Integrative Modeling of 3D Genome Organization by Bayesian Molecular Dynamics Simulations with Hi-C Metainference.

Polymer modeling has been playing an increasingly important role in complementing 3D genome experiments, both to aid their interpretation and to reveal the underlying molecular mechanisms. This chapter illustrates an application of Hi-C metainference, a Bayesian approach to explore the 3D organization of a target genomic region by integrating experimental contact frequencies into a prior model of chromatin. The method reconstructs the conformational ensemble of the target locus by combining molecular dynamics simulation and Monte Carlo sampling from the posterior probability distribution given the data. Using prior chromatin models at both 1 kb and nucleosome resolution, we apply this approach to a 30 kb locus of mouse embryonic stem cells consisting of two well-defined domains linking several gene promoters together. Retaining the advantages of both physics-based and data-driven strategies, Hi-C metainference can provide an experimentally consistent representation of the system while at the same time retaining molecular details necessary to derive physical insights.

Commentary: Best practices for performing olfactory behavioral assays on aquatic animals: A guide for comparative physiologists.

As more physiologists start to incorporate animal behavior into their experiments, especially in the olfactory behavior research field, some considerations are often overlooked, partly due to the inherited way that physiological experiments are traditionally designed and performed. Here we highlight some of these subtle but important considerations and make a case for why these might affect the results collected from behavioral assays. Our aim is to provide useful suggestions for increased standardization of methods so they can be more easily replicated among different experiments and laboratories. We have focused on areas that are less likely to be mentioned in the materials and methods section of a manuscript such as starvation, preliminary experiments, appropriate sample sizes and considerations when choosing an odorant for an assay. Additionally, we are strongly cautioning against the use of alarm cue to generate behavioral responses due to its highly unstable chemical properties/potency. Instead, we suggest using pure chemicals (made up of one known molecule) such as amino acids, bile acids, or polyamines that are commercially available and easier to make up in known concentrations. Lastly, we strongly suggest using environmentally relevant concentrations of these odorants. We believe these guidelines will help standardize these assays and improve replication of experiments within and between laboratories.

Plant extract-mediated biosynthesis of sulphur nanoparticles and their antibacterial and plant growth-promoting activity.

This study reports green synthesis of sulphur nanoparticles using sodium thiosulfate pentahydrate (Na2S2O35H2O) and Cannabis sativa leaf extracts. X-ray diffraction (XRD) pattern and scanning electron microscopy (SEM) was employed to examine the crystallinity of the particles and morphological characteristics, proved both spherical and rod-shaped morphology of the S NPs having porous nature. The FTIR spectra revealed the interaction of the synthesized SNPs with the biomolecules present in the leaf extract. UV-VIS spectral investigations confirmed the production of SNPs from C. sativa leaf extract and that these SNPs can be used for visible region photocatalysis for the removal of pollutants from wastewater. Energy dispersive X-ray (EDX) spectrum of the SNP shows a single peak around 2.4 keV, confirmed S NPs purity. TEM image revealed the formation of mainly nanorods having a width of ∼20-25 nm and a length of 50-100 nm. Furthermore, some spherical particles (∼20-30 nm) were also formed. HRTEM image of the rod-shaped particles clearly shows the crystal fringe spacing of 0.38 nm. Further, disc diffusion method (DDM) was used to check the antibacterial activity of S NPs against gram-positive S. aureus (MTCC737) 18 ± 0.12 mm and gram-negative bacteria against E. coli (MTCC443) 21.5 ± 0.12 mm, A. salmonicida (MTCC1522) 19.1 ± 0.12 mm, K. pneumoniae (MTCC3384) 17.8 ± 0.10 mm. Among all the strains of bacteria, E. coli (MTCC443) showed a maximum zone of inhibition of 21.5 ± 0.12 mm and its antibacterial activity is somewhat like streptomycin sulfate. These SNPs also promote growth of C. sativa in pot experiment, resulting in a 30 % increase in biomass, 90 cm in shoot length and 28 cm in root length and higher fresh and dry weight (50g and 20g, respectively) with 1.0 mg mL-1 NPs treatment. In addition, SEM-EDX confirmed the accumulation of nanomaterial in plant leaves. This environmentally friendly approach to SNP synthesis using C. sativa extracts demonstrates both potent antibacterial properties and plant growth-promoting effects, making it a promising solution for agriculture and biomedicine.

