Thrombospondin-1 Regulates Trophoblast Necroptosis via NEDD4-Mediated Ubiquitination of TAK1 in Preeclampsia.

Preeclampsia (PE) is considered as a disease of placental origin. However, the specific mechanism of placental abnormalities remains elusive. This study identified thrombospondin-1 (THBS1) is downregulated in preeclamptic placentae and negatively correlated with blood pressure. Functional studies show that THBS1 knockdown inhibits proliferation, migration, and invasion and increases the cycle arrest and apoptosis rate of HTR8/SVneo cells. Importantly, THBS1 silencing induces necroptosis in HTR8/SVneo cells, accompanied by the release of damage-associated molecular patterns (DAMPs). Necroptosis inhibitors necrostatin-1 and GSK'872 restore the trophoblast survival while pan-caspase inhibitor Z-VAD-FMK has no effect. Mechanistically, the results show that THBS1 interacts with transforming growth factor B-activated kinase 1 (TAK1), which is a central modulator of necroptosis quiescence and affects its stability. Moreover, THBS1 silencing up-regulates the expression of neuronal precursor cell-expressed developmentally down-regulated 4 (NEDD4), which acts as an E3 ligase of TAK1 and catalyzes K48-linked ubiquitination of TAK1 in HTR8/SVneo cells. Besides, THBS1 attenuates PE phenotypes and improves the placental necroptosis in vivo. Taken together, the down-regulation of THBS1 destabilizes TAK1 by activating NEDD4-mediated, K48-linked TAK1 ubiquitination and promotes necroptosis and DAMPs release in trophoblast cells, thus participating in the pathogenesis of PE.

A fully validated LC­MS/MS method for quantifying bevacizumab in plasma samples from patients with NSCLC and its implications in therapeutic drug monitoring.

Given the increasing use of bevacizumab in combinatorial drug therapy for a multitude of different cancer types, there is a need for therapeutic drug monitoring to analyze the possible correlation between drug trough concentration, and therapeutic effect and adverse reactions. An ultra-performance liquid chromatography tandem-mass spectrometry method was then developed and validated to determine bevacizumab levels in human plasma samples. Chromatographic separation was achieved on a Shimadzu InertSustainBio C18 HP column, whereas subsequent mass spectrometric analysis was performed using a Shimadzu 8050CL triple quadrupole mass spectrometer equipped with an electro-spray ionization source in the positive ion mode. In total, three multiple reaction monitoring transitions of each of the surrogate peptides were chosen with 'FTFSLDTSK' applied as the quantification peptide whereas 'VLIYFTSSLHSGVPSR' and 'STAYLQMNSLR' were designated as the verification peptides using the Skyline software. This analytical method was then fully validated, with specificity, linearity, lower limit of quantitation, accuracy, precision, stability, matrix effect and recovery calculated. The linearity of this method was developed to be within the concentration range 5-400 µg/ml for bevacizumab in human plasma. Subsequently, eight patients with non-small cell lung cancer (NSCLC) were recruited and injected with bevacizumab over three periods of treatment to analyze their steady-state trough concentration and differences. To conclude, the results of the present study suggest that bevacizumab can be monitored in a therapeutic setting in patients with NSCLC.

Effects of Sodium Vacancies and Concentrations in Na3SO4F Solid Electrolyte.

The sodium-rich solid electrolyte, Na3SO4F (NSOF), holds promise for eco-friendly and resource-abundant energy storage. While the introduction of heterovalent dopants has the potential to enhance its suitability for battery applications by creating Na vacancies, the effect of vacancies and sodium concentrations on sodium conduction remains unclear. In this work, Mg2+ was introduced into Na+ sites in Na3SO4F, generating sodium vacancies with different contents by using solid-state synthesis method. Among the resulting materials, Na2.96Mg0.02SO4F exhibited an ionic conductivity that is two-order-of-magnitude higher than NSOF at 298 K. Notably, as the sodium concentration decreased, the ionic conductivity also declined, revealing an equilibrium between Na vacancies and concentrations. To further investigate the influence of sodium concentration, excess Na+ was introduced into NaMgSO4F, which inherently possesses a lower sodium content by using solid-state synthesis method. However, this adjustment only led to an approximately one-order-of-magnitude enhancement in optimal ionic conductivity at 298 K. Combined with an in situ X-ray diffraction analysis, our findings underscore the greater sensitivity of sodium conduction to variations in sodium vacancies. This study paves the way for the development of ultrafast sodium ion conductors, offering exciting prospects for advanced energy storage solutions.

