans . By visualizing neuronal activity patterns, we show that contrary to previous expectations based on its genome sequence, the salt responses of P. pacificus are encoded in a left/right asymmetric manner in the bilateral ASE neuron pair. Our study illustrates patterns of evolutionary stability and change in the gustatory system of nematodes.

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Volumetric muscle loss (VML) injuries are characterized by the traumatic loss of skeletal muscle resulting in permanent damage to both tissue architecture and electrical excitability. To address this challenge, we previously developed a 3D aligned collagen-glycosaminoglycan (CG) scaffold platform that supported in vitro myotube alignment and maturation. In this work, we assessed the ability of CG scaffolds to facilitate functional muscle recovery in a rat tibialis anterior (TA) model of VML. Functional muscle recovery was assessed following implantation of either non-conductive CG or electrically conductive CG-polypyrrole (PPy) scaffolds at 4, 8, and 12 weeks post-injury by in vivo electrical stimulation of the peroneal nerve. After 12 weeks, scaffold-treated muscles produced maximum isometric torque that was significantly greater than non-treated tissues. Histological analysis further supported these reparative outcomes with evidence of regenerating muscle fibers at the material-tissue interface in scaffold-treated tissues that was not observed in non-repaired muscles. Scaffold-treated muscles possessed higher numbers of M1 and M2 macrophages at the injury while conductive CG-PPy scaffold-treated muscles showed significantly higher levels of neovascularization as indicated by the presence of pericytes and endothelial cells, suggesting a persistent wound repair response not observed in non-treated tissues. Finally, only tissues treated with non-conductive CG scaffolds displayed neurofilament staining similar to native muscle, further corroborating isometric contraction data. Together, these findings show that CG scaffolds can facilitate improved skeletal muscle function and endogenous cellular repair, highlighting their potential use as therapeutics for VML injuries.

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OBJECTIVES: This study aimed to assess the validity of external quality assessment (EQA) laboratory results across various cultural and environmental contexts and to identify potential improvement areas. METHODS: The International Federation of Clinical Chemistry and Laboratory Medicine (IFCC) Task Force on Global Laboratory Quality (TF-GLQ) conducted a 2-year study (2022 and 2023) in which EQA materials, related software and online training was provided by a commercial vendor to 100 laboratories in ten IFCC member society countries. The results were analysed on a monthly basis by the TF-GLQ, to show the number of submissions per country, tests per lab, acceptability rates, random failures and to get a measure of which analytes performed poorly. RESULTS: The EQA material was dispatched on a quarterly basis. Some countries had problems with customs releasing the material in a timely manner, resulting in laboratories not receiving them on time leading to no submission. We report here the results for the second year of the survey. The number of examinations varied between laboratories, ranging from seven to 84 analytes. Of the ten countries surveyed, six averaged greater than 90 % acceptable results over the whole 12-months cycle, one had unacceptable results for two of the nine months they returned results and the other four were considered to not perform to an acceptable standard. CONCLUSIONS: All 100 participating laboratories indicated satisfaction with the EQA survey and related services, including on-site training, and report handling. However, specimen receiving issues, suggest benefits in dispatching materials for a full 12-month cycle. Significant discrepancies in EQA performance indicate that four countries require long-term assistance, training and guidance. To ensure reliable patient results, promoting EQA in certain countries is essential to achieve the required level of quality.

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Several clades of luminescent bacteria are known currently. They all contain similar lux operons, which include the genes luxA and luxB encoding a heterodimeric luciferase. The aldehyde oxygenation reaction is presumed to be catalyzed primarily by the subunit LuxA, whereas LuxB is required for efficiency and stability of the complex. Recently, genomic analysis identified a subset of bacterial species with rearranged lux operons lacking luxB. Here, we show that the product of the luxA gene from the reduced luxACDE operon of Enhygromyxa salina is luminescent upon addition of aldehydes both in vivo in Escherichia coli and in vitro. Overall, EsLuxA is much less bright compared with luciferases from Aliivibrio fischeri (AfLuxAB) and Photorhabdus luminescens (PlLuxAB), and most active with medium-chain C4-C9 aldehydes. Crystal structure of EsLuxA determined at the resolution of 2.71 Å reveals a (ß/α)8 TIM-barrel fold, characteristic for other bacterial luciferases, and the protein preferentially forms a dimer in solution. The mobile loop residues 264-293, which form a ß-hairpin or a coil in Vibrio harveyi LuxA, form α-helices in EsLuxA. Phylogenetic analysis shows EsLuxA and related proteins may be bacterial protoluciferases that arose prior to duplication of the luxA gene and its speciation to luxA and luxB in the previously described luminescent bacteria. Our work paves the way for the development of new bacterial luciferases that have an advantage of being encoded by a single gene.

