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Enhanced mixing in microfluidic systems is necessary in many applications such as chemical processing, biological assays, and diagnosis. We are developing a microfluidic system to efficiently mix minute reagents (down to several microliters) using vibration-induced flow (VIF), in which a net flow is generated around a micropillar by applying periodic vibration. In this study, we numerically investigate the enhancement in chaotic mixing using the VIF technique and periodic switching of vibrations. By extending our previous numerical simulation model, we investigate the flow field and trajectories of fluid particles in three-dimensional space. We demonstrate that chaotic advection characteristics can be observed by periodically switching the vibrational direction of a substrate using simple cylindrical pillars. In addition, using an appropriate interval for switching the vibration axes yields better mixing performance. The extent of chaotic advection is evaluated quantitatively using the Lyapunov exponent considering various vibration parameters, such as the vibration amplitude, separation distance between each pillar and pillar shape. The flow field induced by a large-amplitude and sharp-edged wall pillar provides excellent mixing results. Thus, VIF is successfully applied to obtain an efficient mixing strategy with the aid of the chaotic theory. [ABSTRACT FROM AUTHOR]

Detection sensitivity of aquatic species using environmental DNA (eDNA) generally decreases in turbid water but is poorly characterized. In this study, eDNA detection targeted delta smelt (Hypomesus transpacificus), a critically endangered estuarine fish associated with turbid water. eDNA sampling in the field was first paired with a trawl survey. Species-specific detection using a Taqman qPCR assay showed concordance between the methods, but a weak eDNA signal. Informed by the results of field sampling, an experiment was designed to assess how turbidity and filtration methods influence detection of a rare target. Water from non-turbid (5 NTU) and turbid (50 NTU) estuarine sites was spiked with small volumes (0.5 and 1 mL) of water from a delta smelt tank to generate low eDNA concentrations. Samples were filtered using four filter types: cartridge filters (pore size 0.45 μm) and 47 mm filters (glass fiber, pore size 1.6 μm and polycarbonate, pore sizes 5 and 10 μm). Prefiltration was also tested as an addition to the filtration protocol for turbid water samples. eDNA copy numbers were analyzed using a censored data method for qPCR data. The assay limits and lack of PCR inhibition indicated an optimized assay. Glass fiber filters yielded the highest detection rates and eDNA copies in non-turbid and turbid water. Prefiltration improved detection in turbid water only when used with cartridge and polycarbonate filters. Statistical analysis identified turbidity as a significant effect on detection probability and eDNA copies detected; filter type and an interaction between filter type and prefilter were significant effects on eDNA copies detected, suggesting that particulate-filter interactions can affect detection sensitivity. Pilot experiments and transparent criteria for positive detection could improve eDNA surveys of rare species in turbid environments.

mRNA measurement is dominated by RT-PCR, which requires expensive laboratory equipment and personnel with advanced degrees. Loop-mediated isothermal amplification (LAMP) is a versatile technique for detecting target DNA and RNA. The sensitivity of LAMP in early reports has been below that of the standard RT-PCR tests. Here, we report the use of a fluorescence-based RT-LAMP protocol to measure CDX2 expression patterns, which match extremely well to the standards of sophisticated RT-PCR techniques (r = 0.99, p

Cerium oxide (CeO2) and yttrium oxide (Y2O3) nanoparticles possess interesting surface properties and interfacial interactions that make them attractive candidates for various applications. This study presents a comprehensive investigation into the synthesis and toxicity assessment of yttrium-doped cerium oxide (Ce-Y) nanocomposite using a combination of analytical techniques and biological assays. The nanocomposite was characterized using UV–Vis spectroscopy, X-ray diffraction (XRD), Fourier-transform infrared spectroscopy (FTIR), electron microscopy imaging, dynamic light scattering (DLS), and zeta potential measurements to elucidate their structural, optical and physicochemical properties. The synthesized nanocomposite exhibited distinctive absorption spectra and precise alignment of diffraction peaks, confirming the successful incorporation of yttrium ions into the cerium oxide lattice. Electron microscopy images revealed well-dispersed yttrium particles within the ceria matrix, indicating uniform distribution and morphology. DLS and zeta potential analysis provided insights into the size distribution and stability of the nanocomposite. Furthermore, in vivo toxicity assessment using Drosophila melanogaster model revealed no significant toxicity of Ce:Y nanocomposite, as evidenced by survival assay and behavioral assays. Superoxide dismutase (SOD) activity and reactive oxygen species (ROS) levels were also evaluated, demonstrating no discernible nanoparticle-induced toxicity. Overall, this study highlights the potential applications of Ce:Y nanocomposite and underscores the importance of comprehensive toxicity evaluation in nanomaterial development. [ABSTRACT FROM AUTHOR]

The translated protein undergoes a specific modification process, which involves the formation of covalent bonds on lysine residues and the attachment of small chemical moieties. The protein's fundamental physicochemical properties undergo a significant alteration. The change significantly alters the proteins' 3D structure and activity, enabling them to modulate key physiological processes. The modulation encompasses inhibiting cancer cell growth, delaying ovarian aging, regulating metabolic diseases, and ameliorating depression. Consequently, the identification and comprehension of post-translational lysine modifications hold substantial value in the realms of biological research and drug development. Post-translational modifications (PTMs) at lysine (K) sites are among the most common protein modifications. However, research on K-PTMs has been largely centered on identifying individual modification types, with a relative scarcity of balanced data analysis techniques. In this study, a classification system is developed for the prediction of concurrent multiple modifications at a single lysine residue. Initially, a well-established multi-label position-specific triad amino acid propensity algorithm is utilized for feature encoding. Subsequently, PreMLS: a novel ClusterCentroids undersampling algorithm based on MiniBatchKmeans was introduced to eliminate redundant or similar major class samples, thereby mitigating the issue of class imbalance. A convolutional neural network architecture was specifically constructed for the analysis of biological sequences to predict multiple lysine modification sites. The model, evaluated through five-fold cross-validation and independent testing, was found to significantly outperform existing models such as iMul-kSite and predML-Site. The results presented here aid in prioritizing potential lysine modification sites, facilitating subsequent biological assays and advancing pharmaceutical research. To enhance accessibility, an open-access predictive script has been crafted for the multi-label predictive model developed in this study. Author summary: Proteins undergo a variety of post-translational modifications (PTMs) after synthesis, such as lysine modifications, which significantly influence their structure and function. These modifications of lysine are known to regulate physiological processes, including the inhibition of cancer cell growth, the delay of aging, the regulation of metabolic diseases, and the improvement of depressive disorders. Abnormal modifications are closely associated with the occurrence and progression of a multitude of diseases. Therefore, the identification and comprehension of these modifications are of paramount importance for biological research and drug development. A multitude of studies have focused on a single type of lysine modification, with prediction methods for multiple lysine modification sites being relatively scarce. In this research, a novel multi-label prediction model named PreMLS has been developed for the simultaneous identification of four lysine modifications: methylation, acetylation, crotonylation, and succinylation. The imbalance issue in the dataset was addressed utilizing the ClusterCentroids undersampling algorithm, following which a predictive model, PreMLS, was constructed using a CNN to forecast multiple lysine modification sites. Compared to existing models, this new approach has significantly enhanced the accuracy and reliability of the predictions. [ABSTRACT FROM AUTHOR]

Numerous clinical disorders have been linked to the etiology of dysregulated NLRP3 (NACHT, LRR, and PYD domain‐containing protein 3) inflammasome activation. Despite its potential as a pharmacological target, modulation of NLRP3 activity remains challenging. Only a sparse number of compounds have been reported that can modulate NLRP3 and none of them have been developed into a commercially available drug. In this research, we identified three potent NLRP3 inflammasome inhibitors, gymnoasins A‐C (1–3), with unprecedented pentacyclic scaffolds, from an Antarctic fungus Pseudogymnoascus sp. HDN17‐895, which represent the first naturally occurring naphthopyrone‐macrolide hybrids. Additionally, biomimetic synthesis of gymnoasin A (1) was also achieved validating the chemical structure and affording ample amounts of material for exhaustive bioactivity assessments. Biological assays indicated that 1 could significantly inhibited in vitro NLRP3 inflammasome activation and in vivo pro‐inflammatory cytokine IL‐1β release, representing a valuable new lead compound for the development of novel therapeutics with the potential to inhibit the NLRP3 inflammasome. [ABSTRACT FROM AUTHOR]

Currently, research in the development of high-performance sensing platforms has increased to fulfill the needs of analysis and detection. In this study, we developed a novel type of surface-enhanced Raman scattering (SERS) chip composed of a covalent organic framework (COF)-silver nanoparticles (AgNPs) nanocomposite, and this nanocomposite was fabricated by a one-step method of ultrasonically mixing the obtained COF and AgNPs. The fabricated chip exhibited high sensitivity and repeatable SERS effects. Practical application results showed that the chip was highly sensitive and reliable and capable of detecting DNA bases (adenine) to fit an equation in the range from 0.01 pM to 1 nM, with an R-square of 0.97253 and a detection limit of ~0.026 pM (signal-to-noise ratio (S/N) = 3). Therefore, the proposed SERS system has potential applications in biological assays. Schematic of AgNP@COF as SERS substrates for the detection of DNA bases. [ABSTRACT FROM AUTHOR]

