A natural experiment is a causal inference study design grounded in real-world research. Its core concept is to use quasi-random variation in exposure generated by external factors to evaluate the effects of interventions or exposures on outcomes. It is particularly suitable for research settings in which randomized controlled trials （RCTs） are infeasible or ethically constrained， and it combines causal inference capacity with relatively strong external validity. This article provides an overview of the definition， key features， and current application status of natural experiments， and proposes the application scenarios for applying natural experiment strategies to the evaluation of therapeutic effects in traditional Chinese medicine （TCM）， including policy implementation or policy change evaluation， assessment of variations in clinical practice within hospitals， and evaluation of differences in resource allocation across regions or institutions. The design pathway includes starting from clinical or policy-related TCM questions， systematically collecting relevant data， performing statistical modeling and effect estimation， and interpreting effect estimates in the context of clinical practice. It further analyzes the advantages and challenges of using natural experiments in TCM.

A natural experiment is a causal inference study design grounded in real-world research. Its core concept is to use quasi-random variation in exposure generated by external factors to evaluate the effects of interventions or exposures on outcomes. It is particularly suitable for research settings in which randomized controlled trials （RCTs） are infeasible or ethically constrained， and it combines causal inference capacity with relatively strong external validity. This article provides an overview of the definition， key features， and current application status of natural experiments， and proposes the application scenarios for applying natural experiment strategies to the evaluation of therapeutic effects in traditional Chinese medicine （TCM）， including policy implementation or policy change evaluation， assessment of variations in clinical practice within hospitals， and evaluation of differences in resource allocation across regions or institutions. The design pathway includes starting from clinical or policy-related TCM questions， systematically collecting relevant data， performing statistical modeling and effect estimation， and interpreting effect estimates in the context of clinical practice. It further analyzes the advantages and challenges of using natural experiments in TCM.

Objective To investigate the potential protective mechanism of tanshinone ⅡA against acetaminophen （APAP）-induced acute liver injury. Methods Thirty mice were randomly divided into the normal group, model group, low-dose tanshinone ⅡA group （5 mg/kg）, medium-dose tanshinone ⅡA group （10 mg/kg） and high-dose tanshinone ⅡA group （20 mg/kg）. All groups were intragastrically administered once daily for 7 consecutive days. On the 7th day, except for the normal group, mice in the remaining groups were intraperitoneally injected with 400 mg/kg APAP to establish the acute lung injury （ALI） model. The protective effect of tanshinone ⅡA was evaluated based on the liver weight ratio, the levels of aspartate aminotransferase （AST） and alanine aminotransferase （ALT）, and HE staining. Targets of tanshinone ⅡA were predicted by the Traditional Chinese Medicine Systems Pharmacology （TCMSP）, and a shared target protein-protein interaction （PPI） network was constructed by Search Tool for the Retrieval of Interacting Genes/Proteins （STRING） 11.5 in combination with disease targets from GeneCards. The Gene Ontology （GO） and Kyoto Encyclopedia of Genes and Genomes （KEGG） enrichment analysis was performed by Database for Annotation, Visualization and Integrated Discovery （DAVID）, and the component-target-pathway network was constructed. The molecular docking between tanshinone ⅡA and the core targets was performed by AutoDock and visualised. Results Compared with the normal group, the liver weight ratio and serum aminotransferase level of mice with acute liver injury were significantly elevated （P<0.05）, and the pathological injury of liver tissue was obvious. After tanshinone ⅡA treatment, the above index were significantly reduced （P<0.05）, and the pathological injury was significantly improved. Further network pharmacological analysis was carried out and found that KEGG pathway enrichment analysis focused on IL-17 signaling pathway and tumor necrosis factor （TNF） signaling pathway, etc. In addition, the PPI core network showed that the key targets of tanshinone ⅡA to ameliorate acute liver injury mainly included TP53, AKT1, SRC, TNF and JUN. The molecular docking results showed that tanshinone ⅡA had high binding scores with the targets of MMP9, NFKB1, TNF, EP300 and SMAD3; and was able to bind to the targets of TP53, AKT1, SRC and JUN. Conclusion Tanshinone ⅡA may play a protective role against acute liver injury by regulating genes such as AKT1, JUN and TNF, and participating in signaling pathways such as TNF and IL-17.

