Probiotics are natural systems bridging synthetic biology, physical biotechnology, and immunology, initiating innate and adaptive anti-tumor immune activity. We previously constructed an all-in-one engineered food-grade probiotic Lactococcus lactis (FOLactis) which could boost the crosstalk among different immune cells such as dendritic cells (DCs), natural killer cells, and T cells. Herein, considering the limited clinical efficacy of naked personalized neoantigen peptide vaccines, we decorate FOLactis with tumor antigens by employing a Plug-and-Display system comprising membrane-inserted peptides. Intranodal injection of FOLactis coated with neoantigen peptides (Ag-FOLactis) induces robust DCs presentation and neoantigen-specific cellular immunity. Notably, Ag-FOLactis not only triggers a 45-fold rise in the quantity of locally reactive neoantigen-specific T cells but also induces epitope spreading in both subcutaneous and metastatic tumor-bearing models, leading to potent inhibition of tumor growth. These findings imply that Ag-FOLactis represents a powerful platform to rapidly and easily display antigens, facilitating the development of a bio-activated platform for personalized therapy.

Objective:To develop and assess a three-class machine learning model for predicting wild-type, 19 del, and 21 L858R mutations of the epidermal growth factor receptor (EGFR) in lung adenocarcinoma using 18F-FDG PET/CT radiomic features and clinical features. Methods:The retrospective data was collected from 703 patients (346 males, 357 females; age (64.3±9.0) years) with lung adenocarcinoma at the First People′s Hospital of Changzhou from January 2018 to June 2023. Patients were divided into the training set (563 cases) and test set (140 cases) at the ratio of 8∶2. Clinical features were selected using recursive feature elimination (RFE). Radiomic features were extracted from PET and CT images, and the optimal feature sets were selected using minimum redundancy maximum relevance (mRMR) and least absolute shrinkage and selection operator (LASSO) methods. Base models were constructed by using random forest (RF), logistic regression (LR), support vector machine (SVM), K-nearest neighbors (KNN), and multi-layer perceptron (MLP), and the stacking method was applied to establish the CT and PET ensemble models. Delong test was used to compare the AUC differences between the PET/CT combined model and the clinical + PET/CT integrated model.Results:Among 703 patients, 273 were with EGFR wild-type, 202 were with 19 del mutation, and 228 were with 21 L858R mutation. In the single-modal analysis, the AUCs of CT ensemble model in the training and test sets were 0.893 and 0.667, respectively, while the AUCs of PET ensemble model were 0.692 and 0.660. The AUC of PET/CT combined model were 0.897 in training set and 0.672 in test set. The AUC of clinical + PET/CT integrated model showed further improvement, with AUCs of 0.902 and 0.721 in training and test sets, respectively. Notably, the clinical + PET/CT integrated model outperformed PET/CT combined model in predicting wild-type EGFR (test set AUC: 0.784 vs 0.707; Z=3.28, P=0.001). Conclusion:The three-class model (clinical + PET/CT integrated model) based on 18F-FDG PET/CT radiomics and clinical features effectively predicts EGFR mutation subtypes in lung adenocarcinoma.