Quality by Design for Preclinical In Vitro Assay Development.

Quality by Design (QbD) is an approach to assay development to determine the design space, which is the range of assay variable settings that should result in satisfactory assay quality. Typically, QbD is applied in manufacturing, but it works just as well in the preclinical space. Through three examples, we illustrate the QbD approach with experimental design and associated data analysis to determine the design space for preclinical assays.

Thoughts and suggestions about ethical requirement in acupuncture animal experiments. / 针刺动物实验中的伦理要求思考及建议.

The animal experiment is an important form of acupuncture research, which is of great significance for revealing the regularities and mechanisms of acupuncture effects. The ethical principle of animal welfare is the important guarantee of animal rights and experimental scientificity in the process of research. Starting from the "3R principle" and "five welfare" rules, and the specific operation process of acupuncture animal experiments, we, in this paper, proposed the implementation methods of animal ethics in acupuncture experiments, including ethical execution before acupuncture experiments, and ethical requirements during the experimental stage (such as animal fixation method that meets the needling needs, selection of acupuncture apparatus, acupuncture manipulations, acupuncture stimulation intensity, and sampling of animal tissues). These proposed methods may provide some appropriate references for the guarantee of animal ethics in acupuncture research, and provide ideas for establishing a new paradigm of animal welfare ethics in acupuncture animal experiments, and finally promote the standardization and scientificity process of the implementation of animal welfare ethics in acupuncture animal experiments.

What Would Miffy Do? Applying Informed Consent by Proxy to All Sentient Animals.

If we want to take sentient non-human animals and their interests seriously, we can try to ask for their consent before using them for human purposes. With mentally competent humans, we speak of informed consent: for them to participate in scientific studies, for example, it is required that they consent explicitly, in full understanding of the risks and benefits. This full understanding cannot be expected from non-human animals. We must therefore look for ways to know what they want and to estimate what they would do if they had a deep understanding of their options and the consequences of these options for themselves and others. This concept is explored by applying it to animal experiments. The most promising method is to gain informed consent by proxy from thoroughly informed competent humans, in combination with seeking assent where possible and being alert to dissent.

A Lab-Scale Evaluation of Parameters Influencing the Mechanical Activation of Kaolin Using the Design of Experiments.

This research investigates the mechanical activation of kaolin as a supplementary cementitious material at the laboratory scale, aiming to optimize milling parameters using the response surface methodology. The study evaluated the effects of rotation speed and milling time on the amorphous phase content, the reduction in crystalline kaolinite, and impurity incorporation into the activated clay through the Rietveld method. The results demonstrated that adjusting milling parameters effectively enhanced clay activation, which is crucial for its use in low-carbon cements. High rotation speeds (300/350 rpm) and prolonged grinding times (90/120 min) in a planetary ball mill increased the pozzolanic activity by boosting the formation of amorphous phases from kaolinite and illite and reducing the particle size. However, the results evidenced that intermediate milling parameters are sufficient for reaching substantial degrees of amorphization and pozzolanic activity, avoiding the need for intensive grinding. Exceedingly aggressive milling introduced impurities like ZrO2 from the milling equipment wear, underscoring the need for a balanced approach to optimizing reactivity while minimizing impurities, energy consumption, and equipment wear. Achieving this balance is essential for efficient mechanical activation, ensuring the prepared clay's suitability as supplementary cementitious materials without excessive costs or compromised equipment integrity.

Exploring the key flavor compounds and aroma profiles in traditionally fermented golden pompano (Trachinotus ovatus): Perspective from molecular sensory science.

Fermented golden pompano (Trachinotus ovatus) has a distinctive flavor, but the key flavor compounds and aroma profiles remains unclear. Thus, a molecular sensory science approach was used to investigate flavor and key aromatic compounds. The fermentation process enhanced the overall flavor, as evidenced by sensory evaluation and electronic nose (E-nose) analyses. A total of 48 aroma compounds were identified at different fermentation stages. Among them, 11 key aroma compounds were identified by flavor dilution (FD) factors ≥8 and odor activity values (OAVs) ≥ 1. Aroma recombination model successfully reproduces the characteristic floral and fruity aromas of fermented golden pompano. Omission experiments identified hexanal, decanal, 1-octen-3-ol, limonene, and isovaleric acid as significant contributors to the overall aroma profile. This study elucidates flavor dynamic modulation and key aromatic compounds during golden pompano fermentation, to provide a theoretical reference for the targeting process regulation of the product.