Arterial thrombosis is a leading cause of death and disability worldwide with no effective bioassay for clinical prediction. As a symbolic feature of arterial thrombosis, severe stenosis in the blood vessel creates a high-shear, high-gradient flow environment that facilitates platelet aggregation towards vessel occlusion. Here, we present a thrombus profiling assay that monitors the multi-dimensional attributes of thrombi forming in such biomechanical conditions. Using this assay, we demonstrate that different receptor–ligand interactions contribute distinctively to the composition and activation status of the thrombus. Our investigation into hypertensive and older individuals reveals intensified biomechanical thrombogenesis and multi-dimensional thrombus profile abnormalities, endorsing the diagnostic potential of the assay. Furthermore, we identify the hyperactivity of GPIbα-integrin αIIbβ3 mechanosensing axis as a molecular mechanism that contributes to hypertension-associated arterial thrombosis. By studying drug-disease interactions and inter-individual variability, our work reveals a need for personalized anti-thrombotic drug selection that accommodates each patient's pathological profile. Arterial thrombosis is a leading cause of death and disability with no clinical bioassay for prediction. Here, the authors develop an assay that monitors the multi-dimensional attributes of thrombi forming in its physiologically mimicking rheological condition to predict arterial thrombotic risks [ABSTRACT FROM AUTHOR]

Introduction: Prostate cancer (PCa) is the second most frequent cancer diagnosis in men and the fifth leading cause of death worldwide. Prostate Specific Membrane Antigen (PSMA) and Gastrin Releasing Peptide (GRP) receptors are overexpressed in PCa. In this study, we have developed iron oxide nanoparticles (IONs) functionalized with the Prostate Specific Membrane Antigen (PSMA) and Gastrin Releasing Peptide (GRP) ligands for dual targeting of Prostate cancer. Methods: IONs were developed with a thin silica layer on their surface with MPTES (carrying -SH groups, IONs-SH), and they were coupled either with a pharmacophore targeting PSMA (IONs-PSMA) or with bombesin peptide (IONs-BN), targeting GRP receptors, or with both (IONs-PSMA/BN). The functionalized IONs were characterized for their size, zeta potential, and efficiency of functionalization using dynamic light scattering (DLS) and Fourier-Transform Infrared Spectroscopy (FT-IR). All the aforementioned types of IONs were radiolabeled directly with Technetium-99m (99mTc) and evaluated for their radiolabeling efficiency, stability, and binding ability on two different PCa cell lines (PC3 and LNCaP). Results and Discussion: The MTT assay demonstrated low toxicity of the IONs against PC3 and LNCaP cells, while the performed wound-healing assay further proved that these nanostructures did not affect cellular growth mechanisms. The observed hemolysis ratio after co-incubation with red blood cells was extremely low. Furthermore, the 99mTc-radiolabeled IONs showed good stability in human serum, DTPA, and histidine, and high specific binding rates in cancer cells, supporting their future utilization as potential diagnostic tools for PCa with Single Photon Emission Computed Tomography (SPECT) imaging. [ABSTRACT FROM AUTHOR]

Image-based models that use features extracted from cell microscopy images can estimate the activity of small molecules in various biological assays. Typically, models are trained on images stained by an optimized protocol (e.g. Cell Painting) after exposure to a fairly high small molecule concentration (referred to as 'image concentration') of 10 μ M or higher. Low concentration images (e.g. 0.16 μM, 0.8 μM, 4 μM) tend to yield models with worse performance. In this work, we nevertheless report a practical use for low image concentration data. We propose the combination of well-performing models trained at higher image concentrations, with lower image concentration for inference to identify more potent compounds. We show that this approach improves on the conventional method (directly training a high-potency model) in 65 % of assays investigated in terms of AUC-ROC, and 75 % of assays in terms of RIPtoP-corrected AUC-PR. [ABSTRACT FROM AUTHOR]

SUMMARY: Presilphiperfolane‐type sesquiterpenes represent a unique group of atypical sesquiterpenoids characterized by their distinctive tricyclic structure. They have significant potential as lead compounds for pharmaceutical and agrochemical development. Herein, we utilized a transcriptomic approach to identify a terpene synthase (TPS) gene responsible for the biosynthesis of rare presilphiperfolane‐type sesquiterpenes in Inula lineariifolia, designated as IlTPS1. Through phylogenetic analysis, we have identified the evolutionary conservation of key motifs, including RR(x)8W, DDxxD, and NSE/DTE in IlTPS1, which are shared with other tricyclic sesquiterpene synthases in the TPS‐a subfamily of Asteraceae plants. Subsequent biochemical characterization of recombinant IlTPS1 revealed it to be a multiproduct enzyme responsible for the synthesis of various tricyclic sesquiterpene alcohols from farnesyl diphosphate (FPP), resulting in production of seven distinct sesquiterpenes. Mass spectrometry and nuclear magnetic resonance (NMR) spectrometry identified presilphiperfolan‐8β‐ol and presilphiperfol‐7‐ene as predominant products. Furthermore, biological activity assays revealed that the products from IlTPS1 exhibited a potent antifungal activity against Nigrospora oryzae. Our study represents a significant advancement as it not only functionally identifies the first step enzyme in presilphiperfolane biosynthesis but also establishes the heterologous bioproduction of these unique sesquiterpenes. Significance Statement: Our research has uncovered the molecular mechanism responsible for the synthesis of rare, bridged tricyclic sesquiterpenes by IlTPS1. Furthermore, we have established their bioproduction and assessed their antimicrobial properties. This study serves as a promising example of how unique sesquiterpene natural products can be harnessed and leveraged in the pharmaceutical and agrochemical sectors for various applications. [ABSTRACT FROM AUTHOR]

In this study, new peripherally substituted symmetric zinc and magnesium phthalocyanines (4 and 5) were successfully prepared by cyclotetramerization of the tetrahydropyrimidone (THPM)-linked phthalonitrile 3. The identity of the compounds were confirmed primarily through spectroscopic analysis including NMR, FT-IR, UV-Vis and MALDI-TOF mass spectroscopy. The photophysical and photochemical properties of the synthesized phthalocyanines (Pcs) were examined using UV–Vis absorption and fluorescence emission spectroscopy techniques. The quantum yields of singlet oxygen were found to be 0.50 and 0.33 for compounds 4 and 5 in DMSO, respectively. In addition to photo-physicochemical properties, the enhanced biological activities of compounds 4 and 5 were investigated using a range of biological assays, namely, antibiofilm, microbial cell viability, antioxidant, DNA cleavage, antimicrobial and photodynamic antimicrobial assays. The maximum DPPH inhibition of 4 and 5 was detected as 40.46% and 25.76% at 100 mg L−1, respectively. Fragmentation of the DNA molecule was observed at concentrations of 25 mg L−1, 50 mg L−1 and 100 mg L−1 for 4 and 5. Additionally, effective inhibition of microbial cell viability was observed with the targeted Pcs. The antibiofilm properties of these compounds were found to be concentration-dependent. The biofilm inhibition activities of 4 and 5 were found to be 96.01% and 92.04% for S. aureus, while they were 95.42% and 91.27%, for P. aeruginosa, respectively. The antimicrobial activities of 4 and 5 on different microorganisms were evaluated using the microdilution assay. In the case of photodynamic antimicrobial treatment, the newly synthesized Pcs showed more effective antimicrobial inhibition compared to the control. These findings suggest that compounds 4 and 5 can be used as promising photodynamic antimicrobial agents for the treatment of many diseases, particularly infectious diseases. [ABSTRACT FROM AUTHOR]

ABSTRACT A thiophosphoric triamide (SPT) ligand P(S)[NH‐2Py]3 was synthesized and utilized as a ligand to make novel binuclear and mononuclear Co2+ and Cd2+ complexes. The compounds are the first examples of discrete chelate SPT complexes with a [N]3P(S)‐based backbone in which the SPT acts as a flexible tridentate N,N,N‐donor or N,N,O‐donor ligand. The metal cation features a hexacoordinate environment M(N)3(Cl)3/M(N)4(O)2 adopting a distorted octahedral geometry. Hirshfeld surface analysis affirms that the pyridinyl nitrogen is a more dominant H‐bond acceptor than the thiophosphoryl sulfur atom. The inhibitory activity against the MDA‐MB‐231 cancer cell line in vitro indicated a good inhibitory effect for all compounds. The mononuclear Co2+ complex showed a stronger inhibitory activity against cancer cells than other studied compounds with an IC50 of 62 ± 1.4 nM. Antioxidant and antihemolytic activity of the compounds showed appreciable performance with the highest activities for the Cd2+ complex. A computational DNA binding study by molecular docking indicated a good affinity of the compounds with the DNA receptor by a mixed mode of interactions. Docking was also performed on SARS‐CoV‐2 and monkeypox (Mpox), resulting in binding energies (−5.5 to −8.9 kcal/mol) that are competitive with current effective drugs. [ABSTRACT FROM AUTHOR]

