We investigated whether pyrroloquinoline quinone (PQQ) could improve cognitive performance in twenty-month-old naturally aged mice and explored the potential mechanisms involved. Results showed that PQQ supplementation improved spatial working memory and recognition memory without inducing anxiety-like behavior, and was associated with better preservation of hippocampal neuronal integrity. In HT-22 hippocampal neuronal cells, PQQ reduced reactive oxygen species (ROS) accumulation, restored mitochondrial membrane potential, and enhanced mitochondrial respiratory capacity. Hippocampal transcriptomic analysis and upstream regulator prediction identified sirtuin 1 (SIRT1) as a major regulator associated with the PQQ-induced transcriptional response, while uncoupling protein 2 (UCP2) emerged as a candidate downstream mitochondrial effector. Consistently, PQQ increased hippocampal SIRT1 protein expression and downregulated UCP2 at both mRNA and protein levels. Pharmacological inhibition of SIRT1 attenuated the PQQ-induced increase in ATP production and partially weakened the regulatory effect of PQQ on UCP2, supporting the involvement of SIRT1 in PQQ-associated mitochondrial bioenergetic regulation. Collectively, these findings indicate that PQQ improves cognitive-related behavioral performance in naturally aged mice and is associated with mitochondrial bioenergetic regulation, and suggest that modulation of a SIRT1-UCP2-associated pathway may contribute to its neuroprotective effects.

Background:Dementia associates with long-term progression, but the health status of individuals years before diagnosis remains poorly understood. We therefore conducted a large retrospective cohort study to examine the association between early-life morbidity and incident dementia. Methods:We designed a large-scale retrospective matched-cohort study using data from the National Health Insurance Research Database of Taiwan. Individuals newly diagnosed with dementia (2010-2013) were identified as the pre-dementia cohort and matched 1:3 with non-dementia controls. We assessed prevalence and outpatient visits for 30 common conditions (infectious to various of cancers.) between 2000 and 2013, with a backtracking time of 10 years for each individual. Conditional logistic regression was used to estimate odds ratios (ORs) for disease prevalence, while Negative Binomial regression was applied to calculate Incidence Rate Ratios (IRRs) for outpatient visit. Results:From 806,292 individuals, 19 of 30 diseases showed significant associations. Notably, the pre-dementia cohort exhibited a significantly lower prevalence of and fewer hospital visits for 13 conditions compared to controls. These included acute respiratory infections, peripheral inflammatory diseases, and certain digestive system malignancies. Conversely, a significantly higher prevalence was observed in only six conditions, predominantly those associated with direct or indirect brain injury, as well as cancers of the respiratory and urinary systems. Conclusion:Pre-dementia patients showed lower prevalence of most correlated common diseases. These unexpected findings are consistent with the hypothesis that altered peripheral immune function may be involved in the etiology and pathogenesis of dementia, which is beneficial for normal health but detrimental to the brain. Meanwhile, conditions that directly or indirectly contribute to brain damage are more likely to be positively associated with dementia. However, given the observational nature of our study and the absence of immunological biomarkers, these mechanistic interpretations remain speculative and require further validation.

Human Umbilical Cord-derived Mesenchymal Stromal Cells (hUC-MSCs) represent a promising candidate for regenerative medicine, though their therapeutic potential is constrained by replicative senescence. Pyrroloquinoline quinone (PQQ), a redox-active coenzyme, has been reported to protect against cellular aging. However, its precise role and mechanism of action in mitigating replicative senescence of hUC-MSCs remain to be elucidated. This study employed an integrated approach of phenotypic screening and transcriptomic profiling to systematically evaluate the anti-senescence effects of PQQ on replicatively senescent hUC-MSCs. Our results indicated that PQQ treatment enhanced proliferative capacity, reduced senescence-associated β-galactosidase (SA-β-gal) activity, and attenuated G1 phase cell cycle arrest. Moreover, PQQ improved mitochondrial membrane potential, reduced intracellular reactive oxygen species (ROS) accumulation, and attenuated telomere attrition. RNA sequencing analysis suggests that PQQ treatment appears to alleviate senescence-related transcriptional features, which is consistent with the observed phenotypic improvements. Gene Set Enrichment Analysis (GSEA) revealed a significant upregulation of pathways governing cell cycle progression and DNA replication following PQQ intervention. Key Driver Analysis (KDA) further identified regulators within these pathways, including PLK1, MCM5, and CDC6. Subsequent qPCR validation showed that the expression of these genes, which are critical for DNA replication initiation and mitotic progression, was downregulated in senescent cells and increased following PQQ treatment. In conclusion, the effect of PQQ on the replicative senescence of hUC-MSCs may be related to the upregulation of genes associated with the cell cycle and DNA replication.

