Self-Healable, High-Stability Anode for Rechargeable Magnesium Batteries Realized by Graphene-Confined Gallium Metal.

In this work, a self-healable, high-stability anode material for rechargeable magnesium batteries (RMBs) has been developed by introducing a core-shell structure of Ga confined by reduced graphene oxide (Ga@rGO). Via this Ga@rGO anode, a specific capacity of 150 mAh g-1 at a current of 0.5 A g-1 stable up to 1200 cycles at room temperature and a specific capacity of 100 mAh g-1 under an ultrahigh current of 1 A g-1 stable up to 700 cycles at a slightly elevated temperature of 40 °C have been achieved. Additionally, the ultrahigh rate, high-cycling stability, and long-cycle life of the anode are attributed to the stabilized structure; such a low-cost, simple, and environmentally friendly direct drop coating (DDC) method is developed to maximize the original state of the active materials. Remarkably, the self-healing ability of anodes is still presented under the ultrahigh charging current. This anode is promising for the development of high rate and high stability RMBs.

Association between polycyclic aromatic hydrocarbon exposure and cognitive performance in older adults: a cross-sectional study from NHANES 2011-2014.

Background: polycyclic aromatic hydrocarbons (PAHs) are classified as neurotoxins, but the relationship between exposure to PAHs and cognition in adults is unclear, and their non-linear and mixed exposure association hasn't been explored. Objective: to evaluate the non-linear and joint association between co-exposure to PAHs and multiple cognitive tests in U.S. older people. Methods: restricted cubic spline (RCS) and Bayesian kernel machine regression (BKMR) were conducted to evaluate the non-linear and mixed exposure association, based on the cross-sectional data from NHANES 2011-2014: 772 participants over 60 years old, 4 cognitive test scores, including the Immediate Recall Test (IRT), Delayed Recall Test (DRT), Animal Fluency Test (AFT), and Digit Symbol Substitution test (DSST), and 5 urinary PAH metabolites. Results: a V-shaped nonlinear relationship was found between 3-hydroxyfluorene (3-FLUO), 2-hydroxyfluorene (2-FLUO), and DRT. Negative trends between mixed PAH exposure and IRT, DRT, and DSST scores were observed. 2-FLUO contributed the most to the negative association of multiple PAHs with IRT and DRT scores and 2-hydroxynaphthalene (2-NAP) played the most important role in the decreasing relationship between mixed PAH exposure and DSST scores. Conclusion: our study suggested that PAH exposure in the U.S. elderly might be related to their poor performances in IRT, DRT and DSST. Further prospective studies are needed to validate the association.

Alcohol intake exacerbates experimental autoimmune prostatitis through gut microbiota driving cholesterol biosynthesis-mediated Th17 differentiation.

BACKGROUND: Chronic prostatitis/chronic pelvic pain syndrome (CP/CPPS) is very common worldwide, and alcohol consumption is a notable contributing factor. Researches have shown that gut microbiota can be influenced by alcohol consumption and is an important mediator in regulating Th17 cell immunity. However, it is still unclear the exact mechanism by which alcohol exacerbates the CP/CPPS and the role of gut microbiota in this process. METHOD: We first constructed the most-commonly used animal model for CP/CPPS, the experimental autoimmune prostatitis (EAP) model, through immunoassay. Based on this, mice were divided into EAP group and alcohol-consuming EAP group. By 16S rRNA sequencing and non-targeted metabolomics analysis, differential gut microbiota and their metabolites between the two groups were identified. Subsequently, metabolomics detection targeting cholesterols was carried out to identify the exact difference in cholesterol. Furthermore, multiple methods such as flow cytometry and immunohistochemistry were used to detect the differentiation status of Th17 cells and severity of prostatitis treated with 27-hydroxycholesterol (the differential cholesterol) and its upstream regulatory factor-sterol regulatory element-binding protein 2 (SREBP2). Lastly, fecal transplantation was conducted to preliminary study on whether alcohol intake exacerbates EAP in immune receptor mice. RESULTS: Alcohol intake increased the proportion of Th17 cells and levels of related inflammatory factors. It also led to an altered gut bacterial richness and increased gut permeability. Further metabolomic analysis showed that there were significant differences in a variety of metabolites between EAP and alcohol-fed EAP mice. Metabolic pathway enrichment analysis showed that the pathways related to cholesterol synthesis and metabolism were significantly enriched, which was subsequently confirmed by detecting the expression of metabolic enzymes. By targeting cholesterol synthesis, 27-hydroxycholesterol was significantly increased in alcohol-fed EAP mice. Subsequent mechanistic research showed that supplementation with 27-hydroxycholesterol could aggravate EAP and promote Th17 cell differentiation both in vivo and in vitro, which is regulated by SREBP2. In addition, we observed that fecal transplantation from mice with alcohol intake aggravated EAP in immunized recipient mice fed a normal diet. CONCLUSION: Our study is the first to show that alcohol intake promotes Th17 cell differentiation and exacerbates EAP through microbiota-derived cholesterol biosynthesis.

