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28 December 2024, Volume 44 Issue 12 Previous Issue   
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Innovative research team achievement column
Basic research
Clinical research
Public health
Techniques and methods
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Innovative research team achievement column

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Comparison of human-induced pluripotent stem cell-derived macrophages with peripheral blood-derived macrophages using single-cell genomics ZHANG Yutong, HOU Guojun, SHEN Nan 2024, 44 (12):  1477-1489.  doi: 10.3969/j.issn.1674-8115.2024.12.001 Abstract ( 37 )   HTML ( 5 )   PDF (5900KB) ( 63 )   Objective ·To explore the heterogeneity in single-cell genomics between human-induced pluripotent stem cell (iPSC)-derived macrophages (IPSDM) and human peripheral blood-derived macrophages (PBDM). Methods ·iPSCs were differentiated into IPSDMs in vitro using a feeder-free and serum-free protocol. The expression of cluster of differentiation antigen 14 (CD14) and monocyte-macrophage marker genes in IPSDMs was analyzed using flow cytometry and real-time reverse transcription quantitative polymerase chain reaction (RT-qPCR), respectively. Single-cell sequencing was then performed on IPSDMs. Simultaneously, the single-cell sequencing dataset GSE126085 was downloaded from the Gene Expression Omnibus database as a reference dataset for PBDMs. Sequencing data for both IPSDMs and PBDMs were processed and analyzed using the seurat package in R software, with PBDMs annotated using the singleR package. A reference dataset was constructed with highly variable genes from PBDMs, and the highly variable genes of IPSDMs were projected onto the PBDM dataset using the scmap package to infer IPSDMs cell identities based on variable gene similarity. IPSDMs were annotated using cell-type annotation tools and referenced against relevant studies. The expression distribution of macrophage marker genes was compared between IPSDMs and PBDMs. Differentially expressed genes (DEGs) between IPSDMs and PBDMs were identified using the seurat package, and their potential biological functions were explored through Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses. Results ·Suspended IPSDMs were obtained after 29 d of in vitro differentiation. Flow cytometry and RT-qPCR confirmed that approximately 23.1% of IPSDMs expressed CD14, and IPSDMs exhibited higher expression of monocyte-macrophage marker genes compared to the U-937 cell line. All cells in the PBDM dataset were annotated as macrophages. After constructing a scmap reference dataset using PBDMs, 59.8% of IPSDMs were annotated as macrophages through mapping their highly variable genes to the PBDM dataset. The remaining 40.2% of IPSDM cells could not be matched to the variable genes of PBDMs. Further manual annotation of IPSDMs revealed a composition of 97.15% macrophages, 2.71% hematopoietic precursor cell-like cells, and 0.14% dendritic cells. When comparing the expression of macrophage markers, both IPSDMs and PBDMs highly expressed the classical macrophage marker CD68 gene, while IPSDMs exhibited higher expression of markers associated with tissue-resident macrophages. GO analysis of DEGs showed enrichment in the molecular functions such as ubiquitin-like protein ligase binding, cellular components such as the nuclear speck and nuclear envelope, and biological processes such as the regulation of translation. KEGG pathway enrichment indicated that the DEGs between IPSDMs and PBDMs might be related to various intracellular pathogen infections. Conclusion ·Human IPSDMs and PBDMs exhibit certain similarities and heterogeneity at the single-cell transcriptional level. Transcriptomic analysis indicates that IPSDMs display more characteristics of tissue-resident macrophages. The DEGs between IPSDMs and PBDMs are potentially associated with intracellular infection immunity. Figures and Tables | References | Related Articles | Metrics