Objective assessment of insomnia remains challenging, and most physiological investigations have focused on nocturnal sleep. Whether daytime electrophysiological signals contain discriminative information related to insomnia remains insufficiently explored. This exploratory cross-sectional study aimed to investigate the feasibility of using daytime multimodal electrophysiological signals for insomnia assessment and to identify the physiological features contributing most strongly to machine-learning classification. A total of 152 participants were included. Daytime multimodal electrophysiological recordings comprising electrocardiography (ECG), right-frontal electroencephalography (EEG; F4–M1), and bilateral pulse-wave signals were collected. Candidate digital biomarkers characterizing waveform morphology, temporal characteristics, variability, and nonlinear complexity were extracted. Support vector machine (SVM), logistic regression, random forest, and k-nearest neighbors models were developed and evaluated using AUC, accuracy, sensitivity, specificity, precision, and F1-score.SHapley Additive exPlanations (SHAP) analysis was applied to interpret the optimal model. The top SHAP-ranked features were further compared between the insomnia and normal-control groups, with false discovery rate (FDR) correction and effect-size estimation. SVM demonstrated the best overall classification performance among the evaluated models [AUC = 0.87; accuracy = 0.80]. Among the 10 most influential SHAP features, six were derived from pulse signals, two from ECG, and two from right-frontal EEG. Nine of these features showed lower values in the insomnia group and negative SHAP associations, whereas one pulse systolic-wave-height feature was higher and showed a positive SHAP association. All 10 features differed significantly between groups after FDR correction (FDR-adjusted P < 0.05), with effect sizes ranging from r = 0.22 to 0.57. Several highly ranked features were related to multiscale entropy, indicating that altered multiscale organization of electrophysiological signals contributed importantly to insomnia classification. Daytime EEG, ECG, and bilateral pulse-wave signals contain complementary physiological information associated with insomnia. The concordance between SHAP-derived feature contributions and group-level statistical differences suggests that insomnia may be characterized by altered multiscale and dynamic organization across central and peripheral physiological systems. These findings provide exploratory evidence supporting interpretable multimodal electrophysiological approaches for objective daytime assessment of insomnia, although external and longitudinal validation is required.

Objective The prevalence of metabolic obesity continues to rise, yet its pathogenesis remains controversial. This study focuses on the central neural regulatory differences underlying phenotypic divergence between diet-induced obese (OB) and obesity-resistant (OR) rats following long-term high-fat and high-sucrose diet (HFHSD) exposure. Methods Forty male Wistar rats were randomized to normal diet group (ND,n=8) or HFHSD group (n=32). After 8 weeks, HFHSD rats were split into OB (n=7) and OR (n=8) by weight. OB/OR continued HFHSD for 8 weeks with food intake recorded. At week 15, OGTT and functional magnetic resonance imaging (fMRI)-derived amplitude of low-frequency fluctuations (ALFF) were performed. At week 16, metabolic and somatic parameters were measured. Results Compared to the OR group, OB rats exhibited persistent hyperphagia, elevated triglycerides, TyG index, and leptin levels, significantly increased liver, heart, and kidney masses, and impaired glucose tolerance (all P<0.05). fMRI revealed significantly reduced ALFF in the hindbrain (bilateral brainstem and cerebellar molecular layers) and increased ALFF in the right_ dysgranular_retrosplenial cortex (RSC) of OB rats. Metabolic load parameters correlated negatively with hindbrain ALFF and positively with RSC ALFF, with a significant negative correlation between hindbrain and RSC activity. Conclusion Long-term HFHSD induces functional remodeling of the hindbrain–RSC circuit in obese rats, and this neuroplastic alteration is closely associated with systemic metabolic burden, suggesting its involvement in the pathophysiology of obesity.