ObjectiveTo investigate the correlation between iron metabolism parameters and various syndrome types of unstable angina pectoris （UAP）. MethodsA cross-sectional study was conducted from October 2021 to October 2023， encompassing 213 patients diagnosed with UAP at Xiyuan Hospital of Chinese Academy of Chinese Medical Sciences. Additionally， 30 healthy individuals were selected as control cases. Single-factor analysis was used to investigate the differences in clinical data among different Traditional Chinese Medicine （TCM） syndrome types of UAP and their correlation with iron metabolism indices. The study conducted a comparative analysis of the aforementioned clinical data among patients with and without heat-toxic and blood-stasis syndrome. Logistic regression was used to analyze the correlation between TCM syndrome types and related factors. The receiver operator characteristic （ROC） curve was employed to assess the predictive value of iron metabolism indices， along with their sensitivity and specificity. ResultsCompared to those in the control group， serum iron （SI） and serum ferritin （SF） levels were significantly increased in the UAP group （P<0.01）， while transferrin （TRF） and total iron binding capacity （TIBC） levels were decreased （P<0.01）. However， there was no significant difference in unsaturated iron binding capacity （UIBC）. Multivariate binary Logistic regression analysis identified apolipoprotein A1 （ApoA1）， homocysteine （HCY）， high-sensitivity C-reactive protein （hs-CRP）， and SF as independent influencing factors for the UAP patients （P<0.05， P<0.01）. Additionally， statistically significant differences were observed in SI， SF， TRF， and TIBC among 213 patients with different TCM types （P<0.01）. Patients with heat-toxic and blood-stasis syndrome had higher SI and SF values than those without the syndrome （P<0.01）， while their TIBC and TRF values were lower （P<0.01）. Multivariate binary logistic regression analysis showed that SI and LDL-C levels were closely associated with the differentiation of heat-toxic and blood-stasis syndrome. ConclusionUAP patients often experience iron metabolism disorders， and the heat-toxic and blood-stasis syndrome are significantly correlated with iron metabolism parameters. The SI and LDL-C levels have high specificity and sensitivity in diagnosing heat-toxic and blood-stasis syndrome.

Objective To conduct a unique and pioneering molecular epidemiological investigation of a case of Eurasian avian-like H1N1 swine influenza identified in Yunnan Province in 2022(the fourth such case in the province)and to understand its genetic characteristics so as to reveal its potential impact on human health.Methods Real-time fluorescent quantitative PCR detection technology was used for the nucleic acid testing of the case's pharyngeal swab samples,close contacts,and environmental samples from the living area.Positive samples were subjected to virus isolation using MDCK cells.Cell cultures were authenticated using erythrocyte agglutination assay with guinea pig blood and real-time fluorescence quantitative RT-PCR.Whole genome sequencing was performed using the Illumina MiseqNext-generation sequencing platform,and a phylogenetic tree was constructed using MEGA 7.0 software to analyze the genetic molecular characteristics.Results The first G5 genotype Eurasian avian-like H1N1 swine influenza virus in Yunnan Province was successfully isolated,and the whole genome sequence of the virus was obtained.This virus possessed the molecular characteristics associated with increased adaptability,virulence,or transmissibility in mammals and had a nucleotide consistency of 99.2％～99.7％with a porcine strain isolated in Jiangsu province.These findings underscored the potential threat this virus poses to human health.Conclusion The study underscores the importance of further monitoring swine influenza in preventing new influenza virus subtypes that can infect humans.

OBJECTIVE: To carry out pathological and genetic analyses on a fetus with intrauterine growth restriction and death during second trimester after induced abortion. METHODS: A fetus undergone induced abortion due to intrauterine growth restriction and death during second trimester at the the Seventh Affiliated Hospital of Sun Yat-Sen University in 2024 was selected as the study subject. Clinical data of the pregnancy were collected. DNA was extracted from tissues from the aborted fetus and peripheral blood samples from its parents. Chromosomal microarray analysis and whole exome sequencing were carried out. Candidate variants were verified by Sanger sequencing. Following abortion, routine autopsy and pathological analysis were conducted. This study was approved by the Medical Ethics Committee of the hospital (Ethics No.: KY-2025-334-01). RESULTS: The aborted fetus was a male and harbored compound heterozygous nonsense variants of the TH gene (c.457C>T/p.Arg153* and c.694C>T/p.Gln232*), for which both parents were heterozygous carriers. Autopsy and pathological analysis revealed that the fetus had pathological features including loose arrangement of myocardial fibers and congestion in the liver. CONCLUSION Biallelic null variants of the TH gene may cause heart failure by affecting the development of cardiovascular system, which in turn may lead to intrauterine death. This study has provided new clues for the molecular diagnosis of stillbirth and recurrent pregnancy loss.
Humans ; Female ; Pregnancy ; Male ; Heterozygote ; Codon, Nonsense/genetics* ; Fetal Growth Retardation/pathology* ; Adult ; Stillbirth/genetics*