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Background: Tranexamic acid has been increasingly used in facial plastic surgery to improve perioperative hemostasis. While subcutaneous tranexamic acid has been found to not significantly decrease postoperative ecchymoses following upper blepharoplasty, systemic administration has not previously been studied. Methods: A total of 325 patients undergoing upper blepharoplasty were randomly assigned to either receive intravenous tranexamic acid or serve as a control. Patients in the experimental group were administered 1 g of tranexamic acid intravenously 10 minutes before surgical incision. A similar upper blepharoplasty technique was performed by two American Society of Ophthalmic Plastic and Reconstructive Surgery-trained surgeons. Follow-up was conducted at a median of 8 days postsurgery. Patient photographs were evaluated by two independent graders to rate ecchymoses on a scale of 0 (least) to 10 (most). Results: Of the 325 included patients, 138 patients received intravenous tranexamic acid and 187 patients did not. The average ecchymosis rating for the control group at day 8 was 5.8 ± 1.7, while the average rating for the tranexamic acid group at the same time point was 4.1 ± 1.6 (P < 0.0001). There was a trend toward decreased ecchymoses in the tranexamic acid group at earlier and later postoperative timepoints that did not reach statistical significance. No hemorrhagic or systemic embolic complications occurred. Conclusions: Systemic tranexamic acid may reduce postoperative ecchymoses after upper blepharoplasty surgery, reaching significance at the eighth postoperative day, which may lead to improved patient satisfaction and decreased occupational downtime.

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PURPOSE: The purpose of this study is to evaluate the usage of oxymetazoline hydrochloride ophthalmic solution, 0.1% in the treatment of Graves' disease to improve lid symmetry. METHODS: Fourteen patients were identified with unilateral upper eyelid retraction, secondary to Graves' disease. A phenylephrine test was performed to evaluate whether treating the unaffected (non-retracted) side would result in improved symmetry using Hering's law. Patients were prescribed oxymetazoline hydrochloride ophthalmic solution, 0.1% to instill into the unaffected eye. Marginal reflex distance 1 (MRD1) measurements were taken pre- and post-treatment and patient satisfaction surveys were completed. RESULTS: The average pre-treatment MRD1 difference between the right eye (OD) and the left eye (OS) was 2.46 [range 1.50-4.00]. The average post-treatment MRD1 difference between OD and OS was 0.39 [range 0.00-1.00]. One hundred percent of patients were satisfied or very satisfied with the improvement in symmetry. CONCLUSIONS: Oxymetazoline hydrochloride ophthalmic solution, 0.1% is a potential method to improve symmetry in patients with Graves' disease and unilateral upper eyelid retraction via Hering's law, especially for non-surgical candidates or those who prefer non-surgical treatment. Our results indicate improved MRD1 differences between fellow eyes and patient satisfaction regarding the symmetry of the upper eyelids.

