OBJECTIVE To investigate the effects of total flavonoids from Carthamus tinctorius L. （TFCTL） on hepatic stellate cell （HSC） activation based on the microRNA （miRNA）-204/NUAK family SNF1-like kinase 1 （NUAK1）/Hippo signaling axis， thereby elucidating the potential mechanism underlying their antifibrotic effects. METHODS The HSC-T6 cells were divided into control group， model group， TFCTL low-concentration group （20 μg/mL）， TFCTL medium-concentration group （40 μg/mL）， and TFCTL high-concentration group （60 μg/mL）. Except for control group， the remaining groups were treated with 5 ng/mL of transforming growth factor-β to induce the activation of hepatic stellate cells， followed by the addition of corresponding drug solutions/culture medium and incubation for 24 hours. Cell apoptosis was assessed， the expression levels of α-smooth muscle actin （α-SMA）， type Ⅰ collagen （Collagen Ⅰ） and proteins associated with the Hippo/Yes-associated protein （YAP） pathway [YAP， large tumor suppressor kinase 1 （LATS1）， and mammalian STE20-like kinase 1 （MST1）] were detected. Additionally， cell transfection was used to investigate the activity of the miRNA-204/NUAK1/Hippo signaling axis at both the genetic and protein levels. RESULTS After intervention with TFCTL， the apoptosis rate of HSC-T6 cells and the protein expressions of MST1 （except for the TFCTL high-concentration group） and LATS1 were significantly increased （P＜0.05）， while the protein expressions of α-SMA， CollagenⅠ， and YAP （except for the TFCTL medium-concentration group） were significantly decreased （P＜0.05）. Further results from cell transfection experiments revealed that after transfection with miRNA-204 mimics， the mRNA it’s protein expressions of α-SMA， CollagenⅠ， NUAK1， and YAP in HSC-T6 cells were significantly decreased （P＜0.05）， while the mRNA and protein expressions of LATS1 and the mRNA expression of MST1 were significantly increased （P＜0.05）. Conversely， the results were opposite following transfection with miRNA-204 inhibitors. CONCLUSIONS TFCTL can exert anti-hepatic fibrosis effects by up-regulating the expression of miRNA-204， thereby down- regulating the expressions of NUAK1， inactivating the Hippo/YAP pathway， which in turn suppresses the activation of HSC and promotes their apoptosis.

BackgroundIn recent years， the incidence of depression among adolescents has been increasing steadily， posing a serious threat to their physical and mental health and even leading to severe consequences such as self-harm and suicide. At the same time， the detection rate of subclinical depression symptoms among adolescents is even higher. Although these symptoms do not meet the clinical diagnostic criteria， they have significantly affected their quality of life， and their persistence over time may further develop into depression. Therefore， in-depth exploration of adolescent depression symptoms and the predictive factors holds significant practical significance and research value. However， up to now， no large-scale investigation and research on depression symptoms among children and adolescents has been conducted in Inner Mongolia Autonomous Region. ObjectiveTo understand the prevalence of depressive symptoms among children and adolescents in Inner Mongolia Autonomous Region， in order to provide references for formulating scientific and effective prevention strategies and intervention measures. MethodsBy using the cluster stratified random sampling method， 6 281 students from the third grade of primary school to the second grade of high school in 12 leagues and cities of Inner Mongolia Autonomous Region were selected in March 2024. A self-designed questionnaire and the Self-rating Depression Scale （SDS） were used for on-site investigation. ResultsA total of 6 058 （96.45%） children and adolescents completed the valid questionnaire survey， and 2 728 cases （45.03%） were found to have depressive symptoms. There were statistically significant differences in the detection rates of depressive symptoms among children and adolescents of different genders， ages， whether they were only children， different family types， family monthly income， parents' educational levels， and whether the mother was employed （χ²=33.769， 40.618， 48.593， 29.972， 142.648， 195.999， 168.190， 5.445， P<0.05 or 0.01）.The results of the Logistic regression analysis showed that