Objective To explore the construction of α-1,3-galactosyltransferase (GGTA1) gene-knockout (GTKO) Diannan miniature pigs and the kidney xenotransplantation from pigs to rhesus macaques, and to assess the effectiveness of GTKO pigs. Methods The GTKO Diannan miniature pigs were constructed using the CRISPR/Cas9 gene-editing system and somatic cell cloning technology. The phenotype of GTKO pigs was verified through polymerase chain reaction, Sanger sequencing and immunofluorescence staining. Flow cytometry was used to detect antigen-antibody (IgM) binding and complement-dependent cytotoxicity. Kidney xenotransplantation was performed from GTKO pigs to rhesus macaques. The humoral immunity, cellular immunity, coagulation and physiological indicators of the recipient monkeys were monitored. The function and pathological changes of the transplanted kidneys were analyzed using ultrasonography, hematoxylin-eosin staining, immunohistochemical staining and immunofluorescence staining. Results Single-guide RNA (sgRNA) targeting exon 4 of the GGTA1 gene in Diannan miniature pigs was designed. The pGL3-GGTA1-sgRNA1-GFP vector was transfected into fetal fibroblasts of Diannan miniature pigs. After puromycin selection, two cell clones, C59# and C89#, were identified as GGTA1 gene-knockout clones. These clones were expanded to form cell lines, which were used as donor cells for somatic cell nuclear transfer. The reconstructed embryos were transferred into the oviducts of trihybrid surrogate sows, resulting in 13 fetal pigs. Among them, fetuses F04 and F11 exhibited biallelic mutations in the GGTA1 gene, and F04 had a normal karyotype. Using this GTKO fetal pig for recloning and transferring the reconstructed embryos into the oviducts of trihybrid surrogate sows, seven surviving piglets were obtained, all of which did not express α-Gal epitope. The binding of IgM from the serum of rhesus monkey 20# to GTKO pig PBMC was reduced, and the survival rate of GTKO pig PBMC in the complement-dependent cytotoxicity assay was higher than that of wild-type pig. GTKO pig kidneys were harvested and perfused until completely white. After the left kidney of the recipient monkey was removed, the pig kidney was heterotopically transplanted. Following vascular anastomosis and blood flow restoration, the pig kidney rapidly turned pink without hyperacute rejection (HAR). Urine appeared in the ureter 6 minutes later, indicating successful kidney transplantation. The right kidney of the recipient was then removed. Seven days after transplantation, the transplanted kidney had good blood flow, the recipient monkey's serum creatinine level was stable, and serum potassium and cystatin C levels were effectively controlled, although they increased 10 days after transplantation. Seven days after transplantation, the levels of white blood cells, lymphocytes, monocytes and eosinophils in the recipient monkey increased, while platelet count and fibrinogen levels decreased. The activated partial thromboplastin time, thrombin time and prothrombin time remained relatively stable but later showed an upward trend. The recipient monkey survived for 10 days. At autopsy, the transplanted kidney was found to be congested, swollen and necrotic, with a small amount of IgG deposition in the renal tissue, and a large amount of IgM, complement C3c and C4d deposition, as well as CD68⁺ macrophage infiltration. Conclusions The kidneys of GTKO Diannan miniature pigs may maintain normal renal function for a certain period in rhesus macaques and effectively overcome HAR, confirming the effectiveness of GTKO pigs for xenotransplantation.

