ObjectiveThis study systematically analyzed the arginine metabolic dysregulation in the rat model of heart failure （HF）， providing a modern scientific basis for elucidating the pathogenesis of HF and offering new insights for the prevention and treatment of HF with traditional Chinese medicine （TCM）. MethodsA thoracotomy was performed to ligate the left anterior descending coronary artery of rats， which induced acute myocardial ischemia and thus led to the development of post-myocardial infarction heart failure. The rats were divided into a sham surgery group and a model group， with eight rats in each group. Serum targeted metabolomics analysis was performed using ultra-performance liquid chromatography-triple quadrupole mass spectrometry （UPLC-TQ-S）， and the spatial distribution of metabolites in cardiac tissue was observed using airflow-assisted desorption electrospray ionizationmass spectrometry imaging （AFADESI-MSI）. Targets associated with HF and arginine metabolism were screened from databases including GeneCards and the Gene Expression Omnibus （GEO）， a protein-protein interaction （PPI） network was constructed， and enrichment analysis of the Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway and Gene Ontology （GO） was performed. Finally， molecular docking was conducted to verify the binding between core metabolic components and key targets， and potential TCMs were predicted based on the core pathways and targets. ResultsCompared with the sham surgery group， the levels of arginine and citrulline in the serum of model rats were significantly decreased （P<0.01）， while those of proline， ornithine， creatine， creatinine and glutamate were significantly increased （P<0.05， P<0.01）. Cardiac mass spectrometry imaging showed a decreased abundance of arginine in the local myocardial tissue. Bioinformatics analysis identified 24 core functional targets， such as the angiotensin-converting enzyme （ACE）， neuronal nitric oxide synthase （NOS1）， 5-hydroxytryptamine receptor 2A （HTR2A）， and epidermal growth factor receptor （EGFR）， and enrichment analysis indicated that these targets were significantly involved in the calcium signaling pathway， neuroactive ligand-receptor interactions， and phosphatidylinositol signaling pathway. Molecular docking confirmed strong binding activities between arginine， citrulline and HTR2A， as well as between creatine， creatinine and EGFR. Based on pathway-target prediction， potential TCM interventions， such as ginseng and magnolia， were identified. ConclusionThis study revealed characteristic arginine metabolic disorder in HF， and the core targets of HF were closely associated with the phosphatidylinositol signaling pathway. It provides a modern biological interpretation of the pathogenesis of HF in TCM from the perspectives of metabolites and signaling pathways， and offers valuable insights for targeted therapy of HF and the development of TCM.

ObjectiveThis study systematically analyzed the arginine metabolic dysregulation in the rat model of heart failure （HF）， providing a modern scientific basis for elucidating the pathogenesis of HF and offering new insights for the prevention and treatment of HF with traditional Chinese medicine （TCM）. MethodsA thoracotomy was performed to ligate the left anterior descending coronary artery of rats， which induced acute myocardial ischemia and thus led to the development of post-myocardial infarction heart failure. The rats were divided into a sham surgery group and a model group， with eight rats in each group. Serum targeted metabolomics analysis was performed using ultra-performance liquid chromatography-triple quadrupole mass spectrometry （UPLC-TQ-S）， and the spatial distribution of metabolites in cardiac tissue was observed using airflow-assisted desorption electrospray ionizationmass spectrometry imaging （AFADESI-MSI）. Targets associated with HF and arginine metabolism were screened from databases including GeneCards and the Gene Expression Omnibus （GEO）， a protein-protein interaction （PPI） network was constructed， and enrichment analysis of the Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway and Gene Ontology （GO） was performed. Finally， molecular docking was conducted to verify the binding between core metabolic components and key targets， and potential TCMs were predicted based on the core pathways and targets. ResultsCompared with the sham surgery group， the levels of arginine and citrulline in the serum of model rats were significantly decreased （P<0.01）， while those of proline， ornithine， creatine， creatinine and glutamate were significantly increased （P<0.05， P<0.01）. Cardiac mass spectrometry imaging showed a decreased abundance of arginine in the local myocardial tissue. Bioinformatics analysis identified 24 core functional targets， such as the angiotensin-converting enzyme （ACE）， neuronal nitric oxide synthase （NOS1）， 5-hydroxytryptamine receptor 2A （HTR2A）， and epidermal growth factor receptor （EGFR）， and enrichment analysis indicated that these targets were significantly involved in the calcium signaling pathway， neuroactive ligand-receptor interactions， and phosphatidylinositol signaling pathway. Molecular docking confirmed strong binding activities between arginine， citrulline and HTR2A， as well as between creatine， creatinine and EGFR. Based on pathway-target prediction， potential TCM interventions， such as ginseng and magnolia， were identified. ConclusionThis study revealed characteristic arginine metabolic disorder in HF， and the core targets of HF were closely associated with the phosphatidylinositol signaling pathway. It provides a modern biological interpretation of the pathogenesis of HF in TCM from the perspectives of metabolites and signaling pathways， and offers valuable insights for targeted therapy of HF and the development of TCM.