Objective: To understand the current status and related factors of oral health behavior in preschool children, so as to provide scientific basis for improving parents oral health care abilities. Methods: From March 5 to 12, 2024, using a stratified cluster sampling method, an online questionnaire survey was conducted among parents of 1 381 preschool children aged 3-6 from 6 selected kindergartens of a district in Beijing. A structural equation model (SEM) was constructed based on the Theory of Planned Behavior framework to analyze the factors associated with preschool children s oral health behaviors. Results: The rates of parental assistance in brushing teeth, regular oral examination, and postprandial rinsing were 44.4%, 18.6% and 23.6%, respectively. Attitude, subjective norms were positively correlated with oral health behavior intention ( r =0.61, 0.55); attitude, perceived behavioral control, and behavioral intention were positively correlated with oral health behavior ( r =0.14, 0.13, 0.16) (all P < 0.01 ). The results of the SEM showed that attitude and subjective norms had a significant direct effect on oral health behavior intention ( β = 0.42 , 0.40), and the direct effect of perceived behavioral control on oral health behavior was statistically significant ( β =0.42) (all P <0.01). Conclusions The overall level of oral health behavior among preschool children needs to be improved. Perceived behavioral control is the most direct positive related factor of oral health behavior. In the future, intervention measures should focus on perceived behavioral control as the entry point and provide systematic intervention guidance in combination with planned behavior theory.

Background/Aims: Acetyl coenzyme A (acetyl-CoA) is one of the most essential metabolites in cell metabolism but its function and concentration in hepatocellular carcinoma (HCC) remain elusive and controversial. Methods: A comprehensive analysis of acetyl-CoA levels and acetyl-CoA synthetase 2 (ACSS2) expression across a range of samples, including patient specimens from both hepatitis B virus (HBV) positive and HBV negative HCC individuals, HBV-transgenic mouse HCC models, and multiple cell lines. Furthermore, to evaluate the functional significance of ACSS2 in HBV-related HCC, we implemented both genetic and pharmacological inhibition strategies targeting ACSS2. Molecular mechanism and mitophagy assessment were revealed by cleavage under target and tagmentation sequencing, RNA sequencing, bioinformatic analyses, transmission electron microscopy and JC-1 staining. Results: Our study revealed a distinct metabolic signature of HBV-related HCC, marked by elevated acetyl-CoA, which was driven by ACSS2. ACSS2 was upregulated by the carbohydrate response element-binding protein in HBV-related HCC. Furthermore, ACSS2 improved tumor cell proliferation, an effect that was dependent on its enzymatic activity. Mechanistically, ACSS2-induced acetyl-CoA accumulation activated voltage-dependent anion channels 1 transcription through increased H3K27ac occupancy, which subsequently promoted mitophagy and HBV-related HCC tumorigenesis. Notably, targeting ACSS2 by depletion or inhibition with a catalytic inhibitor significantly suppressed tumor growth. Conclusions These findings not only illustrate the interplay between metabolic reprogramming, epigenetic modification, and tumorigenesis in the context of HBV infection, but also highlight ACSS2 as a novel metabolic vulnerability in HBV-related HCC. Therefore, targeting ACSS2 could be a novel strategy against HBV-related HCC.

Objective:To explore the influence of artificial intelligence(AI)-assisted compressive sensing(ACS)technique with different acceleration factors on the image quality and scan time of three dimensional time-of-flight magnetic resonance angiography(3D-TOF-MRA)for brain.Methods:Thirty participants who underwent brain magnetic resonance imaging(MRI)at Tongzhou Branch of Beijing Friendship Hospital were recruited.All subjects underwent imaging scans about four different parameters:a non-accelerated technique(control group),ACS technique integrated with acceleration factor of 4.03(ACS4 group),ACS technique integrated with acceleration factor of 5.02(ACS5 group),and ACS technique integrated with acceleration factor of 6.06(ACS6 group).The image clarity,ranking of imaging capabilities of distal branch blood vessels and the ratio of pseudo-stenosis were qualitatively analyzed.The signal-to-noise ratio(SNR),contrast-to-noise ratio(CNR),edge sharpness,and scan time were quantitatively analyzed.Results:There was not difference in vessel clarity among three ACS groups at the proximal and middle intracranial segments.For distal segments of blood vessel,the ACS4 group[3.0(3.0,3.0)]and ACS5 group[3.0(3.0,3.0)]were better than ACS6[2.5(2.0,3.0)](q=29.800,27.500,P<0.05).The imaging capabilities of distal-branch vessel of ACS4 group and ACS5 group were better than ACS6 group.There was no stenosis in the proximal and middle segments of the images of the three ACS group,and there were no stenosis in the images of distal vessels of ACS4 group and ACS5 group.A total of 5 cases were pseudo-stenosis in the distal vessels of ACS6 group.Compared with the control group,the incidence of pseudo-stenosis in the distal vessels of ACS6 group was 16.7%.The SNR and CNR values of quantitative analysis for proximal vessels in ACS6 group were higher than them in ACS4 group(q=27.800,26.200,P<0.05),and there was not significant difference in them among ACS4 group,ACS6 group and ACS5 group(P>0.05).The differences of SNR and CNR values in the middle and distal segments of blood vessels among different groups were not significant(P>0.05).There was not significant difference in the edge sharpness of blood vessels among ACS4 group,ACS5 group,and ACS6 group(P>0.05),while all of them were higher than those of control group,and the differences were significant(q=48.150,53.367,44.883,P<0.001).Compared with control group,the scan-time of ACS4 group was reduced by 55.19%,and that of ACS5 group was reduced by 64.07%,and that of ACS6 group was reduced by 70%.Conclusion:ACS technique can accelerate the imaging speed of brain 3D-TOF-MRA and ensure image quality.It is clinically recommended to set the ACS acceleration factor as 5.02 to undergo brain 3D-TOF-MRA scans.