Increased cTnI Predicts Early Death in Patients with Severe Fever with Thrombocytopenia: A Multicenter Study in North China.

Background: Myocardial injury is common in severe fever with thrombocytopenia syndrome (SFTS) patients. Currently, research on the prognostic value of cardiac troponin I (cTnI) for predicting the mortality of SFTS patients, especially death within 7 days is limited. Methods: Between May 2011 and October 2022, clinical and laboratory data on admission of consecutive SFTS cases were collected from six medical centres in China. The clinical endpoint was in-hospital all-cause death within seven days. Risk factors of myocardial injury and death were analysed using multivariable regression models. Prognostic models were established using Cox regression and performance of indicators was evaluated in terms of calibration, discrimination. Results: A total of 1379 laboratory-confirmed patients were enrolled, in which 686 subjects were included for analysis. The median age was 66 years, with 48.1% of male. Eighty-seven patients died within seven days and 396 patients diagnosed with myocardial injury during hospitalization. Non-survivors had significant higher levels of cardiac indices than survivors, including cTnI, aspartic transaminase (AST) and lactate dehydrogenase (LDH). Elevated levels of cTnI (HR = 1.058, 95% CI:1.032-1.085), AST (HR = 1.191, 95% CI:1.150-1.234) and LDH (HR = 1.019, 95% CI:1.009-1.029) predicted risk of early in-hospital mortality. cTnI model performed best, with area under curve of 0.850 (0.774-0.926) and concordance index of 0.842, respectively. Statistical differences were found between high and low levels of cTnI for mortality (P<0.001) using 0.35 ng/mL as the optimal cut-off. Conclusion: The risk of early in-hospital death can be predicted by cTnI. Clinical doctors should remind vigilant concerning the elevation of cardiac enzyme as soon as possible.

Targeting ONECUT2 inhibits tumor angiogenesis via down-regulating ZKSCAN3/VEGFA.

OC-2 plays a vital role in tumor growth, metastasis and angiogenesis, but molecular mechanism how OC-2 regulates angiogenic factors is unclear. We found that OC-2 was highly expressed in HepG2, COLO, MCF-7, SKOV3 cells and rectum carcinoma tissues, and angiogenic factors levels were positively related to OC-2. Then OC-2 KD inhibited the tumor growth, metastasis and angiogenesis process in vitro and vivo. ChIP-Seq showed that 228 target genes of OC-2 were identified and they were associated with tumor growth, metastasis, angiogenesis and signal transduction; OC-2 bound to ZKSCAN3 at promoter region. Luciferase assays showed that ZKSCAN3 was identified as target gene of OC-2 and VEGFA was identified as target gene of ZKSCAN3; OC-2 promoted VEGFA expression via activating ZKSCAN3 transcriptional program. Importantly, OC-2 KD down-regulated VEGFA secretion to suppress tumor angiogenesis of HUVECs. Besides VEGFA, OC-2 was positively correlated with other angiogenic factors HIF-1α, FGF2, EGFL6 and HGF. Meanwhile, ERK1/2 and Smad1 signaling pathways might be related to function of OC-2 driving tumor aggressiveness. We revealed that OC-2 might regulate tumor growth, metastasis, angiogenesis via ERK1/2, Smad1 signaling pathways and regulate VEGFA expression for tumor angiogenesis via activating ZKSCAN3 transcriptional program, indicating that OC-2 was a convincing target to develop novel anti-tumor drugs based on angiogenesis.