Objective To establish an epirubicin (EPI)-resistant murine triple-negative breast cancer (TNBC) (4T1/EPI) cell line and evaluate its biological characteristics and drug resistance. Methods The EPI-resistant cell line 4T1/EPI was developed through intermittent induction with gradually increasing EPI concentrations in vitro. Morphological changes were observed under an inverted microscope. Drug resistance index (MTT assay), cell doubling time (CCK-8 assay), and migration ability (wound healing assay) were evaluated. Western blot was used to detect the expression of drug resistance-related proteins. Transcriptome sequencing and KEGG pathway enrichment analysis were performed to identify the pathways and targets involved in EPI resistance, followed by experimental validation. Results The 4T1 cells eventually grew normally in a medium containing 100 ng/mL EPI, confirming the establishment of the 4T1/EPI resistant cell line. After stable resistance was acquired, morphological alterations were observed. Compared with their parental 4T1 cells, 4T1/EPI cells showed significantly prolonged doubling time (P<0.01) and enhanced migration ability (P<0.05). Expression levels of drug resistance-related proteins MDR1, MRP1 (P<0.01), and ABCG2 (P<0.05) were elevated in 4T1/EPI cells. In vivo models also demonstrated significant EPI resistance in 4T1/EPI tumors in terms of tumor weight and volume. Transcriptome sequencing highlighted the involvement of the PI3K/Akt signaling pathway and ABC transporter pathway. Validation experiments showed the upregulation of Erbb3, Egfr, PI3K, and Akt (P<0.05) and significant downregulation of Fgfr1 (P<0.01) in 4T1/EPI cells. Conclusion The EPI-resistant TNBC cell line 4T1/EPI was successfully established, exhibiting significant resistance in vitro and in vivo. The mechanism may involve the EPI-induced upregulation of Egfr and Erbb3, activating the PI3K/Akt pathway and subsequently enhancing ABC transporter expression.