Factors influencing the accuracy and safety of preoperative computed tomography (CT)-guided soft hook-wire localization for pulmonary nodules: a comprehensive analysis.

Background: The necessity of localization of pulmonary nodules lies in ensuring the ability to locate the nodule quickly and accurately during surgery, thereby improving the success rate of the operation. The accuracy and risk of preoperative localization of pulmonary nodules need further exploration. Therefore, the purpose of this study was to investigate the factors of accuracy and safety of computed tomography (CT)-guided localization of pulmonary nodules using a flexible wire hook positioner. Methods: In this retrospective cross-sectional analysis, 281 patients with a single pulmonary nodule underwent video-assisted thoracoscopic surgery (VATS) following localization with a soft hook-wire guided by CT scan from January 2021 to July 2022 at Nanjing Drum Tower Hospital. The patients underwent VATS to remove pulmonary nodules within 24 hours after localization. The demographic, pulmonary nodule, and technical factors were analyzed retrospectively. Univariate and multivariate analysis were used to analyze the identified factors that influence pulmonary nodule localization accuracy and complications. Results: Localization was successfully performed in 280 patients, with only 1 patient being excluded due to a displaced positioner and the hook wire failing to enter the lung parenchyma as a result of pneumothorax. Out of the total cases, 191 (68.2%) were accurately positioned in group G0, whereas 89 cases (31.7%) were inaccurately positioned in group G1. Hemorrhage and self-limited hemoptysis were observed in 64 patients (22.8%), whereas pneumothorax was observed in 84 patients (29.9%). There were no serious complications such as air embolism or death. The accuracy of localization was found to be influenced by both the depth of pulmonary nodules [odds ratio (OR) =22.610, 95% confidence interval (CI): 10.351-49.391, P=0.001] and the depth of the needle used (OR =0.322, 95% CI: 0.136-0.765, P=0.010). Additionally, postoperative hemorrhage was found to be affected by several important factors, including the diameter (P=0.036) and depth of the nodule (P=0.011), as well as the thickness of the chest wall (P=0.043) and the depth of the needle used (P=0.005). Conclusions: The CT-guided flexible wire hook positioner has been found to be a safe and effective device for locating pulmonary nodules. The depth of pulmonary nodules and needle penetration are key factors affecting the accuracy of lung nodule localization under CT guidance and are important factors affecting postoperative bleeding.

Prediction of DNA i-motifs via machine learning.

i-Motifs (iMs), are secondary structures formed in cytosine-rich DNA sequences and are involved in multiple functions in the genome. Although putative iM forming sequences are widely distributed in the human genome, the folding status and strength of putative iMs vary dramatically. Much previous research on iM has focused on assessing the iM folding properties using biophysical experiments. However, there are no dedicated computational tools for predicting the folding status and strength of iM structures. Here, we introduce a machine learning pipeline, iM-Seeker, to predict both folding status and structural stability of DNA iMs. The programme iM-Seeker incorporates a Balanced Random Forest classifier trained on genome-wide iMab antibody-based CUT&Tag sequencing data to predict the folding status and an Extreme Gradient Boosting regressor to estimate the folding strength according to both literature biophysical data and our in-house biophysical experiments. iM-Seeker predicts DNA iM folding status with a classification accuracy of 81% and estimates the folding strength with coefficient of determination (R2) of 0.642 on the test set. Model interpretation confirms that the nucleotide composition of the C-rich sequence significantly affects iM stability, with a positive correlation with sequences containing cytosine and thymine and a negative correlation with guanine and adenine.