Thermodynamic Assessment of the P2O5-Na2O and P2O5-MgO Systems.

Knowledge about the thermodynamic equilibria of the P2O5-Na2O and P2O5-MgO systems is very important for controlling the phosphorus content of steel materials in the process of steelmaking dephosphorization. The phase equilibrium and thermodynamic data of the P2O5-Na2O and P2O5-MgO systems were critically evaluated and re-assessed by the CALPHAD (CAlculation of PHAse Diagram) approach. The liquid phase was described by the ionic two-sublattice model for the first time with the formulas (Na+1)P(O-2, PO3-1, PO4-3, PO5/2)Q and (Mg+2)P(O-2, PO3-1, PO4-3, PO5/2)Q, respectively, and the selection of the species constituting the liquid phase was based on the structure of the phosphate melts. A new and improved self-consistent set of thermodynamic parameters for the P2O5-Na2O and P2O5-MgO systems was finally obtained, and the calculated phase diagram and thermodynamic properties exhibited excellent agreement with the experimental data. The difference in the phase composition of invariant reactions from the experimentally determined values reported in the literature is less than 0.9 mol.%. The present thermodynamic modeling contributes to constructing a multicomponent oxide thermodynamic database in the process of steelmaking dephosphorization.

Diffusivities and Atomic Mobilities in BCC Ti-Fe-Cr Alloys.

In this research, the diffusion behaviors within the Ti-Fe-Cr ternary system were examined at the temperatures of 1273 K and 1373 K through the diffusion couple technique. This study led to the determination of both ternary inter-diffusion and impurity diffusion coefficients in the body-centered cubic (bcc) phase for the Ti-Fe-Cr alloy, utilizing the Whittle-Green and Hall methods. The statistics show that the average diffusion coefficients D˜FeFeTi and D˜CrCrTi measured at 1273 K were 1.34 × 10-12 and 3.66 × 10-13, respectively. At 1373 K, the average values of D˜FeFeTi and D˜CrCrTi were 4.89 × 10-12 and 1.43 × 10-12. By adopting the CALPHAD method, a self-consistent database for atomic mobility in the bcc phase of the Ti-Fe-Cr system was established. This database underwent refinement by comparing the newly acquired diffusion coefficients with data from the existing literature. Diffusion simulations for the diffusion couples were performed, drawing on the established database. The error between the simulated diffusion coefficient and the experimental measurement data is within 15%, and the simulated data of the component distance distribution and diffusion path are in good agreement with the experimental data. The simulations generated results that aligned well with the observed experimental diffusion characteristics, thereby affirming the reliability and accuracy of the database.

A Morpholine Derivative N-(4-Morpholinomethylene)ethanesulfonamide Induces Ferroptosis in Tumor Cells by Targeting NRF2.

Small molecule drugs containing morpholine-based moieties have become crucial candidates in the tumor targeted therapy strategies, but the specific molecular mechanisms of these drugs causing tumor cell death require further investigation. The morpholine derivative N-(4-morpholinomethylene)ethanesulfonamide (MESA) was used to stimulate prostate and ovarian cancer cells and we focused on the ferroptosis effects, including the target molecule and signal pathways mediated by MESA. The results showed that MESA could induce ferroptosis to cause the proliferation inhibition and apoptosis effects of tumor cells according to the identification of ferroptosis inhibitor fer-1 and other cell death inhibitors. Further MESA could significantly increase the intracellular malondialdehyde (MDA), reactive oxygen species (ROS) and Fe2+ levels in tumor cells and mediate the dynamic changes of ferroptosis-relative molecules GPX4, nuclear factor erythroid2-related factor 2 (NRF2), ACSL4, SLC7A11 and P62-Kelch-like ECH-associated protein 1 (KEAP1)-NRF2-antioxidant response element (ARE) signal pathways. Further, NRF2 overexpression could reduce the tumor cell death and ROS levels exposure to MESA. Most importantly, it was confirmed that MESA could bind to NRF2 protein through molecular docking and thermal stability assays and NRF2 was a target molecule of MESA for inducing ferroptosis effects in tumor cells. Collectively, our findings indicated the ferroptosis effects of the morpholine derivative MESA in prostate and ovarian cancer cells and its function mechanism including targeted molecule and signal pathways, which would be helpful for developing MESA as a prospective small molecule drug for cancer therapy based on cell ferroptosis.

miR-129-2-3p mediates LPS-induced macrophage polarization and ferroptosis by targeting the SMAD3-GPX4 axis.