OBJECTIVE To explore the intervention mechanism of Bushen anzhi decoction （BSAZD） in rats with insomnia induced by kidney failing to store spirit. METHODS A total of 36 SD rats were randomly divided into normal group （normal saline）， model group （normal saline）， eszopiclone group （positive control group， 0.27 mg/kg）， and low-， medium- and high-dose BSAZD groups （5.85， 11.70， 23.40 g/kg）， with 6 rats in each group. Except for the normal group， rats in the remaining groups were subjected to combined administration of D-galactose and DL-4-chlorophenylalanine to establish the rat model of insomnia due to kidney failing to store spirit. After successful modeling， corresponding liquid medicine or normal saline was intragastrically administered once daily for 14 consecutive days. After the last administration， behavioral indicators and hippocampal histopathological morphology of rats were detected. Untargeted metabolomics was adopted to screen serum differential metabolites and enrich relevant signaling pathways. RESULTS Compared with the model group， the high-dose BSAZD group exhibited significantly increased open arm entries ， open arm time and sleep time （ P ＜0.05）， accompanied by markedly decreased closed arm entries， closed arm time and sleep latency （ P ＜0.05）. Hippocampal neurons in the high-dose BSAZD group were arranged regularly， and the size， morphology and location of cell nuclei were nearly normal. Untargeted metabolomics analysis identified oxidized glutathione， glutamate and other differential metabolites in serum of rats in the high-dose BSAZD group； these differential metabolites were mainly enriched in GABAergic synapse， glutamatergic synapse， cAMP signaling pathway and 2-oxocarboxylic acid metabolism pathway. CONCLUSIONS BSAZD can improve sleep quality， relieve anxiety-like behaviors in rats with insomnia due to kidney failing to store spirit. Its therapeutic mechanism may be associated with regulating GABAergic and glutamatergic synaptic pathways to restore neurotransmitter balance and further alleviate oxidative stress injury， modulating the cAMP signaling pathway to facilitate neural repair， and intervening the 2-oxocarboxylic acid metabolism pathway to optimize energy supply.

OBJECTIVE To explore the intervention mechanism of Bushen anzhi decoction （BSAZD） in rats with insomnia induced by kidney failing to store spirit. METHODS A total of 36 SD rats were randomly divided into normal group （normal saline）， model group （normal saline）， eszopiclone group （positive control group， 0.27 mg/kg）， and low-， medium- and high-dose BSAZD groups （5.85， 11.70， 23.40 g/kg）， with 6 rats in each group. Except for the normal group， rats in the remaining groups were subjected to combined administration of D-galactose and DL-4-chlorophenylalanine to establish the rat model of insomnia due to kidney failing to store spirit. After successful modeling， corresponding liquid medicine or normal saline was intragastrically administered once daily for 14 consecutive days. After the last administration， behavioral indicators and hippocampal histopathological morphology of rats were detected. Untargeted metabolomics was adopted to screen serum differential metabolites and enrich relevant signaling pathways. RESULTS Compared with the model group， the high-dose BSAZD group exhibited significantly increased open arm entries ， open arm time and sleep time （ P ＜0.05）， accompanied by markedly decreased closed arm entries， closed arm time and sleep latency （ P ＜0.05）. Hippocampal neurons in the high-dose BSAZD group were arranged regularly， and the size， morphology and location of cell nuclei were nearly normal. Untargeted metabolomics analysis identified oxidized glutathione， glutamate and other differential metabolites in serum of rats in the high-dose BSAZD group； these differential metabolites were mainly enriched in GABAergic synapse， glutamatergic synapse， cAMP signaling pathway and 2-oxocarboxylic acid metabolism pathway. CONCLUSIONS BSAZD can improve sleep quality， relieve anxiety-like behaviors in rats with insomnia due to kidney failing to store spirit. Its therapeutic mechanism may be associated with regulating GABAergic and glutamatergic synaptic pathways to restore neurotransmitter balance and further alleviate oxidative stress injury， modulating the cAMP signaling pathway to facilitate neural repair， and intervening the 2-oxocarboxylic acid metabolism pathway to optimize energy supply.