BACKGROUND:Liver carcinoma, as a major global health concern due to its high incidence and mortality rates. Despite advancements in diagnostic and treatment methodologies, outcomes for hepatocellular carcinoma remain unsatisfactory. In response to these limitations, patients increasingly turn to alternative therapies such as traditional Chinese medicine, which has demonstrated potential in enhancing quality of life and prolonging survival in combination with conventional treatments. Triptolide (TP) and quercetin, as two broad-spectrum antitumor activities, act through multiple mechanisms, and whether the combination of them can provide a synergistic effect to improve the treatment effect. To optimize the dosage combination of TP and quercetin to maximize their therapeutic benefits in treating liver cancer, potentially advancing the field of drug combination therapy. This approach seeks to explore new treatment strategies and elucidate the underlying mechanisms that could lead to improved outcomes for hepatocellular carcinoma patients facing limited effective treatment options. AIM:To investigate the synergistic anti-hepatoma effect of TP and quercetin and elucidate the underlying molecular mechanism involving the Janus kinase (JAK)-signal transducer and activator of transcription (STAT) and mammalian target of rapamycin (mTOR) signaling pathways. METHODS:The study utilized 5-week-old female BALB/c-nu mice for establishing a liver cancer subcutaneous transplant tumor model. TP and quercetin were administered intraperitoneally over 21 days to evaluate the effectiveness of the combination, with monitoring of tumor growth. IncuCyte Zoom and CompuSyn software were employed to analyze drug effects for different dose combination on cell proliferation and synergy. Various assays such as CCK-8 cell proliferation analysis, plate cell clone formation, cell scratch experiments, Transwell migration and invasion assays, Annexin V-FITC flow cytometry, and western blotting using specific antibodies were employed to assess cell apoptosis, migration, invasion. Then transcriptome analysis was used RNA sequencing to find the potential synergistic mechanisms and proved by western blotting. RESULTS:In vivo, the combination therapy significantly slowed down tumor growth compared to the control group, quercetin alone group, and TP alone group. The tumor inhibition rates were 28.91% (quercetin), 28.8% (TP), and 59.3% (combination therapy), respectively. The determination of IncuCyte Zoom and CCK-8 confirmed that there is a concentration gradient and time gradient effect on tumor inhibition, with the synergistic effect of 25 nmol/L TP and 100 μmol/L quercetin being the best. Platelet cell clone formation and cell wound scratch assay showed that the combination group had better inhibitory effects. Transwell analysis showed a decrease in migration and invasion in the combination therapy group. Flow cytometry showed that over time, cell apoptosis increased after combination therapy. Transcriptome analysis emphasizes unique pathways influenced by the combination (JAK-STAT and mTOR signaling pathways) and has been validated at the protein level. CONCLUSION:Compared with a single drug, the specific metering combination of TP and quercetin has enhanced anti-tumor effects, mediated by inhibition of cell proliferation, inducing cell apoptosis and inhibiting migration/invasion. This synergistic effect is closely related to the simultaneous inhibition of signaling pathways JAK-STAT and mTOR concurrently.

ETHNOPHARMACOLOGICAL RELEVANCE:Chronic hepatitis B virus (HBV) infection is still a widespread global health issue. HuaganJiedu Decoction (HGJDD) is a common prescription for treating HBV in China, which has the effect of enhancing antiviral efficacy and improving clinical efficacy. However, its precise mechanism of action remains unclear, warranting further investigation to elucidate its therapeutic potential and integration into standard medical practices. AIM OF THE STUDY:This study aims to explore the therapeutic mechanism of HuaganJiedu Decoction (HGJDD) in HBV. MATERIALS AND METHODS:We investigated the therapeutic potential of HGJDD, and LC-MS analysis characterized the chemical profile of HGJDD. In vitro, we utilized HepG2.2.15 cell line to assess cytotoxicity and treatment efficacy of HGJDD compared to Entecavir controls. In vivo, assessments included monitoring HBV-related biomarkers and viral load. Network pharmacology and RNA-seq analyses identified molecular pathways and targets influenced by HGJDD treatment. Immunofluorescence and Western blotting provided further insights into the therapeutic mechanisms underlying HGJDD for HBV. RESULTS:HGJDD showed no toxicity on HepG2.2.15 cells at 10%, 20%, 40%, and 80% serum concentrations. In vitro, HGJDD reduced HBsAg, HBeAg, and HBV DNA levels by dose-dependently and time-dependently. HGJDD can decrease the levels of HBsAg, HBeAg, and HBV DNA in serum and liver levels, meanwhile the therapeutic effect of high-dose HGJDD approach to EVT's in HBV Tg mice. According to intersection of network pharmacology and transcriptome, FOXO signal pathway was highlighted as potential targets and Immunofluorescence find that FOXO4D protein expression lever was increased in three HGJDD group, especially in high-dose HGJDD group. Western blotting confirmed increased level of FOXO4, ERK, and p-ERK and decreased levels of HNF4α, which reflected that the therapeutic effect was closely to FOXO4/ERK/HNF4α signal pathway. CONCLUSIONS:Traditional Chinese medicine (TCM) offers diverse herbal treatments for HBV, with HGJDD showing efficacy in reducing HBsAg, HBeAg, and HBV DNA levels at cellular and animal levels. This study identified that FOXO4/ERK/HNF4α signal pathway played an important role in HGJDD's therapeutic effects. These findings support HGJDD's potential in HBV treatment, providing a scientific basis for clinical use.