Macrophages has become a promising target of sepsis treatment because macrophages dysfunction contributes to the progress of sepsis. The targeted therapy of sepsis based on macrophages ferroptosis is drawing more and more attention, but the molecular mechanism involved is poorly understood. In this study, Mus musculus-derived macrophages were used for in-vitro experiments. We found that LPS could induce ferroptosis in macrophages via the detection of apoptosis, GSH, lipid peroxide and GPX4 levels. Meanwhile, miR-129-2-3p was up-regulated in macrophages exposure to LPS. Next, we confirmed that miR-129-2-3p promoted the LPS-induced polarization of M1 phenotype in macrophages via the detection of Arg-1 and iNOS levels; miR-129-2-3p promoted the LPS-induced ferroptosis in macrophages. Further, luciferase assay showed that SMAD3 was identified as a target gene of miR-129-2-3p and its expression was negatively regulated by miR-129-2-3p and LPS. SMAD3 could inhibit the LPS-induced polarization of M1 phenotype and ferroptosis in macrophages by targeting GPX4. Collectively, we demonstrated the target gene and molecular mechanism of miR-129-2-3p mediating LPS-induced polarization and ferroptosis in macrophages by targeting the SMAD3-GPX4 axis, which would provide a novel strategy for sepsis targeted therapy based on macrophages polarization and ferroptosis.

Efficient Pt/KFI zeolite catalysts for the selective catalytic reduction of NOx by hydrogen.

Aiming at purification of NOx from hydrogen internal combustion engines (HICEs), the hydrogen selective catalytic reduction (H2-SCR) reaction was investigated over a series of Pt/KFI zeolite catalysts. H2 can readily reduce NOx to N2 and N2O while O2 inhibited the deNOx efficiency by consuming the reductant H2. The Pt/KFI zeolite catalysts with Pt loading below 0.1 wt.% are optimized H2-SCR catalysts due to its suitable operation temperature window since high Pt loading favors the H2-O2 reaction which lead to the insufficient of reactants. Compared to metal Pt0 species, Ptδ+ species showed lower activation energy of H2-SCR reaction and thought to be as reasonable active sites. Further, Eley-Rideal (E-R) reaction mechanism was proposed as evidenced by the reaction orders in kinetic studies. Last, the optimized reactor was designed with hybrid Pt/KFI catalysts with various Pt loading which achieve a high NOx conversion in a wide temperature range.

An online soft sensor method for biochemical reaction process based on JS-ISSA-XGBoost.

BACKGROUND: A method combining offline techniques and the just-in-time learning strategy (JITL) is proposed, because the biochemical reaction process often encounters changing features and parameters over time. METHODS: Firstly, multiple sub-databases in the fermentation process are constructed offline by an improved fuzzy C-means algorithm and the sample data are adaptively pruned by a similarity query threshold. Secondly, an improved eXtreme Gradient Boosting (XGBoost) method is used on the online modeling stage to build soft sensor models, and the multi-similarity-driven just-in-time learning strategy is used to increase the diversity of the model. Finally, to improve the generalization of the whole algorithm, the output of the base learner is fused by an improved Stacking integration model and then the predictive output is performed. RESULTS: Applying the constructed soft sensor model to the problem of predicting cell concentration and product concentration in Pichia pastoris fermentation process. The experimental results show that the root mean square error of the cell concentration is 0.0260, the coefficient of determination is 0.9945, the root mean square error of the product concentration is 2.6688, and the coefficient of determination is 0.9970. It shows that the proposed method has the advantages of timely prediction and high prediction accuracy, which validates the effectiveness and practicality of the method. CONCLUSION: The JS-ISSA-XGBoost is an extensive and excellent soft measurement model that meets the practical needs for real-time monitoring of parameters and prediction of control in biochemical reactions.

Establishing entrustable professional activities for psychiatry residents in China.