Objective:To explore the molecular mechanism by which interferon regulatory factor 1 (IRF1) affects hepatic ischemia-reperfusion injury (HIRI) by regulating the polarization of Kupffer cells.Methods:Twelve male healthy C57BL/6 wild-type mice weighing 20-25 g and aged 6-8 weeks were divided into a sham operation group ( n=6) and a HIRI group ( n=6); Twelve male healthy C57BL/6 IRF1 gene knockout (IRF1 -/-) mice weighing 20-25 g and aged 6-8 weeks were divided into a sham operation IRF1 -/- group ( n=6) and a HIRI IRF1 -/- group ( n=6). The levels of serum alanine transaminase (ALT) and aspartate transaminase (AST) in mice were measured, and hematoxylin-eosin (HE) staining of liver tissues was performed for Suzuki scoring to evaluate liver injury. Fluorescence quantitative polymerase chain reaction (qRT-PCR) was used to evaluate the mRNA levels of IRF1 and tumor necrosis factor α (TNFα) in liver tissues. Flow cytometry and qRT-PCR were used to detect the proportion and functional changes of M1/M2-type Kupffer cells in liver tissues. IRF1 was overexpressed or knocked down in the mononuclear macrophage cell line ANA1, and a co-culture and hypoxia-reoxygenation system with the hepatocyte cell line AML12 was established. Flow cytometry was used to detect the apoptosis of AML12 cells. Results:At 12 hours after hepatic ischemia-reperfusion in wild-type mice, the liver tissue injury was the most severe. Compared with the sham operation group, the levels of serum ALT [(8 073±83) U/L vs. (81±19) U/L, q=13.59] and AST [(11 170±2 890) U/L vs. (412±210) U/L, q=13.77] in the HIRI group were significantly higher, and the differences were statistically significant (both P<0.001). The Suzuki score reached 5-6 points. At 12 hours after hepatic ischemia-reperfusion in IRF1 gene knockout mice, the liver tissue injury was not obvious. There were no significant differences in the levels of serum ALT [668 (514, 2 344) U/L vs. 254 (147, 285) U/L, q=2.52, P=0.348] and AST [1 936 (1 262, 2 003) U/L vs. 628 (423, 759) U/L, q=1.22, P=0.824] between the HIRI IRF1 -/- group and the sham operation IRF1 -/- group. Compared with the HIRI group, the ratio of M1/M2-type Kupffer cells in the liver of the HIRI IRF1 -/- group decreased [(0.958±0.090) vs. (2.788±0.258), q=2.06, P<0.0001], and the mRNA expression of TNFα decreased [(4.363±0.393) vs. (12.900±5.504), q=5.59, P=0.018], and the differences between the two groups were statistically significant. In the co-culture and hypoxia-reoxygenation experiment using ANA1 cells overexpressing IRF1 and AML12 cells, the proportion of AML12 hepatocytes in late apoptosis was higher than that in the control group [(14.05±4.25) vs. (3.15±1.16), t=2.85, P=0.047], and the difference was statistically significant. In contrast, when the expression of IRF1 was knocked down, the proportion of apoptotic AML12 cells decreased [(9.26±3.04) vs. (13.36±4.64), t=2.15, P=0.098], but the difference was not statistically significant. Conclusion:The IRF1 protein can regulate the polarization of Kupffer cells into M1-type macrophages, promote the inflammatory injury of the liver tissue after ischemia-reperfusion, and increase the apoptosis of hepatocytes.

This paper presents an automatic acquisition and analytic procedure of acupuncture manipulation based on optical navigation, aiming at solving the shortcomings of existing acquisition methods of acupuncture manipulation. An acquisition holder installed at the handle tail of filiform needle was designed to display the movement trajectory of the needle during acupuncture delivery by collecting the movement trajectory of holder. The 3-month old male Bama miniature pig was selected as the experimental subject, and 6 points, "Bojian" "Qiangfeng" "Housanli" "Xiaokua" "Huiyang" (BL35) and "Baihui" (GV20), were selected during acupuncture manipulation. The optical navigation system was used to collect the real-time data, and these data were per-processed and analyzed using mean filtering and Fourier transform. The acupuncture procedure was divided into 3 stages, inserting, lifting-thrusting, and twisting. The results showed that the accuracy was 96.3% at lifting-thrusting stage, and that was 100.0% at twisting stage. The decomposition effect of the entire procedure was satisfactory. This study provides a new approach to the quantitative analysis of acupuncture manipulation. In the future, it needs to further optimize the algorithm and expand the sample size so as to improve the accuracy of this analytic technique.