for children and adolescents， being female， aged between 12 and 16， over 16 years old， not being an only child， living in a reconstituted family， having a monthly family income of less than 5 000 yuan， and having parents with an education level of primary school or below were predictors of depressive symptoms （OR=1.241， 1.427， 1.273， 1.177， 1.549， 1.278， 1.462， 1.417， 1.514， 1.929， 1.660， 1.528， P<0.05 or 0.01）. ConclusionThe detection rate of depressive symptoms among children and adolescents in Inner Mongolia Autonomous Region is relatively high. Factors that may predict depressive symptoms in children and adolescents include female gender， ages between 12 and 16， ages over 16 years old， non-only children， families with a restructured structure， monthly family income of less than 5 000 yuan， and parents with an education level of primary school or below. ［Funded by Science and Technology Planning Project of the Inner Mongolia Autonomous Region （number， 2022YFSH0119）］

ObjectiveTo investigate the molecular mechanisms by which Wenyang Jiedu granules （WYJD） alleviate influenza A virus （IAV）-induced pneumonia based on single-cell transcriptome sequencing. MethodsThirty female BALB/c mice were randomly divided into a blank control group （Control）， IAV group， and WYJD low-， medium-， and high-dose groups （WYJD-L， WYJD-M， WYJD-H； 2.925， 5.85， 11.7 g·kg^-1， n=6）. Except for the Control group， all other groups were intranasally inoculated with IAV subtype H1N1 （A/PR/8/34） to establish an infection model. Two hours after modeling， drug administration was initiated and continued for 5 consecutive days， with daily monitoring of body weight and general condition. On day 6， mice were sacrificed and samples were collected. Lung index was calculated， and histopathological examination of lung tissue was performed. Lung tissues from the Control， IAV， and WYJD-H groups were subjected to single-cell transcriptome sequencing （n=3）， focusing on type I alveolar epithelial cells （AT1） to analyze changes in gene expression and signaling pathways. Western blot was used to detect the expression changes of relevant proteins to validate the single-cell sequencing results. ResultsCompared with the Control group， the IAV group exhibited significantly decreased body weight （P<0.05） and significantly increased lung index （P<0.05）. Compared with the IAV group， all WYJD-treated groups exhibited significantly increased body weight （P<0.01） and significantly decreased lung index （P<0.01）. Single-cell sequencing analysis revealed that WYJD inhibited overactivation of interferon and inflammatory signaling pathways in AT1 cells after IAV infection， including interferon-γ response， interferon-α response， tumor necrosis factor-α/nuclear factor-κB （TNF-α/NF-κB）， and interleukin-6/Janus kinase/signal transducer and activator of transcription 3 （IL-6/JAK/STAT3） pathways. Compared with the Control group， the number of AT1 cells in the IAV group showed a decreasing trend. Compared with the IAV group， the WYJD-H group showed an increasing trend， although neither difference was statistically significant. Further analysis of AT1 cell subpopulation gene expression showed that， compared with the Control group， the IAV group exhibited increased expression of pro-apoptotic genes FAS cell surface death receptor （FAS） and cyclin-dependent kinase inhibitor 1A （CDKN1A）， a significant increase in tumor protein p53 （Tp53） expression （P<0.05）， and significant decreases in expression of the AT1 marker gene advanced glycosylation end-product-specific receptor （AGER） and membrane structural gene caveolin1 （CAV1） （P<0.05， P<0.01）. Compared with the IAV group， the WYJD-H group showed significantly decreased expression of FAS， CDKN1A， and Tp53 （P<0.05， P<0.01）， and significantly increased expression of AGER and CAV1 （P<0.05， P<0.01）. Regarding interferon response-related genes， compared with the Control group， the IAV group showed increased expression of interferon-stimulated gene 15 （ISG15）， interferon-induced protein with tetratricopeptide repeats 3 （IFIT3）， signal transducer and activator of transcription 2 （STAT2）， bone marrow stromal cell antigen 2 （BST2）， and C-X-C motif chemokine ligand 10 （CXCL10）， with a significant increase in 2′，5′-oligoadenylate synthetase-like protein 1 （OASL1） （P<0.05）. Compared with the IAV group， the WYJD-H