Chondroitin Sulfate for Cartilage Regeneration, Administered Topically Using a Nanostructured Formulation.

In the pharmaceutical sector, solid lipid nanoparticles (SLN) are vital for drug delivery incorporating a lipid core. Chondroitin sulfate (CHON) is crucial for cartilage health. It is often used in osteoarthritis (OA) treatment. Due to conflicting results from clinical trials on CHON's efficacy in OA treatment, there has been a shift toward exploring effective topical systems utilizing nanotechnology. This study aimed to optimize a solid lipid nanoparticle formulation aiming to enhance CHON permeation for OA therapy. A 3 × 3 × 2 Design of these experiments determined the ideal parameters: a CHON concentration of 0.4 mg/mL, operating at 20,000 rpm speed, and processing for 10 min for SLN production. Transmission electron microscopy analysis confirmed the nanoparticles' spherical morphology, ensuring crucial uniformity for efficient drug delivery. Cell viability assessments showed no significant cytotoxicity within the tested parameters, indicating a safe profile for potential clinical application. The cell internalization assay indicates successful internalization at 1.5 h and 24 h post-treatment. Biopharmaceutical studies supported SLNs, indicating them to be effective CHON carriers through the skin, showcasing improved skin permeation and CHON retention compared to conventional methods. In summary, this study successfully optimized SLN formulation for efficient CHON transport through pig ear skin with no cellular toxicity, highlighting SLNs' potential as promising carriers to enhance CHON delivery in OA treatment and advance nanotechnology-based therapeutic strategies in pharmaceutical formulations.

Can We Simplify Liposome Manufacturing Using a Complex DoE Approach?

Microfluidic liposome production presents a streamlined pathway for expediting the translation of liposomal formulations from the laboratory setting to clinical applications. Using this production method, resultant liposome characteristics can be tuned through the control of both the formulation parameters (including the lipids and solvents used) and production parameters (including the production speed and mixing ratio). Therefore, the aim of this study was to investigate the relationship between not only total flow rate (TFR), the fraction of the aqueous flow rate over the organic flow rate (flow rate ratio (FRR)), and the lipid concentration, but also the solvent selection, aqueous buffer, and production temperature. To achieve this, we used temperature, applying a design of experiment (DoE) combined with machine learning. This study demonstrated that liposome size and polydispersity were influenced by manipulation of not only the total flow rate and flow rate ratio but also through the lipids, lipid concentration, and solvent selection, such that liposome attributes can be in-process controlled, and all factors should be considered within a manufacturing process as impacting on liposome critical quality attributes.

Sustainable application of modified Luffa cylindrica biomass for removal of trimethoprim in water by adsorption with process optimization.

The present study describes a set of methodological procedures (seldom applied together), including (i) development of an alternative adsorbent derived from abundant low-cost plant biomass; (ii) use of simple low-cost biomass modification techniques based on physical processing and chemical activation; (iii) design of experiments (DoE) applied to optimize the removal of a pharmaceutical contaminant from water; (iv) at environmentally relevant concentrations, (v) that due to initial low concentrations required determination by ultra-performance liquid phase chromatography coupled to mass spectrometry (UPLC-MS/MS). A central composite rotational design (CCRD) was employed to investigate the performance of vegetable sponge biomass (Luffa cylindrica), physically processed (crushing and sieving) and chemically activated with phosphoric acid, in the adsorption of the antibiotic trimethoprim (TMP) from water. The optimized model identified pH as the most significant variable, with maximum drug removal (91.1 ± 5.7%) achieved at pH 7.5, a temperature of 22.5 °C, and an adsorbent/adsorbate ratio of 18.6 mg µg-1. The adsorption mechanisms and surface properties of the adsorbent were examined through characterization techniques such as scanning electron microscopy (SEM), point of zero charge (pHpzc) measurement, thermogravimetric analysis (TGA), specific surface area, and Fourier-transform infrared spectroscopy (FTIR). The best kinetic fit was obtained by the Avrami fractional-order model. The hypothesis of a hybrid behavior of the adsorbent was suggested by the equilibrium results presented by the Langmuir and Freundlich models and reinforced by the Redlich-Peterson model, which achieved the best fit (R2 = 0.982). The thermodynamic study indicated an exothermic, spontaneous, and favorable process. The maximum adsorption capacity of the material was 2.32 × 102 µg g-1 at an equilibrium time of 120 min. Finally, a sustainable and promising adsorbent for the polishing of aqueous matrices contaminated by contaminants of emerging concern (CECs) at environmentally relevant concentrations is available for future investigations.