The escalating incidence of hospital infections due to antibiotic resistance necessitates the identification of alternative therapeutic agents such as probiotics. This study was designed to isolate and evaluate the efficacy of probiotics against Staphylococcus saprophyticus, a prevalent etiological agent of urinary tract infections (UTIs). A total of 100 S. saprophyticus strains were isolated from clinical samples and subjected to antibiotic susceptibility testing via the disc diffusion method. Concurrently, probiotic bacteria were isolated from Bulgarian cheese and shallot yogurt, and their antibacterial activity against S. saprophyticus strains was assessed. The inhibitory potential of probiotic supernatants was evaluated using microtiter plate assays, with the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) determined at a 1/2 dilution. Cytotoxicity was evaluated using the MTT assay, and high‐performance liquid chromatography (HPLC) was employed to analyze the concentrations of organic acids produced by the probiotics. The results revealed that all S. saprophyticus strains were resistant to tetracycline and doxycycline but susceptible to other antibiotics. Lactobacillus rhamnosus strains M and B demonstrated notable antibacterial and antibiofilm activity against S. saprophyticus isolates. These probiotics exhibited susceptibility to most antibiotics and lacked virulence factors, suggesting their safety for therapeutic use. The organic acids produced by the probiotics were identified as lactic acid, acetic acid, and formic acid. In conclusion, L. rhamnosus strains M and B exhibit potent antimicrobial properties against S. saprophyticus, indicating their potential as therapeutic agents for UTIs. Further research is warranted to validate these findings and explore the possibility of these probiotics in clinical applications. [ABSTRACT FROM AUTHOR]

ABSTRACT Aromia bungii is a serious wood borer damaging Prunus persica and other numerous economic tree species. Host plant volatiles play a key role in pest population monitoring and trapping. To ascertain the influence of volatile organic compounds emitted by P. persica at varying physiological statuses on the host‐searching behaviour of A. bungii adults, we initially collected and identified the volatiles emitted by P. persica leaves infested with A. bungii using gas chromatography–mass spectrometry. Subsequently, we evaluated the behavioural response of A. bungii adults to the crude extracts, single compounds and blends of these compounds by Y‐tube olfactometer bioassay. Results showed that volatiles emitted from P. persica infested with A. bungii larvae at Grade I were more attractive to females, but males were more attracted to volatiles from P. persica infested with A. bungii larvae at Grade IV. Five compounds were identified and showed relatively high levels from both infested plants at Grade I and Grade IV. Among them, 3‐carene, cis‐3‐hexen‐1‐ol and cis‐3‐hexenyl acetate were attractive to females, and a blend of the three volatiles in a ratio of 3:91:6 for Grade I infestation was more attractive to females than a ratio of 3:83:14 from Grade IV infestation. The compounds, 3‐carene, cis‐3‐hexenyl acetate, cis‐3‐hexene‐1‐ol and benzaldehyde, were attractive to males and a blend in a ratio of 3:81:13:3 from Grade IV infestation was more attractive to males than a ratio of 3:89:6:2 from Grade I infestation. The results show that the kinds and ratios of volatiles emitted from P. persica infested with A. bungii larvae play important roles in host‐searching behaviour of A. bungii adults. [ABSTRACT FROM AUTHOR]

Background: Neuroinflammation is widely recognized as a key factor in the pathogenesis of Alzheimer's disease (AD), alongside ß-amyloid deposition and the formation of neurofibrillary tangles. The NLR family pyrin domain containing 3 (NLRP3) inflammasome, part of the innate immune system, has been implicated in the neuropathology of both preclinical amyloid and tau transgenic models. Activation of the NLRP3 pathway involves an initial priming step, which increases the expression of Nlrp3 and interleukin (IL)-1ß, followed by the assembly of the NLRP3 inflammasome complex, comprising NLRP3, ASC, and caspase-1. This assembly leads to the proteolytic maturation of the pro-inflammatory cytokines IL-1ß and IL-18. Additionally, the NLRP3 inflammasome induces Gasdermin D (GSDMD) cleavage, forming membrane pores through which IL-1ß and IL-18 are secreted. Inhibition of NLRP3 has been shown to enhance plaque clearance by modulating microglial activation. Furthermore, blocking NLRP3 in tau transgenic mice has been found to reduce tau phosphorylation by affecting the activity of certain tau kinases and phosphatases. Methods: In this study, organotypic brain slice cultures from P301S transgenic mice were treated with lipopolysaccharide (LPS) plus nigericin as a positive control or exposed to tau seeds (K18) to evaluate NLRP3 inflammasome activation. The effect of tau seeding on NLRP3 activity was further examined using Meso Scale Discovery (MSD) assays to measure IL1ß secretion levels in the presence and absence of NLRP3 inhibitors. The role of NLRP3 activity was investigated in fullbody Nlrp3 knockout mice crossbred with the tau transgenic P301S model. Additionally, full-body and microglia-selective Gsdmd knockout mice were crossbred with P301S mice, and tau pathology and neurodegeneration were evaluated at early and late stages of the disease using immunohistochemistry and biochemical assays. Results: Activation of the NLRP3 pathway was observed in the mouse organotypic slice culture (OSC) model following stimulation with LPS and nigericin or exposure to tau seeds. However, Nlrp3 deficiency did not mitigate tauopathy or neurodegeneration in P301S mice in vivo, showing only a minor effect on plasma neurofilament (NF-L) levels. Consistently, Gsdmd deficiency did not alter tau pathology in P301S mice. Furthermore, neither full-body nor microglia-selective Gsdmd deletion had an impact on neuronal pathology or the release of pro-inflammatory cytokines. Conclusion: The absence of key components of the NLRP3 inflammasome pathway did not yield a beneficial effect on tau pathology or neurodegeneration in the preclinical Tau-P301S mouse model of AD. Nonetheless, organotypic slice cultures could serve as a valuable ex vivo mechanistic model for evaluating NLRP3 pathway activation and pharmacological inhibitors. [ABSTRACT FROM AUTHOR]

Background: Many diseases are increasingly recognized as public health concerns worldwide because of the increasing incidence of multidrug-resistant bacteria. Recently, interest in the use of indigenous medicinal plants to treat infectious illnesses has increased, highlighting the need to find new bioactive phytochemicals. Ajuga integrifolia is a plant commonly utilized in traditional drugs to treat a wide range of diseases, although its effectiveness has not been scientifically validated. The present study aimed to evaluate the total phenolic and flavonoid contents and assess the biological activities of A. integrifolia leaf extracts produced via different solvent systems. Methods: Soxhlet extraction was employed to obtain crude extracts from different solvents (methanol, ethanol and water). The 2,2-diphenyl-1-picrylhydrazyl (DPPH) and ferric-reducing power assays were used to measure the antioxidant activity, and the antibacterial activity of the extract was evaluated on the basis of its minimum inhibitory concentration (MIC) against two gram-negative bacteria (Escherichia coli (ATCC-25922) and Pseudomonas aeruginosa (ATCC-43495)) and two gram-positive bacteria (Staphylococcus aureus (ATCC-25923) and Streptococcus pyogenes (ATCC-19615)) via the agar disk-diffusion technique. Results: A significant amount of total phenolic content (TPC) and flavonoid content (TFC) were present in all the extracts. The extracts presented powerful antioxidant activity in all the assays. The disc diffusion and MIC results revealed the ability of the methanol and ethanol extracts of A. integrifolia leaves to inhibit S. aureus growth at a concentration of 3.125 mg/mL. However, the water extracts were ineffective against E. coli and P. aeruginosa. Conclusions: These findings indicate that A. integrifolia leaf extracts have reasonable biological activities. These findings underscore the importance of A. integrifolia leaves as a source of health benefits. [ABSTRACT FROM AUTHOR]

Purpose: Ketogenic diets are proposed as a therapeutic approach for type 1 and type 2 diabetes due to their low glucose intake. However, their potential effects on the immune system need investigation. This study aims to explore how glucose concentration and beta-hydroxybutyrate (BHB) impact T cell phenotype, metabolism, and function, with a focus on systemic inflammatory response (T2D) and autoimmunity (T1D). Methods: T cells from healthy donors were cultured in vitro under varying glucose concentrations with or without BHB. Flow cytometry was employed to analyze changes in T cell phenotype, while proliferation was evaluated through a CFSE dilution assay. Additionally, we used a novel flow cytometry method allowing a direct assessment of T cell metabolism. Results: Culturing T cells in low glucose concentrations revealed their dependency on glucose metabolism, leading to reduced proliferation rates, overexpression of exhaustion markers and increased susceptibility to Treg suppression and the influence of immune-modulating drugs such as rapamycin, FK506, and MMF. Notably, T cells cultured in low glucose concentrations increased the expression of BDH1 to utilize BHB as an alternative fuel source. Finally, the addition of BHB to the culture effectively rescued T cell impairments caused by insufficient glucose levels. Conclusions: T cells display limited capacity to adapt to low glucose levels, resulting in profound functional impairment. However, T cell functions can be efficiently recovered by the presence of 2mM BHB. [ABSTRACT FROM AUTHOR]

Behavior and fitness are important ecological traits frequently measured in insect bioassays. A common method to measure them in soft‐bodied herbivorous insects involves confining individuals to plant leaves using clip cages. Although studies have previously highlighted the negative effects of clip cages on leaf physiology, little is known about the impact that using this confinement method has on insect fitness. The responses of different aphid genotypes/clones to different containment methods have not previously been investigated. Here we measured key fitness traits (intrinsic rate of natural increase, mean relative growth rate, time to reach reproductive adulthood and population doubling time) in the potato aphid, Macrosiphum euphorbiae Thomas (Hemiptera: Aphididae), when confined to plants using two methods: (1) clip cages to confine aphids to individual strawberry leaves and (2) a mesh bag to confine aphids to whole strawberry plants. Our study identified a strong negative impact on all the measured aphid fitness traits when using clip cages instead of mesh bags. We also identified genotype‐specific differences in response to confinement method, where clip cage confinement differentially affected the fitness of a given aphid genotype compared to the same genotype on whole plants. These results suggest that clip cage use should be carefully considered when experiments seek to quantify insect fitness and that whole plants should be used wherever possible. Given the prevalence of clip cage use in insect bioassays, our results highlight the need for caution when interpreting the existing literature as confinement method significantly impacts aphid fitness depending on their genotype. [ABSTRACT FROM AUTHOR]