group showed significantly decreased expression of all the above genes， with highly significant differences for ISG15， IFIT3， STAT2， BST2， and OASL1 （P<0.01）， and a significant difference for CXCL10 （P<0.05）. Among inflammation-related genes， compared with the Control group， the IAV group showed significantly increased expression of intercellular adhesion molecule 1 （ICAM1）， tumor necrosis factor alpha-induced protein 3 （TNFAIP3）， keratin 8 （KRT8）， tumor necrosis factor receptor superfamily member 1A （TNFRSF1A）， and TNFRSF1B （P<0.01）， and increased expression of NFKBIA， a negative regulator of NF-κB （P<0.05）. Compared with the IAV group， the WYJD-H group showed significantly decreased expression of KRT8 and TNFRSF1B （P<0.05）， while ICAM1， NFKBIA， TNFAIP3， and TNFRSF1A showed decreasing trends without statistical significance. Western blot validation showed that， compared with the Control group， protein expression levels of ISG15， FAS， p53， and phosphorylated p65 （p-p65） in lung tissue of the IAV group were significantly increased （P<0.05， P<0.01）. Compared with the IAV group， the WYJD-H group showed significantly decreased expression of these proteins （P<0.05， P<0.01）. ConclusionWYJD may alleviate viral pneumonia by targeting gene expression in AT1 cells， inhibiting overactivated interferon and inflammatory signaling pathways after IAV infection， and downregulating pro-apoptotic signaling， thereby reducing alveolar epithelial injury.

ObjectiveTo establish a mouse model of hypoxic pulmonary hypertension (HPH) through intermittent hypoxia induction, and to develop a comprehensive and reliable evaluation system for the HPH model, thereby providing experimental evidence for mechanistic studies and translational applications related to this disease. MethodsTwenty-four male specific pathogen-free (SPF) BALB/c mice were randomly divided into a control group and a model group, with 12 mice per group. Mice in the model group were placed in a hypobaric oxygen chamber control system to establish the HPH model, whereas mice in the control group received no intervention. After 28 days of modeling, a comprehensive evaluation of the pathophysiological characteristics of both groups was performed using a general condition scoring scale, echocardiography, hemodynamic measurements, blood gas analysis, hematological tests, organ coefficient determination, histopathological examination, and enzyme-linked immunosorbent assay (ELISA). In addition, correlation analyses were conducted among echocardiographic parameters, the contents of endothelin‑1 (ET‑1) and N‑terminal pro‑B‑type natriuretic peptide (NT‑proBNP) in lung tissue, and other measured indices. ResultsWith prolonged modeling duration time, body weight, water intake, and activity level of mice in the model group were significantly decreased compared with the control group (P<0.05). Echocardiography showed that, compared with the control group, the model group exhibited increased right ventricular dimensions (P<0.01), right ventricular anterior wall thickening (P<0.01), and widened main pulmonary artery diameter (P<0.01), whereas the peak systolic velocity across the pulmonary valve, tricuspid annular plane systolic excursion, and peak systolic velocity of the tricuspid annulus were significantly decreased (P<0.001); tricuspid regurgitation was observed in some model animals. Hemodynamic results revealed that right ventricular systolic pressure was elevated in the model group compared with the control group (P<0.001). Blood gas analysis showed that pH, partial pressure of oxygen, oxygen saturation, actual bicarbonate, and total carbon dioxide were all decreased in the model group compared with the control group (P<0.01). Hematological indices demonstrated that lymphocyte counts (P<0.05) and reticulocyte counts (P<0.001) were decreased in the model group compared with the control group. Compared with the control group, the organ coefficients of heart and lung in the model group were significantly increased (P<0.05 and P<0.001). Pathological examination revealed that the right ventricular hypertrophy index was significantly increased in the model group (P<0.001), with varying degrees of damage to the right ventricle, pulmonary artery, and pulmonary vessels; both the pulmonary artery wall thickness percentage and the pulmonary wall area percentage