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Tracking cell death in vivo can enable a better understanding of the biological mechanisms underlying tissue homeostasis and disease. Unfortunately, existing cell death labeling methods lack compatibility with in vivo applications or suffer from low sensitivity, poor tissue penetration, and limited temporal resolution. Here, we fluorescently labeled dead cells in vivo with Trypan Blue (TBlue) to detect single scattered dead cells or to generate whole-mount three-dimensional maps of large areas of necrotic tissue during organ regeneration. TBlue effectively marked different types of cell death, including necrosis induced by CCl4 intoxication in the liver, necrosis caused by ischemia-reperfusion in the skin, and apoptosis triggered by BAX overexpression in hepatocytes. Moreover, due to its short circulating lifespan in blood, TBlue labeling allowed in vivo "pulse and chase" tracking of two temporally spaced populations of dying hepatocytes in regenerating mouse livers. Additionally, upon treatment with cisplatin, TBlue labeled dead cancer cells in livers with cholangiocarcinoma and dead thymocytes due to chemotherapy-induced toxicity, showcasing its utility in assessing anticancer therapies in preclinical models. Thus, TBlue is a sensitive and selective cell death marker for in vivo applications, facilitating the understanding of the fundamental role of cell death in normal biological processes and its implications in disease.

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The isothiourea derivative NT-1505 is known as a neuroprotector and cognition enhancer in animal models of neurodegenerative diseases. Bearing in mind possible relation of the NT-1505-mediated neuroprotection to mitochondrial uncoupling activity, here, we examine NT-1505 effects on mitochondria functioning. At concentrations starting from 10 µM, NT-1505 prevented Ca2+-induced mitochondrial swelling, similar to common uncouplers. Alongside the inhibition of the mitochondrial permeability transition, NT-1505 caused a decrease in mitochondrial membrane potential and an increase in respiration rate in both isolated mammalian mitochondria and cell cultures, which resulted in the reduction of energy-dependent Ca2+ uptake by mitochondria. Based on the oppositely directed effects of bovine serum albumin and palmitate, we suggest the involvement of fatty acids in the NT-1505-mediated mitochondrial uncoupling. In addition, we measured the induction of electrical current across planar bilayer lipid membrane upon the addition of NT-1505 to the bathing solution. Importantly, introduction of the palmitic acid into the lipid bilayer composition led to weak proton selectivity of the NT-1505-mediated BLM current. Thus, the present study revealed an ability of NT-1505 to cause moderate protonophoric uncoupling of mitochondria, which could contribute to the neuroprotective effect of this compound.

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This study aimed to develop a fast procedure for caffeine extraction from roasted coffee beans. The microwave-assisted extraction was carried out in the microwave oven with an operating frequency of 2450 MHz. The response surface methodology based on a Box-Behnken design was used to model and optimize the extraction process. Among the analyzed extraction parameters (factors), the influence of extraction time (2-6 min), liquid-to-solid ratio (5-15 mL/g), and microwave power (336-595 W) were considered, while the yield of extracted caffeine was observed as the response of the system. Water was used as the solvent of choice for the extraction of caffeine. The optimum conditions were as follows: extraction time, 2 min; liquid-to-solid ratio, 15 mL/g; and microwave power, 500 W. In this optimized condition, the expected extraction yield of caffeine was 1.01 g/100 g dry weight (value confirmed by experimental assays). The total energy consumed of 1.7 kWh/100 g of purified caffeine indicated a more energy-efficient procedure by about 1200-15,000 times than the reported procedures. This study showed that caffeine can be quantitatively extracted from roasted coffee beans through a green approach and that the isolated caffeine has a high purity degree, which was confirmed by the UHPLC-ESI-MS/MS method. With this quality, isolated caffeine could be further used as an active ingredient in the food industry, while for pharmaceutical purposes, it must be further purified.