Simple Summary: Elevated levels of the mitotic checkpoint kinase BUB1 are frequently observed in most solid cancers, including lung cancer. High BUB1 expression is linked to poorer patient survival. In the present study, we aimed to evaluate if a BUB1 targeting agent could enhance the effectiveness of chemotherapy and chemoradiation in lung cancer. Our findings suggest that BUB1 inhibitor sensitizes lung cancer cell lines to chemotherapy, radiotherapy, and chemoradiation and thus validate BUB1 as a novel molecular target. Background: Despite advances in targeted therapies and immunotherapies, traditional treatments like microtubule stabilizers (paclitaxel, docetaxel), DNA-intercalating platinum drugs (cisplatin), and radiation therapy remain essential for managing locally advanced and metastatic lung cancer. Identifying novel molecular targets could enhance the efficacy of these treatments. Hypothesis: We hypothesize that BUB1 (Ser/Thr kinase) is overexpressed in lung cancers and its inhibition will sensitize lung cancers to chemoradiation. Methods: BUB1 inhibitor (BAY1816032) was combined with cisplatin, paclitaxel, a PARP inhibitor olaparib, and radiation in cell proliferation and radiation-sensitization assays. Biochemical and molecular assays evaluated the impact on DNA damage signaling and cell death. Results: Immunostaining of lung tumor microarrays (TMAs) confirmed higher BUB1 expression in non-small cell lung cancer (NSCLC) and small cell lung cancer (SCLC) compared to normal tissues. In NSCLC, BUB1 overexpression correlated directly with the expression of TP53 mutations and poorer overall survival in NSCLC and SCLC patients. BAY1816032 synergistically sensitized lung cancer cell lines to paclitaxel and olaparib and enhanced cell killing by radiation in both NSCLC and SCLC. Molecular analysis indicated a shift towards pro-apoptotic and anti-proliferative states, evidenced by altered BAX, BCL2, PCNA, and Caspases-9 and -3 expressions. Conclusions: Elevated BUB1 expression is associated with poorer survival in lung cancer. Inhibiting BUB1 sensitizes NSCLC and SCLC to chemotherapies (cisplatin, paclitaxel), targeted therapy (olaparib), and radiation. Furthermore, we present the novel finding that BUB1 inhibition sensitized both NSCLC and SCLC to radiotherapy and chemoradiation. Our results demonstrate BUB1 inhibition as a promising strategy to sensitize lung cancers to radiation and chemoradiation therapies. [ABSTRACT FROM AUTHOR]

Simple Summary: Our research integrates two key areas of focus. We first focus on a comparative biostatistical analysis of the agreement between three laboratory methods for genotyping gene copy number variations, and then we explore new characteristics of the distribution of this type of genetic aberration among cancer-associated genes. The results could be used to help molecular oncologists pick the smallest set of methods that will allow for accurate CNV genotyping in personalized cancer prognosis and drug-response prediction. We see the following strengths of our research: (1) We employ various methods for the detection of CNVs, such as CoreExome microarrays, nCounter v2 Cancer CN Assay panel, and digital droplet PCR; (2) This study sets out an interpretable comprehensive biostatistical analysis of the agreement using the PABAK score and Passing–Bablok regression; (3) This study presents the detection of CNVs in 12 paired tumor ovarian cancer/normal samples with clinical data; and (4) The research has translational potential. Objective: The goal of this study was to compare the results of CNV detection by three different methods using 13 paired carcinoma samples, as well as to perform a statistical analysis of the agreement. Methods: CNV was studied using NanoString nCounter v2 Cancer CN Assay (Nanostring), Illumina Infinium CoreExome microarrays (CoreExome microarrays) and digital droplet PCR (ddPCR). Results: There was a good level of agreement (PABAK score > 0.6) between the CoreExome microarrays and the ddPCR results for finding CNVs. There was a moderate level of agreement (PABAK values ≈ 0.3–0.6) between the NanoString Assay results and microarrays or ddPCR. For 83 out of 87 target genes studied (95%), the agreement between the CoreExome microarrays and NanoString nCounter was characterized by PABAK values < 0.75, except for MAGI3, PDGFRA, NKX2-1 and KDR genes (>0.75). The MET, HMGA2, KDR, C8orf4, PAX9, CDK6, and CCND2 genes had the highest agreement among all three approaches. Conclusions: Therefore, to get a better idea of how to genotype an unknown CNV spectrum in tumor or normal tissue samples that are very different molecularly, it makes sense to use at least two CNV detection methods. One of them, like ddPCR, should be able to quantitatively confirm the results of the other. [ABSTRACT FROM AUTHOR]

Chlorpromazine (CPZ) is a first-generation neuroleptic with well-established antitumor and antiviral properties. Currently, numerous studies are focused on developing new methods for CPZ delivery; however, the knowledge regarding its conjugates with metal nanoparticles remains limited. The aim of this study was to prepare CPZ conjugates with gold nanoparticles (AuNPs) and evaluate their biological activity on human lymphocytes (HUT-78 and COLO 720L), as well as human (COLO 679) and murine (B16-F0) melanoma cells, in comparison to the effects induced by unconjugated CPZ molecules and AuNPs with well-defined properties. During the treatment of cells with CPZ, AuNPs, and CPZ-AuNP conjugates, changes in mitochondrial activity, membrane integrity, and the secretion of lipid peroxidation mediators were studied using standard biological assays such as MTT, LDH, and MDA assays. It was found that positively charged CPZ-AuNP conjugates more effectively reduced cell viability compared to AuNPs alone. The dose-dependent membrane damage was correlated with oxidative stress resulting from exposure to CPZ-AuNP conjugates. The activity of the conjugates depended on their composition and the size of the AuNPs. It was concluded that conjugating CPZ to AuNPs reduced its biological activity, while the cellular response to the treatment varied depending on the specific cell type. [ABSTRACT FROM AUTHOR]

The Centers for Disease Control and Prevention (CDC) Radiation Laboratory's primary mission is to provide laboratory support for an effective and efficient response to public health radiological emergencies. The laboratory has developed methods for several radiological threat agents, including Iridium-192 (Ir-192). Ir-192 can be analyzed via its gamma energy through analytical methods such as High Purity Germanium (HPGe) and its beta energy through Liquid Scintillation Counting (LSC). In this work, we present and compare HPGe and LSC rapid response methods for Ir-192 quantification. Both methods show the reasonable results and can be used in emergency situations. [ABSTRACT FROM AUTHOR]

Objective: The biochip method allows microscale multiparametric analysis of macromolecular samples using a matrix of immobilized molecular probes. Selection of materials for biochip fabrication, functionalization of the carrier surface, and the method of immobilization of molecular probes are the key tasks of biochip technology. Methods: Methods of obtaining polymer coating from polyvinyl acetate on the surface of polyethylene terephthalate polymer substrates and subsequent production of brush polymers by photoinduced radical copolymerization of acrylate monomers have been studied. Cell matrices with numerous reactive chemical groups for subsequent immobilization of proteins were formed by photolithography method. Methods of activation of carboxyl groups on brush polymers attached to the surface of polyethylene terephthalate were tested. Immobilization of model protein streptavidin labeled with fluorescent dye Su3 was performed to test the method of activation of carboxyl groups. Results and Discussion: A variant of the immunofluorescence assay in a biological microarray format was tested on the model "streptavidin–biotinylated immunoglobulin." Conclusions: Streptavidin immobilized in brush polymer cells retains functionality and spatial accessibility for binding to biotinylated immunoglobulin and subsequent manifestation by antibodies fluorescently labeled with Cy5 dye, which opens prospects for the use of biological microarrays with brush polymer cells on polyethylene terephthalate substrates for immunofluorescence analysis of various protein targets. [ABSTRACT FROM AUTHOR]

Regarding economic significance, the Dioscoreaceae family, which includes the well-known Dioscorea (yam) species, is the fourth most important tuber crop globally. A basic meal found all across the world, yams encourage and ensure the food security of households. Although they are challenging to preserve and store however a post-harvest deterioration control must be developed. Over the years, Nigerian farmers have employed medicinal herbs to manage post-harvest pest invasion; nevertheless, the scientific basis for this fungal management has not yet been well documented. As a result, Landolphia owarensis leaf extract was used as a biological control, and fungi were isolated from Dioscorea cayenensis, Dioscorea alata, and Discorea rotundata. Samples of yam tubers – both rotten and healthy – were procured in the markets and brought to the lab for fungus isolation and identification. Leaves of Landolphia oweriensis were collected, air-dried, milled, and macerated in pure methanol for 72 h, extracted and concentrated with a rotary evaporator. The agar-well diffusion method was used for the biological control assay against the fungal isolates. The biological control results were presented as mean ± SD (standard deviation) while analysis of variance (Anova) was used to deduce least significant difference (LSD). A total number of 605 fungi isolates were observed in which D. cayenensis had the highest, 50.58 %, D. alata and Discorea rotundata 24.71 % respectively. Aspergillus niger and Aspergillus fumigatus were found in the three species while Aspergillus flavus, Rhizopus stolonifer, Collentotricum spp. and Penicillium spp. in either of the species. L. owarensis methanolic leaves extracts possesses biological potentials against Colletotrium spp., A. fumigatus, A. niger and A. flavus only. The problem of postharvest fungal invasion of Dioscorea species can be solved with the inhibition observed with L. owarensis methanolic leaves extracts as a biological control measure thereby sustaining food security in the world. [ABSTRACT FROM AUTHOR]