were significantly elevated (P<0.001). ELISA results showed that the levels of ET‑1 and NT‑proBNP in lung tissues were significantly increased in the model group compared with the control group (P<0.05 and P<0.001, respectively). Correlation analysis indicated that some echocardiographic parameters were highly correlated with multiple indices in the development of HPH (P<0.05). ConclusionEchocardiography can accurately assess a series of hemodynamic changes in HPH, including right ventricular structural, functional impairment, and pulmonary hypertension. Laboratory tests not only help verify whether the model has been successfully established, but also provide deeper insights into the pathogenesis of HPH, evaluate the effects of interventions, and offer scientific evidence for clinical outcomes. Pathological examination can further confirm the alterations in pulmonary vascular remodeling and increased right heart load. This multimodal analysis provides a reliable animal model and evaluation paradigm for both basic and translational research on HPH, and is of great significance for exploring disease mechanisms and developing precision therapeutic strategies.

Background: Alzheimer’s disease (AD) is the most prevalent neurodegenerative disorder. Cellular drug screening represents a key approach in the preclinical discovery of AD therapeutics. Moreover, selecting an appropriate cellular model is essential for exploring the mechanisms underlying AD pathogenesis. Objective: This study aimed to establish a progressive strategy integrating cell model construction, monomer evaluation, and effective component group-based drug discovery for the development of novel therapeutics targeting AD. Methods: A novel AD12 cell line stably expressing the double-transgenic genes amyloid precursor protein (APP)695/presenilin 1(PS1)-ΔE9 was first constructed and subjected to lipopolysaccharide (LPS) stimulation. The resulting LPS-induced neuroinflammatory response was characterized using metabolomic analysis. Subsequently, 16 alkaloids derived from traditional Chinese medicine(TCM) were screened with the LPS-AD12 cell model. Molecular docking was then performed to assess the binding affinity of benzoylaconitine (BAC) with APP and PS1 proteins. A targeted metabolomics approach focusing on amino acids and the tricarboxylic acid (TCA) cycle was applied to BAC-treated LPS-AD12 cells. Finally, aconite extract was administered to five-familial-AD-mutations(5xFAD) transgenic mice, and therapeutic efficacy was evaluated through phenotypic and metabolomic analyses. Results: LPS stimulation induced neuroinflammation in AD12 cells, characterized by elevated levels of interleukin-6, amyloid-β (Aβ ), and PS1. The primary metabolic pathways affected in AD12 cells included alanine, aspartate, and glutamate metabolism, as well as arginine biosynthesis. Notably, BAC significantly reduced the levels of Aβ and interleukin-6 in LPS-AD12 cells. Furthermore, molecular docking analyses revealed a strong binding affinity of BAC with APP and PS1 proteins. Targeted metabolomics analysis indicated that the pharmacological activity of BAC was primarily associated with its anti-inflammatory effects, modulation of the cholinergic signaling pathway, and regulation of energy metabolism. Behavioral experiments in 5xFAD mice, together with immunofluorescence assessments of brain tissues (amyloid-β, glial fibrillary acidic protein, and ionized calcium-binding adapter molecule 1), demonstrated that aconite extract treatment significantly alleviated AD-related symptoms. In addition, metabolomics-based mechanistic analyses revealed that serine, acetylcholine, ornithine, and gamma-aminobutyric acid exhibited consistent trends in both brain tissue and serum samples from treated mice. Conclusion: This study established a novel AD12 cell model and an effective, reliable LPS-induced cell-based screening system. Additionally, we proposed a new drug discovery strategy integrating cell model construction, monomer evaluation, and effective component group-based drug discovery. This screening system has the potential to accelerate the identification of anti-AD agents and provides a feasible framework for subsequent in-depth investigations into new drug development from TCM, thereby contributing to the modernization of TCM-derived therapeutics.