Basic research

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Association of genetic variations in the NOS1 gene with insomnia, sleep duration and obstructive sleep apnea-related clinical quantitative traits YUAN Haolin, LI Niannian, HU Junhui, SHEN Jinhong, GAO Zhenfei, GUAN Jian, LIU Feng, YIN Shankai 2024, 44 (12):  1490-1503.  doi: 10.3969/j.issn.1674-8115.2024.12.002 Abstract ( 25 )   HTML ( 2 )   PDF (2602KB) ( 39 )   Objective ·To explore the correlation between the genetic variations rs7305526 and rs11615756 of nitric oxide synthase 1 (NOS1) and the human sleep traits, including insomnia, sleep duration, and clinical quantitative traits related to obstructive sleep apnea (OSA). Methods ·The NOS1 gene expression pattern at the whole-brain level using the Allen Human Brain Atlas dataset was analyzed. Subsequently, we performed expression quantitative trait locus (eQTL) analysis to investigate the impact of rs7305526 and rs11615756 on NOS1 gene expression. Regression analysis was conducted to assess the associations between rs7305526 and rs11615756 with insomnia and sleep duration based on the United Kingdom Biobank (UKB) Genome-Wide Association Study (GWAS) dataset. Furthermore, we explored the relationships between rs7305526 and rs11615756 with clinical quantitative traits of OSA, such as respiratory events, oxygen levels, and sleep traits, using clinical monitoring data from the Shanghai Sleep Health Study Cohort (SSHS) based on standard polysomnography (PSG). Results ·The NOS1 genedemonstrated elevated levels of expression in various brain regions crucial for regulating sleep, namely the amygdala, basal forebrain, striatum, and thalamus, as well as in the respiratory center, including the mesencephalon and pontine tegmentum. In contrast, the expression level of NOS1 gene was significantly reduced or absent in areas such as the cerebral cortex and cerebellum. Variants rs7305526 and rs11615756 were significantly negatively correlated with the expression levels of NOS1 in the cerebral cortex. Additionally, rs11615756 was also significantly negatively correlated with the expression level of NOS1 in the amygdala. Analysis of the UKB GWAS data revealed that the variant rs7305526 was not significantly associated with either insomnia or sleep duration, while rs11615756 demonstrated a noteworthy negative correlation specifically with sleep duration. Data obtained from the SSHS indicated a significant association between rs7305526 and alterations in clinical quantitative traits of OSA, including lowest pulse blood oxygen saturation (LSpO2), apnea-hypopnea index (AHI), and the ratio of non-rapid eye movement (NREM) stage 2 sleep duration. Although rs11615756 showed a notable negative correlation solely with the quantity of NREM stages 2 and 3, both rs11615756 and rs7305526 showed significant correlations with some respiratory events and oxygen traits after stratification according to the severity of OSA. Conclusion ·Genetic variants of NOS1 gene are respectively associated with human sleep duration traits and OSA-related variables, suggesting that NOS1 gene plays a crucial regulatory role in human sleep and clinical quantitative traits of OSA. The regulation of sleep traits by rs7305526 (C>A) is independent of its regulation of respiratory events and oxygen traits. Figures and Tables | References | Related Articles | Metrics

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Mechanistic studies on regulation of the activity of GPI-anchored serine protease testisin WAN Jiahui, ZHOU Aiwu 2024, 44 (12):  1504-1513.  doi: 10.3969/j.issn.1674-8115.2024.12.003 Abstract ( 24 )   HTML ( 3 )   PDF (2871KB) ( 34 )   Objective ·To explore the activation methods and activity regulation mechanisms of the testisin zymogen in vitro, laying a foundation for further research on the physiological functions of testisin in organisms. Methods ·The eukaryotic expression plasmid for the mouse-derived testisin (mTN) zymogen was constructed through full-gene synthesis and subsequently transfected into eukaryotic HEK293S cells for expression. Purified recombinant testisin protein was obtained through methods such as nickel ion affinity chromatography. Site-directed mutagenesis was performed on the activation site and active center of the zymogen, and the activation mechanism of testisin was analyzed through enzymatic activity assays. Additionally, the activation rate of the protein was determined by altering the incubation conditions of the zymogen. The self-cleavage site of the protease was identified by using Edman degradation and site-directed mutagenesis. Finally, the regulatory mechanism of protease activity was examined by analyzing the proteolytic rate of the protein towards its substrates. Results ·The recombinant mTN zymogen was expressed in eukaryotic HEK293S cells, and high-purity, uniform recombinant protein was obtained through a two-step purification process. This protein was found to remain stable under acidic conditions but underwent self-activation in neutral or alkaline environments. The rate of activation was influenced by factors such as pH value and temperature. The self-activation process required on the integrity of the enzyme's activation site, Arg46, and its active center, Ser240. Additionally, the self-activation of the recombinant mTN zymogen was often accompanied by the self-cleavage at the 170/175 loop on the molecular surface, but this self-cleavage did not affect the activity of testisin. Furthermore, the optimal reaction pH for activated mTN was 8.0, with the protein remaining relatively stable near this pH. The optimal reaction temperature was 50 ℃, while the protein was best stored at temperatures below 30 ℃. Zn2+ and Ca2+ were found to significantly inhibit mTN activity. Conclusion ·An effective method for the preparation of recombinant mTN zymogen is successfully established, and it is discovered that the recombinant mTN undergoes self-activation and self-cleavage in vitro. Figures and Tables | References | Related Articles | Metrics