Background Traditional Chinese medicine (TCM), with its extensive historical background, provides herbal remedies such as HuaganJiedu Decoction (HGJDD) that may serve as adjuncts to conventional strategies for hepatitis B virus (HBV). HGJDD has been demonstrated effectiveness in enhancing clinical outcomes for patients with HBV. Purpose Uncovering the mechanism by which HGJDD acts in HBV treatment by Multi-omics analysis of Network Pharmacology, Metabolomics, and Transcriptomics. Method In this research, the composition of HGJDD was identified by LC-MS. Evaluate the effects of HGJDD through metabolomics analysis. We conducted in vitro and in vivo studies using HepG2.2.15 cells and HBV-TG mice, and detected HBsAg, HBeAg, and HBV DNA in different levels. Multi-omics analyses were used to identify the key mechanism influenced by HGJDD treatment. Molecular docking evaluated the components' interaction with the FXR/RXRα-SHP1-HNF1α axis. Immunohistochemistry, co-immunoprecipitation, and Western blotting verified the mechanisms underlying the action of HGJDD. Result Using LC-MS, the study identified compounds within HGJDD. HGJDD exhibited the ability to lower HBsAg, HBeAg, and HBV DNA in a dose- and duration-dependent manner. Multi-omics analyses identified that FXR/RXRα-SHP1-HNF1α axis is the potential therapeutic target for its therapeutic effect. Meanwhile, multiple components in HGJDD have good interactions with the proteins on this axis. Immunofluorescence, co-immunoprecipitation, and Western blotting demonstrated the influence of FXR/RXRα-SHP1-HNF1α axis. Conclusion FXR/RXRα-SHP1-HNF1α axis is main mechanism for HGJDD against HBV.

BackgroundIschemic stroke (IS), the leading stroke subtype (∼87%), arises from vascular occlusions, triggering brain necrosis through ischemia-reperfusion injury. Ferroptosis, an iron-driven cell death via Fe2+-mediated lipid peroxidation, is implicated in IS pathology. This study demonstrates that enoyl-coA hydrolase 1 (ECH1) may serve as a peripheral biomarker and therapeutic target for IS based on ferroptosis signaling.MethodsWe integrated transcriptome data from the GEO database with preprocessing and normalization. Hub genes were screened using differential expression analysis and machine learning algorithms. Subsequently, genes were further filtered by mendelian randomization and also validated using transient middle cerebral artery occlusion (tMCAO) model.ResultsFerroptosis-related genes in the IS group showed higher expression compared with the healthy control group. Using differential expression analysis and machine learning algorithms, 12 potential hub genes were successfully screened. Mendelian randomization analysis further confirmed the causal association between ECH1 and stroke. In the tMCAO mouse model, ECH1 mRNA levels were down-regulated, consistent with the results of the clinical samples.ConclusionIn this study, taking ferroptosis as an entry point, ECH1 may serve as a potential peripheral blood biomarker and therapeutic target for IS through multidimensional validation, providing a basis for the development of relevant precision diagnostic strategies.