PURPOSE: The authors established entrustable professional activities for psychiatry residents in China. METHODS: The authors conducted a literature research and two expert consultation rounds following the Delphi method in 2022 to screen and optimize entrustable professional activities for psychiatry residents. RESULTS: The effective questionnaire recovery rate in the two consultation rounds was 100% (44/44). The expert authority coefficients of the first and second consultation rounds were 0.861 and 0.881, respectively. The Kendall harmony coefficients of the first and second expert consultation rounds were 0.279 (χ2 = 405.43, P < .001) and 0.389 (χ2 = 3456.83, P < .001), respectively. The arithmetic means of the various indicators' evaluation results in the two consultation rounds ranged between 3.61 and 4.93, and the full score rates were between 13.6% and 93.2%. The authors established 17 entrustable professional activities for psychiatry residents and their contents with phase-based modularization and formulated the entrustable level of each at various stages. CONCLUSIONS: Combined with standardized psychiatry training characteristics, the authors preliminarily established phase-specific and modular entrustable professional activities for psychiatry residents. The formulated entrustable professional activities are suitable for the practice and clinical environment of standardized psychiatry training in China. The devised system has good observability and measurability and provides a simple and feasible competency evaluation method for standardized psychiatry resident training.

New Frontiers in the Phase Structure and Functional Properties of Materials.

Materials' functional properties are strongly related to their phase structures [...].

Functionalization of pine kernel protein by pH-shifting combined with ultrasound treatments: Further improvement with increasing acidity.

As a novel plant protein, developing various aspects of pine kernel protein (PKP) functionality is essential to meet the demand for protein-rich foods. To achieve this, the PKP was functionalized by a combination of pH-shifting and ultrasound techniques. The solubility, emulsification and droplet stability of the PKP in the pH range suitable to food (pH 3 to 7) were further investigated. The pH 12-shifting was an effective strategy to increase the solubility of PKP under extreme acidic and neutral conditions, characterized by a higher content of ß-sheets and random coils, a greater exposure of free sulfhydryl and hydrophobic groups. Furthermore, appropriate ultrasonic power (250 W) further improved the solubility of PKPs by disrupting intermolecular hydrogen and hydrophobic bonds. As the ambient acidity increased, the emulsions exhibited higher viscoelasticity and stronger protein interactions. Especially at pH 3, the oil droplets stabilized by U250-PKP-12 (PKP treated with 250 W ultrasound-assisted pH 12-shifting) were homogeneously dispersed and surrounded by dense protein, maintaining small particle size and large electrostatic repulsion, and there was no apparent creaming or phase separation in the emulsions after 10 days of storage. Thus, the functionality of PKP after pH-shifting combined with ultrasonic treatments is further enhanced by increasing the environmental acidity.

The Role of ElastPQ in Assessing Liver Stiffness for Non-Alcoholic Fatty Liver Disease in Patients Treated with Atypical Antipsychotic Drugs.

Objective: To evaluate the role of elastography point quantification (ElastPQ) for the quantitative assessment of stiffness in the fatty liver disease in mental disorder patients and to provide a noninvasive detection method for non-alcoholic fatty liver (NAFLD) caused by atypical antipsychotics drugs (AAPDs). Methods: A total number of 168 mental disorder patients treated with AAPDs and 58 healthy volunteers were enrolled in this study. All the subjects underwent ultrasound and ElastPQ tests. The basic data of the patients were analyzed. Results: BMI, liver function, and the value of ElastPQ were considerably higher in the patient group than that in the healthy volunteers. The values of liver stiffness obtained by ElastPQ were increased gradually from 3.48(3.14-3.81) kPa in the normal liver to 8.15(6.44-9.88) in the severe fatty liver. The receiver operating characteristic (ROC) for the diagnosis of fatty liver with ElastPQ were 0.85, 0.79, 0.80, and 0.87 for the diagnosis of normal, mild, moderate, and severe steatosis, respectively, with a sensitive/specificity of 79%/76.4%, 85.7%/78.3%, 86.2%/73%, and 81.3%/82.1%, correspondingly. Moreover, ElastPQ in the olanzapine group was higher than those in the risperidone and aripiprazole groups (5.11(3.83-5.61) kPa vs 4.35(3.63-4.98) kPa, P < 0.05; 5.11(3.83-5.61) kPa vs 4.79(4.18-5.24) kPa, P < 0.05). After one-year treatment, the value of ElastPQ was 4.43(3.85-5.22) kPa, but it was 5.81(5.09-7.33) kPa in patients treated for more than three years. This value increased with treatment prolongation (P < 0.05). Conclusion: ElastPQ is a real-time, quantitative method for assessing the stiffness of NAFLD. The liver stiffness value could be varied in the different stages of fatty liver. Olanzapine has a considerable influence on liver stiffness. The long-term use of AAPDs can increase the stiffness value of fatty liver.