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Wastewater based epidemiology (WBE) has been used worldwide to estimate drug consumption routinely. Even though WBE provides valuable data to support legal and health interventions associated to drug use, monitoring studies in Portuguese wastewaters are scarce. Hence, this work aimed to estimate the consumption of some conventional abuse and illicit drugs such as amphetamine (AMP), methamphetamine (MAMP), 3,4-methylenedioxymethamphetamine (MDMA), and the synthetic cathinones buphedrone (BPD), butylone (BTL), 3,4-dimethylmethcathinone (3,4-DMMC) and 3-methylmethcathinone (3-MMC), considering not only the liquid phase, but also the suspended particulate matter (SPM). Moreover, the enantiomeric profiling of the samples was studied, exploring for the first time the possible enantioselective sorption of these drugs onto SPM. For that, 24 h composite raw wastewaters were collected from a conventional wastewater treatment plant (WWTP) in Portugal. After extraction, the liquid phase and SPM extracts were derivatized with an enantiomerically pure reagent and then, analysed using a gas chromatography-mass spectrometry (GC-MS) analytical method. The results showed a low and non-enantioselective adsorption to SPM at environmental relevant levels. Only (S)-AMP was detected in two SPM samples, whereas AMP, MAMP, MDMA, BPD, and 3,4-DMMC were detected in the liquid phase. AMP was the most frequently found drug with an estimated load up to 166.0 mg day-1 1000 people-1 and mostly found with enrichment of (S)-AMP. Nevertheless, (R)-AMP was also determined, which may be related to the consumption of either the illicit racemic AMP or the medicine (R)-deprenyl. The use of MDMA, MAMP and synthetic cathinones (BPD and 3,4-DMMC) was also suggested in Portugal. Nevertheless, the levels and the consumption estimate of the target chemicals were lower than in other European countries or worldwide. These findings provide the first step to the implementation of WBE monitoring campaigns to assess the status of drug consumption in Portuguese communities, contributing to the understanding of drug use patterns and trends worldwide and helping enforce preventive measures.

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PURPOSE: Based on considerable interest to enlarge the experimental database of radioresistant cells after their irradiation with helium ions, HTB140, MCF-7 and HTB177 human malignant cells are exposed to helium ion beams having different linear energy transfer (LET). MATERIALS AND METHODS: The cells are irradiated along the widened 62 MeV/u helium ion Bragg peak, providing LET of 4.9, 9.8, 23.4 and 36.8 keV/µm. Numerical simulations with the Geant4 toolkit are used for the experimental design. Cell survival is evaluated and compared with reference γ-rays. DNA double strand breaks are assessed via γ-H2AX foci. RESULTS: With the increase of LET, surviving fractions at 2 Gy decrease, while RBE (2 Gy, γ) gradually increase. For HTB140 cells, above the dose of 4 Gy, a slight saturation of survival is observed while the increase of RBE (2 Gy, γ) remains unaffected. With the increase of LET the increase of γ-H2AX foci is revealed at 0.5 h after irradiation. There is no significant difference in the number of foci between the cell lines for the same LET. From 0.5 to 24 h, the number of foci drops reaching its residual level. For each time point, there are small differences in DNA DSB among the three cell lines. CONCLUSION: Analyses of data acquired for the three cell lines irradiated by helium ions, having different LET, reveal high elimination capacity and creation of a large number of DNA DSB with respect to γ-rays, and are between those reported for protons and carbon ions.

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OBJECTIVE: Aim: The aim is to study the motivation of students towards Internet dependent behavior and develop practical recommendations for improving a set of measures for its prevention. PATIENTS AND METHODS: Materials and Methods: The research involved 154 students of the National Academy of Internal Affairs. Research methods: analysis and generalization of literature sources, questionnaire, statistical methods. RESULTS: Results: The priority motives of students who manifest Internet dependence behavior were identif i ed. The motivational orientations of students determine their systematic stay in the virtual environment and include, first of all, compliance with modern world trends; accessibility of content; the need for recognition of personal results by other users; satisfaction with virtual communication with the social environment. CONCLUSION: Conclusions: The practical recommendations for improving a set of measures to counteract the spread of Internet dependence among students were developed. Overcoming Internet dependence involves influencing a person to change his or her motivational and value as well as communication spheres. Prevention of Internet dependence involves public health professionals conducting awareness-raising as well as psychological and correctional work with the most vulnerable categories of people.

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This study delves into the intriguing properties of the 1H/1T-TaS2 van der Waals heterostructure, focusing on the transparency of the 1H layer to the charge density wave of the underlying 1T layer. Despite the sizable interlayer separation and metallic nature of the 1H layer, positive bias voltages result in a pronounced superposition of the 1T charge density wave structure on the 1H layer. The conventional explanation relying on tunneling effects proves insufficient. Through a comprehensive investigation combining low-temperature scanning tunneling microscopy, scanning tunneling spectroscopy, non-contact atomic force microscopy, and first-principles calculations, we propose an alternative interpretation. The transparency effect arises from a weak yet substantial electronic coupling between the 1H and 1T layers, challenging prior understanding of the system. Our results highlight the critical role played by interlayer electronic interactions in van der Waals heterostructures to determine the final ground states of the systems.