Photochemical-promoted ZVI reduction for highly efficient removal of 4-chlorophenol and Cr(VI): Catalytic activity, performance and electron transfer mechanisms.

Zero-valent iron (ZVI) reduction represents a promising methodology for water remediation, but its broad application is limited by two critical challenges (i.e., aggregation and passivation). Here, we report a hybrid strategy of photochemical-promoted ZVI reduction with high efficiency and reduction capacity for removing coexisting refractory pollutants in water. A composite material with Pd/Fe bimetallic nanoparticles supported onto semiconducting metal oxide (Pd/Fe@WO3-GO) was prepared and subsequently used as the model catalyst. By using the developed strategy with visible light as light source, this catalyst showed a remarkable catalytic performance for simultaneously eliminating 4-chlorophenol (4-CP) and Cr(VI), with dehalogenation rate as high as 0.43 min-1, outperforming the reported ZVI-based catalysts. A synergistic interaction of photocatalysis and ZVI reduction occurred in this strategy, where the interfacial electron transfer on particles surface were greatly strengthened with light irradiation. The activation was attributed to the dual functions of semiconducting material as support to disperse Pd/Fe nanoparticles and as (photoexcited) electron donor to directly trigger reduction reactions and/or indirectly inhibit the formation of oxides passivation layer. Both direct electron transfer and H*-mediated indirect electron transfer mechanisms were confirmed to participate in the reduction of pollutants, while the later was quantitatively demonstrated as the predominant reaction route. Importantly, this strategy showed a wide pH applicability, long-term durability and excellent catalytic performance in different real-water systems. This work provides new insights into ZVI reduction and advances its applications for the removal of combined organic and inorganic pollutants. The developed photochemical-promoted ZVI reduction strategy holds a great potential for practical applications.

Potential roles of the interactions between gut microbiota and metabolites in LPS-induced intrauterine inflammation (IUI) and associated preterm birth (PTB).

BACKGROUND: Prenatal exposure to intrauterine inflammation (IUI) is a crucial event in preterm birth (PTB) pathophysiology, increasing the incidence of neurodevelopmental disorders. Gut microbiota and metabolite profile alterations have been reported to be involved in PTB pathophysiology. METHOD AND RESULTS: In this study, IUI-exposed PTB mouse model was established and verified by PTB rate and other perinatal adverse reactions; LPS-indued IUI significantly increased the rates of PTB, apoptosis and inflammation in placenta tissue samples. LPS-induced IUI caused no significant differences in species richness and evenness but significantly altered the species abundance distribution. Non-targeted metabolomics analysis indicated that the metabolite profile of the preterm mice was altered, and differential metabolites were associated with signaling pathways including pyruvate metabolism. Furthermore, a significant positive correlation between Parasutterella excrementihominis and S4572761 (Nb-p-coumaroyltryptamine) and Mreference-1264 (pyruvic acid), respectively, was observed. Lastly, pyruvic acid treatment partially improved LPS-induced IUI phenotypes and decreased PTB rates and decreased the apoptosis and inflammation in placenta tissue samples. CONCLUSION: This study revealed an association among gut microbiota dysbiosis, metabolite profile alterations, and LPS-induced IUI and PTB in mice models. Our investigation revealed the possible involvement of gut microbiota in the pathophysiology of LPS-induced IUI and PTB, which might be mediated by metabolites such as pyruvic acid. Future studies should be conducted to verify the findings through larger sample-sized animal studies and clinical investigations.

Polyolefin-derived substrate-grown carbon nanotubes as binder-free electrode for hydrogen evolution in alkaline media.