[Objective] To explore the feasibility of using whole blood as a source of NK cells for allogeneic CAR NK cell therapy and activated NK cell reinfusion therapy, and initially construct a technical system for the separation and purification of NK cells from whole blood. [Methods] All peripheral blood mononuclear cells (PBMCs) were enriched from 400 mL of whole blood by manual separation and machine separation, respectively. The erythrocyte loss rate, PBMCs number, NK cell purity of the two methods were compared. NK cells were sorted from PBMCs by three separation and enrichment methods as immunomagnetic bead negative selection method, platelet lysate culture expansion and PERCOLL density gradient separation method, and the purity and yield of NK cells, the activity of NK cells and the tumor-killing ability of the three separation and enrichment methods were compared. [Results] The proportion of NK cells in the lymphocyte population was higher in the manual separation method than in the machine separation method[(13.16±5.16)% vs (8.56±3.92)%, P<0.05]; the number PBMCs was lower in the manual separation method than in the machine separation method[(4.09±1.80)×10⁸vs (6.49±2.16)×10⁸, P<0.05], and there was no difference in the red blood cell loss between the two methods (P>0.05). The purity of NK cells isolated and enriched from PBMCs by manual separation method using immunomagnetic was (96.77±2.31)%; the yield was (56.27±10.47)%; the inhibition of tumor proliferation was (38.67±14.05)%; and the tumor killing rate was (19.90±8.05)%. The purity of NK cells isolated and enriched from PBMCs by manual separation method using platelet lysis culture expansion method was the highest at day 7, which was (54.84±15.80)%; the cell expansion multiple could reach 16.92±6.28 at day 7; the in vitro tumor killing rate of NK cells was (15.83±5.5)%; the tumor inhibition rate was (44.33±13.5)%; and there was no difference in the toxicity and activity of NK cells between the two methods (P>0.05). The purity of NK cells isolated and enriched by PERCOLL density gradient separation method was (15.83±5.82)%, and the yield was (14±6.25)%, which was significantly lower than the other two methods. [Conclusion] PBMCs isolated from whole blood by manual separation and NK cells enriched by negative selection with immunomagnetic beads have the potential to provide NK cell materials for CAR-NK cell therapy, and NK cells enriched by platelet lysate-conditioned medium have the potential to provide NK cells for large-scale NK cell activation reinfusion therapy.

Radiomics represents a non-invasive, quantitative image analysis method, aimed at correlating high-throughput quantitative radiomics features with clinical and biological endpoints. This method enables comprehensive, multiple, and non-invasive tumor assessment based on different types of images in various stages of radiotherapy. Advancements have been made in the applications of radiomics combined with machine learning-based statistical method in multiple fields including individualized target delineation, the prediction of local recurrence and radiation-induced injuries, the assessment of distant metastasis risks, the prediction of tumor motion, adaptive radiotherapy (ART), and multi-omics analysis. These advancements provide more possibilities for personalized radiotherapy. This study presents a brief introduction to the applications of radiomics in personalized radiotherapy.