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GPR87 promotes invasion and migration through the RHO/ROCK pathway in non-small cell lung cancer LIU Chenxi, HAN Lin, YANG Yi, ZHOU Han, LIU Yayun, SHENG Deqiao 2024, 44 (12):  1514-1525.  doi: 10.3969/j.issn.1674-8115.2024.12.004 Abstract ( 30 )   HTML ( 3 )   PDF (7860KB) ( 41 )   Objective ·To explore the role and molecular mechanism of GPR87 in regulating the invasion and migration of non-small cell lung cancer (NSCLC). Methods ·Bioinformatics methods, including GEO, UALCAN, KM Plotter and other public database analysis platforms, were used to screen candidate genes related to NSCLC invasion and predict their clinical relevance to NSCLC. Eighty NSCLC clinical patient samples and corresponding clinical pathological data were collected from Yichang Central People's Hospital from January 2018 to August 2020. Immunohistochemistry was used to analyze the expression of GPR87 in tumor tissues and the clinical relevance of GPR87 was analyzed. siRNA-GPR87 and pCMV-GPR87-his were transfected into the human lung adenocarcinoma cell line A549 and the human lung squamous cell carcinoma cell line SK-MES-1, to construct cell lines with low and high expression of GPR87. Transwell assay was used to investigate the effect of GPR87 expression on the migration and invasion ability of NSCLC cells. ELISA was used to detect the secretion of MMP7 in the culture supernatant. RT-qPCR was used to detect the mRNA expression levels of GPR87, MMP2, MMP7, MMP9, E-cadherin,N-cadherin, vimentin,snail,twist, RHOA, RHOC, and ROCK1. ELISA was used to detect the secreted protein MMP7. Western blotting was used to detect the protein expression levels ofGPR87, MMP9, E-cadherin, vimentin, RHOA, and ROCK1. Results ·Bioinformatics analysis of clinical sample data showed that GPR87 was highly expressed in NSCLC. Patients with higher expression of GPR87 had worse clinical stage and were more prone to lymph node metastasis, suggesting that GPR87 might be a key gene for the high invasiveness of NSCLC. Downregulation of GPR87 expression significantly reduced the invasion and migration ability of A549 and SK-MES-1 cells, while overexpression of GPR87 enhanced the invasion and migration ability of A549 and SK-MES-1 cells. Further detection revealed that downregulation of GPR87 led to decreased mRNA expression levels of MMP2, MMP7, MMP9, RHOA, RHOC, and ROCK1, as well as a reduction in the secretion of MMP7 and the protein expression levels of MMP9, RHOA, and ROCK1 in A549 and SK-MES-1 cells. Overexpression of GPR87 increased the mRNA expression levels of MMP2, MMP7, MMP9, RHOA, RHOC, and ROCK1, as well as the secretion of MMP7 and the protein expression levels of MMP9, RHOA, and ROCK1. Regardless of GPR87 knockdown or overexpression, the expression of genes and proteins related to epithelial-mesenchymal transition (EMT) in the cells did not change significantly. Conclusion ·High expression of GPR87 is closely related to the high invasiveness of NSCLC. In SK-MES-1 and A549 cells, GPR87 can activate the RHOA/ROCK1 signaling pathway, promote the expression of MMPs, and ultimately promote the invasion and migration of NSCLC. Figures and Tables | References | Related Articles | Metrics

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Function of vasohibin-2 and the mechanism of alternative splicing in triple-negative breast cancer WANG Wei, WANG Hongli, ALIBIYATI·i Ain, YILIYAER· Rousu, AYI NUER, YANG Liang 2024, 44 (12):  1526-1535.  doi: 10.3969/j.issn.1674-8115.2024.12.005 Abstract ( 23 )   HTML ( 1 )   PDF (4754KB) ( 41 )   Objective ·To explore the role of vasohibin-2 (VASH2) in the regulation of proliferation and metastasis of triple-negative breast cancer (TNBC) cells, and explore the mechanism of VASH2 in the occurrence and development of TNBC through regulation of gene expression and alternative splicing. Methods ·TCGA-GTEx was used to analyze the expression of VASH2 in TNBC. VASH2 methylation levels in TNBC were also analyzed. VASH2 was overexpressed in the MDA-MB-231 human TNBC cell line and transcriptome sequencing was performed. Differentially expressed genes and alternatively spliced genes regulated by VASH2 were analyzed to explore the mechanism of action of VASH2 in TNBC. Results ·VASH2 was significantly overexpressed in TNBC compared to the normal tissues. Hypomethylation of the VASH2 gene was implicated in the upregulation of VASH2 expression in TNBC. Overexpression of VASH2 caused significant differential expression of 81 genes, of which 23 genes were up-regulated and 58 genes were down-regulated. Genes with significantly altered alternative splicing levels due to VASH2 overexpressed were enriched in cell cycle and p53 signaling pathways. Conclusion ·VASH2 regulates the alternative splicing of TNBC oncogenes and promotes TNBC occurrence and development. Figures and Tables | References | Related Articles | Metrics