Switching from a linear mode of waste management to a circular loop by transforming plastic waste into carbon nanotubes (CNTs) is a promising approach to current plastic waste treatment. One of the many applications of CNTs is its use for electrocatalytic water splitting for hydrogen evolution. Existing methods of CNTs-based hydrogen evolution reaction (HER) electrode fabrication involve additives like polymeric binders and additional steps to improve CNT dispersion, which are detrimental to the CNT structure and properties. The in-situ fabrication approach can potentially be a one-pot solution to HER electrode synthesis. In this study, polyolefins pyrolysis gas and a Co:Ni:Mg catalyst were used to fabricate binder-free CNTs-based electrodes on different substrates for HER. The study assessed CNT quality on conductive carbon paper, semiconductive silicon, and dielectric glass substrates, evaluating their HER performance in 1 M KOH. A mixture of hollow-core, bamboo-like, and cup-stacked arrangement nanotubes were synthesized on the substrates, with CNTs on glass and carbon paper substrates possessing better graphitization than CNTs grown on silicon. This is in agreement with HER performance, whereby the as-prepared electrodes required overpotentials of 267 mV, 241 mV, and 216 mV for silicon, glass, and carbon paper, respectively, to achieve 10 mA/cm2. Despite being poorly conductive, the glass substrate electrode achieved a lower overpotential than the silicon electrode. Additionally, the as-prepared silicon electrode faced a delamination issue likely attributed to the lower surface energy of the silicon substrate surface, demonstrating the weaker adhesion between the CNTs and silicon surface. The proposed approach thus showed that the in-situ fabricated electrodes performed better than separately synthesized CNTs prepared into electrodes by 27.4% and 14.2% for carbon paper and glass substrates, respectively. The improved performance of the as-prepared, binder-free electrodes can be linked to the lower charge-transfer resistance and reduced contact resistance between the CNTs and substrate.

Synthesis of TiOxNy oxynitrides with a tunable nitrogen content.

TiOxNy is a solid solution of cubic TiN and cubic TiO, with the ability to adjust its physical and chemical properties by modifying the O/N ratio, thus making it suitable for various applications. However, the synthesis of TiOxNy remains a significant challenge for inorganic chemists. In this work, we have successfully synthesized TiOxNy nanoparticles using the layered oxide Cs0.68Ti1.83O4 as the precursor and urea as the nitrogen source. The synthesis process was conducted within a temperature range of 800 to 1200 °C, leading to the transformation of the nitrided products from a two-dimensional layered precursor structure to a three-dimensional structure as the temperature increased. By varying the reaction temperature, the O/N ratio can be controlled. The experimental findings demonstrate that the nitrogen content in TiOxNy nanoparticles increases with rising temperature, ranging from TiO0.8N0.2 at 800 °C to TiO0.25N0.75 at 1200 °C. This work highlights the potential of the solid-state method in tailoring the properties of TiOxNy nanoparticles and presents a novel approach for synthesizing oxynitrides.

Alterations and potential roles of microbial population of pregnant mouse saliva and amniotic fluid.

PROBLEM: Prenatal exposure to intrauterine inflammation (IUI) is a crucial event in PTB pathophysiology. However, the relationship between microflora and PTB is not fully elucidated. METHOD OF STUDY: In this study, we established an intrauterine inflammation mouse model via LPS intrauterine injection. The saliva and amniotic fluid were collected for 16s RNA gene sequencing. The levels of TNF-α and IL-1ß in mouse amniotic fluid were determined by ELISA assays. RESULTS: Up to 60% of the operational taxonomic units (OTUs) in the saliva and amniotic fluid of PBS-treated mice were overlapped. LPS treatment-induced changes in the abundance of oral and amniotic fluid microorganisms. Both immune-associated probiotics, salivarius and mastitidis, were still detected in saliva (at significantly increased levels) after LPS-induced intrauterine inflammation and almost no probiotics of any type were detected in amniotic fluid, suggesting that the uterine cavity seems to be more susceptible to LPS compared to the oral cavity. Moreover, the abundance of pathogenic bacteria Escherichia coli was increased in both saliva and amniotic fluid after LPS treatment. The level of TNF-α and IL-1ß in amniotic fluid is positively related to the amniotic fluid E. coli abundance. CONCLUSIONS: The microbial composition of saliva and amniotic fluid of pregnant mice was similar. LPS-induced intrauterine inflammation decreased the consistency of microbial composition in mouse saliva and amniotic fluid, increased the abundance of E. coli in saliva and amniotic fluid, and decreased the abundance of immune-associated probiotics, especially in amniotic fluid.

