OBJECTIVE: To explore the genetic etiology of 46 Chinese pedigrees affected with Hereditary multiple exostoses (HME) and provide genetic counseling and reproductive intervention. METHODS: Whole-exome sequencing and Sanger sequencing were carried out on 87 patients from the 46 pedigrees to analyze the variants of EXT1 and EXT2 genes. Pathogenicity of the variants was assessed based on the guidelines from the American College of Medical Genetics and Genomics and Association for Molecular Pathology (ACMG/AMP). Prenatal diagnosis and preimplantation genetic testing (PGT) were provided for couples with identified pathogenic mutations. This study was approved by the Medical Ethics Committee of the hospital (Ethics No.: LL-SC-SG-2014-010). RESULTS: In total 17 and 22 pathogenic variants were respectively identified in the EXT1 and EXT2 genes, among which 5 EXT1 and 12 EXT2 variants were unreported previously. Three patients with no family history were found to harbor de novo variants of the EXT1 gene. Twenty nine couples had opted for PGT or underwent prenatal diagnosis following natural conception, and 17 healthy babies were born. CONCLUSION This study has clarified the genetic etiology of 45 HME pedigrees and identified 17 novel variants, which has enriched the mutational spectrum of the EXT1 and EXT2 genes. Reproductive intervention through PGT and prenatal diagnosis have prevented the recurrence of HME in these families.
Humans ; Female ; Male ; Pedigree ; Exostoses, Multiple Hereditary/diagnosis* ; N-Acetylglucosaminyltransferases/genetics* ; Adult ; Exostosin 1 ; Asian People/genetics* ; Genetic Testing ; Exostosin 2 ; Mutation ; China ; Prenatal Diagnosis ; Pregnancy ; Genetic Counseling ; Preimplantation Diagnosis ; Exome Sequencing ; East Asian People

Objective To identify immune cell-related biomarkers in lung adenocarcinoma (LUAD) using weighted gene co-expression network analysis (WGCNA). Methods Based on data from The Cancer Genome Atlas (TCGA) database, a gene co-expression network was constructed for the TCGA-LUAD dataset using the "WGCNA" R package, and genes were clustered into different modules. Concurrently, the Estimation of STromal and Immune cells in MAlignant Tumours using Expression data (ESTIMATE) algorithm was applied to the tumor samples in the TCGA-LUAD dataset. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were performed to evaluate the biological functions of genes within the most significantly correlated module. Candidate hub genes from the key module were intersected with a protein-protein interaction (PPI) network to identify the final hub genes. The prognostic performance of these hub genes and their correlation with immune cell infiltration were validated using Kaplan-Meier curves and the Tumor IMmune Estimation Resource (TIMER) algorithm. Finally, a multivariate Cox regression analysis was conducted on the identified hub genes to construct a prognostic risk model. Results In the co-expression network, the brown module was found to be highly correlated with the ImmuneScore, StromalScore, and ESTIMATE Score. Five immune-related hub genes were identified: CD53, PLEK, SPI1, IL10RA, and C3AR1. Enrichment analysis of the brown module revealed that its genes were primarily enriched in GO terms such as "regulation of innate immune response" and KEGG pathways like the "NF-kappa B signaling pathway". Furthermore, the expression levels of these five hub genes were significantly and positively correlated with the infiltration abundance of various immune cells. The immune relevance of the model was validated by the Immunophenoscore (IPS) and the Tumor Immune Dysfunction and Exclusion (TIDE) score. Moreover, the established RiskScore demonstrated significant potential in predicting the response to immunotherapy. Conclusion These five immune-related key genes may serve as novel and effective potential therapeutic targets for LUAD immunotherapy, facilitating the development of personalized diagnosis and treatment strategies for patients with LUAD.