Feasibility and safety of bipolar-plasmakinetic transurethral enucleation and resection of the prostate in day surgery mode. / åŒæžç­‰ç¦»å­ç»å°¿é“å‰åˆ—è ºå‰œåˆ‡æ—¥é—´æ‰‹æœ¯ä¸´åºŠå®žè·µ.

OBJECTIVES: To evaluate the feasibility and safety of bipolar-plasmakinetic transurethral enucleation and resection of the prostate (B-TUERP) in day surgery. METHODS: From January 2021 to August 2022, 34 patients with benign prostatic hyperplasia (BPH) underwent B-TUERP in day surgery in the First Affiliated Hospital of Anhui Medical University. Patients completed the screening and anesthesia evaluation before admission and received the standard surgery which implements "anatomical enucleation of the prostate" and "absolute bleeding control" on the same day of admission, and by the same doctor. Bladder irrigation was stopped, catheter was removed and the discharge evaluation was performed on the first day after operation. The baseline data, perioperative conditions, time of recovery, treatment outcomes, hospitalization costs, and postoperative complications were analyzed. RESULTS: All operations were successfully conducted. The average age of the patients was (62.2±7.8) years, average prostate volume was (50.2±29.3) mL. The average operation time was (36.5±19.1) min, the average hemoglobin and blood sodium were decreased by (16.2±7.1) g/L and (2.2±2.0) mmol/L, respectively. The average postoperative length of hospital stay, and total length of hospital stay were (17.7±2.2) and (20.8±2.1) h, respectively, and the average hospitalization cost was (13 558±2320) CNY. All patients were discharged on the day after surgery except for one patient who was transferred to a general ward. Three patients received indwelling catheterization after catheter removal. The 3-month follow-up results showed a substantial improvement in the International Prostate Symptom Score, quality of life score and maximum urinary flow rate (all P<0.01). Three patients experienced temporary urinary incontinence, 1 patient experienced urinary tract infection, 4 patients were diagnosed with urethral stricture and 2 patients experienced bladder neck contracture. No complications above Clavien grade Ⅱ occurred. CONCLUSIONS: The preliminary results showed that B-TUERP ambulatory surgery is a safe, feasible, economical and effective treatment for appropriately selected patients with BPH.

Elastin-like recombinamer-mediated hierarchical mineralization coatings on Zr-16Nb-xTi (x = 4,16 wt%) alloy surfaces improve biocompatibility.

The biocompatibility of biomedical materials is vital to their applicability and functionality. However, modifying surfaces for enhanced biocompatibility using traditional surface treatment techniques is challenging. We employed a mineralizing elastin-like recombinamer (ELR) self-assembling platform to mediate mineralization on Zr-16Nb-xTi (x = 4,16 wt%) alloy surfaces, resulting in the modification of surface morphology and bioactivity while improving the biocompatibility of the material. We modulated the level of nanocrystal organization by adjusting the cross-linker ratio. Nanoindentation tests revealed that the mineralized configuration had nonuniformity with respect to Young's modulus and hardness, with the center areas having higher values (5.626 ± 0.109 GPa and 0.264 ± 0.022 GPa) compared to the edges (4.282 ± 0.327 GPa and 0.143 ± 0.023 GPa). The Scratch test results indicated high bonding strength (2.668 ± 0.117 N) between the mineralized coating and the substrate. Mineralized Zr-16Nb-xTi (x = 4,16 wt%) alloys had higher viability compared to untreated alloys, which exhibited high cell viability (>100 %) after 5 days and high alkaline phosphatase activity after 7 days. Cell proliferation assays indicated that MG 63 cells grew faster on mineralized surfaces than on untreated surfaces. Scanning electron microscopy imaging confirmed that the cells adhered and spread well on mineralized surfaces. Furthermore, hemocompatibility test results revealed that all mineralized samples were non-hemolytic. Our results demonstrate the viability of employing the ELR mineralizing platform to improve alloy biocompatibility.