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Inhibition of Tannerpin-M encoded by periodontal pathogens on serine proteases released by granulocytes PAN Zihao, XU Jiawei, ZHOU Aiwu 2024, 44 (12):  1536-1544.  doi: 10.3969/j.issn.1674-8115.2024.12.006 Abstract ( 18 )   HTML ( 1 )   PDF (3082KB) ( 12 )   Objective ·To prepare a serine protease inhibitor (Serpin) derived from Tannerella which is associated with periodontosis, and analyze its specificity in inhibiting target proteases and its structural characteristics. Methods ·Through amino acid sequence analysis, a Serpin from the human oral microbiome database (eHOMD) was selected and expressed in Escherichia coli. The recombinant protein was purified using methods such as nickel ion affinity chromatography. Its specificity in inhibiting serine proteases was analyzed, followed by an analysis of its three-dimensional spatial structure using structural biology methods. Results ·A novel Serpin, named Tannerpin-M, with methionine as the active center P1 residue, was identified, and a high-purity recombinant protein was successfully prepared from Escherichia coli BL21 (DE3). Further activity testing demonstrated that recombinant Tannerpin-M could effectively form SDS-stable covalent complexes with proteases derived from granulocytes (human neutrophil elastase, cathepsin G, and proteinase 3), as well as with other proteases including kallikrein 1 (KLK1), KLK7, and elastase. Tannerpin-M inhibited KLK7 with a second-order association rate constant of 4.12×104 L/(mol·s). The crystal structure of Tannerpin-M in its relaxed state conformation was resolved at a resolution of 2.4 ? (1 ?=0.1 nm). It revealed that Tannerpin-M possessed a significantly elongated reactive center loop and could undergo the classical conformational transition from a stressed to a relaxed state. Conclusion ·Tannerpin-M, derived from oral pathogenic bacteria, is a typical inhibitory Serpin, and can effectively inhibit the serine protease released by granulocytes, by which it may protect the oral pathogenic bacteria from attacks of the human immune system. Figures and Tables | References | Related Articles | Metrics

Clinical research

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Timing of ultrasonography in the diagnosis of fallopian tubal heterotopic pregnancy HUANG Qin, HUANG Ying, LI Wen 2024, 44 (12):  1545-1551.  doi: 10.3969/j.issn.1674-8115.2024.12.007 Abstract ( 24 )   HTML ( 1 )   PDF (2036KB) ( 11 )   Objective ·To assess the value of ultrasonography in the diagnosis of fallopian tubal heterotopic pregnancy (THP). Methods ·From January 2010 to December 2022, among the 13 082 cases of ectopic pregnancy diagnosed and discharged from the International Peace Maternity ＆Child Health Hospital, Shanghai Jiao Tong University School of Medicine, 88 cases suspected of having THP were selected as the study objects. Pre-admission ultrasound images and clinic data from these cases were collected. Then these cases were divided into three groups according to their gestational age at the time of ultrasonography: Group Ⅰ, gestational age within 38 d; Group Ⅱ, gestational age of 39?45 d; Group Ⅲ, gestational age of 46?73 d. The incidence of THP, pregnancy outcomes, the sensitivity and specificity of ultrasound diagnostic results, and the consistency between postoperative pathological results, discharge diagnosis and preoperative ultrasound diagnosis results (i.e. accuracy) were retrospective analyzed. Receiver operating characteristic (ROC) curve analysis was used to evaluate the diagnostic value of ultrasonography for THP in these three groups. Results ·In all the ectopic pregnancy cases, 72 THP cases were confirmed by postoperative pathology or further ultrasonography. The incidence rate of THP was 0.5% among the 13 082 patients with ectopic pregnancy. There were two types of preoperative ultrasound images of these 72 THP cases: Type Ⅰ, echogenic mass in gestational sac, accounting for 30.6%; Type Ⅱ, hypoechoic and mixed echogenic mass, accounting for 69.4%. In addition, 54.2% of TPH patients underwent salpingectomy, and the intrauterine live birth rate in TPH patients was 59.7%. There were significant differences between groups in the overall sensitivity (91.7%), specificity (81.3%), and accuracy (89.8%) of ultrasonography in the diagnosis of THP (all P<0.05). Sensitivity and specificity in Group Ⅰ were significantly lower than those in Group Ⅲ; the accuracy of Group Ⅰ was significantly lower than that of Group Ⅱ and Group Ⅲ (all P<0.05). ROC curve analysis further showed that the diagnostic value of ultrasonography for THP was low for cases within 38 d of pregnancy, whereas for cases with gestational ages of 39?45 d and 46?73 d, ultrasonography demonstrated notable diagnostic value. Conclusion ·Ultrasonography has high diagnostic value for THP. The gestational age at the time of ultrasonography is closely related to the sensitivity, specificity, and accuracy of diagnosing THP. Special attention should be paid to ultrasonography for pregnant women with a gestational age over 46 d to assist in the diagnosis of THP. Figures and Tables | References | Related Articles | Metrics