Background/Aims: Lenvatinib resistance remains a critical barrier in advanced hepatocellular carcinoma (HCC) therapy. However, the underlying mechanisms and strategies for reversing resistance remain incompletely understood. Methods: Integrated transcriptomics of lenvatinib-resistant patient tumors and an acquired-resistance murine model identified a novel macrophage subpopulation. Functional validation employed CRISPR-SAM screening, conditioned medium (CM) assays, subcutaneous/orthotopic xenografts, patient-derived organoids (PDOs), and patient-derived xenografts (PDXs). Mechanistic studies included ChIP-qPCR, co-immunoprecipitation, and pharmacologic targeting. Clinical relevance was assessed in a retrospective cohort. Results: Resistant HCC exhibited significant enrichment of a COLEC12high TAM subset , which correlated with poor survival and treatment response. These TAMs secreted neuregulin-1 (NRG1) , activating HER2/HER3-AKT signaling in tumor cells to drive cancer stemness and lenvatinib resistance. Mechanistically, in TAMs COLEC12 sequestered STAT1 in the cytoplasm, preventing its phosphorylation, and thereby derepressing STAT3-mediated NRG1 transcription. Depletion of NRG1 reversed the stemness phenotypes and resensitized tumors to lenvatinib both in vitro and in vivo. Clinically, high NRG1 expression predicted an inferior lenvatinib response and shorter survival. Crucially, the bispecific anti-HER2/HER3 antibody zenocutuzumab restored lenvatinib efficacy in PDOs, PDXs, and murine models. Conclusions Our work establishes the COLEC12high TAM/NRG1 axis as a master regulator of therapeutic resistance and identifies NRG1 as a predictive biomarker, providing a clinically actionable strategy to overcome lenvatinib resistance in HCC.

ObjectiveTo investigate whether modified Taohe Chengqitang （JTCT） treats liver fibrosis by inhibiting hepatic sinusoidal capillarization， and to verify whether its specific mechanism is related to the Homologous genes responsible for the notches along the edges of Drosophila wings（Notch） signaling pathway and endothelial nitric oxide synthase （eNOS） signaling pathway. MethodsFifty C57BL/6 mice were randomized into 5 groups： Control， model （5 mL·kg^-1·3 d^-1）， low-dose JTCT， medium-dose JTCT， and high-dose JTCT （JTCT-L，JTCT-M，JTCT-H）. Except the control group， the other groups were intraperitoneally injected with 20% carbon tetrachloride （CCl₄） olive oil solution every 3 days for 4 weeks to induce liver fibrosis. Meanwhile， JTCT-L， JTCT-M， and JTCT-H groups were administrated with JTCT by gavage at doses of 16.6， 33.3， and 66.6 g·kg^-1·d^-1， respectively， for 4 consecutive weeks （the control and model groups received equal volumes of normal saline by gavage）. Primary liver sinusoidal endothelial cells （LSECs） were isolated from rat livers through in situ perfusion of collagenase， Percoll density gradient centrifugation， and selective removal of Kupffer cells. After verification of cell phenotype， LSECs were used for experiments. For cells at the logarithmic growth phase， the small interfering RNA（siRNA） kit was used to knock down the expression of soluble guanylate cyclase （sGC） through transient transfection. Subsequently， the Notch agonist Jagged-1 was applied. Cell counting kit-8 （CCK-8） was used to examine the cytotoxicity of JTCT on LSECs. Real-time PCR and Western blot were employed to measure the expression of molecules in the Notch and eNOS signaling pathways at mRNA and protein levels， respectively. Intracellular Ca²⁺ was labeled with the fluorescent probe Fluo-4 acetoxymethyl ester（Fluo-4/AM）. ResultsThe animal experiment showed that compared with the control group， the model group exhibited hepatic steatosis， inflammatory cell infiltration， hepatocyte degeneration and necrosis， lobular structural disorder， and fibrous tissue hyperplasia in the mouse liver tissue， significantly elevated serum levels of hydroxyproline （Hyp）， alanine transaminase （ALT）， and aspartate transaminase （AST） （P<0.01）， significantly upregulated expression levels of Notch1， Hes family bHLH transcription factor 1（Hes1）， Cluster of Differentiation 31（CD31）， and von Willebrand factor（vWF） （P<0.01）， and significantly downregulated phosphorylate（p）-eNOS/eNOS ratio and sGC expression （P<0.01）. Compared with the model group， all the JTCT groups showed significantly reduced serum Hyp， ALT， and AST levels （P<0.01）， downregulated expression of Notch1， Hes1， CD31， and vWF （P<0.05， P<0.01）， and upregulated p-eNOS/eNOS ratio and sGC expression （P<0.05， P<0.01） in the liver tissue， with the JTCT-H group showing the most significant changes （P<0.01）. The cell experiment showed that compared with the LSEC-1day group， the LSEC-5day group exhibited significantly increased expression of Notch1， Hes1， CD31， and vWF （P<0.01）， significantly decreased p-eNOS/eNOS ratio and sGC expression （P<0.01）， and elevated intracellular Ca²⁺ concentration （P<0.01）. Compared with the LSEC-5day group， the JTCT group showed decreased expression of Notch1， Hes1， CD31， and vWF （P<0.01）， significantly increased p-eNOS/eNOS ratio （P<0.01）， and significantly reduced intracellular Ca²⁺ concentration （P<0.01）. No significant differences were observed in the expression of Notch1， Hes1， eNOS， p-eNOS， CD31， vWF， sGC， or intracellular Ca²⁺ concentration between the Jagged1 group and the Jagged1+JTCT group. Compared with the LSEC-5day group， the sGC knockout group （sGC-KO） showed significantly increased expression of CD31 and vWF （P<0.01）. There was no significant difference in CD31 or vWF expression between the sGC-KO group and the sGC-KO+JTCT group. ConclusionJTCT inhibits the activation of the Notch signaling pathway and restores the activity of the eNOS signaling pathway to suppress sinusoidal capillarization of LSECs， thereby treating liver fibrosis.