Lipid droplets (LDs) are dynamic organelles that are ubiquitous across most organisms, including animals, plants, protists, and microorganisms. Their core consists of neutral lipids, surrounded by a phospholipid monolayer adorned with a specific set of proteins. As critical intracellular hubs of metabolic regulation, lipid droplets play essential roles in maintaining physiological homeostasis and contributing to the progression of various pathological processes. They store neutral lipids for energy production during periods of starvation or for membrane biosynthesis, and they sequester fatty acids to mitigate lipotoxicity. Clinically, dysregulation of lipid droplet function is associated with a wide range of diseases, including metabolic dysfunction-associated steatotic liver disease (MASLD), obesity, type 2 diabetes mellitus (T2DM), neurodegenerative disorders, and cancer. Research into the biological functions of lipid droplets—as dynamic organelles and their links to multiple diseases—has emerged as a cutting-edge focus in cell biology. In recent years, significant advances have been made in understanding lipid droplet biogenesis. Researchers have developed a more refined framework that elucidates how LDs are assembled in the endoplasmic reticulum (ER). Triacylglycerols and sterol esters are synthesized between the inner and outer leaflets of the ER bilayer, and when they exceed the critical nucleation concentration (CNC), they coalesce to form neutral lipid lenses. These then bud from the ER under the coordinated action of key proteins such as Seipin, fat storage-inducing transmembrane protein 2 (FIT2), and the peroxisomal membrane protein Pex30. This budding process is driven by changes in membrane curvature and surface tension, induced by the asymmetric distribution of phospholipids. Nascent lipid droplets recruit lipid-synthesizing enzymes via ER-LD bridging structures, enabling localized lipid production and surface expansion, ultimately resulting in the formation of mature LDs. Biochemical and biophysical approaches have revealed important features of this process, underscoring the critical roles of ER membrane biophysical properties and specific phospholipids. Structural biology and proteomic studies have identified key regulators—particularly Seipin and FIT2—as central players in LD biogenesis. This review systematically summarizes recent advances in the molecular mechanisms of LD biogenesis. It delves into the processes of LD nucleation, membrane budding, and expansion in eukaryotic cells, with a special focus on how core factors such as Seipin and FIT2 dynamically regulate LD morphology. In addition, it examines the mechanisms and pathways by which class I and class II proteins are targeted to LDs, compares LD biogenesis involving different neutral lipid cores, and discusses the disease relevance of specific regulatory proteins. Finally, the review outlines critical unresolved questions in the field of LD biogenesis, offering clear directions for future research and providing a comprehensive framework for deepening our understanding of LD formation and its implications for disease intervention.

OBJECTIVE: To investigate effectiveness of the bridge combined fixation system (BCFS) for Bado typeⅠchronic Monteggia fractures (CMF) in children. METHODS: A clinical data of 8 children with Bado type ⅠCMF, who were treated with the BCFS between November 2023 and February 2025, was retrospectively analyzed. There were 6 boys and 2 girls, with a mean age of 7.0 years (range, 4-12 years). The time from injury to operation ranged from 29 to 370 days (median, 68.5 days). Preoperative elbow range of motion was (111.3±17.9)° in flexion, (13.1±13.9)° in extension, (71.9±14.6)° in pronation, and (75.6±13.5)° in supination. Fracture healing time and postoperative complications were observed, and clinical outcomes were evaluated using the Mayo elbow performance score. RESULTS: All incisions healed by primary intention without infection, non-healing of the incision, or iatrogenic nerve injury. All children were followed up 4-18 months (mean, 10.3 months). At last follow-up, the elbow range of motion significantly improved to (142.5±2.7)° in flexion, (2.5±2.7)° in extension, (87.5±2.7)° in pronation, and (88.8±2.3)° in supination ( P<0.05). According to the postoperative Mayo elbow performance score, all cases were rated as excellent. Radiographic review showed no radial head dislocation, nonunion at the ulnar osteotomy site, or elbow stiffness, and no breakage of the BCFS or screw loosening. The fracture healing time ranged from 3 to 6 months, with a median of 4 months. CONCLUSION The BCFS was confirmed to be effective in the treatment of pediatric Bado type Ⅰ CMF, with good restoration of elbow function and the advantage of avoiding secondary implant removal surgery.
Humans ; Child ; Monteggia's Fracture/surgery* ; Male ; Female ; Child, Preschool ; Range of Motion, Articular ; Retrospective Studies ; Fracture Fixation, Internal/instrumentation* ; Elbow Joint/physiopathology* ; Bone Plates ; Treatment Outcome ; Fracture Healing ; Bone Screws ; Elbow Injuries