Clostridioides difficile infection（CDI）is a common cause of antibiotic-associated diarrhea，posing a significant burden on global healthcare systems.Clostridioides difficile（C.difficile）spreads through spores outside the body，germinates and grows into vegetative cells in the intestines，and releases toxins to cause disease.Bile acids are crucial metabolites in the gut，where primary and secondary bile acids play important roles throughout the lifecycle of C.difficile.This article reviews bile acids metabolism，their impact on C.difficile，and how they influence CDI treatment through antibiotics and fecal microbiota transplantation（FMT）.The aim is to provide new insights and reference for clinical management and research of CDI.

Objective To investigate the relationship between the levels of soluble FMS-like tyrosine ki-nase-1(sFlt-1),monocyte chemoatgulant protein-1(MCP-1),and soluble lectin-like oxidized low-density lipo-protein receptor 1(sLOX-1)in the serum of patients with coronary heart disease and the disease state and prognosis.Methods A total of 126 patients with coronary heart disease treated in Zhongshan Hospital,Dalian University from May 2022 to May 2024 were selected as study objects.The patients were divided into good prognosis group(n=78)and poor prognosis group(n=48)according to prognostic results.The serum sFlt-1,MCP-1 and sLOX-1 levels were detected by enzyme-linked immunosorbent assay.The degree of coronary artery stenosis in patients was measured by coronary angiography,as indicated by the Gensini score.Spearman correlation analysis was used to analyze the correlation between Gensini score and serum sFlt-1,MCP-1 and sLOX-1 levels.Multivariate Logistic regression was used to analyze the factors affecting the prognosis of pa-tients with coronary heart disease.The predictive efficacy of serum sFlt-1,MCP-1 and sLOX-1 levels for the prognosis of patients with coronary heart disease was evaluated by receiver operating characteristic(ROC)curve.Results The age in the poor prognosis group was older than that in the good prognosis group(P＜0.05),Gensini score was higher than that in the good prognosis group(P＜0.05).Compared with the poor prognosis group,the serum sFlt-1,MCP-1 and sLOX-1 levels in the good prognosis group were lower(P＜0.05).Gensini score was positively correlated with the serum sFlt-1,MCP-1 and sLOX-1 levels.Multivariate Logistic regression analysis showed that old age,high Gensini score,high sFlt-1 level,high MCP-1 level and high sLOX-1 level were all risk factors effecting the prognosis of patients with coronary heart disease(P＜0.05).ROC curve analysis showed that the areas under the curve(AUC)of serum sFlt-1,MCP-1,and sLOX-1 for predicting the prognosis of coronary heart disease patients were 0.787(95％CI:0.703-0.870),0.815(95％CI:0.741-0.890)and 0.795(95％CI:0.715-0.876),respectively.The AUC for predicting the prognosis of coronary heart disease patients using a combination of three was 0.923(95％CI:0.876-0.970).Conclusion The serum levels of sFlt-1,MCP-1 and sLOX-1 in patients with coronary heart disease are signifi-cantly increased,and are positively correlated with the degree of coronary artery stenosis.The levels of sFlt-1,MCP-1 and sLOX-1 in serum have a certain predictive effect on the prognosis of patients with coronary heart disease.