Ash characteristics investigation from co-firing of Calliandra calothyrsus and Gliricidia sepium with a high-sulfur and -iron coal in drop tube furnace.

Calliandra calothyrsus and Gliricidia sepium ash characteristics as co-firing fuel to high-sulfur and -iron coal were investigated through theoretical prediction, FactSage modelling, and combustion experiment in drop tube furnace. The results show that the addition of Calliandra and Gliricidia were able to mitigate the slagging risk according to theoretical prediction and FactSage modelling. Slag formations were predicted to occur at higher combustion temperature compared to coal which was reflected in higher ash fusion temperature. The results of combustion experiment showed an agreement with less ash deposition and cleaner probe surface. The addition of Calliandra and Gliricidia increased the high-melting minerals and decreased Fe-based minerals. However, domination of Ca2SO4 in addition of 25 wt% Gliricidia can be detrimental with presence of strongly adhered ash and material degradation. This research provides an important insight of Calliandra and Gliricidia utilization as co-firing fuel that improve the ash characteristics of high-sulfur and -iron coal.

Inheritance patterns of male asexuality in hybrid males of a water frog Pelophylax esculentus.

Gametogenesis produces gametes as a piece of genetic information transmitted to the offspring. While during sexual reproduction, progeny inherits a mix of genetic material from both parents, asexually reproducing organisms transfer a copy of maternal or paternal DNA to the progeny clonally. Parthenogenetic, gynogenetic and hybridogenetic animals have developed various mechanisms of gametogenesis, however, their inheritance is not fully understood. Here, we focused on the inheritance of asexual gametogenesis in hybrid Pelophylax esculentus (RL), emerging after crosses of P. lessonae (LL) and P. ridibundus (RR). To understand the mechanisms of gametogenesis in hybrids, we performed three-generation experiments of sexual P. ridibundus females and hybrids from all-male hybrid populations. Using fluorescent in situ hybridization, micronuclei analysis, flow cytometry and genotyping, we found that most adult hybrid males simultaneously produced two types of clonal sperm. Also, most male tadpole progeny in two successive backcrossed generations simultaneously eliminated L and R parental genomes, while some progeny produced only one type of sperm. We hypothesize that the reproductive variability of males producing two kinds of sperm is an adaptive mechanism to reproduce in mixed populations with P. ridibundus and may explain the extensive distribution of the all-male lineage across the European River Basin.

Lignin extraction from acacia wood: Crafting deep eutectic solvents with a systematic D-optimal mixture-process experimental design.

Lignin is a complex biopolymer whose efficient extraction from biomass is crucial for various applications. Deep eutectic solvents (DES), particularly natural-origin DES (NADES), have emerged as promising systems for lignin fractionation and separation from other biomass components. While ternary DES offer enhanced fractionation performance, the role of each component in these mixtures remains unclear. In this study, the effects of adding tartaric acid (Tart) or citric acid (Cit) to a common binary DES mixture composed of lactic acid (Lact) and choline chloride (ChCl) were investigated for lignin extraction from acacia wood. Ternary Cit-based DES showed superior performance compared to Tart-based DES. Using a combined mixture-process D-Optimal experimental design, the Lact:Cit:ChCl DES composition and extraction temperature were optimized targeting maximum lignin yield and purity. The optimal conditions (i.e., Lact:Cit:ChCl, 0.6:0.3:0.1 M ratio, 140 °C) resulted in a lignin extraction yield of 99.63 ±â€¯1.24 % and a lignin purity of 91.45 ±â€¯1.03 %. Furthermore, this DES exhibited feasible recyclability and reusability without sacrificing efficiency.