AIM To investigate the impact of varying dosages of Supplemented Wendan Decoction on the PI3K/Akt/FOXO1 glycolipid metabolic pathway in 3T3-L1 adipocytes.METHODS The CCK-8 assay was used to determine the concentration of Supplemented Wendan Decoction-medicated serum.The mature adipocytes differentiated from 3T3-L1 preadipocytes after induction were further divided into the blank control group,the model group,the rosiglitazone group(10 mg/L),and the Supplemented Wendan Decoction groups(5％,10％,and 20％),followed by the sample collections after 48 hours of treatment.Oil red O staining quantified lipid accumulation in 3T3-L1 adipocytes;extracellular glucose levels were measured using glucose oxidase(GOD)assay;RT-qPCR analyzed mRNA expressions of IRS-1,PI3K,Akt,GLUT4,IL-6,TNF-α and IL-1β;Western blot assessed protein expressions of INSR,IRS-1,PI3K-p85,Akt,FOXO1 and GLUT4.RESULTS No significant changes in cell viability(P＞0.05)were observed in 3T3-L1 preadipocytes exposed to serum containing supplemented Wendan Decoction at different concentrations for 24,48,or 72 hours.The 3T3-L1 preadipocytes held the capacity to differentiate into mature adipocytes within a 14-day induction period.Compared to the model group,all supplemented Wendan Decoction groups exhibited reduced lipid accumulation in adipocytes and downregulated mRNA expression of IRS-1,IL-6,TNF-α and IL-1β(P＜0.01);the low-dose group demonstrated increased mRNA expressions of PI3K and GLUT4(P＜0.05,P＜0.01),alongside elevated protein expressions of INSR,IRS-1,PI3K-p85,Akt and GLUT4(P＜0.05,P＜0.01);the medium-dose group showed enhanced GLUT4 mRNA expression,and upregulated protein expressions of INSR and FOXO1(P＜0.01).After 24 hours intervention,the high-dose Supplemented Wendan Decoction group exhibited increased glucose consumption in adipocytes(P＜0.01),and elevated protein expression of INSR,Akt and FOXO1(P＜0.05,P＜0.01).CONCLUSION Supplemented Wendan Decoction reduces lipid accumulation in adipocytes,regulates glucose and lipid metabolism,and promotes metabolic homeostasis through PI3K/Akt/FOXO1 signaling pathway.

Molecularly imprinted polymers(MIPs)of colchicine(COL)were prepared by precipitation polymerization with COL as template,methacrylic acid(MAA)as monomer,ethylene glycol dimethacrylate(EGDMA)as crosslinking agent,azobisisobutyronitrile(AIBN)as initiator and chloroform as porogen.The basic structure of the prepared MIPs was characterized by scanning electron microscopy(SEM),transmission electron microscopy(TEM)and Fourier transform infrared(FT-IR)spectrometry,and the adsorption properties of MIPs were evaluated by kinetic adsorption,isothermal adsorption and selective adsorption experiments.The results indicated that the prepared MIPs had uniform particles size,with a maximum adsorption capacity of 62.61 mg/g for COL and an imprinting factor of 3.74,and was suitable for sample pretreatment in determination of COL blood concentration,enabling the enrichment and separation of trace or even ultra-trace target components,thereby improving analytical capabilities.