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This study aimed to improve the conventional procedure of alginate isolation from the brown seaweed (Laminaria digitata L.) biomass and investigate the possibility of further valorization of the ethanolic fraction representing the byproduct after the degreasing and depigmentation of biomass. The acid treatment of biomass supported by ultrasound was modeled and optimized regarding the alginate yield using a response surface methodology based on the Box-Behnken design. A treatment time of 30 min, a liquid-to-solid ratio of 30 mL/g, and a treatment temperature of 47 °C were proposed as optimal conditions under which the alginate yield related to the mass of dry biomass was 30.9%. The use of ultrasonic radiation significantly reduced the time required for the acid treatment of biomass by about 4 to 24 times compared to other available conventional procedures. The isolated alginate had an M/G ratio of 1.08, which indicates a greater presence of M-blocks in its structure and the possibility of forming a soft and elastic hydrogel with its use. The chemical composition of the ethanolic fraction including total antioxidant content (293 mg gallic acid equivalent/g dry weight), total flavonoid content (14.9 mg rutin equivalent/g dry weight), contents of macroelements (the highest content of sodium, 106.59 mg/g dry weight), and microelement content (the highest content of boron, 198.84 mg/g dry weight) was determined, and the identification of bioactive compounds was carried out. The results of ultra high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry analysis confirmed the presence of 48 compounds, of which 41 compounds were identified as sugar alcohol, phenolic compounds, and lipids. According to the 2,2-diphenyl-1-picrylhydrazyl assay, the radical scavenging activity of the ethanolic fraction (the half-maximal inhibitory concentration of 42.84 ± 0.81 µg/mL) indicated its strong activity, which was almost the same as in the case of the positive control, synthetic antioxidant butylhydroxytoluene (the half-maximal inhibitory concentration of 36.61 ± 0.79 µg/mL). Gram-positive bacteria (Staphylococcus aureus, Enterococcus faecalis, and Bacillus cereus) were more sensitive to the ethanolic fraction compared to Gram-negative bacteria (Escherichia coli, Pseudomonas aeruginosa, and Shigella sonnei). The obtained results indicated the possibility of the further use of the ethanolic fraction as a fertilizer for plant growth in different species and antifouling agents, applicable in aquaculture.

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Macrophytes and cladocerans represent the main antagonistic groups that regulate phytoplankton biomass; however, the mechanism behind this interaction is unclear. In laboratory conditions, we separately evaluated the effects of three submerged macrophytes (Ceratophyllum demersum, Myriophyllum aquaticum, and Stuckenia pectinata), as well as their exudates, and plant-associated microbiota (POM < 25 µm) + exudates on the population growth of Daphnia cf. pulex and Simocephalus cf. mixtus. Living Ceratophyllum, exudates, and POM < 25 µm + exudates exhibited the most robust positive effects on Simocephalus density and the rate of population increase (r). Subsequently, we examined the effects of Ceratophyllum on the filtration and feeding rates of Simocephalus and Daphnia, revealing significant (p < 0.001) promotion of filtration and feeding in Simocephalus but not in Daphnia. To elucidate the specific effects of this macrophyte on Simocephalus demography, we assessed selected life table variables across the same treatments. The treatments involving exudates and living Ceratophyllum resulted in approximately 40% longer survivorship and significantly (p < 0.01) enhanced fecundity. Our findings indicate that exudates from submerged macrophytes positively influence Simocephalus demography by increasing filtration rates, survivorship, and fecundity. This synergy suggests a substantial impact on phytoplankton abundance.