To analyze voice signals and identify asthma patients using voice signal analysis and machine learning techniques, we collected clear, low-noise fixed-pattern voice signals from 50 asthma patients and 50 healthy controls to build an analysis database. The research conducted multi-dimensional voice signal analysis based on MATLAB and selected voice feature indicators with significant differences between asthma patients and healthy controls. After dimensionality reduction analysis on differential phonetic features, the processed features were incorporated into subsequent SVM and RF modeling and classification research. The study established over 400 voice feature indicators related to diagnosis, of which 20 indicators showed significant differences between asthma patients and healthy controls (P < 0.01). In the classification study, both the SVM and RF models achieved identical accuracy rates of 87% on the test set, with AUC values of 0.95 for SVM and 0.93 for RF. This demonstrates their comparable performance in terms of overall classification accuracy, while the disparity in AUC values suggests that the SVM model may achieve a better trade-off between sensitivity and specificity. Thus, this paper not only provides a new method for non-invasive early detection of asthma but also lays the foundation for further application and optimization of this method in real-world settings.

The objective of this study was to develop and evaluate a non-invasive method for distinguishing patients with chronic obstructive pulmonary disease (COPD) from those with respiratory tract infections (RTI) using voice signal analysis and machine learning. Fixed-pattern voice signals were collected from 25 COPD patients and 25 RTI patients (serving as the control/comparison group). Multi-dimensional voice feature analysis was performed to identify features significantly differentiating the two groups. Statistically significant features were selected and subjected to dimensionality reduction. Logistic Regression (LR) and Random Forest (RF) models were then trained and evaluated for classification performance in distinguishing COPD from RTI. Over 400 voice features were initially analyzed. Eighteen features showed highly significant differences between COPD and RTI patients (P <; 0.05). In the task of distinguishing COPD patients from RTI patients, the LR model achieved a test set area under the curve AUC of 0.95, significantly outperforming the RF model (AUC = 0.76). This study demonstrates the feasibility of using voice analysis and machine learning, particularly the LR model, as a promising non-invasive tool for differentiating COPD from RTI. It provides a foundation for the practical application and further optimization of this voice-based approach in clinical settings requiring differential diagnosis of respiratory conditions.

This study investigated the mechanism of hydroxysafflor yellow A (HSYA) on senescent human umbilical cord mesenchymal stem cells (hUC-MSCs) through transcriptome sequencing. HSYA treatment identified 2377 differentially expressed genes (DEGs). Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses revealed that these DEGs were primarily enriched in cell adhesion regulation and the extracellular matrix (ECM)–receptor interaction pathway. Gene Set Enrichment Analysis (GSEA) and protein–protein interaction (PPI) network analysis corroborated the central role of ECM–receptor interaction signaling, and Key Driver Analysis (KDA) revealed 10 core regulatory genes (e.g., ID1, SMAD3, TGFB3). SA-β-gal staining showed that HSYA significantly reduced senescence-associated β-galactosidase activity. Flow cytometry showed no significant changes in cell cycle distribution. Western blot analysis indicated that HSYA treatment reduced the protein expression level of p16 without significantly altering p53 levels. Furthermore, HSYA significantly attenuated intracellular reactive oxygen species (ROS) accumulation. qPCR validation demonstrated that HSYA significantly upregulated ID1, GDF5, SMAD3, and TGFB3 while downregulating BMP4, TGFB2, and CCN2. These findings indicate that HSYA modulates genes associated with the ECM–receptor interaction pathway, potentially contributing to improved ECM homeostasis in senescent hUC-MSCs.

Objective: Morusin (Mor), a prenylated flavonoid isolated from the root bark of Morus alba L., exhibits potent anti-tumour effects; however, the molecular target of Mor is still not entirely clear. This study aimed to elucidate the mechanism of Mor against hepatocellular carcinoma (HCC) and identify potential molecular targets. Methods: Mitochondrial function was assessed by measuring the mitochondrial membrane potential, mitochondrial ultrastructure, oxygen consumption, and ATP levels. Mor-induced mitophagy was confirmed using western blotting, immunofluorescence, and fluorescent probes. Transcriptomics, flow cytometry, western blotting, qRT-PCR and biochemical assays were used to reveal the molecular mechanisms and targets of Mor against HCC. We further validated the interaction between Mor and the target proteins using molecular docking and biolayer interferometry (BLI). The inhibitory effect of Mor in vivo was evaluated using a Hep3B murine xenograft model. Results: Mor significantly reduced the ATP citrate lyase (ACLY) expression and inhibited ACLY activity in HCC cells. BLI analysis demonstrated a direct interaction between Mor and the ACLY active domain. Mor-induced ACLY inhibition led to ROS accumulation in HCC cells, which caused mitochondrial damage, triggered PINK1/Parkin-mediated mitophagy, and ultimately induced mitochondrial apoptosis. We further verified that ROS is crucial in the apoptotic action of Mor through experiments regarding an ROS scavenger. Mor also significantly inhibited tumour xenograft growth in vivo. In addition, analysis of human liver cancer clinical samples revealed elevated ACLY levels positively correlated with histologic grade. Conclusion: Collectively, our findings highlight Mor as a potent bioactive inhibitor of ACLY and a promising candidate for HCC therapy.