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Development of indicators and analysis of barriers for assessment and prevention of cutaneous graft-versus-host disease after haematopoietic stem cell transplantation in children YANG Liling, ZHAN Sha, XUE Xiaoyan, CHEN Kai, LIU Qing, WANG Li, WANG Yiyi, WU Meng, LU Qunfeng 2024, 44 (12):  1552-1560.  doi: 10.3969/j.issn.1674-8115.2024.12.008 Abstract ( 19 )   HTML ( 2 )   PDF (1384KB) ( 8 )   Objective ·To gain a comprehensive understanding of the current clinical status of assessment and prevention of cutaneous graft-versus-host disease (GVHD) after haematopoietic stem cell transplantation in children, construct review indicators, analyze obstacles and facilitators in the process of conducting the study in light of clinical reality, and formulate strategies for change. Methods ·Utilizing the JBI Evidence-Based Health Care Model as a theoretical guiding framework, the clinical problem was clarified. An evidence-based practice team was established, and a systematic literature search was conducted. The evidence was then evaluated and summarized. Additionally, review indicators were constructed, and review methods were clarified in relation to the evidence. Using the Ottawa Model of Research Use, the three dimensions of evidence change, potential adopters, and practice environment were analyzed for barriers and facilitators, and corresponding strategies were developed. Results ·A total of 24 pieces of best evidence were included in the study, comprising five dimensions: skin assessment, assessment of protection, management of adverse skin reactions, medication guidance and follow-up and screening. Twenty-two review indicators were constructed on this basis, of which 14 had an implementation rate of <60% and 10 had an implementation rate of 0%; 8 had an implementation rate of >60% and 6 had an implementation rate of 100.00%. The main obstacles were the lack of relevant training and the absence of a rational management mechanism. The main facilitating factors were reliable sources of evidence and a high level of cooperation from potential stakeholders. Considering the results of the analysis of the implementation rate of the indicators under review and the obstacles, alternative measures were formulated. Conclusion ·There is a large gap between the evidence and clinical practice for the assessment and prevention of cutaneous GVHD after paediatric haematopoietic stem cell transplantation. Clinical change should be effectively implemented based on barriers and facilitators. Figures and Tables | References | Related Articles | Metrics

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Research on the characteristics of 18F-FDG PET/CT in mantle cell lymphoma and the discrimination between cellular morphological variants REN Yixuan, CHEN Cheng, CAI Mingci, CHEN Jiamin, YANG Xinxin, WANG Chao, LIN Xiaozhu, CHENG Shu, JIANG Xufeng, CHEN Dongxu 2024, 44 (12):  1561-1569.  doi: 10.3969/j.issn.1674-8115.2024.12.009 Abstract ( 25 )   HTML ( 1 )   PDF (4578KB) ( 8 )   Objective ·To analyze the imaging characteristics and diagnostic value of 18F-fluorodeoxyglucose positron emission tomography-computed tomography (18F-FDG PET/CT) in mantle cell lymphoma (MCL) and explore its application to distinguishing between classic and aggressive variants of MCL. Methods ·A retrospective analysis was conducted on the 18F-FDG PET/CT images and clinical data of 116 pathologically confirmed, newly diagnosed MCL patients. The imaging features of intra- and extra-nodal lesions were summarized. The accuracy of 18F-FDG PET/CT in diagnosing bone marrow and gastrointestinal involvement in MCL was evaluated. Furthermore, differences in 18F-FDG PET/CT findings and clinical characteristics between the classic and aggressive variants of MCL were analyzed. Results ·Among the 116 patients, 100.0% showed positive findings on 18F-FDG PET/CT, with 99.1% exhibiting abnormal lymph nodes and 85.3% having extra-nodal involvement. The most common extra-nodal sites were the spleen, Waldeyer's ring, bone marrow, and gastrointestinal tract. Compared with bone marrow aspiration results, the sensitivity, specificity, and accuracy of 18F-FDG PET/CT for detecting bone marrow involvement in MCL were 43.4%, 91.5%, and 66.0%, respectively. When compared with endoscopic biopsy results, the sensitivity of 18F-FDG PET/CT for detecting gastric and intestinal involvement was 100.0% and 94.1%, respectively, with specificity of 75.0% and 100.0%, and accuracy of 92.9% and 94.7%, respectively. There were significant differences in the highest maximum standardized uptake value (SUVmax) and Ki-67 index between the classic and aggressive variants of MCL, with SUVmax positively correlated with Ki-67 index. By using SUVmax > 10.4 as the diagnostic threshold, the sensitivity and specificity for differentiating between the classic and aggressive variants of MCL were 73.9% and 77.4%, respectively, with an AUC value of 0.797. Conclusion ·18F-FDG PET/CT demonstrates a high detection rate for both intra- and extra-nodal lesions in MCL patients. It exhibits high specificity in diagnosing bone marrow involvement and high sensitivity and specificity in diagnosing gastrointestinal involvement, providing reliable non-invasive diagnostic information for MCL bone marrow and gastrointestinal involvement. However, it is not a substitute for pathological examination. Additionally, the positive correlation between SUVmax and Ki-67 index allows SUVmax to effectively differentiate between the classic and aggressive variants of MCL, with a higher SUVmax (>10.4) indicating a higher likelihood of the aggressive variant. These findings have clinical implications for treatment planning and prognosis assessment. Figures and Tables | References | Related Articles | Metrics