Atherosclerosis, a chronic inflammatory disease, is markedly influenced by both immune and inflammatory reactions throughout its progression. Dendritic cells, as pivotal antigen-presenting entities, play a crucial role in the initiation of immune responses and the preservation of immunological homeostasis. Accumulating data indicates that dendritic cells are present in healthy arteries and accumulate significantly in atherosclerotic plaques. Novel immunotherapeutic approaches and vaccination protocols have yielded substantial clinical advancements in managing chronic inflammatory diseases, with dendritic cell-centric modalities emerging for atherosclerotic management. In this review, we delineate the essential functions and underlying mechanisms of dendritic cells and their subsets in the modulation of atherosclerotic inflammation and immune responses. We underscore the immense promise of dendritic cell-based immunotherapeutic strategies, including vaccines and innovative combinations with nanotechnological drug delivery platforms for atherosclerosis treatment. We also discuss the challenges associated with dendritic cell immunotherapy and provide perspectives on the future direction of this field.

T-cell acute lymphoblastic leukemia (T-ALL) is a highly aggressive hematologic malignancy with a poor prognosis, despite advancements in treatment. Many patients struggle with relapse or refractory disease. Investigating the role of the super-enhancer (SE) regulated gene ubiquitin-specific protease 20 (USP20) in T-ALL could enhance targeted therapies and improve clinical outcomes. Analysis of histone H3 lysine 27 acetylation (H3K27ac) chromatin immunoprecipitation sequencing (ChIP-seq) data from six T-ALL cell lines and seven pediatric samples identified USP20 as an SE-regulated driver gene. Utilizing the Cancer Cell Line Encyclopedia (CCLE) and BloodSpot databases, it was found that USP20 is specifically highly expressed in T-ALL. Knocking down USP20 with short hairpin RNA (shRNA) increased apoptosis and inhibited proliferation in T-ALL cells. In vivo studies showed that USP20 knockdown reduced tumor growth and improved survival. The USP20 inhibitor GSK2643943A demonstrated similar anti-tumor effects. Mass spectrometry, RNA-Seq, and immunoprecipitation revealed that USP20 interacted with hypoxia-inducible factor 1 subunit alpha (HIF1A) and stabilized it by deubiquitination. Cleavage under targets and tagmentation (CUT&Tag) results indicated that USP20 co-localized with HIF1A, jointly modulating target genes in T-ALL. This study identifies USP20 as a therapeutic target in T-ALL and suggests GSK2643943A as a potential treatment strategy.

Objective To establish a method for the simultaneous determination of peimine,peiminine,and hupehenine in Juhong pills,to investigate the raw material usage of Fritillaria thunbergii Miq.,and to preliminarily develop a detection approach for identifying adulteration with Fritillaria hupehensis Hsiao et K.C.Hsia.Methods The high-performance liquid chromatography-triple quadruple mass spectrometry(HPLC-MS/MS)was performed on an Agilent Poroshell 120 SB C18 column(2.1 mm×100 mm,2.7 μm)with a mobile phase consisting of acetonitrile-methanol(1∶1)and 0.1%formic acid under gradient elution.The flow rate was set at 0.3 mL·min-1.The column temperature was maintained at 35℃,and the injection volume was 2 μL.Electrospray ionization(ESI)in positive ion mode and multiple reaction monitoring were employed to quantify the three components in 170 batches of Juhong pills.Simulated positive samples with varying adulteration ratios of Fritillaria hupehensis Hsiao et K.C.Hsia were prepared to establish a simple yet efficient detection limit for adulteration.Results Three components showed good linear correlation within their ranges(r≥0.997 5),and the averaged recoveries ranged from 92.0%-106.2%.A total of 61 batches were suspected of Fritillaria thunbergii Miq.adulteration with Fritillaria hupehensis Hsiao et K.C.Hsia in raw material inputs,with the peimine to peiminine ratio below 1.15.Among these samples,27 batches were large honey-bound pills with hupehenine levels of 2.636-9.939 μg·g-1;34 batches were water-honeyed pills showing significantly higher hupehenine contamination at 6.752-48.137 μg·g-1.Conclusion The established method is simple,reliable,and accurate for the quality control of Fritillaria thunbergii Miq.adulteration in Juhong pills without imposing significant additional costs.