The theory of critical flicker fusion frequency and its application in cataracts.

Background: Due to media opacity, it is usually difficult to accurately evaluate the postoperative visual acuity in cataracts patients. As a small and portable tool, the critical flicker fusion frequency (CFF) device reflects the temporal resolution of visual function and has been widely used in clinical research. However, poor understanding of the technique and equipment limitations have restricted its clinical application in China. Main text: There was a decrease in the CFF value in various ophthalmic diseases, indicating that the CFF is sensitive to detect visual functional changes. A number of studies have shown that the CFF test can accurately distinguish patients with simple cataracts from those with cataracts combined with fundus disease, and, as a visual test, it can more accurately predict postoperative visual acuity without being affected by media opacity. This study comprehensive reviews the basic principles of CFF and its application in ophthalmology, especially in cataracts. Conclusions: As one of the tools for dynamic visual function detection, the CFF test could help doctors to assess the possible presence of fundus disease in cataracts patients, especially in eyes with dense cataracts, and more precisely provide a reasonable visual prognosis than other available visual tests.

Specific Recognition and Adsorption of Volatile Organic Compounds by Using MIL-125-Based Porous Fluorescence Probe Material.

The severity of the volatile organic compounds (VOCs) issue calls for effective detection and management of VOC materials. Metal-organic frameworks (MOFs) are organic-inorganic hybrid crystals with promising prospects in luminescent sensing for VOC detection and identification. However, MOFs have limitations, including weak response signals and poor sensitivity towards VOCs, limiting their application to specific types of VOC gases. To address the issue of limited recognition and single luminosity for specific VOCs, we have introduced fluorescent guest molecules into MOFs as reference emission centers to enhance sensitivity. This composite material combines the gas adsorption ability of MOFs to effectively adsorb VOCs. We utilized (MIL-125/NH2-MIL-125) as the parent material for adsorbing fluorescent molecules and selected suitable solid fluorescent probes (FGFL-B1) through fluorescence enhancement using thioflavin T and MIL-125. FGFL-B1 exhibited a heightened fluorescence response to various VOCs through charge transfer between fluorescent guest molecules and ligands. The fluorescence enhancement effect of FGFL-B1 on tetrahydrofuran (THF) was particularly pronounced, accompanied by a color change from yellow to yellowish green in the presence of CCl4. FGFL-B1 demonstrated excellent adsorption properties for THF and CCl4, with saturated adsorption capacities of 655.4 mg g-1 and 811.2 mg g-1, respectively. Furthermore, FGFL-B1 displayed strong luminescence stability and reusability, making it an excellent sensing candidate. This study addresses the limitations of MOFs in VOC detection, opening avenues for industrial and environmental applications.

A fully validated method for simultaneous determination of icotinib, osimertinib, gefitinib and O-desmethyl gefitinib in human plasma using UPLC-MS/MS for therapeutic drug monitoring.