Osteogenic differentiation of human bone marrow-derived mesenchymal stem cells (hBMSCs) is important for human bone formation. Long non-coding RNAs (lncRNAs) are critical regulators in osteogenic differentiation. This study aimed to explore the function and mechanisms of long intergenic non-protein coding RNA 963 (LINC00963) in affecting osteogenesis. Cell differentiation was assessed by alkaline phosphatase (ALP) activity detection and ALP staining assay. Meanwhile, levels of osteogenic marker genes, including RUNX family transcription factor 2 (RUNX2), osteocalcin (OCN), and osteopontin (OPN), were detected by RT-qPCR and western blot. Cell proliferation and apoptosis were measured using CCK-8 assay and flow cytometry analysis. RNA immunoprecipitation (RIP), RNA pull-down and luciferase reporter assays were used to investigate the interaction between genes. LINC00963 expression was down-regulated in hBMSCs treated with osteogenic induction. LINC00963 overexpression inhibited hBMSCs differentiation, proliferation, and elevated apoptosis. LINC00963 acted as a competing endogenous RNA (ceRNA) to interact with miR-10b-5p and thereby regulated the expression level of Ras-related protein Rap-2a (RAP2A). LINC00963 regulated RAP2A to inhibit the level of phosphorylated AKT (p-AKT). LINC00963 inhibited hBMSCs differentiation, proliferation, and elevated apoptosis via the miR-10b-5p/RAP2A/AKT signaling, which might help improve the treatment of osteoporosis. [ABSTRACT FROM AUTHOR]

The COVID-19 pandemic has prompted the medical systems of many countries to develop effective treatments to combat the high rate of infection and death caused by the disease. Within the array of proteins found in SARS-CoV-2, the 3 chymotrypsin-like protease (3CLpro) holds significance as it plays a crucial role in cleaving polyprotein peptides into distinct functional nonstructural proteins. Meanwhile, RNA-dependent RNA polymerase (RdRp) takes center stage as the key enzyme tasked with replicating the viral genomic RNA within host cells. These proteins, 3CLpro and RdRp, are deemed optimal subjects for QSAR modeling due to their pivotal functions in the viral lifecycle. In this study, SMILES-based QSAR classification models were developed for a dataset of 2377 compounds that were defined as either active or inactive against 3CLpro and RdRp. Pharmacophore (PH4) and QSAR modeling were used for the virtual screening on 60.2 million compounds including ZINC, ChEMBL, Molport, and MCULE databases to identify new potent inhibitors against 3CLpro and RdRp. Then, a filter was established based on typical molecular characteristics to identify drug-like molecules. The molecular docking was also performed to evaluate the binding affinity of 156 AND 51 potential inhibitors to 3CLpro and RdRp, respectively. Among the 15 hits identified based on molecular docking scores, M3, N2, and N4 were identified as promising inhibitors due to their good synthetic accessibility scores (3.07, 3.11, and 3.29 out of 10 for M3, N2, and N4 respectively). These compounds contain amine functional groups, which are known for their crucial role in the binding interactions between drugs and their targets. Consequently, these hits have been chosen for further biological assay studies to validate their activity. They may represent novel 3CLpro and RdRp inhibitors possessing drug-like properties suitable for COVID-19 therapy. [ABSTRACT FROM AUTHOR]

Objectives This study looked at the role of anti-carbamylated protein (anti-CarP) antibodies in contributing to lung fibrosis in CTD-associated interstitial lung disease (ILD) in an autoantigen-dependent manner. Methods ELISA was used to test serum samples, including 89 from the CTD-ILD group and 170 from the non-CTD-ILD group, for anti-CarP levels. Male C57BL/6 mice were used for the pulmonary fibrosis model and anti-CarP treatment in vivo (n  = 5) and patient serum-derived or commercialized anti-CarP was used for cell treatment. We identified the carbamylated membrane protein via immunofluorescence (IF) and co-immunoprecipitation followed by mass spectrometry (MS) analysis. Quantitative RT-PCR, IF and western blot were performed to explore the antigen-dependent role of anti-CarP. A native electrophoretic mobility shift assay and MS analysis were used to verify direct interaction and carbamylation sites. Results A significantly higher serum anti-CarP level was observed in CTD with ILD than without ILD. In vivo , intrapulmonary delivery of anti-CarP induces epithelial–mesenchymal transition (EMT) and microfibrotic foci. Carbamylation was enriched in type II alveolar epithelial cells (AEC II). A novel carbamylated membrane receptor, specifically recognized by anti-CarP, was identified as toll-like receptor 5 (TLR5). We found anti-CarP induces the nuclear translocation of NF-κB and downstream events, including EMT and expression of inflammatory cytokines in AEC II, which were reversed by TLR5 blocking or TLR5 knockdown. Moreover, up to 12 lysine carbamylation sites were found in TLR5 ectodomain, allowing the interaction of anti-CarP with carbamylated TLR5. Conclusions Overall, we found anti-CarP drives aberrant AEC II activation by interacting with carbamylated TLR5 to promote ILD progression. [ABSTRACT FROM AUTHOR]

Context.--Sinonasal tract malignancies are rare cancers with frequent morphologic overlap. Given the similar histologic profiles seen in many of these entities, they often present a diagnostic challenge to the practicing pathologist. Objective.--To provide a streamlined algorithm using histologic clues, immunohistochemical profiles, and molecular assays to aid in diagnosis of these lesions. Data Sources.--Sources were the World Health Organization Tumor Classification, literature review, and institutional experience. Conclusions.--Although many sinonasal tract malignancies show similar histology, distinct immunohistochemical and molecular profiles can help parse out differences, thereby facilitating diagnosis for the pathologist. [ABSTRACT FROM AUTHOR]

The methanolic extract of Indigofera dosua leaves was subjected to GC-MS analysis, its antioxidant and antimicrobial potential. Sixteen compounds were identified having resorcinol TMS derivatives, propanoic acid, dimethyl(ethenyl)silyl ester, 4-O-methylmannose, hexadecanoic acid, trimethylsilyl ester, 9,12,15-octadecatriene-1-ol, and phytol as major components. FRAP content was 40.24 ± 0.54AAE/g, and the IC50 values were 8.25 ± 0.04 µg mL−1 and 19.05 ± 0.15 µg mL−1 when employing DPPH and ABTS, respectively. Total phenolics (50.2 ± 0.48 mg GAE/g) and total flavonoid content (5.33 ± 0.40 mg QE/g) were estimated. Antibacterial activity was observed, showing significant inhibition against Pseudomonas aeruginosa and Proteus mirabilis with MIC values 12.5 and 25 mg mL−1, respectively. [ABSTRACT FROM AUTHOR]

Introduction: Propylene glycol (PG) is frequently used as a solvent for various medications. However, there is substantial evidence of Propylene glycol toxicity, such as depression, agitation, and seizures, particularly when used in combination with other drugs. Here, we aimed to study the effect of Propylene glycol administration in combination with amyloid β1-40 injection on hippocampal neurons. Materials & Methods: Thirty-six male Wistar rats were randomly divided into four groups: sham, amyloid β1-40 injection group, Propylene glycol group, and amyloid β1-40+ Propylene glycol group. Alternation behavior, number of neurons in the hippocampus, lipid peroxidation markers, and superoxide dismutase levels were analyzed in all rats. Results: When Propylene glycol was co-administered with amyloid β1-40, a notable reduction in the mean neuronal count was observed in the CA1, CA3, and DG regions compared with the amyloid β1-40 only injected animals (P < 0.05). Furthermore, Propylene glycol induced an increase in lipid peroxidation markers (10.78±0.4) and a decrease in antioxidant content (2.8 ± 0.17) when administered with amyloid β1-40s compared to the animals that received only amyloid β1-40 (P<0.05). A similar pattern was found in alternation behavior compared with the group with amyloid β1-40 injection (P<0.001). Conclusion: Propylene glycol could produce excessive neurotoxicity in regions of the hippocampus when co-administered with amyloid β1-40. It likely increases lipid peroxidation and reduces superoxide dismutase in the rat brain. The use of different agents as a vehicle should be considered, especially in the elderly. [ABSTRACT FROM AUTHOR]

Background: Galatella is a genus in the family Asteraceae, represented by 35-45 species. Considering the high effectiveness of the ethyl acetate (EtOAc) fraction of G. grimmii against Mycobacterium tuberculosis (MIC = 0.5 µg/mL), a bioassay-directed fractionation of this extract was carried out. Methods: The methanolic extract of the aerial parts of G. grimmii was obtained using maceration, then it was suspended in water and partitioned with petroleum ether, dichloromethane (CH2Cl2), EtOAc, and n-butanol (n-BuOH), successively. The most potent fraction (EtOAc), was selected for further isolation by Sephadex LH–20 and semi-preparative HPLC to obtain active compounds. Results: Fractionation of the EtOAc solvent fraction resulted in the characterization of five compounds, among them, compounds 1 and 2 showed the highest anti-mycobacterial effects with MICs of 0.062 and 1.00 µg/mL against H37Rv M. tuberculosis, respectively, which were higher than those of rifampin (MIC of 1.25 µg/mL) and isoniazid (MIC of 0.31 µg/mL), as positive controls. Also, compound 1 ​​inhibited all tested strains of drug-resistant Mycobacterium (MDR and XDR). Notably, the isolated compounds have been reported for the first time from G. grimmii. Conclusion: Due to the potent anti-mycobacterial effect of isolated compounds from G. grimmii, this study could pave the way for developing a novel class of natural anti-tuberculosis compounds. [ABSTRACT FROM AUTHOR]