Public health

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Relationship between prenatal mixed heavy metal/metalloid exposure and offspring cognitive and temperament development XU Zujing, JIANG Yining, XU Jian 2024, 44 (12):  1570-1578.  doi: 10.3969/j.issn.1674-8115.2024.12.010 Abstract ( 21 )   HTML ( 6 )   PDF (2576KB) ( 19 )   Objective ·To analyze the relationship between prenatal mixed heavy metal/metalloid exposure and the cognitive and temperament development of the offspring. Methods ·A prospective birth cohort study was conducted in Shanghai from 2010 to 2012. Maternal venous blood samples were collected at 28?36 weeks of gestation to measure prenatal maternal blood heavy metal/metalloid concentrations. At 24?36 months of age, the cognitive and temperament development of the offspring were assessed by using Gesell Developmental Scale and Toddler Temperament Scale, respectively. Demographic and clinical information of mothers and children was collected via questionnaires and hospital medical records. Bayesian kernel machine regression (BKMR) models were employed to assess the effects of prenatal mixed heavy metal/metalloid exposure on toddlers' cognitive and temperament development. Results ·A total of 139 mother-child pairs with complete data were included in the study. At enrollment, the mean maternal age was (29.39±3.41) years, and the mean birth weight of the infants was (3.47±0.42) kg, with 59.71% female infants. At follow-up, the average age of the toddlers was (32.91±2.69) months. Based on prior research and multivariable linear regression models, four heavy metals/metalloid—chromium (Cr), manganese (Mn), arsenic (As), and lead (Pb)—were identified as potentially associated with cognitive and temperament development in children. The median concentrations of Mn, Pb, As, and Cr in the maternal venous blood during pregnancy were 3.32, 3.60, 2.03, and 1.78 μg/dL, respectively. After adjusting for related confounders, the BKMR analyses found that when the overall concentrations of the four heavy metals/metalloid (Cr, Mn, As, and Pb) were higher than the 50th percentile, children's fine motor development quotients showed a decreasing trend as the overall concentrations of the four heavy metals/metalloid increased. Scores for adaptability, persistence, and threshold of response on the temperament scale showed a decreasing trend as well. Prenatal Mn exposure levels made the greatest contribution to the effects on children's fine motor development [PIP (posterior inclusion probability)=0.617], prenatal As exposure levels made the greatest contribution to the effects on children's persistence trait (PIP=0.656), and prenatal Cr exposure levels made the greatest contribution to the effects on children's reaction threshold trait (PIP=0.447). The contribution of all four heavy metals/metalloid to the effects on children's adaptive dimension was similar. Conclusion ·Children's fine motor development and the adaptability, persistence, and threshold traits of temperament may be related to prenatal mixed exposure to heavy metals/metalloid (Mn, As, Pb, and Cr). Particular attention should be paid to the relationship between prenatal Mn exposure and children's fine motor development, and between prenatal As and Cr exposure and children's temperament development. Figures and Tables | References | Related Articles | Metrics