Due to the multitudinous structural types of glycosylated com-ponents,accurate identification of glycosylation modifications and secondary metabolite structures in herbs remains a challenge for natural drug analysis and new drug discovery[1].

BackgroundCardioembolic Stroke (CS) and Atrial Fibrillation (AF) are prevalent diseases that significantly impact the quality of life and impose considerable financial burdens on society. Despite increasing evidence of a significant association between the two diseases, their complex interactions remain inadequately understood. We conducted bioinformatics analysis and employed machine learning techniques to investigate potential shared biomarkers between CS and AF.MethodsWe retrieved the CS and AF datasets from the Gene Expression Omnibus (GEO) database and applied Weighted Gene Co-Expression Network Analysis (WGCNA) to develop co-expression networks aimed at identifying pivotal modules. Next, we performed Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis on the shared genes within the modules related to CS and AF. The STRING database was used to build a protein-protein interaction (PPI) network, facilitating the discovery of hub genes within the network. Finally, several common used machine learning approaches were applied to construct the clinical predictive model of CS and AF. ROC curve analysis to evaluate the diagnostic value of the identified biomarkers for AF and CS.ResultsFunctional enrichment analysis indicated that pathways intrinsic to the immune response may be significantly involved in CS and AF. PPI network analysis identified a potential association of 4 key genes with both CS and AF, specifically PIK3R1, ITGAM, FOS, and TLR4.ConclusionIn our study, we utilized WGCNA, PPI network analysis, and machine learning to identify four hub genes significantly associated with CS and AF. Functional annotation outcomes revealed that inherent pathways related to the immune response connected to the recognized genes might could pave the way for further research on the etiological mechanisms and therapeutic targets for CS and AF.

Background: Parkinson’s disease (PD) is a common movement disorder characterized by bradykinesia, rigidity, and resting tremors. Fatigue is a common disabling symptom but is easily ignored in PD. Half of the PD patients were influenced by fatigue. Acupuncture is one of the conservative treatments for fatigue related to other conditions, especially in China. Therefore, we perform a systematic review and meta-analysis to evaluate the evidence for acupuncture’s effectiveness, safety, and cost benefits for the treatment. Methods: This protocol is based on the previously published randomized controlled trial (RCT) studies. A literature search will be performed on the following database: PubMed, the Cochrane Library, Chinese BioMedical Literature Database, China National Knowledge Infrastructure (CNKI), China Science and Technology Journal Database (VIP), and Wanfang Data. According to the Cochrane Risk of Bias Tool and the level of evidence for results, we will assess the quality of the included studies by using the Grading of Recommendations Assessment, Development, and Evaluation (GRADE) method. The Review Manager (v5.3) software will be applied to statistical analysis.   Results: From the study, we will assess the effectiveness, safety, and cost-benefit of acupuncture on fatigue relief and functional improvement in patients with Parkinson’s Disease.### Competing Interest StatementThe authors have declared no competing interest.### Funding StatementYes### Author DeclarationsI confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained.YesThe details of the IRB/oversight body that provided approval or exemption for the research described are given below:Not applicable.I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals.YesI understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance).YesI have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable.YesNo datasets were generated or analysed during the current study. All relevant data from this study will be made available upon study completion.

Background The surge in multi-drug resistant organisms (MDROs) poses a dire threat to global health, necessitating novel antibacterial strategies. Traditional Chinese medicine (TCM) offers a multi-level approach potentially mitigating resistance development. Methods We evaluated the Sang Shen Pu Gong Ying (SSPGY) beverage’s antibacterial activity through in-vitro tests, real sputum samples, and a clinical case study. The beverage, rooted in TCM, was prepared using a blend of medicinal and food homologous herbs and tested for its effect on MDROs, including minimal inhibitory concentration (MIC) determination. Results In-vitro bacteriostatic assays indicate that SSPGY beverage can significantly reduce MDROs, including Pseudomonas aeruginosa, carbapenem-resistant Acinetobacter baumannii, and penicillin-resistant Streptococcus pneumoniae with distinct antibacterial effects from conventional antibiotics. However, it showed ineffective inhibitory activity in sputum samples. Amazingly, by a intermittent 165-d intervention of SSPGY beverage to a clinical case of severe Klebsiella pneumoniaeinfection, it manifests the effectiveness against MDROs and play an essential role in combating antibiotic resistance. Conclusion Our findings indicate that SSPGY, with its unique antibacterial profile, could offer a promising alternative or intervention to traditional antibiotics. Further research is warranted to elucidate its mechanisms and optimize its therapeutic potential in the context of MDROs.