Objective To explore the protective effect and possible mechanism of modified Shenqi Dihuang Decoction on podocytes in db/db mice based on ROS/NLRP3/GSDMD signaling pathway.Methods Fifty 8-week-old male db/db mice(SPF grade)were randomly divided into the model group,losartan group and TCM low-,medium-and high-dosage groups,with 10 mice in each group.Ten heterozygous db/m mice served as the blank group.Interventions were administered respectively for 12 weeks.The body mass,random blood glucose,serum creatinine,blood urea nitrogen and 24 h urinary protein content were detected,HE,PAS,PASM,Masson and Sirius red staining was used to observe the morphology of renal tissue,transmission electron microscopy was used to observe the ultrastructure of renal tissue,fluorescent probes were used to observe the release of ROS in renal tissue,immunofluorescence staining was used to detect the expression of Nephrin,NLRP3,Cleaved Caspase-1 and GSDMD-N in renal tissue,and Western blot was used to detect the expression of NLRP3,Cleaved Caspase-1,GSDMD-N,IL-1β,IL-18,Nephrin,Podocin,PODXL,WT-1 and Desmin proteins.Results Compared with the blank group,the body mass and random blood glucose of the model group mice significantly increased(P<0.05),and the contents of serum creatinine,blood urea nitrogen and 24 h urinary protein were significantly increased(P<0.05);glomerular hypertrophy,dilation of renal glomeruli and tubules,thickening of basement membrane,matrix proliferation in mesangial area,abnormal deposition of collagen fibers in renal interstitium,accompanied by damage to renal tubular epithelial structure and focal glomerulosclerosis,significant increase in type Ⅰ collagen deposition,extensive fusion of podocyte processes,and scattered electron dense material in the basement membrane or subepithelial layer;the ROS content in renal tissue significantly increased(P<0.05),and the protein expression of NLRP3,Cleaved Caspase-1,GSDMD-N,IL-1β,IL-18 and Desmin significantly increased(P<0.05),the protein expression of Nephrin,Podocin,PODXL and WT1 significantly decreased(P<0.05).Compared with the model group,the body mass and random blood glucose of mice in each dosage of TCM group were relatively stable,the contents of serum creatinine,blood urea nitrogen and 24 h urinary protein decreased;the pathological damage to renal tissue was reduced,the ultrastructure of podocytes was improved,and the density of podocytes increased;the ROS content decreased,and the protein expression of NLRP3,Cleaved Caspase-1,GSDMD-N,IL-1β,IL-18 and Desmin decreased,while the protein expression of Nephrin,Podocin,WT1 and PODXL increased.With the dosage of modified Shenqi Dihuang Decoction increased,the improvement effect gradually strengthened,and the differences in TCM high-dose group was statistically significant(P<0.05).Conclusion Modified Shenqi Dihuang Decoction can protects podocytes in db/db mice,potentially by modulating the ROS/NLRP3/GSDMD signaling pathway.

BACKGROUND:The incidence of fragility fractures caused by osteoporosis is increasing year by year,and it is urgent to explore its pathophysiology,potential biomarkers,therapeutic targets and effective drugs.There are multiple cells in the bone microenvironment,which are crucial for maintaining bone metabolism.In recent years,single-cell RNA sequencing technology has been developed to characterize the transcriptome of single cells,and has been gradually applied to the study of osteoporosis prevention and treatment.OBJECTIVE:To review the application and progress of single-cell transcriptome sequencing technology in osteoporosis research.METHODS:The first author used a computer search of Web of Science,PubMed database,and CNKI from 2009 to 2023.Chinese and English search terms were "single-cell RNA sequencing,bone marrow derived stromal cells,osteoblasts,osteocytes,osteoclasts,immune cells,bone marrow vascular endothelial cells." Through reading and screening of relevant literature,89 articles were finally included in the review analysis.RESULTS AND CONCLUSION:(1) Single-cell transcriptome sequencing technology can gain an in-depth understanding of the trajectory and regulatory mechanism of cell development,proliferation,differentiation.(2) Candidate genes affecting bone mineral density are mainly concentrated in bone marrow mesenchymal stem cell subpopulation,in which Sox9 is a key transcription factor.(3) The differential expression of Runx2 in different subsets of osteoblasts controls the differentiation of bone marrow mesenchymal stem cells into osteoblasts.(4) RAB38 is related to histone modification and transcriptional regulation during osteoclast differentiation.(5) Studies at the single-cell level have confirmed that there are many kinds of intercellular communication in the bone microenvironment,and osteoclast may conduct intercellular communication through CD160-TNFRSF14 ligand-receptor binding.The neutrocyte/monocyte may interact with osteoblasts through the RESISTIN pathway.Plasma dendritic cells communicate with osteoblast cell lines through the epidermal growth factor pathway.Both can provide directions for target prediction and drug development.(6) An unrecognized capillary subset is called S-type endothelial cells,which originates entirely from the secondary ossification center of the epiphysis,and is even more malleable than H-type blood vessels.(7) This paper reviews the progress of single-cell transcriptome sequencing in characterizing the differentiation fate and differentiation trajectory of different bone tissue cells,identifying new cell subsets,identifying cell regulatory factors,and clarifying intercellular communication from a cellular perspective,so as to provide cell-level information for exploring the pathogenesis of osteoporosis,screening potential diagnostic markers,predicting therapeutic targets,and exploring the mechanism of drug action.(8) In the future,single-cell sequencing technology will provide more valuable information for changes in the bone microenvironment in the direction of single-cell multiomics (genomics,proteomics,and metabolomics) and other technologies such as spatial transcriptome technology.