Objective:To explore the role and possible mechanism of ACOT11 in renal fibrosis model mice.Methods:A mouse model of renal fibrosis was established by unilateral ureteral obstruction(UUO)(Sham group and UUO7 group),and the expression of ACOT11 in the kidneys of UUO induced fibrosis mouse models was detected by protein immunoblotting and real-time fluorescence quantitative PCR(qRT-PCR).Subsequently,immunohistochemistry,Masson staining,H&E staining,PAS staining,and other experimental methods were used to detect the expression levels of fibrosis biomarkers fibronectin,α-SMA,and COL-1 in the kidneys of control and experimental group mice.In addition,by constructing ACOT11 gene knockout model mice and using the gene knockout model mice to construct a renal fibrosis model,the expression levels of fibrosis biomarkers such as fibronectin,α-SMA,COL-1,as well as fibrosis mechanism pathway related indicators TGF-β and Smad2 in the kidneys of each group of mice were further detected.Results:The results of WB and qRT-PCR experiments showed that the expression of ACOT11 in the kidney tissue of UUO model mice was significantly reduced compared to the Sham group.After knocking out the ACOT11 gene,H&E staining,PAS staining,and Masson staining showed that pathological inflammatory reactions such as abnormal glomerular and tubular structures,inflammatory cell infiltration and interstitial fibrous tissue proliferation in mice were significantly aggravated compared to the control group,and the expression of fibrosis markers Fibronectin,α-SMA,and COL-1 was significantly higher than that of the control group.Conclusion:ACOT11 plays a protective role in mice with unilateral ureteral obstruction model.After ACOT11 gene knockout,the fibrosis biomarkers of the mouse kidney increases and the degree of fibrosis worsens.

Coronavirus disease 2019 (COVID-19) is an acute infectious respiratory disease that has been prevalent since December 2019. Chinese medicine (CM) has demonstrated its unique advantages in the fight against COVID-19 in the areas of disease prevention, improvement of clinical symptoms, and control of disease progression. This review summarized the relevant material components of CM in the treatment of COVID-19 by searching the relevant literature and reports on CM in the treatment of COVID-19 and combining with the physiological and pathological characteristics of the novel coronavirus. On the basis of sorting out experimental methods in vivo and in vitro, the mechanism of herb action was further clarified in terms of inhibiting virus invasion and replication and improving related complications. The aim of the article is to explore the strengths and characteristics of CM in the treatment of COVID-19, and to provide a basis for the research and scientific, standardized treatment of COVID-19 with CM.
Humans ; Drugs, Chinese Herbal/pharmacology* ; COVID-19 Drug Treatment ; SARS-CoV-2/drug effects* ; COVID-19/therapy* ; Medicine, Chinese Traditional/methods* ; Antiviral Agents/pharmacology* ; Animals