Background: Decellularized extracellular matrix (dECM) has been proposed as a useful source of biomimetic materials for regenerative medicine due to its biological properties that regulate cell behaviors. The present study aimed to investigate the influence of decellularized ECM derived from dental pulp stem cells (DPSCs) on gingival fibroblast (GF) cell behaviors. Cells were isolated from dental pulp and gingival tissues. ECM was derived from culturing dental pulp stem cells in growth medium supplemented with ascorbic acid. A bioinformatic database of the extracellular matrix was constructed using Metascape. GFs were reseeded onto dECM, and their adhesion, spreading, and organization were subsequently observed. The migration ability of the cells was determined using a scratch assay. Protein expression was evaluated using immunofluorescence staining. Results: Type 1 collagen and fibronectin were detected on the ECM and dECM derived from DPSCs. Negative phalloidin and nuclei were noted in the dECM. The proteomic database revealed enrichment of several proteins involved in ECM organization, ECM–receptor interaction, and focal adhesion. Compared with those on the controls, the GFs on the dECM exhibited more organized stress fibers. Furthermore, cultured GFs on dECM exhibited significantly enhanced migration and proliferation abilities. Interestingly, GFs seeded on dECM showed upregulation of FN1, ITGB3, and CTNNB1 mRNA levels. Conclusions: ECM derived from DSPCs generates a crucial microenvironment for regulating GF adhesion, migration and proliferation. Therefore, decellularized ECM from DPSCs could serve as a matrix for oral tissue repair. [ABSTRACT FROM AUTHOR]

Circumventing the limitations of current bioassays, we introduce a light-controlled assay, OptoAssay, toward wash-and pump-free point-of-care diagnostics. Extending the capabilities of standard bioassays with light-dependent and reversible interaction of optogenetic switches, OptoAssays enable a bidirectional movement of assay components, only by changing the wavelength of light. Demonstrating exceptional versatility, the OptoAssay showcases its efficacy on various substrates, delivering a dynamic bioassay format. The applicability of the OptoAssay is successfully demonstrated by the calibration of a competitive model assay, resulting in a superior limit of detection of 8 pg ml-1, which is beyond those of conventional ELISA tests. In the future, combined with smartphones, OptoAssays could obviate the need for external flow control systems such as pumps or valves and signal readout devices, enabling on-site analysis in resource-limited settings. [ABSTRACT FROM AUTHOR]

Backgrounds: Various physiological functions and reaction cascades, as well as disease progression in the living systems, are controlled by the activity of specific proteolytic enzymes. We conducted the study to evaluate protease activity by assessing peptide fragments from either conserved or labeled red blood cells (RBCs) with aminofluorescein (AF) in the reaction media. Methods: RBCs were incubated in media containing trypsin. Subsequently, the concentration of peptide fragments in the reaction media, resulted by the digestion with trypsin from conserved cells, was estimated by 3‐(4‐carboxybenzoyl)quinoline‐2‐carboxaldehyde (CBQCA) as an amine‐reactive fluorogenic reagent. In a second approach, we conjugated AF to the conserved RBCs and then exposed AF‐labeled RBCs to trypsin. This was followed by directly measuring the fluorescence intensity (FI) in the reaction media to estimate the concentration of AF‐labeled peptide fragments resulting from the enzyme's activity. Results: Show a concentration‐ and time‐dependent increase in FIs, reflecting the activity of trypsin as a proteolytic enzyme. The FIs increased significantly by 4 to 5 folds in samples treated with different enzyme concentrations, and by over 11 folds after 2 h incubation in media containing a 50 μL trypsin, as evidenced by CBQCA assays. Conclusion: These fast and affordable approaches could be applied with high reliability for the general estimation of protease activity in samples and customized for diagnostic purposes and prognostic evaluation in various diseases. [ABSTRACT FROM AUTHOR]

Thermal proteome profiling (TPP) is a proteome wide technology that enables unbiased detection of protein drug interactions as well as changes in post-translational state of proteins between different biological conditions. Statistical analysis of temperature range TPP (TPP-TR) datasets relies on comparing protein melting curves, describing the amount of non-denatured proteins as a function of temperature, between different conditions (e.g. presence or absence of a drug). However, state-of-the-art models are restricted to sigmoidal melting behaviours while unconventional melting curves, representing up to 50% of TPP-TR datasets, have recently been shown to carry important biological information. We present a novel statistical framework, based on hierarchical Gaussian process models and named GPMelt, to make TPP-TR datasets analysis unbiased with respect to the melting profiles of proteins. GPMelt scales to multiple conditions, and extension of the model to deeper hierarchies (i.e. with additional sub-levels) allows to deal with complex TPP-TR protocols. Collectively, our statistical framework extends the analysis of TPP-TR datasets for both protein and peptide level melting curves, offering access to thousands of previously excluded melting curves and thus substantially increasing the coverage and the ability of TPP to uncover new biology. Author summary: Proteins interactions with other proteins, nucleic acids or metabolites, are key to all biological processes. Being able to detect these interactions is essential to understand biological systems. Thermal proteome profiling is a proteome-wide biological assay able to capture these interactions. It consists in analysing the effect of heat treatment on proteins. Indeed, proteins, under physiological conditions, are folded. This folding gets disrupted as the temperature increases. The way this unfolding happens, called the melting profile of the protein, informs on the interactions of proteins. For example, the interaction of a protein with another protein can increase (thermally stabilise) or decrease (thermally destabilise) the temperature at which this protein starts unfolding. In this work, we present a new statistical method, named GPMelt, to analyse these melting profiles. Notably, GPMelt allows to analyse any melting profiles, independently of their shapes. The proposed improvements over previously published methods allow to investigate more robustly the melting profiles of more proteins, hence increasing the ability of thermal proteome profiling assays to discover new protein interactions. We anticipate that these advancements will aid in unravelling complex biological phenomena. [ABSTRACT FROM AUTHOR]

Traditional antibiotics have specific intracellular targets and disinfect in chemical ways, and multi-drug resistance induced by antibiotics has become an emerging threat to human health. Therefore, it is urgent to develop novel antibacterial agents to combat multi-drug resistance. Herein, we report a new antibacterial nanocomposite (PGH@AgNPs), which was prepared by decorating polymeric guanidine hydrochloride with silver nanoparticles (AgNPs). The Ce(IV)/pentaerythritol redox system was used to initiate the polymerization of a vinyl monomer, guanidine hydrochloride (GH), to prepare star-shaped polymerized GH (PGH), which was then decorated with silver nanoparticles (AgNPs). The physicochemical properties of the nanocomposite were characterized using UV, SEM, XPS and XRD. The biological assays revealed that the nanocomposite possessed effective broad-spectrum antibacterial properties, showing the minimum inhibitory concentrations (MICs) ranging from 6 to 12 µg mL−1, a sterilization rate of higher than 90%, and MIC values of 8, 12 and 12 µg mL−1 against the clinically isolated drug-resistant bacteria MRSA (TACC43300), VRE and P. aeruginosa, respectively. Particularly, S. aureus and P. aeruginosa treated with PGH@AgNPs did not develop drug resistance even after 21 passages because PGH@AgNPs could irreversibly damage the bacterial membrane structure and induce the generation of reactive oxygen species (ROS), ultimately killing the bacteria. In addition, PGH@AgNPs exhibited high blood biocompatibility in mouse fibroblast cells (L929). Overall, PGH@AgNPs has great potential to be used as an antibacterial material with high cytocompatibility for the control of multi-drug-resistant bacterial infections. [ABSTRACT FROM AUTHOR]

Early‐stage disease detection, particularly in Point‐Of‐Care (POC) wearable formats, assumes pivotal role in advancing healthcare services and precision‐medicine. Public benefits of early detection extend beyond cost‐effectively promoting healthcare outcomes, to also include reducing the risk of comorbid diseases. Technological advancements enabling POC biomarker recognition empower discovery of new markers for various health conditions. Integration of POC wearables for biomarker detection with intelligent frameworks represents ground‐breaking innovations enabling automation of operations, conducting advanced large‐scale data analysis, generating predictive models, and facilitating remote and guided clinical decision‐making. These advancements substantially alleviate socioeconomic burdens, creating a paradigm shift in diagnostics, and revolutionizing medical assessments and technology development. This review explores critical topics and recent progress in development of 1) POC systems and wearable solutions for early disease detection and physiological monitoring, as well as 2) discussing current trends in adoption of smart technologies within clinical settings and in developing biological assays, and ultimately 3) exploring utilities of POC systems and smart platforms for biomarker discovery. Additionally, the review explores technology translation from research labs to broader applications. It also addresses associated risks, biases, and challenges of widespread Artificial Intelligence (AI) integration in diagnostics systems, while systematically outlining potential prospects, current challenges, and opportunities. [ABSTRACT FROM AUTHOR]