BACKGROUND AND AIMS: A few researches have reported the exposure-efficacy/toxicity relationships of epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKIs). On account of the large interpatient pharmacokinetic variability, therapeutic drug monitoring (TDM) seems promising for optimizing dosage regimen and improving treatment efficacy and safety. Therefore, a rapid and convenient ultrahigh performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method was developed and validated for the determination of icotinib, osimertinib, gefitinib and O-demesthyl gefitinib in human plasma for TDM. MATERIALS AND METHODS: Icotinib-D4 and osimertinib-13CD3 were used as the internal standards (ISs). The samples were prepared by protein precipitation using acetonitrile. Chromatographic separation was achieved on a 40 â„ƒ Shimadzu Shim-pack Scepter C18-120 column (2.1 ×50 mm, 3.0 µm, Japan) by a Shimadzu 30 A solvent management system. Detection was carried out using a Shimadzu LC-MS 8050CL triple quadrupole mass spectrometer coupled with an electrospray ionization source in positive mode. RESULTS: This analytical method was fully validated with selectivity, carry-over, linearity, lower limit of quantification, accuracy (from 92.68% to 106.62%) and precision (intra- and inter-day coefficients of variation ranged from 0.92% to 9.85%), matrix effect, extraction recovery, stability and dilution integrity. The calibration curves were developed to be within the concentration ranges of 200-4000 ng/mL for icotinib, 50-1000 ng/mL for osimertinib, gefitinib and O-desmethyl gefitinib in human plasma which meet the needs of routine TDM. CONCLUSIONS: The proposed method was used in 100 patients with non-small cell lung cancer for monitoring plasma concentration of the mentioned EGFR-TKIs. The trough concentrations of ICO were distributed between 226.42 ng/mL and 3853.36 ng/mL, peak concentrations were between 609.20 ng/mL and 2191.54 ng/mL. The trough concentrations of OSI were distributed between 110.48 ng/mL and 1183.13 ng/mL. The trough concentrations of GEF were distributed between 117.71 ng/mL and 582.74 ng/mL, while DeGEF was distributed from 76.21 ng/mL to 1939.83 ng/mL with two less than 20 ng/mL. The results of therapeutic drug monitoring aimed to investigate exposure-efficacy/toxicity relationship and improve the efficacy and safety of targeted therapies.

BNTA attenuates temporomandibular joint osteoarthritis progression by directly targeting ALDH3A1: An in vivo and in vitro study.

BNTA is known to have a therapeutic effect on knee osteoarthritis and inflammatory osteoclastogenesis. However, the protective effect of BNTA regarding temporomandibular mandibular joint osteoarthritis (TMJOA) and its underlying mechanism and physiological target remains unclear. In the present study, BNTA ameliorated cartilage degradation and inflammation responses in monosodium iodoacetate (MIA)-induced TMJOA in vivo. In IL-1ß-induced condylar chondrocytes, BNTA prevents oxidative stress, inflammatory responses and increasing synthesis of cartilage extracellular matrix through activating nuclear factor-E2-related factor 2 (NRF2) signaling. Suppression of NRF2 signaling abolishes the protective effect of BNTA in TMJOA. Notably, BNTA may bind directly to ALDH3A1 and act as a stabilizer, as evidenced by drug affinity responsive target stability assay (DARTS), cellular thermal shift assay (CETSA) and molecular docking results. Further investigation of the underlying molecular and cellular mechanism infers a positive correlation of ALDH3A1 regulating NRF2 signaling. In conclusion, BNTA may attenuate TMJOA progression via the ALDH3A1/NRF2 axis, inferring that BNTA is a therapeutic target for treating temporomandibular mandibular joint osteoarthritis.

Fabrication of Monolayer Graphene-Coated Grids for Cryoelectron Microscopy.

Cryogenic electron microscopy (cryoEM) has emerged as a powerful technique for probing the atomic structure of macromolecular complexes. Sample preparation for cryoEM requires preserving specimens in a thin layer of vitreous ice, typically suspended within the holes of a fenestrated support film. However, all commonly used sample preparation approaches for cryoEM studies expose the specimen to the air-water interface, introducing a strong hydrophobic effect on the specimen that often results in denaturation, aggregation, and complex dissociation. Further, preferred hydrophobic interactions between regions of the specimen and the air-water interface impact the orientations adopted by the macromolecules, resulting in 3D reconstructions with anisotropic directional resolution. Adsorption of cryoEM specimens to a monolayer of graphene has been shown to help mitigate interactions with the air-water interface while minimizing the introduction of background noise. Graphene supports also offer the benefit of substantially lowering the required concentration of proteins required for cryoEM imaging. Despite the advantages of these supports, graphene-coated grids are not widely used by the cryoEM community due to the prohibitive expense of commercial options and the challenges associated with large-scale in-house production. This paper describes an efficient method for preparing batches of cryoEM grids that have nearly full coverage of monolayer graphene.