OBJECTIVE: To explore the preventive and therapeutic effects of Dahuang Zhechong Pill (DZP) on pulmonary fibrosis and the underlying mechanisms. METHODS: The first key rate-limiting enzyme hexokinase 2 (HK2) of glycolysis was silenced and over-expressed through small interfering RNA and lentivirus using lung fibroblast MRC-5 cell line, respectively. The cell viability, migration, invasion and proliferation were detected by cell counting kit-8, wound healing assay, transwell assay, and flow cytometry. The mRNA and protein expression levels of HK2 were detected by RT-PCR and Western blotting, respectively. The contents of glucose, adenosine triphosphate (ATP) and lactate in MRC-5 cells were determined by enzyme-linked immunosorbnent assay (ELISA). Then, the relationship between miR-29b-2-5p and HK2 was explored by luciferase reporter gene assay. Pulmonary fibrosis cell model was induced by transforming growth factor-β 1 (TGF-β 1) in MRC-5 cells, and the medicated serum of DZP (DMS) was prepared in rats. MRC-5 cells were divided into control, TGF-β 1, TGF-β 1+10% DMS, TGF-β 1+10% DMS+miR-29b-2-5p inhibitor, TGF-β 1+10% DMS+inhibitor negative control, TGF-β 1+10% DMS+miR-29b-2-5p mimic and TGF-β 1+10% DMS+mimic negative control groups. After miR-29b-2-5p mimics and inhibitors were transfected into MRC-5 cells, all groups except control and model group were treated with DMS. The effect of DMS on MRC-5 cells were detected using aforementioned methods and immunofluorescence. Similarly, the contents of glucose, ATP and lactate in each group were measured by ELISA. RESULTS: The mRNA and protein expressions of HK2 in MRC-5 cells were successfully silenced and overexpressed through si-HK2-3 and lentiviral transfection, respectively. After silencing HK2, the mRNA and protein expressions of HK2 were significantly decreased (P<0.01), and the concentrations of glucose, ATP and lactate were also significantly decreased (P<0.05). The proliferation, migration and invasion of MRC-5 cells were significantly declined (P<0.05 or P<0.01), while the apoptosis of MRC-5 cells was significantly increased (P<0.01). After overexpressing HK2, the mRNA and protein expressions of HK2 were significantly increased (P<0.05), and the concentrations of glucose, ATP and lactate were also significantly increased (P<0.05 or P<0.01). The proliferation, migration and invasion of MRC-5 cells were significantly increased (P<0.05 or P<0.01), while the apoptosis of MRC-5 cells was significantly decreased (P<0.05). The relative luciferase activity of 3'UTR-WT+hsa-miR-29b-2-5p transfected with HK2 was significantly decreased (P<0.01). After miR-29b-2-5p mimic and inhibitor were transfected into the MRC-5 cells, DMS intervention could significantly reduce the concentration of glucose, ATP and lactate, and the mRNA and proteins expressions of HK2, phosphofructokinase and pyruvate kinase isoform M2 (P<0.05 or P<0.01). The proliferation, migration and invasion of MRC-5 cells were alleviated (P<0.05 or P<0.01), and the deposition of fibronectin, α-smooth muscle actin, and collagen I were significantly decreased (P<0.05 or P<0.01). CONCLUSIONS Glycolysis is closely related to pulmonary fibrosis. DZP reduced glycolysis and inhibited fibroblasts' excessive differentiation and abnormal collagen deposition through the miR-29b-2-5p/HK2 pathway, which played a role in delaying the process of pulmonary fibrosis.
MicroRNAs/genetics* ; Glycolysis/genetics* ; Animals ; Pulmonary Fibrosis/metabolism* ; Humans ; Drugs, Chinese Herbal/therapeutic use* ; Hexokinase/genetics* ; Cell Line ; Cell Proliferation/drug effects* ; Rats, Sprague-Dawley ; Rats ; Cell Movement/drug effects* ; Male ; Cell Survival/drug effects* ; Signal Transduction/drug effects*