Acupuncture Therapy/methods* ; Male ; Animals ; Swine ; Acupuncture Points ; Humans ; Swine, Miniature ; Needles

Objective : To investigate the impact of fecal microbiota transplantation (FMT) on the composition of 15 intestinal-derived estrogens and their metabolites (EMs) and its correlation with non-alcoholic fatty liver disease (NAFLD) . Methods: Thirty male C57BL/6J mice were divided into a normal control group (Control) , a high- sugar high-fat diet combined with low-dose CCl4 -induced NAFLD model group ( Model) , and a group of model mice treated with fecal microbiota from normal female mice (FMT) . After 17 weeks of modeling , liver pathology in each group was observed using HE staining , biochemical methods were used to measure serum alanine aminotrans- ferase (ALT) and aspartate aminotransferase (AST) levels , as well as hepatic triglyceride (TG) and total choles- terol (TC) levels. and the content of 15 EMs in portal vein serum was detected using ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS) . The correlation between disease phenotype and intesti- nal EMs was analyzed using Pearson ′s method. Results: The NAFLD model was successfully established , and the FMT group showed improved liver structure and morphology , with significant decreases in liver function and hepatic lipids compared to the Model group. In NAFLD mice , the contents of E1 , E2 , and their 2- and 4-position metabo- lites in portal vein blood serum was reduced compared to normal mice , while the content of most 16- and 17-posi- tion metabolites ( except 16α-OHE1) increased compared to normal mice. Correlation analysis showed that ALT was strongly positively correlated with E3 and 17-epiE3 , and strongly negatively correlated with E1 , E2 , 4- MeOE1 , and 16α-OHE1 . The TC was strongly positively correlated with 17-epiE3 and strongly negatively correla- ted with E1 , 4-MeOE1 , and 16α-OHE1 . Conclusion FMT restores the disrupted composition of intestinal EMs and improves NAFLD.

Objective To investigate the effects of acupoint application at Tianshu(ST25)combined with kidney-warming and spleen-invigorating therapy on clinical efficacy,intestinal function,and the expressions of Toll-like receptor 4(TLR4)and nuclear factor kappa-B(NF-κB)in patients with ulcerative colitis(UC).Methods A total of 100 UC patients with depletion and deficiency of spleen-kidney complicated with internal accumulation of pathogenic dampness syndrome,treated in the Department of Proctology,Shanghai Municipal Hospital of Traditional Chinese Medicine Affilated to Shanghai University of Traditional Chinese Medicine from June 2022 to June 2024,were selected and randomly divided into four groups(Groups A,B,C,D)using a random number table method,with 25 patients in each group.Group A received standard mesalazine treatment.Group B received Kidney-Warming and Spleen-Invigorating Formula(composed of Psoraleae Fructus,Myristicae Semen,Euodiae Fructus,Schisandrae Chinensis Fructus,Codonopsis Radix,Poria,Atractylodis Macrocephalae Rhizoma,etc.)combined with standard mesalazine treatment.Group C received acupoint application at Tianshu(the medicinal paste for application composed of Caryophylli Flos,Cinnamomi Cortex,Atractylodis Macrocephalae Rhizoma,Euodiae Fructus,Coptidis Rhizoma,etc.)combined with standard mesalazine treatment.Group D received Kidney-Warming and Spleen-Invigorating Formula+acupoint application at Tianshu+standard mesalazine treatment.All groups were treated continuously for 12 weeks.The time for disappearance of clinical symptoms was compared among the four groups.Serum immune cytokines[β-defensin-1,total immunoglobulin A(IgA),interleukin-6(IL-6),tumor necrosis factor-alpha(TNF-α)],peripheral blood levels of TLR4 and NF-κB,protein expression levels of TLR4 mRNA and NF-κB p65 mRNA in intestinal mucosal tissue,and changes in intestinal microbiota were observed before and after treatment.The clinical efficacy of the four groups was evaluated.Results(1)Compared to the other three groups,Group D had the shortest time for disappearance of clinical symptoms such as abdominal pain,diarrhea,mucopurulent bloody stools,and tenesmus(P＜0.05).The time for disappearance of various clinical symptoms in Groups B and C was similar(P＞0.05)but was shorter than that in Group A(P＜0.05).