目的 观察脾氨肽口服液对人肺癌细胞系A549增殖凋亡迁移是否有影响,并进一步探讨脾氨肽口服液的可能作用机制.方法 取对数生长期A549细胞,随机分为四组,1、2、3组分别在含10%胎牛血清的DMEM培养基中加入0.5、1.0及2.0 mg/mL脾氨肽口服液,4组不做任何处理,观察细胞的增殖、凋亡及迁移情况,检测各组细胞线粒体膜电位及活性氧簇(ROS).结果 与4组比较,培养24 h时1、2、3组细胞的细胞活力低,各时间点2、3组细胞的细胞汇合度低、相对划痕区域密度小;培养24 h时1、2、3组细胞的细胞凋亡率高,线粒体膜电位低,培养24 h时2、3组细胞的ROS相对表达量高(P均<0.05).随脾氨肽口服液浓度上升,A549细胞活力逐渐下降、细胞汇合度降低、细胞凋亡率升高、相对划痕区域密度降低、线粒体膜电位下降,ROS相对表达量升高,且呈剂量依赖性(P均<0.05).结论 脾氨肽口服液可抑制A549细胞的增殖迁移,促进细胞凋亡,其中2.0 mg/mL的脾氨肽口服液作用更明显.脾氨肽口服液可能通过降低细胞线粒体膜电位、促进细胞ROS表达抑制A549细胞的增殖迁移、促进细胞凋亡.

Objective:To observe the effect of peritumoral electroacupuncture on the induction of vascular normalization in a mouse breast cancer model.Methods:A subcutaneous graft model of breast cancer was established with 4T1 breast cancer cell line in female BALB/c mice aged 6-8 weeks. The mice were randomly assigned to three groups, a tumor-bearing group (TG), peritumoral electroacupuncture tumor-bearing group (EATG), and bevacizumab tumor-bearing group (BTG), with 18 mice in each group. The TG mice did not receive any intervention, the EATG mice received peritumoral electroacupuncture for 30 minutes, and the BTG mice were intraperitoneally injected with bevacizumab at 10mg/kg. Immunofluorescence was performed to assess the expression of CD31/alpha smooth muscle actin (α-SMA) and hypoxia-inducible factor 1-alpha (HIF-1α) in the tumor tissue at various points of time, including before intervention and 3 days and 5 days after intervention. Then, 3 days after intervention, observation of morphological changes of the microvessels in the tumor tissue was performed through Hematoxylin and Eosin (HE) staining and scanning electron microscope.Results:There was no significant difference in the expression of CD31, α-SMA, and HIF-1α in the tumor tissues of all groups before experimental intervention ( P>0.05). On day 3 of the experimental interventions, the CD31 and HIF-1α expression levels in the tumor tissues of the EATG and BTG mice were significantly reduced ( P<0.01), while α-SMA expression levels were significantly increased ( P<0.01) in both groups. On day 5 of the experimental interventions, the CD31 and HIF-1α expression levels in the tumor tissues of the EATG and BTG mice were still significantly lower than those in the TG mice ( P<0.01), while the α-SMA expression level was significantly higher than that in the TG group ( P<0.05). On day 3 of the experimental interventions, H&E staining showed visible microvessels in the tumor tissues of all 3 groups. In addition, scanning electron microscopic observation showed that the tumor microvessel walls of the TG mice were rough and defective, and that obvious deformities appeared in the lumen. In contrast, the walls of the microvessels of the EATG and BTG mice were generally intact and there was no obvious deformities in the lumen.Conclusion:Peritumoral electroacupuncture may induce microvasculature normalization by decreasing microvascular density and increasing pericyte coverage of the neovasculature, thereby improving hypoxic microenvironment of breast cancer in mice.