A workflow for the development of template-assisted membrane crystallization downstream processing for monoclonal antibody purification.

Monoclonal antibodies (mAbs) are commonly used biologic drugs for the treatment of diseases such as rheumatoid arthritis, multiple sclerosis, COVID-19 and various cancers. They are produced in Chinese hamster ovary cell lines and are purified via a number of complex and expensive chromatography-based steps, operated in batch mode, that rely heavily on protein A resin. The major drawback of conventional procedures is the high cost of the adsorption media and the extensive use of chemicals for the regeneration of the chromatographic columns, with an environmental cost. We have shown that conventional protein A chromatography can be replaced with a single crystallization step and gram-scale production can be achieved in continuous flow using the template-assisted membrane crystallization process. The templates are embedded in a membrane (e.g., porous polyvinylidene fluoride with a layer of polymerized polyvinyl alcohol) and serve as nucleants for crystallization. mAbs are flexible proteins that are difficult to crystallize, so it can be challenging to determine the optimal conditions for crystallization. The objective of this protocol is to establish a systematic and flexible approach for the design of a robust, economic and sustainable mAb purification platform to replace at least the protein A affinity stage in traditional chromatography-based purification platforms. The procedure provides details on how to establish the optimal parameters for separation (crystallization conditions, choice of templates, choice of membrane) and advice on analytical and characterization methods.

OsWRKY97, an Abiotic Stress-Induced Gene of Rice, Plays a Key Role in Drought Tolerance.

Drought stress is one of the major causes of crop losses. The WRKY families play important roles in the regulation of many plant processes, including drought stress response. However, the function of individual WRKY genes in plants is still under investigation. Here, we identified a new member of the WRKY families, OsWRKY97, and analyzed its role in stress resistance by using a series of transgenic plant lines. OsWRKY97 positively regulates drought tolerance in rice. OsWRKY97 was expressed in all examined tissues and could be induced by various abiotic stresses and abscisic acid (ABA). OsWRKY97-GFP was localized to the nucleus. Various abiotic stress-related cis-acting elements were observed in the promoters of OsWRKY97. The results of OsWRKY97-overexpressing plant analyses revealed that OsWRKY97 plays a positive role in drought stress tolerance. In addition, physiological analyses revealed that OsWRKY97 improves drought stress tolerance by improving the osmotic adjustment ability, oxidative stress tolerance, and water retention capacity of the plant. Furthermore, OsWRKY97-overexpressing plants also showed higher sensitivity to exogenous ABA compared with that of wild-type rice (WT). Overexpression of OsWRKY97 also affected the transcript levels of ABA-responsive genes and the accumulation of ABA. These results indicate that OsWRKY97 plays a crucial role in the response to drought stress and may possess high potential value in improving drought tolerance in rice.

Machine learning-based radiomics for predicting BRAF-V600E mutations in ameloblastoma.