Network Pharmacology and in vitro Experimental Verification on Intervention of Oridonin on Non-Small Cell Lung Cancer.

OBJECTIVE: To explore the key target molecules and potential mechanisms of oridonin against non-small cell lung cancer (NSCLC). METHODS: The target molecules of oridonin were retrieved from SEA, STITCH, SuperPred and TargetPred databases; target genes associated with the treatment of NSCLC were retrieved from GeneCards, DisGeNET and TTD databases. Then, the overlapping target molecules between the drug and the disease were identified. The protein-protein interaction (PPI) was constructed using the STRING database according to overlapping targets, and Cytoscape was used to screen for key targets. Molecular docking verification were performed using AutoDockTools and PyMOL software. Using the DAVID database, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis were conducted. The impact of oridonin on the proliferation and apoptosis of NSCLC cells was assessed using cell counting kit-8, cell proliferation EdU image kit, and Annexin V-FITC/PI apoptosis kit respectively. Moreover, real-time quantitative PCR and Western blot were used to verify the potential mechanisms. RESULTS: Fifty-six target molecules and 12 key target molecules of oridonin involved in NSCLC treatment were identified, including tumor protein 53 (TP53), Caspase-3, signal transducer and activator of transcription 3 (STAT3), mitogen-activated protein kinase kinase 8 (MAPK8), and mammalian target of rapamycin (mTOR). Molecular docking showed that oridonin and its key target molecules bind spontaneously. GO and KEGG enrichment analyses revealed cancer, apoptosis, phosphoinositide-3 kinase/protein kinase B (PI3K/Akt), and other signaling pathways. In vitro experiments showed that oridonin inhibited the proliferation, induced apoptosis, downregulated the expression of Bcl-2 and Akt, and upregulated the expression of Caspase-3. CONCLUSION: Oridonin can act on multiple targets and pathways to exert its inhibitory effects on NSCLC, and its mechanism may be related to upregulating the expression of Caspase-3 and downregulating the expressions of Akt and Bcl-2.

An acoustic bellows-type round window transducer for middle-ear implants.

BACKGROUND: This study describes the development of output devices for round window middle-ear. To overcome the problems of output devices that apply sound pressure directly to the round window, an acoustic bellows-type round window transducer was implemented by combining a small bellows, acoustic tube, and balanced armature driver. METHODS: The output characteristics of the proposed acoustic bellows-type round window transducer were confirmed through bench tests and distortion measurements. To compare the vibration transmission characteristics of the proposed transducer with those of sound pressure stimulation devices, an experiment was performed using four human temporal bones. FINDINGS: The average output magnitude of the acoustic bellows-type round window transducer was equivalent to sound pressure levels of 92, 96, and 108 dB for frequency ranges of <1, 1-2, and > 2 kHz, respectively. The results showed that the proposed transducer delivered vibration consistently without reducing the sound pressure level due to leakage, unlike the sound pressure stimulation device. INTERPRETATION: Therefore, the acoustic bellows-type round window transducer is a more stable and suitable output device for round window middle-ear implants than a sound pressure stimulation device. It is expected to overcome the limitations of sound pressure stimulation devices and to contribute to new technical solutions in the field of round window middle-ear implants development.

Comparison of different methods for categorizing the individual susceptibility to motion sickness - A carsickness study.

In studies aimed at developing avoidance strategies to reduce motion sickness (kinetosis) in autonomous vehicles, failing to account for the wide variability in individual kinetosis susceptibility can lead to inaccuracies and disregard effective countermeasures. Three methods for assessing individual susceptibility to carsickness - two questionnaires focusing on kinetosis experiences and a kinetosis-provoking lab test - were compared with the development of kinetosis during real car driving tests. Questions about car-specific kinetosis-provoking situations (MS-C) exhibit stronger correlations with kinetosis in car experiments compared to the commonly used questions about kinetosis experiences across different types of transportation (MS-VD). While lab-based testing remains highly reliable, especially considering men's tendency to underestimate their carsickness susceptibility in questionnaires, MS-C provides a valuable compromise in terms of technical and time expenses. These findings can also be used to assist passengers of autonomous driving cars in accurately assessing their sensitivity and activating customized countermeasure functions.