Cistanche deserticola Ma (CD) has historically been recognized for its dual role as a culinary and therapeutic parasitic herb, offering noteworthy medicinal and nutritional properties. Polysaccharides extracted and purified from CD have received an increasing great quantity of research attention due to their various pharmacological activities. In this study, using ultrasound combined with complex enzyme extraction, ethanol precipitation, DEAE‐cellulose column (26 mm × 400 mm) chromatography, and Sephacryl S‐400 HR column (26 mm × 1000 mm) chromatography, the heteropolysaccharides polysaccharide, named CDP‐2, was separated and purified from the CDP. CDP‐2 was a molecular weight of 65.60 kDa and consisted of Ara, Gal, Glc, Rha, Xyl, Man, Fuc, Gal‐UA, Glc‐UA, Man‐UA, Gul‐UA at a molar ratio of 39.05 : 23.17 : 16.29 : 11.10 : 1.79 : 1.22 : 0.47 : 3.28 : 2.48 : 0.62 : 0.55. The backbones of CDP‐2 contained ⟶4)‐α‐D‐Glcp‐(1⟶, ⟶3,6)‐β‐D‐Galp‐(1⟶, and ⟶5)‐α‐L‐Araf‐(1⟶, with a minor component of ⟶3)‐β‐D‐Galp‐(1⟶. The side chains are mainly formed by α‐L‐Araf‐(1⟶ or α‐L‐Rhap‐(1⟶ linked to the O‐6 position of residue ⟶3,6)‐β‐D‐Galp‐(1⟶. Biological assays revealed CDP‐2's efficacy in augmenting the proliferation of specific Bacteroides strains. Additionally, it was observed to facilitate the production of short‐chain fatty acids by these bacterial strains. Cumulatively, these insights underscore the CDP‐2's prospective role in bolstering gastrointestinal health via fostering Bacteroides colonization. [ABSTRACT FROM AUTHOR]

ABSTRACT Given the widespread and diverse effects of transition metal ions such as copper and zinc in multiple biological processes, this research endeavor addresses herein the microwave‐aided synthesis of two novel tetradentate ONNO (L1) and tridentate NNO (L2) donor Schiff base ligands derived from 4,5‐dimethyl‐1,2‐phenylenediamine and 4‐bromo‐2‐hydroxybenzaldehyde in a 1:2 ratio and from 4,5‐dimethyl‐1,2‐phenylenediamine and 5‐methoxy‐2‐hydroxybenzaldehyde in a 1:1 ratio, with their respective zinc, ZnL1(1) and ZnL2(3), and copper, CuL1(2) and CuL2(4), Schiff base metal complexes, which are comprehensively characterized utilizing physiochemical and analytical techniques, along with geometry optimization using the DFT approach. Spectrophotometric analyses have been used to assess the biomimetic activity in the conversion of 3,5‐DTBC to 3,5‐DTBQ with turnover numbers 53.4 and 78 h−1 in methanol for 2 and 4, whereas the zinc complexes are found to be ineffective. With CT‐DNA, possible modes of interaction with the synthesized complexes have been explored using absorption spectroscopy, where the associated binding constants ranged between 1.541 × 105 and 2.186 × 105 M−1. EtBr‐bound DNA was used in the fluorescence quenching experiments with Stern–Volmer constant (Ksv) values in the range 4.084 × 104 and 3.402 × 104 M−1, showing a stronger association with CT‐DNA. The compounds' ability to inhibit bacterial strains was also examined in vitro, as well as their ability to reduce inflammation, through the protein denaturation method and the DPPH assay for antioxidant activity have been performed. Furthermore, molecular docking simulations have been carried out to obtain a deeper comprehension of the molecular‐level interactions with CT‐DNA and cyclooxygenase‐2 (COX‐2). [ABSTRACT FROM AUTHOR]

AbstractA dose-response curve is essential for determining the safe dosage of a drug and is widely used in bioassay and in phase 1 clinical trials. It is generally accepted that the probability of death or the probability of dose-limiting toxicity is a non-decreasing function of the dose. This paper proposes and develops an effective point-wise confidence interval for an isotonic dose-response curve, a problem without a satisfactory solution so far. We show that one subset of the observations informs the lower limit while the other subset informs the upper limit. A partial likelihood ratio test using these subsets of observations is fully investigated. The resulting confidence interval is compared to three existing methods (Morris, Meyer, and Oron) in an extensive simulation study. The new method produces tight intervals while maintaining coverage. It is therefore preferred when maintaining an actual coverage probability is crucial as often in a regulatory environment. The new method extends to one-way ANOVA of a continuous response variable in a straightforward way. An R function is provided. [ABSTRACT FROM AUTHOR]

Simple Summary: By hypothesizing that molecular hyper-recombination and cellular proliferation are among the major features of multi-factorial cancer proneness, by using skin fibroblasts derived from eight major cancer syndromes, a significant correlation was found between the hyper-recombination rate quantified by plasmid assay, proliferation capacity assessed by flow cytometry, and excess of relative cancer risk (ERR). The product of the hyper-recombination rate and capacity of proliferation described a linear function of ERR. Context: Although carcinogenesis is a multi-factorial process, the mutability and the capacity of cells to proliferate are among the major features of the cells that contribute together to the initiation and promotion steps of cancer formation. Particularly, mutability can be quantified by hyper-recombination rate assessed with specific plasmid assay, hypoxanthine-guanine phosphoribosyltransferase (HPRT) mutations frequency rate, or MRE11 nuclease activities. Cell proliferation can be assessed by flow cytometry by quantifying G2/M, G1 arrests, or global cellular evasion. Methods: All these assays were applied to skin untransformed fibroblasts derived from eight major cancer syndromes characterized by their excess of relative cancer risk (ERR). Results: Significant correlations with ERR were found between hyper-recombination assessed by the plasmid assay and G2/M arrest and described a third-degree polynomial ERR function and a sigmoidal ERR function, respectively. The product of the hyper-recombination rate and capacity of proliferation described a linear ERR function that permits one to better discriminate each cancer syndrome. Conclusions: Hyper-recombination and cell proliferation were found to obey differential equations that better highlight the intrinsic bases of cancer formation. Further investigations to verify their relevance for cancer proneness induced by exogenous agents are in progress. [ABSTRACT FROM AUTHOR]

Tumor organoids and tumors-on-chips can be built by placing patient-derived cells within an engineered extracellular matrix (ECM) for personalized medicine. The engineered ECM influences the tumor response, and understanding the ECM-tumor relationship accelerates translating tumors-on-chips into drug discovery and development. In this work, we tuned the physical and structural characteristics of ECM in a 3D bioprinted soft-tissue sarcoma microtissue. We formed cell spheroids at a controlled size and encapsulated them into our gelatin methacryloyl (GelMA)-based bioink to make perfusable hydrogel-based microfluidic chips. We then demonstrated the scalability and customization flexibility of our hydrogel-based chip via engineering tools. A multiscale physical and structural data analysis suggested a relationship between cell invasion response and bioink characteristics. Tumor cell invasive behavior and focal adhesion properties were observed in response to varying polymer network densities of the GelMA-based bioink. Immunostaining assays and reverse transcription-quantitative polymerase chain reaction (RT-qPCR) helped assess the bioactivity of the microtissue and measure the cell invasion. The RT-qPCR data showed higher expressions of HIF-1α, CD44, and MMP2 genes in a lower polymer density, highlighting the correlation between bioink structural porosity, ECM stiffness, and tumor spheroid response. This work is the first step in modeling STS tumor invasiveness in hydrogel-based microfluidic chips. We optimized an engineering protocol for making tumor spheroids at a controlled size, embedding spheroids into a gelatin-based matrix, and constructing a perfusable microfluidic device. A higher tumor invasion was observed in a low-stiffness matrix than a high-stiffness matrix. The physical characterizations revealed how the stiffness is controlled by the density of polymer chain networks and porosity. The biological assays revealed how the structural properties of the gelatin matrix and hypoxia in tumor progression impact cell invasion. This work can contribute to personalized medicine by making more effective, tailored cancer models. [Display omitted] [ABSTRACT FROM AUTHOR]

Tetrodotoxin (TTX), which is found in various marine organisms, including pufferfish, shellfish, shrimp, crab, marine gastropods, and gobies, is an effective marine toxin and the cause of many seafood poisoning incidents. Owing to its toxicity and threat to public health, the development of simple, rapid, and efficient analytical methods to detect TTX in various food matrices has garnered increasing interest worldwide. Herein, we reviewed the structure and properties, origin and sources, toxicity and poisoning, and relevant legislative measures of TTX. Additionally, we have mainly reviewed the state-of-the-art progress of analytical methods for TTX detection in the past five years, such as bioassays, immunoassays, instrumental analysis, and biosensors, and summarized their advantages and limitations. Furthermore, this review provides an in-depth discussion of the most advanced biosensors, including cell-based biosensors, immunosensors, and aptasensors. Overall, this study provides useful insights into the future development and wide application of biosensors for TTX detection. [ABSTRACT FROM AUTHOR]