Production of MSTN knockout porcine cells using adenine base-editing-mediated exon skipping.

Gene-knockout pigs have important applications in agriculture and medicine. Compared with CRISPR/Cas9 and cytosine base editing (CBE) technologies, adenine base editing (ABE) shows better safety and accuracy in gene modification. However, because of the characteristics of gene sequences, the ABE system cannot be widely used in gene knockout. Alternative splicing of mRNA is an important biological mechanism in eukaryotes for the formation of proteins with different functional activities. The splicing apparatus recognizes conserved sequences of the 5' end splice donor and 3' end splice acceptor motifs of introns in pre-mRNA that can trigger exon skipping, leading to the production of new functional proteins, or causing gene inactivation through frameshift mutations. This study aimed to construct a MSTN knockout pig by inducing exon skipping with the aid of the ABE system to expand the application of the ABE system for the preparation of knockout pigs. In this study, first, we constructed ABEmaxAW and ABE8eV106W plasmid vectors and found that their editing efficiencies at the targets were at least sixfold and even 260-fold higher than that of ABEmaxAW by contrasting the editing efficiencies at the gene targets of endogenous CD163, IGF2, and MSTN in pigs. Subsequently, we used the ABE8eV106W system to realize adenine base (the base of the antisense strand is thymine) editing of the conserved splice donor sequence (5'-GT) of intron 2 of the porcine MSTN gene. A porcine single-cell clone carrying a homozygous mutation (5'-GC) in the conserved sequence (5'-GT) of the intron 2 splice donor of the MSTN gene was successfully generated after drug selection. Unfortunately, the MSTN gene was not expressed and, therefore, could not be characterized at this level. No detectable genomic off-target edits were identified by Sanger sequencing. In this study, we verified that the ABE8eV106W vector had higher editing efficiency and could expand the editing scope of ABE. Additionally, we successfully achieved the precise modification of the alternative splice acceptor of intron 2 of the porcine MSTN gene, which may provide a new strategy for gene knockout in pigs.

Microstructure and Mechanical Properties of Y4Zr3O12-Added Fe-13.5Cr-2W Oxide-Dispersion-Strengthened Steels, Containing High Contents of C and N, Prepared by Mechanical Alloying and Two-Step Spark Plasma Sintering.

Oxide-dispersion-strengthened (ODS) steel is considered as a promising candidate structural material for nuclear applications. In this study, the microstructure and mechanical properties of Y4Zr3O12-added Fe-13.5Cr-2W ODS steels, containing high contents of C and N, prepared by mechanical alloying (MA) and two-step spark plasma sintering (SPS), were investigated. The results showed that pure Y4Zr3O12 powders, with a grain size of 3.5 nm, were well prepared with NH3·H2O addition by the sol-gel method in advance, in order to avoid the formation of some coarse or undesired oxides. W was completely dissolved into the matrix after 48 h of ball milling at 300 rpm, and the main elements were uniformly distributed on the surface of the milled powders. The unexpected face-centered cubic (FCC, γ)/body-centered cubic (BCC, α) dual-phase structure of the sintered specimens, could be explained by the unexpectedly high contents of C and N from the raw powder production process, fast-sintering characteristic of SPS, and inhibitory effect of W on the diffusion of C. The experimental results were approximately consistent with the simulation results from the Thermo Calc software. The temperature combination of 800 °C and 1100 °C during the SPS process, provided a relatively more homogeneous microstructure, while the combination of 750 °C and 1150 °C, provided the highest ultimate tensile strength (UTS), of 1038 MPa, with the highest uniform elongation (UE), of 6.2%. M23C6, Cr2O3, M2(C,N), and other precipitates, were mainly distributed at grain boundaries, especially at the triple junctions, which led to Cr depletion at grain boundaries.