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Glycans of MVs are proposed to be candidates for mediating targeting specificity or at least promoting it. In contrast to exosomes, glycomic studies of MVs are largely absent. We studied the glycoprofile of endothelial cell-derived MVs using 21 plant lectins, and the results show the dominance of oligolactosamines and their α2-6-sialylated forms as N-glycans and low levels of α2-3-sialylated glycans. The low levels of α2-3-sialosides could not be explained by the action of extracellular glycosidases. Additionally, the level of some Man-containing glycans was also decreased in MVs. Spatial masking as the causative relationship between these low level glycans (as glycosphingolipids) by integral proteins or proteoglycans (thus, their lack of interaction with lectins) seems unlikely. The results suggest that integral proteins do not pass randomly into MVs, but instead only some types, differing in terms of their specific glycosylation, are integrated into MVs.

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Background/Objectives: The intranasal delivery of various neurotropic substances is considered a new attractive therapeutic approach for treating neuropathologies associated with neuroinflammation and altered regeneration. Specific language impairment (SLI) that arises as a result of damage to the cortical speech zones during the developmental period is one of the most common problems in preschool children, and it is characterized by persistent difficulties in the acquisition, understanding, and use of language. This study's objective is to evaluate the efficacy and safety of intranasal immunotherapy using the M2 macrophage secretome as a rich source of immunoregulatory and neurotrophic factors for the treatment of severe language impairment in children. Methods: Seventy-one children (54 boys and 17 girls, aged 3 to 13 years) were recruited to participate in a clinical trial (NCT04689282) in two medical centers. The children were examined before, 1 month after, and 6 months after the start of therapy. In the vast majority of children (55/71), language impairment was associated with autistic-like symptoms and attention deficit hyperactivity disorder (ADHD). Results: Daily intranasal inhalations of M2 macrophage-conditioned medium (for 30 days) were well tolerated and led to a decrease in the severity of language impairments, autistic-like behavior, and ADHD symptoms. The clinical effect appeared within a month after the first procedure and persisted or intensified during a 6-month follow-up. Two-thirds of the children showed a clear clinical improvement, while the rest had less pronounced improvement. Conclusions: Thus, the use of the M2 macrophage secretome and its intranasal delivery is safe, well tolerated, and clinically effective in children with severe language impairments.

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In the continuum from 'farm to fork', the proficiency of food handlers in terms of knowledge, attitudes, and practices (KAP) is essential for ensuring improved food safety outcomes. This study evaluated the KAP of fresh Nile perch fish handlers regarding food safety requirements within Uganda. A cross-sectional survey involving fish handlers (n = 466) engaged in handling fresh Nile perch and fisheries products in both local and export market chains. The study was conducted at 30 gazetted fish landing sites on Lake Victoria, 15 licensed fish export factories, and 9 local markets authorized to sell fisheries products. Data on demographic, knowledge, attitude, and practices was collected using structured questionnaire. Demographic data was analysed using frequencies and percentages. Ordinary least squares bivariate and multivariate regression analyses were performed to determine factors associated with knowledge, attitudes, and practices regarding food safety requirements. Fish handlers (49.8 %) demonstrated good knowledge with the majority (74.6 %) from the export market chain. Majority (81.3 %) of market stall handlers had poor knowledge of food safety requirements. Most (68.2 %) fresh Nile perch fish handlers had limited awareness of food-borne pathogens. Only 31.8 % had knowledge about E. coli, Salmonella spp, and Staphylococcus spp. Furthermore, over 63 % believed that consuming contaminated fish could transmit HIV, Covid-19, and Ebola. Male gender, higher income, and advanced education were positively correlated with fish handlers' knowledge of food safety requirements. Fish handlers (57 %) in the local market chain exhibited poor attitudes. Being a male fish handler was significantly (ß = 3.43, 95 % CI: 1.65-5.21, P = 0.001) associated with positive attitudes compared to being female. Education at different levels was significantly (Primary (ß = 3.19, 95 % CI: 1.05-5.33, P = 0.004); Secondary level (ß = 5.883, 95 % CI: 3.52-8.23, P = 0.001); Tertiary (ß = 6.09, 95 % CI: 3.03-9.15, P = 0.001)) associated with positive attitudes compared to no education. Practices ensuring food safety were common in the factories and the export market. The study suggests the need for interventions that promote knowledge transfer and food safety culture, particularly in the local market chain.