Background: Long-term accumulation of misfolded proteins leads to endoplasmic reticulum (ER) stress in colorectal cancer (CRC). However, the precise pathways controlling the decision between survival and apoptosis in CRC are unclear. Therefore, in this study, we investigated the function and molecular mechanism of glucosidase I (GCS1) in regulating ER stress in CRC. Methods: A public database was used to confirm the expression level of GCS1 in CRC and normal tissues. Clinical samples from our center were used to confirm the mRNA and protein expression levels of GCS1. Cell proliferation, migration, invasion, and apoptosis assays revealed the biological role of GCS1. Immunohistochemical techniques were used to evaluate the expression of key proteins in subcutaneous implanted tumors in nude mice, which provided further evidence for the biological function of GCS1 in promoting cancer in vivo. The results of coimmunoprecipitation-mass spectrometry analysis and immunofluorescence colocalization analysis the interaction between GCS1 and GRP78. In addition, the mechanism of action of USP10, GRP78, and GCS1 at the post- translational level was investigated. Finally, a tissue microarray was used to examine the connection between GCS1 and GRP78 expression and intracellular localization of these proteins using immunohistochemistry and immunofluorescence. Results: The experimental results revealed that GCS1 was substantially expressed in CRC, with higher expression indicating a worse prognosis. Thus, GCS1 can enhance the proliferation and metastasis while inhibiting the apoptosis of CRC cells both in vivo and in vitro. Mechanistically, GCS1 binds to GRP78, recruits USP10 for deubiquitination of GRP78 to promote its degradation, and decreases ER stress-mediated apoptosis, increasing CRC cell proliferation and metastasis. Conclusions: In summary, GCS1 stimulates CRC growth and migration and reduces ER stress-mediated apoptosis via USP10-mediated deubiquitination of GRP78. Our findings identify a possible therapeutic target for CRC. [ABSTRACT FROM AUTHOR]

Aconiti Lateralis Radix Praeparata (Fuzi in Chinese) is widely used in the clinical treatment of tumors. This study aims to explore the active fractions and underlying mechanisms of Fuzi in the treatment of non-small cell lung cancer (NSCLC). Fuzi alkaloids (FZA) is prepared and found to inhibit the growth of NSCLC both in vitro and in vivo significantly. A total of 53 alkaloids are identified in FZA by UPLC-Q-TOF-MS. Proteomics experiment show that 238 differentially expressed proteins regulated by FZA are involved in amino acid anabolism, pyrimidine metabolism and PI3K/Akt-mTOR signaling pathway. Metabolomics analyses identify 32 significant differential metabolites which are mainly involved in amino acid metabolism, TCA cycle and other pathways. Multi-omics research combined with molecular biological assays suggest that FZA might regulate glycolysis through PI3K/Akt-mTOR pathway to treat NSCLC. The study lays a foundation for the anti-cancer investigation of Fuzi and provides a possible scientific basis for its clinical application. Proteomics experiments, metabolomics analyses and network pharmacology combined with molecular biology results suggest that alkaloids of Aconiti Lateralis Radix Praeparata might treat NSCLC by regulating PI3K/Akt-mTOR signaling and glycolysis. [ABSTRACT FROM AUTHOR]

Molecular imaging holds the potential for noninvasive and accurate grading of liver fibrosis. It is limited by the lack of biomarkers that strongly correlate with liver fibrosis grade. Here, we discover the grading potential of fibroblast activation protein alpha (FAPα) for liver fibrosis through transcriptional analysis and biological assays on clinical liver samples. The protein and mRNA expression of FAPα are linearly correlated with fibrosis grade (R2 = 0.89 and 0.91, respectively). A FAPα-responsive MRI molecular nanoprobe is prepared for quantitatively grading liver fibrosis. The nanoprobe is composed of superparamagnetic amorphous iron nanoparticles (AFeNPs) and paramagnetic gadoteric acid (Gd-DOTA) connected by FAPα-responsive peptide chains (ASGPAGPA). As liver fibrosis worsens, the increased FAPα cut off more ASGPAGPA, restoring a higher T1-MRI signal of Gd-DOTA. Otherwise, the signal remains quenched due to the distance-dependent magnetic resonance tuning (MRET) effect between AFeNPs and Gd-DOTA. The nanoprobe identifies F1, F2, F3, and F4 fibrosis, with area under the curve of 99.8%, 66.7%, 70.4%, and 96.3% in patients' samples, respectively. This strategy exhibits potential in utilizing molecular imaging for the early detection and grading of liver fibrosis in the clinic. Molecular imaging holds promise for liver fibrosis grading but limited by a lack of effective biomarker. Here, the authors discover the grading potential of fibroblast activation protein alpha and design a responsive MRI nanoprobe that achieves fibrosis grading in mice models and clinical samples. [ABSTRACT FROM AUTHOR]

Introduction: Triple-negative breast cancer (TNBC) represents a significant global health crisis due to its resistance to conventional therapies and lack of specific molecular targets. This study explored the potential of Eriocephalus racemosus (E. racemosus) as an alternative treatment for TNBC. The cytotoxic properties and high-resolution respirometry mitochondrial activities of E. racemosus against the MDA-MB 231 TNBC cell line were evaluated. Methods: Hexane solvent and bioactive fraction extractions of E. racemosus were performed, while mass spectrometry-based metabolite profiling was used to identify the phytochemical constituents of the extracts. The extracts were further tested against MDA-MB 231 TNBC cells to determine their cytotoxicity. The mode of cell death was determined using flow cytometry. The activities of caspases 3, 8, and 9 were assessed using a multiplex activity assay kit. Glycolytic activity and High-resolution respirometry measurements of mitochondrial function in the MDA-MB 231 cell line were conducted using the Seahorse XFp and Oroboros O2K. Results: Metabolite profiling of E. racemosus plant crude extracts identified the presence of coumarins, flavonoids, sesquiterpenoids, triterpenoids, and unknown compounds. The extracts demonstrated promising cytotoxic activities, with a half maximal inhibitory concentration (IC50) of 12.84 µg/mL for the crude hexane extract and 15.49 µg/mL for the bioactive fraction. Further, the crude hexane and bioactive fraction extracts induced apoptosis in the MDA-MB-231 TNBC cells, like the reference drug cisplatin (17.44%, 17.26% and 20.25%, respectively) compared to untreated cells. Caspase 3 activities confirmed the induction of apoptosis in both cisplatin and the plant crude extracts, while caspase 8 and 9 activities confirmed the activation of both the intrinsic and extrinsic apoptosis pathways. Increased levels of glycolytic activity were observed in the hexane crude extract. High-resolution respiratory measurements showed elevated mitochondrial activities in all mitochondrial states except for complex-IV activity. Conclusion: These findings support further exploration of E. racemosus as a potential therapeutic agent for TNBC, offering a promising avenue for the development of targeted treatments with minimal adverse effects. [ABSTRACT FROM AUTHOR]

Layered double hydroxides (LDHs) are biocompatible materials with excellent anion exchange properties, making them suitable for use as drug carriers by incorporating the drug in its anionic form within their layered structure. In this study, an LDH was synthetized through ion exchange with valproate (VAL), an anticonvulsant drug, using an LDH with nitrate as the parent material. The resulting LDH-VAL sample underwent a comprehensive multi-technique characterization using XRD, TG-DSC, TEM, SEM, EDS, FTIR, XPS and zeta potential measurements. Dissolution kinetics and VAL release kinetics in an ample range of pH (3–9), along with biological assays to test biocompatibility, were also performed. All techniques confirmed the successful intercalation of the drug into the LDH structure, displacing all the nitrate ions of the parent material and increasing the basal spacing d 003 from 8.8 Å to 18.3 Å. VAL release in NaCl and physiological buffer solutions was incongruent with LDH dissolution, clearly indicating that VAL release was controlled by an ion exchange process. In 0.9 % NaCl solutions, 90–100 % of VAL was released with only 15–20 % of LDH dissolution, and in buffer solutions 100 % of VAL was released with, at the most, 30 % of LDH dissolution. Biological assays, including lactate dehydrogenase activity, serum calcium concentration and histological analysis of blood samples, confirmed that the prepared LDH-VAL is a biocompatible material with a potential use as a nanodrug carrier. [Display omitted] [ABSTRACT FROM AUTHOR]

[Objectives] The paper aimed to determine the resistance level of Amaranthus retroflexus L. populations to fomesafen collected from soybean fields in Heilongjiang Province, and to study the molecular mechanism of fomesafen resistance and multiple resistance. [Methods]Three populations from Nenjiang County in Heilongjiang Province were evaluated for resistance level to fomesafen by agar-based rapid bioassay. The plants that survived fomesafen or thifensulfuron-methyl treatment in pots at the field recommend dose were supposed to be the resistant ones, and their full-length sequences of protoporphyrinogen Ⅸ oxidase genes and acetolactate synthase gene were sequenced. [Results] Compared with the sensitive population, the relative resistance index(RI)of the three populations to fomesafen were 83.6, 83.8 and 93.4, respectively. It was found that the nucleotides encoding amino acids at 128 in the highly conserved PPO2 region were mutated from AGG to GGG, resulting in the substitution of arginine to glycine(Arg-128-Gly). Additionally, all three populations exhibited multiple resistance to ALS inhibitors. The als gene sequencing showed that single or combined amino acid substitutions at multiple sites(Ala-205-Val, Asp-376-Glu, Trp-574-Leu, and Ser-653-Thr) occurred in the ALS conserved region of the three resistant populations, with a diverse heterozygosity. [Conclusions] Arg-128-Gly amino acid substitution of PPO2 might be the main resistance mechanism of A.retroflexus L. populations to fomesafen. All the three A.retroflexus L. populations showed multiple resistance to ALS inhibitors, and the resistance mechanism was mainly due to target site ALS mutations. [ABSTRACT FROM AUTHOR]