Techniques and methods

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Preliminary application of a cervical vertebra segmentation method based on Transformer and diffusion model for lateral cephalometric radiographs in orthodontic clinical practice LIU Yang, WU Mengyi, HU Yao, QI Kun, WANG Yubin, ZHAO Yue, SONG Jinlin 2024, 44 (12):  1579-1586.  doi: 10.3969/j.issn.1674-8115.2024.12.011 Abstract ( 45 )   HTML ( 4 )   PDF (4502KB) ( 26 )   Objective ·To construct a cervical vertebra image segmentation model by using a diffusion model with the Transformer deep learning algorithm, and evaluate its segmentation performance, to address the clinical challenge of accurately assessing complex changes in skeletal morphology during the growth and developmental peaks of malocclusion. Methods ·Accurate cervical vertebra segmentation was performed on cephalometric radiographs from 185 orthodontic patients (44 cases from the Stomatological Hospital of Chongqing Medical University and 141 cases from the Stomatological Hospital of Xi'an Jiaotong University) by using a method combining Transformer and diffusion models. First, the images were preprocessed to crop out the cervical vertebra region of interest, and all data were randomly divided into a training set (79.6%) and a test set (20.4%). The diffusion model and a conditional model based on U-Net were utilized for feature extraction, with a Transformer module introduced to learn the interaction between noise and semantic features. Multi-scale images were fused to enhance fine structure and boundary texture features in low-contrast images. The proposed method was compared with U-Net and SOLOv2 methods. The segmentation performance was quantitatively evaluated by two metrics, Dice Similarity Coefficient (DSC) and Intersection over Union (IoU), and also qualitatively assessed through physicians' manual annotations and model visualization results. Results ·The cervical vertebra segmentation method based on Transformer and diffusion models achieved DSC and IoU scores of 93.3% and 87.5%, respectively, significantly outperforming the U-Net and SOLOv2 methods (with improvements of 3.0% and 4.1% in DSC, and 5.2% and 7.1% in loU, respectively). Despite the longer processing time for a single image, segmentation accuracy was significantly improved. Compared with U-Net and SOLOv2, the proposed method also showed higher stability and robustness in processing complex, low-contrast and blurred-boundary images, and was able to accurately segment the cervical vertebrae with clear boundaries and complete structures. Conclusion ·The Transformer-based diffusion model for cervical vertebra segmentation can enhance the edge and texture features in cervical vertebra images and recognize the boundaries of different vertebrae more easily. Thus, automatic, accurate, and robust cervical vertebra segmentation results are achieved, which can assist in cervical vertebral maturation analysis. Figures and Tables | References | Related Articles | Metrics

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Construction of fluorescent transgenic zebrafish Tg(amh:mCherry) and tracer analysis of its gonadal development WU Jing, XU Yue, LING Shiying, SONG Huaidong, QIAO Jie, DONG Mei 2024, 44 (12):  1587-1592.  doi: 10.3969/j.issn.1674-8115.2024.12.012 Abstract ( 19 )   HTML ( 1 )   PDF (2359KB) ( 7 )   Objective ·To establish a transgenic zebrafish line Tg(amh:mCherry) for studying gonadal development and related diseases, and to trace and analyze the developmental process of zebrafish gonads. Methods ·A recombinant transgenic expression vector pTol2-amh-mCherry was constructed by using the upstream promoter sequence of the zebrafish anti-Müllerian hormone (amh) gene coding region and the gene coding sequence of the red fluorescent protein, and was validated by using Sanger sequencing. The recombinant transgenic expression vector and mRNA of Tol2 transposase were co-microinjected into zebrafish embryos, and fluorescent zebrafish were selected by using fluorescence microscope to screen for a stable inherited transgenic zebrafish line Tg(amh:mCherry). In situ hybridization was used to detect whether the location of the fluorescence signals was consistent with the location of endogenous expression of amh. The red fluorescence of transgenic zebrafish from generation F3 was observed by fluorescence microscopy, and the process of gonadal development was tracked and analyzed. Results ·The recombinant transgenic expression vector pTol2-amh-mCherry was successfully constructed, and a stable inherited F3 generation transgenic zebrafish line Tg(amh:mCherry) was successfully established. The results of in situ hybridization showed that the location of the red fluorescence signals in the transgenic zebrafish Tg(amh:mCherry) was consistent with the location of endogenous expression of amh. Through observation and analysis of the fluorescence of transgenic zebrafish from F3 generation, five developmental patterns of zebrafish gonads were found. Conclusion ·The successful construction of Tg(amh: mCherry) transgenic zebrafish line facilitates the tracer analysis of zebrafish gonadal development, which provides a better experimental model for the study of gonad-related diseases. Figures and Tables | References | Related Articles | Metrics

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Research progress in the role of β-arrestin 1 in the regulation of inflammatory response YI Wenjing, FAN Yixuan, QIU Jiatai, FU Xiaoyan, LIU Meifang 2024, 44 (12):  1593-1600.  doi: 10.3969/j.issn.1674-8115.2024.12.013 Abstract ( 21 )   HTML ( 1 )   PDF (3422KB) ( 10 )   The inflammatory response is a systemic process produced by the body in response to stimulation such as infection or injury. The inflammatory response is closely related to the pathogenesis of various diseases, and plays an important role in the progression and outcome of diseases. Under normal conditions, the inflammatory response can maintain tissue homeostasis when the body is not seriously damaged. However, an uncontrolled inflammatory response that occurs when the body is subjected to more severe stimuli may often cause serious harm to the body. The termination of the ideal inflammatory response occurs after the elimination of the inflammatory stimulus. Therefore, it is particularly important to effectively modulate the extent and scope of the inflammatory response. β-arrestin 1 (ARRB1) is a multifunctional regulatory protein that plays a key role in G protein-coupled receptor (GPCR) and non-GPCR-mediated signal transduction. Meanwhile, this protein also participates in the regulation of some immune cell development, differentiation and other functions. Although ARRB1 was originally thought to be a terminator of GPCR signal transduction and can inhibit the inflammatory response, recent studies suggest that its role in inflammatory responses is complex, with dual anti-inflammatory and pro-inflammatory effects. Based on this, this article reviews the relevant research on ARRB1 and the inflammatory response in recent years, focusing on the regulatory role of ARRB1 in signaling pathway transduction and immune cell development and differentiation function, as well as its mechanism of action in regulating the progression of inflammatory diseases, in order to provide new insights for the clinical precision treatment of inflammatory diseases and the screening of drug effect targets. Figures and Tables | References | Related Articles | Metrics