Background: Ameloblastoma is a locally invasive and aggressive epithelial odontogenic neoplasm. The BRAF-V600E gene mutation is a prevalent genetic alteration found in this tumor and is considered to have a crucial role in its pathogenesis. The objective of this study is to develop and validate a radiomics-based machine learning method for the identification of BRAF-V600E gene mutations in ameloblastoma patients. Methods: In this retrospective study, data from 103 patients diagnosed with ameloblastoma who underwent BRAF-V600E mutation testing were collected. Of these patients, 72 were included in the training cohort, while 31 were included in the validation cohort. To address class imbalance, synthetic minority over-sampling technique (SMOTE) is applied in our study. Radiomics features were extracted from preprocessed CT images, and the most relevant features, including both radiomics and clinical data, were selected for analysis. Machine learning methods were utilized to construct models. The performance of these models in distinguishing between patients with and without BRAF-V600E gene mutations was evaluated using the receiver operating characteristic (ROC) curve. Results: When the analysis was based on radiomics signature, Random Forest performed better than the others, with the area under the ROC curve (AUC) of 0.87 (95%CI, 0.68-1.00). The performance of XGBoost model is slightly lower than that of Random Forest, and its AUC is 0.83 (95% CI, 0.60-1.00). The nomogram evident that among younger women, the affected region primarily lies within the mandible, and patients with larger tumor diameters exhibit a heightened risk. Additionally, patients with higher radiomics signature scores are more susceptible to the BRAF-V600E gene mutations. Conclusions: Our study presents a comprehensive radiomics-based machine learning model using five different methods to accurately detect BRAF-V600E gene mutations in patients diagnosed with ameloblastoma. The Random Forest model's high predictive performance, with AUC of 0.87, demonstrates its potential for facilitating a convenient and cost-effective way of identifying patients with the mutation without the need for invasive tumor sampling for molecular testing. This non-invasive approach has the potential to guide preoperative or postoperative drug treatment for affected individuals, thereby improving outcomes.

A highly sensitive method for determination of tacrolimus in peripheral blood mononuclear cells by nano liquid chromatography-high resolution accurate mass spectrometry.

The determination of intracellular tacrolimus concentration in peripheral blood mononuclear cells (PBMCs) is crucial for assessing the effect-site concentration of tacrolimus. Analytical methods previously reported required a minimum of 3 mL of whole blood sample for measuring the tacrolimus concentration. In this study, we developed a highly sensitive method using EASY nLC 1200 combined with Q Exactive orbitrap mass spectrometer for detecting tacrolimus in PBMCs, requiring only 0.5-2 mL of sample. Furthermore, we compared two primary normalization methods for PBMCs tacrolimus concentration using Passing-Bablok regression, Bland-Altman analysis, Spearman's rank correlation, and Mountain plot. The newly established method was employed to compare tacrolimus concentrations in whole blood and PBMCs among 194 lung transplant recipients. The developed method exhibited high sensitivity with a lower limit of quantitation at 5 pg/mL, and excellent intra- and inter-days accuracy and precision. The comparison between different normalization methods for PBMCs tacrolimus concentration revealed a strong correlation between PBMCs count and intracellular protein amount within these cells. This finding suggests that both PBMCs count and intracellular protein amount can be used for normalizing intracellular tacrolimus levels and can be mutually converted. However, a weaker correlation was observed between PBMCs and whole-blood tacrolimus concentrations in lung transplant recipients, warranting further investigation. The method reported herein enables the quantification of PBMCs tacrolimus concentration using smaller volumes of whole blood samples, which has significant implications for both patients and laboratory personnel.

DELE1 oligomerization promotes integrated stress response activation.

Mitochondria are dynamic organelles that continually respond to cellular stress. Recent studies have demonstrated that mitochondrial stress is relayed from mitochondria to the cytosol by the release of a proteolytic fragment of DELE1 that binds to the eIF2α kinase HRI to initiate integrated stress response (ISR) signaling. We report the cryo-electron microscopy structure of the C-terminal cleavage product of human DELE1, which assembles into a high-order oligomer. The oligomer consists of eight DELE1 monomers that assemble with D4 symmetry via two sets of hydrophobic inter-subunit interactions. We identified the key residues involved in DELE1 oligomerization, and confirmed their role in stabilizing the octamer in vitro and in cells using mutagenesis. We further show that assembly-impaired DELE1 mutants are compromised in their ability to induce HRI-dependent ISR activation in cell culture models. Together, our findings provide molecular insights into the activity of DELE1 and how it signals to promote ISR activity following mitochondrial insult.