Objective To investigate the mechanism of Wenjing Decoction in treating endometriosis(EMT)using bioinformatics methods and in vitro experiments.Methods The active components and corresponding targets of Wenjing Decoction were obtained from the TCMSP database,while EMT-related targets were identified using the GEO database.Functional enrichment analysis was conducted on the targets to predict core targets for treating EMT with Wenjing Decoction.Molecular docking was performed on core targets-drug ligands,and in vitro experiments validated the findings.Results Through screening the TCMSP database,117 active components of Wenjing Decoction were identified,corresponding to 248 targets;5 312 EMT-related differential genes were gathered from GEO database,identifying 97 potential targets of Wenjing Decoction for treating EMT,with core targets being IL6,TNF and EGFR.Functional enrichment analysis of EMT differential genes showed enrichment in pathways such as neuroactive ligand-receptor interaction,MAPK signaling pathway,endocytosis,calcium signaling pathway,autophagy and PI3K-Akt signaling pathway.Molecular docking showed that IL6,TNF,EGFR bind stably to their corresponding drug ligands.In vitro experiments indicated that Wenjing Decoction could inhibit the PI3K/Akt/mTOR pathway,promote LC3 Ⅰ to LC3 Ⅱ conversion,enhance the expression of Beclin-1,and reduce P62 expression.Moreover,Wenjing Decoction could hinder the expression of the endometriosis-specific biomarker CA125,decrease EGFR,IL-6 and TNF-α expressions in ectopic endothelial cells,inhibiting proliferation.Conclusion Wenjing Decoction can treat EMT through multiple pathways and targets,with the key mechanism being the reversal of autophagy inhibition via down-regulating of the PI3K/Akt/mTOR pathway.

OBJECTIVE: Acupuncture therapies are known for their effectiveness in treating a variety of gastric diseases, although the mechanisms underlying these effects are not fully understood. This study tested the effectiveness of electroacupuncture (EA) at acupoints Zhongwan (RN12) and Weishu (BL21) for managing gastric motility disorder (GMD) and investigated the underlying mechanisms involved. METHODS: A GMD model was used to evaluate the impact of EA on various aspects of gastric function including the amplitude of gastric motility, electrogastrogram, food intake, and the rate of gastric emptying. Immunofluorescence techniques were used to explore the activation of spinal neurons by EA, specifically examining the presence of cholera toxin B subunit (CTB)-positive neurons and fibers emanating from acupoints RN12 and BL21. The stimulation of γ-aminobutyric acid (GABA)-ergic neurons in the spinal dorsal horn, the inhibition of sympathetic preganglionic neurons in the spinal lateral horn, and their collective effects on the activity of sympathetic nerves were examined. RESULTS: EA at RN12 and BL21 significantly improved gastric motility compromised by GMD. Notably, EA activated spinal neurons, with CTB-positive neurons and fibers from RN12 and BL21 being detectable in both the dorsal root ganglia and the spinal dorsal horn. Further analysis revealed that EA at these acupoints not only stimulated GABAergic neurons in the spinal dorsal horn but also suppressed sympathetic preganglionic neurons in the spinal lateral horn, effectively reducing excessive activity of sympathetic nerves triggered by GMD. CONCLUSION EA treatment at RN12 and BL21 effectively enhances gastric motility in a GMD model. The therapeutic efficacy of this approach is attributed to the activation of spinal neurons and the modulation of the spinal GABAergic-sympathetic pathway, providing a neurobiological foundation for the role of acupuncture in treating gastric disorders. Please cite this article as: Zhang MT, Liang YF, Dai Q, Gao HR, Wang H, Chen L, Huang S, Wang XY, Shen GM. A spinal neural circuit for electroacupuncture that regulates gastric functional disorders. J Integr Med. 2025; 23(1): 56-65.
Electroacupuncture ; Animals ; Male ; Acupuncture Points ; Stomach Diseases/physiopathology* ; Rats, Sprague-Dawley ; Gastrointestinal Motility ; Rats ; Gastric Emptying ; Neurons ; Spinal Cord ; Stomach/physiopathology*

Objective:To establish a linear calibration method using two reference substances for seven characteristic peaks of Cassiae Semen under complicated chromatographic condition,and to optimize the method.Methods:Using 15 different types of screened chromatographic columns and 2 components as reference compounds pair,the linear calibration method with 2 reference substances was established to predict the retention time of the other 5 components,and the method was verified by unknown chromatographic columns and unknown samples.Combined with column confirmation number and average coincidence rate of target peaks,the location results were compared comprehen-sively,and the method was optimized according to the defect under the influence of complicated chromatographic condition.Results:The average conformity rate of the target peak of the method before optimization was 73.3％,and the average conformity rate of the target peak of the optimized method was 98.7％.The optimized method has a high-er average peak coincidence rate and a wider range of applicability for the chromatographic column.Conclusion:The optimized linear calibration method using two reference substances can assist the localization analysis of chromato-graphic peaks in the characteristic chromatogram under complicated chromatographic condition.