BACKGROUND: Lung adenocarcinoma is an important pathohistologic subtype of non-small cell lung cancer (NSCLC). Invasive non-mucinous pulmonary adenocarcinomas (INMA) tend to have a poor prognosis due to their significant heterogeneity and diverse histologic components. Establishing a histologic grading system for INMA is crucial for evaluating its malignancy. In 2021, the International Association for the Study of Lung Cancer (IASLC) proposed that a new histological grading system could better stratify the prognosis of INMA patients. The aim of this study was to establish a visualized nomogram model to predict INMA IASLC grading preoperatively by means of dual-energy computed tomography (DECT), fractal dimension (FD), clinical features and conventional CT parameters. METHODS: A total of 112 patients with INMA who underwent preoperative DECT were retrospectively enrolled from March 2021 to January 2025. Patients were categorized into low-intermediate grade and high grade groups based on IASLC grading. The clinical characteristics and conventional CT parameters, including baseline features, biochemical markers, and serum tumor markers, were collected. DECT-derived parameters, including iodine concentration (IC), effective atomic number (eff-Z), and normalized IC (NIC), were collected and determined as NIC ratio (NICr) and fractal dimension (FD). Univariate analysis was employed to compare differences in conventional characteristics and DECT parameters between the two groups. Variables demonstrating statistical significance were subsequently incorporated into a multivariate Logistic regression analysis. A nomogram model integrating clinical data, conventional CT parameters, and DECT parameters was developed to identify independent predictors for IASLC grading of INMA. The discriminatory performance of the model was evaluated using receiver operating characteristic (ROC) curve analysis. RESULTS: Multivariate analysis identified smoking history [odds ratio (OR)=2.848, P=0.041], lobulation sign (OR=2.163, P=0.004), air bronchogram (OR=7.833, P=0.005), eff-Z in arterial phase (OR=4.266, P<0.001), and IC in arterial phase (OR=1.290, P=0.012) as independent and significant predictors for IASLC grading of INMA. The nomogram model constructed based on these indicators demonstrated optimal predictive performance, achieving an area under the curve (AUC) of 0.804 (95%CI: 0.725-0.883), with specificity and sensitivity of 85.3% and 65.7%, respectively. CONCLUSIONS The nomogram model based on clinical features, imaging features and spectral CT parameters have a large potential for application in the preoperative noninvasive assessment of INMA IASLC grading.
Humans ; Nomograms ; Female ; Male ; Middle Aged ; Tomography, X-Ray Computed/methods* ; Lung Neoplasms/pathology* ; Aged ; Retrospective Studies ; Adenocarcinoma of Lung/pathology* ; Neoplasm Grading ; Adult

Traditional development of small protein scaffolds has relied on display technologies and mutation-based engineering, which limit sequence and functional diversity, thereby constraining their therapeutic and application potential. Protein design tools have significantly advanced the creation of novel protein sequences, structures, and functions. However, further improvements in design strategies are still needed to more efficiently optimize the functional performance of protein-based drugs and enhance their druggability. Here, we extended an evolution-based design protocol to create a novel minibinder, BindHer, against the human epidermal growth factor receptor 2 (HER2). It not only exhibits super stability and binding selectivity but also demonstrates remarkable properties in tissue specificity. Radiolabeling experiments with ^99mTc, ⁶⁸Ga, and ¹⁸F revealed that BindHer efficiently targets tumors in HER2-positive breast cancer mouse models, with minimal nonspecific liver absorption, outperforming scaffolds designed through traditional engineering. These findings highlight a new rational approach to automated protein design, offering significant potential for large-scale applications in therapeutic mini-protein development.

Extensive epigenetic reprogramming involves in memory CD8+ T-cell differentiation. The elaborate epigenetic rewiring underlying the heterogeneous functional states of CD8+ T cells remains hidden. Here, we profile single-cell chromatin accessibility and map enhancer-promoter interactomes to characterize the differentiation trajectory of memory CD8+ T cells. We reveal that under distinct epigenetic regulations, the early activated CD8+ T cells divergently originated for short-lived effector and memory precursor effector cells. We also uncover a defined epigenetic rewiring leading to the conversion from effector memory to central memory cells during memory formation. Additionally, we illustrate chromatin regulatory mechanisms underlying long-lasting versus transient transcription regulation during memory differentiation. Finally, we confirm the essential roles of Sox4 and Nrf2 in developing memory precursor effector and effector memory cells, respectively, and validate cell state-specific enhancers in regulating Il7r using CRISPR-Cas9. Our data pave the way for understanding the mechanism underlying epigenetic memory formation in CD8+ T-cell differentiation.
CD8-Positive T-Lymphocytes/metabolism* ; Cell Differentiation ; Chromatin/immunology* ; Animals ; Mice ; Immunologic Memory ; Epigenesis, Genetic ; SOXC Transcription Factors/immunology* ; NF-E2-Related Factor 2/immunology* ; Mice, Inbred C57BL ; Gene Regulatory Networks ; Enhancer Elements, Genetic