BACKGROUND:Carnosic acid,a bioactive compound found in rosemary,has been shown to reduce inflammation and reactive oxygen species(ROS).However,its mechanism of action in osteoclast differentiation remains unclear. OBJECTIVE:To investigate the effects of carnosic acid on osteoclast activation,ROS production,and mitochondrial function. METHODS:Primary bone marrow-derived macrophages from mice were extracted and cultured in vitro.Different concentrations of carnosic acid(0,10,15,20,25 and 30 μmol/L)were tested for their effects on bone marrow-derived macrophage proliferation and toxicity using the cell counting kit-8 cell viability assay to determine a safe concentration.Bone marrow-derived macrophages were cultured in graded concentrations and induced by receptor activator of nuclear factor-κB ligand for osteoclast differentiation for 5-7 days.The effects of carnosic acid on osteoclast differentiation and function were then observed through tartrate-resistant acid phosphatase staining,F-actin staining,H2DCFDA probe and mitochondrial ROS,and Mito-Tracker fluorescence detection.Western blot and RT-PCR assays were subsequently conducted to examine the effects of carnosic acid on the upstream and downstream proteins of the receptor activator of nuclear factor-κB ligand-induced MAPK signaling pathway. RESULTS AND CONCLUSION:Tartrate-resistant acid phosphatase staining and F-actin staining showed that carnosic acid dose-dependently inhibited in vitro osteoclast differentiation and actin ring formation in the cell cytoskeleton,with the highest inhibitory effect observed in the high concentration group(30 μmol/L).Carnosic acid exhibited the most significant inhibitory effect during the early stages(days 1-3)of osteoclast differentiation compared to other intervention periods.Fluorescence imaging using the H2DCFDA probe,mitochondrial ROS,and Mito-Tracker demonstrated that carnosic acid inhibited cellular and mitochondrial ROS production while reducing mitochondrial membrane potential,thereby influencing mitochondrial function.The results of western blot and RT-PCR revealed that carnosic acid could suppress the expression of NFATc1,CTSK,MMP9,and C-fos proteins associated with osteoclast differentiation,and downregulate the expression of NFATc1,Atp6vod2,ACP5,CTSK,and C-fos genes related to osteoclast differentiation.Furthermore,carnosic acid enhanced the expression of antioxidant enzyme proteins and reduced the generation of ROS during the process of osteoclast differentiation.Overall,carnosic acid exerts its inhibitory effects on osteoclast differentiation by inhibiting the phosphorylation modification of the P38/ERK/JNK protein and activating the MAPK signaling pathway in bone marrow-derived macrophages.

Surgical methods for varicocele remain controversial. This study intends to evaluate the efficacy and safety of different surgical approaches for treating varicocele through a network meta-analysis (NMA). PubMed, Embase, Cochrane, and Web of Science databases were thoroughly searched. In total, 13 randomized controlled trials (RCTs) and 24 cohort studies were included, covering 9 different surgical methods. Pairwise meta-analysis and NMA were performed by means of random-effects models, and interventions were ranked based on the surface under the cumulative ranking curve (SUCRA). According to the SUCRA, microsurgical subinguinal varicocelectomy (MSV; 91.6%), microsurgical retroperitoneal varicocelectomy (MRV; 78.2%), and microsurgical inguinal varicocelectomy (MIV; 76.7%) demonstrated the highest effectiveness in reducing postoperative recurrence rates. In this study, sclerotherapy embolization (SE; 87.2%), MSV (77.9%), and MIV (67.7%) showed the best results in lowering the risk of hydrocele occurrence. MIV (82.9%), MSV (75.9%), and coil embolization (CE; 58.7%) were notably effective in increasing sperm motility. Moreover, CE (76.7%), subinguinal approach varicocelectomy (SV; 69.2%), and SE (55.7%) were the most effective in increasing sperm count. SE (82.5%), transabdominal laparoscopic varicocelectomy (TLV; 76.5%), and MRV (52.7%) were superior in shortening the length of hospital stay. The incidence rates of adverse events for MRV (0), SE (3.3%), and MIV (4.1%) were notably low. Cluster analyses indicated that MSV was the most effective in the treatment of varicocele. Based on the existing evidence, MSV may represent the optimal choice for varicocele surgery. However, selecting clinical surgical strategies requires consideration of various factors, including patient needs, surgeon experience, and the learning curve.
Humans ; Male ; Embolization, Therapeutic/methods* ; Microsurgery/methods* ; Randomized Controlled Trials as Topic ; Sclerotherapy/methods* ; Treatment Outcome ; Urologic Surgical Procedures, Male/methods* ; Varicocele/surgery*