Development and integration of a continuous horizontal belt filter into drug production procedure.

In the pharmaceutical industry, filtration is traditionally carried out in batch mode. However, with the spread of continuous technologies, there is an increasing demand for robust continuous filtration strategies suitable for processing suspensions produced in continuous crystallizers. Accordingly, this study aimed to investigate a lab-scale horizontal conveyor belt filtration approach for pharmaceutical separation purposes for the first time. The newly developed continuous horizontal belt filter (CHBF) was tested under different systems (microcrystalline cellulose (MCC)/water, lactose/ethanol and acetylsalicylic acid (ASA)/water) and diverse conditions. Filtration was robust using a well-defined unimodal particle size distribution MCC in water system, where the residual moisture content varied within narrow limits of 45-52% independently from the process conditions. Besides, the residual moisture content highly depended on the applied solvent and particle size. It could be reduced to below 2% by processing the suspensions of either a volatile solvent (lactose in ethanol) or an aqueous slurry of a large particle size ASA. Finally, the CHBF was connected to a mixed suspension mixed product removal (MSMPR) or a plug flow crystallizer (PFC). The residual moisture content of the CHBF-filtered ASA product and operation characteristics (onset of steady-state) were evaluated in both continuous crystallizer-filter systems. The MSMPR-CHBF system operated with a longer startup period. The size of the in situ-produced crystals was of a similar order magnitude in both systems, resulting in a similar residual moisture content (around 20%). Overall, the tested continuous filter was robust, did not modify the crystal morphology in the examined experimental range, and could be effectively integrated with continuous crystallizers.

Unraveling the complexities of antifungal susceptibility testing in Candida spp.: Insights from design of experiments.

Our study delved into the intricate dynamics of antifungal susceptibility testing for Candida spp., employing a Design of Experiments approach. We systematically investigated the influence of pH, temperature, inoculum size, and glucose concentration on both growth patterns and inhibitory concentrations of Candida spp. Our findings underscore the nuanced interplay between these factors, revealing significant impacts on susceptibility outcomes. Notably, even minor adjustments in these parameters yielded substantial variations in growth and inhibitory concentrations, underscoring the critical importance of meticulous control over growth conditions in antifungal susceptibility testing protocols. Each Candida isolates exhibited unique susceptibility profiles, necessitating tailored culture conditions for accurate testing. Our study sheds light on the variability inherent in Candida spp. growth patterns and emphasizes the need for standardized protocols to ensure consistency across laboratories. By leveraging the design of experiments, our research provides a systematic framework for unraveling the complexities of antifungal susceptibility testing, offering valuable insights for optimizing testing protocols and informing clinical decision-making in antifungal treatment. These findings represent a significant step towards enhancing the efficacy and reliability of antifungal susceptibility testing in clinical practice.

Unlocking DOE potential by selecting the most appropriate design for rAAV optimization.

Producing recombinant adeno-associated virus (rAAV) for gene therapy via triple transfection is an intricate process involving many cellular interactions. Each of the different elements encoded in the three required plasmids-pHelper, pRepCap, and pGOI-plays a distinct role, affecting different cellular pathways when producing rAAVs. The required expression balance emphasizes the critical need to fine-tune the concentration of all these different elements. The use of design of experiments (DOE) to find optimal ratios is a powerful method to streamline the process. However, the choice of the DOE method and design construction is crucial to avoid misleading results. In this work, we examined and compared four distinct DOE approaches: rotatable central composite design (RCCD), Box-Behnken design (BBD), face-centered central composite design (FCCD), and mixture design (MD). We compared the abilities of the different models to predict optimal ratios and interactions among the plasmids and the transfection reagent. Our findings revealed that blocking is essential to reduce the variability caused by uncontrolled random effects and that MD coupled with FCCD outperformed all other approaches, improving volumetric productivity 109-fold. These outcomes underscore the importance of selecting a model that can effectively account for the biological context, ultimately yielding superior results in optimizing rAAV production.