Objective:To explore the value of 18F-fluorodeoxyglucose ( 18F-FDG) PET-CT dual-modality habitat imaging technology in predicting the epidermal growth factor receptor (EGFR) mutation status in lung adenocarcinoma. Methods:This study was designed as a cross-sectional study. Clinical and imaging data of 403 patients with lung adenocarcinoma who underwent 18F-FDG PET-CT imaging with definitive EGFR results from January 2018 to April 2022 at the Third Affiliated Hospital of Soochow University were retrospectively analyzed.The patients were divided into a development set (282 cases) and a validation set (121 cases) using a stratified random sampling method at a 7∶3 ratio. An adaptive clustering algorithm was used to segment the regions of interest, forming different habitats and obtaining derived parameters. Independent samples t-test or Mann-Whitney U test were used to compare clinical, imaging indicators, and habitat-derived parameters between EGFR mutant and wild-type patient. The clinical, imaging indicators, and habitat-derived parameters that showed statistically significant differences in univariate analysis were included in multivariate logistic regression to construct clinical and clinical-habitat combined models, respectively. The receiver operating characteristic curve and area under the curve (AUC) were used to evaluate the model′s ability to predict EGFR mutations in lung adenocarcinoma. Additionally, the net reclassification index (NRI) was employed to assess the model′s classification improvement capability. Results:There were 249 cases of EGFR mutation and 154 cases of wild type. The optimal number of habitats was two, namely Habitat 1 and Habitat 2. The parameters included in the clinical model were smoking history, bronchial sign, pleural indentation sign, and tumor diameter. The parameters incorporated into the clinical-habitat combined model were smoking history, bronchial sign, pleural indentation sign, Habitat 2, and Habitat 1 voxel count. In the development set, the AUCs for predicting EGFR mutations in lung adenocarcinoma using the clinical model and the clinical-habitat combined model were 0.723 and 0.733, respectively, with no statistically significant difference ( Z=0.60, P=0.549); In the validation set, the AUCs were 0.684 and 0.715, respectively, with no statistically significant difference ( Z=1.32, P=0.186). The accuracy (0.694) and specificity (0.609) of the clinical-habitat combined model in the validation set were slightly higher than those of the clinical model (0.686 and 0.565, respectively). NRI analysis confirmed that the clinical-habitat combined model improved the correct classification of EGFR wild-type lung adenocarcinoma by 10.9% compared to the clinical model ( P=0.018). Conclusion:18F-FDG PET-CT dual-modality habitat imaging technology can be used to analyze the microenvironment of lung adenocarcinoma and has the potential in non-invasively predicting EGFR mutation status, providing an important basis for personalized and accurate treatment of patients with lung adenocarcinoma.