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Research progress in the correlation between chronic stress and ovarian dysfunction WEI Liutong, LAI Dongmei 2024, 44 (12):  1601-1606.  doi: 10.3969/j.issn.1674-8115.2024.12.014 Abstract ( 17 )   HTML ( 1 )   PDF (1820KB) ( 7 )   The ovary is an important reproductive organ for women. Decreased ovarian function not only affects the reproductive capabilities, but also leads to the decline of women′s quality of life. Recent studies have shown that chronic stress affects the hypothalamic-pituitary-ovary axis directly or indirectly and leads to ovarian dysfunction-including excessive activation of primordial follicles, a reduction in preantral and preovulatory follicles, decline of oocyte quality, and granulosa cell apoptosis, through corticotropin-releasing hormone, corticosterone and cortisol secreted by the hypothalamic-pituitary-adrenal axis, as well as epinephrine and norepinephrine released by the sympathetic-adrenal-medullary axis and sympathetic nervous system. This article reviews the correlation between chronic stress and ovarian dysfunction, along with the potential mechanisms. Figures and Tables | References | Related Articles | Metrics

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Clinical advances in antibody-drug conjugates for hematological malignancies TANG Sijie, MI Jianqing 2024, 44 (12):  1607-1614.  doi: 10.3969/j.issn.1674-8115.2024.12.015 Abstract ( 17 )   HTML ( 1 )   PDF (1296KB) ( 5 )   An antibody-drug conjugate (ADC) is a targeted therapeutic drug composed of a monoclonal antibody linked to a small-molecule cytotoxic drug via a linker. Once administered, ADCs bind to tumor-specific antigens, forming ADC-antigen complexes, which are internalized through endocytosis. The linkers are then cleaved via endosomal-lysosome pathway and the cytotoxic drug is released, which induces apoptosis in the target cells. ADCs combine the advantages of monoclonal antibody drugs and cytotoxic drugs. They are able to reduce damage to normal cells while killing target cells, thus exhibiting higher anti-tumor efficiency. As the treatment of hematological malignancies gradually advances into the era of targeted immunotherapy, ADCs, as one of the hot spots, have shown broad prospects and also face many challenges in drug development and clinical application. Currently marketed ADCs in China include brentuximab vedotin (anti-CD30), inotuzumab ozogamicin (anti-CD22) and polatuzumab vedotin (anti-CD79B), while those marketed abroad include gemtuzumab ozogamicin (anti-CD33) and loncastuximab tesirine (anti-CD19), all demonstrating good efficacy and safety in clinical practice. Additionally, ADCs targeting different antigens such as CD123, CD19, CD20, receptor tyrosine kinase-like orphan receptor 1 (ROR1), and CD38 are undergoing clinical studies. Globally, there are over a hundred ADCs in development, and it is hoped that more breakthroughs will be achieved in the future to further optimize the treatment strategies for hematologic malignancies. References | Related Articles | Metrics

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Heterogeneous nuclear ribonucleoprotein H1 and its function in tumor development and metastasis ZHANG Zongwen, FENG Li, FAN Zhisong 2024, 44 (12):  1615-1620.  doi: 10.3969/j.issn.1674-8115.2024.12.016 Abstract ( 18 )   HTML ( 3 )   PDF (1212KB) ( 11 )   Heterogeneous nuclear ribonucleoprotein (hnRNP) and heterogeneous nuclear RNA (hnRNA) form complexes that regulate RNA splicing and transportation. hnRNPH1 is a member of the hnRNP family of proteins and plays an important role in physiological processes, including embryonic development of the nervous and reproductive systems. hnRNPH1 is highly expressed in various tumor tissues and tumor cells, and its overexpression correlates with poor prognosis in patients. hnRNPH1 enhances tumor cell proliferation and metastasis through the regulation of alternative splicing of genes, modulation of gene expression, formation of fusion genes with other genes, and involvement in epigenetic regulation.Recent studies have shown that hnRNPH1 could serve as a potential biomarker for certain tumors, and hnRNPH1-targeting drugs show preliminary efficacy in a mouse model of melanoma. This article reviews the role and research progress of hnRNPH1 in the initiation and progression of malignant tumors. References | Related Articles | Metrics