Accurate prediction of drug-target interactions (DTIs) plays a pivotal role in drug discovery, facilitating optimization of lead compounds, drug repurposing and elucidation of drug side effects. However, traditional DTI prediction methods are often limited by incomplete biological data and insufficient representation of protein features. In this study, we proposed KG-CNNDTI, a novel knowledge graph-enhanced framework for DTI prediction, which integrates heterogeneous biological information to improve model generalizability and predictive performance. The proposed model utilized protein embeddings derived from a biomedical knowledge graph via the Node2Vec algorithm, which were further enriched with contextualized sequence representations obtained from ProteinBERT. For compound representation, multiple molecular fingerprint schemes alongside the Uni-Mol pre-trained model were evaluated. The fused representations served as inputs to both classical machine learning models and a convolutional neural network-based predictor. Experimental evaluations across benchmark datasets demonstrated that KG-CNNDTI achieved superior performance compared to state-of-the-art methods, particularly in terms of Precision, Recall, F1-Score and area under the precision-recall curve (AUPR). Ablation analysis highlighted the substantial contribution of knowledge graph-derived features. Moreover, KG-CNNDTI was employed for virtual screening of natural products against Alzheimer's disease, resulting in 40 candidate compounds. 5 were supported by literature evidence, among which 3 were further validated in vitro assays.
Alzheimer Disease/drug therapy* ; Biological Products/therapeutic use* ; Humans ; Neural Networks, Computer ; Machine Learning ; Drug Discovery/methods* ; Algorithms ; Drug Evaluation, Preclinical/methods*

Objective To investigate the relationship between sleep disorders and coronary heart disease through big data combined with Mendelian randomization analysis.Methods Data from 2005 to 2018 National Health and Nutrition Examination Survey in the United States were utilized.Logistic regression analysis was employed to evaluate the association between sleep disorders and coronary heart disease,while analyzing relevant influencing factors.A two-sample Mendelian randomization approach was implemented using Genome-Wide Association Studies to establish causal relationships.Results Logistic regression analysis demonstrated a significant association between sleep disorders and coronary heart disease(P＜0.001),with the neutrophil-to-lymphocyte ratio serving as a mediating factor in this relationship(P＜0.001).Mendelian randomization analysis revealed a positive correlation between sleep disorders and coronary heart disease(OR=1.030,95％CI:1.01-1.04).Conclusion Sleep disorders can increase the risk of coronary heart disease by activating inflammatory factors.

Objective:To investigate the correlation between frailty and telomere length in the elderly.Methods:Cross-sectional study.A total of 128 elderly patients aged ≥65 years who were hospitalized in the Second Hospital of Shanxi Medical University from January to June 2024 were selected.Frailty assessment was conducted using the Edmonton Scale, and patients were divided into the non-frail group(n=64)and the frail group(n=64)based on frailty score.The telomere length of peripheral blood leukocyte was detected by real-time fluorescent quantitative polymerase chain reaction(qPCR), and the correlation analysis between frailty and telomere length was conducted.Results:Compared with the non-frailty group, the frailty group had older age, higher body mass index(BMI), higher American Society of Anesthesiologists(ASA)grade, an increased proportion of diabetes and hypertension, and shorter telomere length( P<0.05). Logistic regression analysis: ageing, increased BMI, hypertension and diabetes were risk factors for frailty in the elderly( OR=1.280, 1.135, 1.543, 1.081, P=0.001, 0.036, 0.010, 0.021), while ASA grade and telomere length were not related to the occurrence of frailty.After stratification, frailty scores were weakly correlated with telomere length in the age ≤73 years old ( r=-0.344, P=0.005)and the BMI ≥24 kg/m 2( r=-0.336, P=0.001). The correlation between frailty scores and telomere length was strengthened in the poorly controlled hypertension group( r=-0.571, P=0.042)and the group with poorly controlled diabetes( r=-0.613, P=0.045)showed a stronger correlation between frailty scores and telomere length. Conclusions:In the overall analysis, there was no definitive association between telomere length and frailty.After stratification, it was found that telomere length was weakly to moderately correlated with frailty, suggesting that telomere length may interact with other factors, and strong risk factors such as age, BMI, hypertension and diabetes may mask the weak effect of telomere length on frailty.