(2)After treatment,the serum levels of β-defensin-1,IgA,IL-6,and TNF-α in all four groups were significantly lower than those before treatment(P＜0.05).Post-treatment intergroup comparisons showed statistically significant differences(P＜0.01),with Group D exhibiting the greatest reduction,significantly lower than the other three groups(P＜0.05).The levels in Groups B and C were similar(P＞0.05)but were lower than those in Group A(P＜0.05).(3)After treatment,the peripheral blood levels of TLR4 and NF-κB,and the protein expression levels of TLR4 mRNA and NF-κB p65 mRNA in intestinal mucosal tissue showed a decreasing trend in all four groups(P＜0.05).Post-treatment intergroup comparison results were statistically significant(P＜0.01),with Group D showing the greatest reduction,significantly lower than the other three groups(P＜0.05).All above levels in Groups B and C were similar(P＞0.05)but were lower than those in Group A(P＜0.05).(4)After treatment,the relative contents of Bifidobacterium and Lactobacillus increased(P＜0.05),while those of Saccharomyces and Clostridium decreased(P＜0.05)in Groups B,C,and D.Post-treatment intergroup comparison results were statistically significant(P＜0.01).Group D showed a significant increase in Bifidobacterium and Lactobacillus and a significant decrease in Saccharomyces and Clostridium,with the magnitude of change significantly greater than that in the other three groups(P＜0.05).The quantities of these bacteria in Groups B and C were similar(P＞0.05),but the improvement was significantly greater than that in Group A(P＜0.05).(5)After 12 weeks of treatment,Group D had the highest total effective rate at 100.00％(25/25).The total effective rates of Groups B and C were similar,both at 84.00％(21/25),but were higher than that of Group A(56.00％,14/25).The intergroup difference was statistically significant(x2=16.310,P＜0.01).Conclusion The comprehensive treatment regimen of acupoint application at Tianshu combined with Kidney-Warming and Spleen-Invigorating Formula,on the basis of standard mesalazine treatment,has a positive impact on UC patients with depletion and deficiency of spleen-kidney complicated with internal accumulation of pathogenic dampness syndrome.This treatment method can effectively improve clinical symptoms,regulate β-defensin-1 and IgA levels,enhance immune defense function,downregulate the expression levels of TLR4 and NF-κB,and alleviate inflammatory response.Compared to monotherapies,it is more effective on improving treatment outcomes.

Objective To investigate the underlying mechanisms of vitamin D treatment improves early-stage renal damage in hypertensive mice.Methods From December 2022 to December 2023,peripheral blood samples were collected from 20 hypertensive patients with early-stage renal injury and healthy controls.ELISA was used to detect interleukin-6(IL?6),C?reactive protein(CRP)and reactive oxygen species(ROS).RT?qPCR and Western blot were utilized to measure the expression of Klotho and disulfidptosis?related proteins[glucose transporter 1(GLUT1),glucose transporter 3(GLUT3)and solute carrier family 7 member 11(SLC7A11)].A mouse model of hypertension with early?stage renal damage was established and treated with vitamin D.Kidney tissue samples were collected for HE staining and ELISA to assess inflammatory factors,and RT?qPCR and Western blot were used to examine the expression of Klotho and disulfidptosis?related proteins.Results Compared to the control group,the level of IL?6,CRP and ROS was significantly increased in the hypertensive patients with early?stage renal damage(P<0.05).The expression of GLUT1 and GLUT3 was significantly increased,while the expression of Klotho and SLC7A11 was significantly decreased(P<0.05).In the mouse model of hypertension with early?stage renal dam?age,HE staining showed that the level of inflammatory factors in the model group was significantly higher than that in control group.Vitamin D suppressed the level of inflammatory factors,increased the expression of Klotho and SLC7A11 and inhibited the expression of GLUT1 and GLUT3 in the model group(P<0.05).Conclusions Vitamin D treatment effectively improves inflammatory response in hypertensive mice with early renal injury.