[Efficacy and mechanism of Xuefu Zhuyu Decoction on model rats of coronary heart disease with heart blood stasis syndrome based on metabolomics].

This study investigated the effects of Xuefu Zhuyu Decoction on myocardial metabolites in a rat model of coronary heart disease with heart blood stasis syndrome and explored the therapeutic mechanism of blood circulation-promoting and blood stasis-removing therapy. SD rats were randomly divided into a sham operation group, a model group, a Xuefu Zhuyu Decoction group(14.04 g·kg~(-1)), and a trimetazidine group(5.4 mg·kg~(-1)). The sham operation group underwent thread insertion without ligation, while the other groups underwent coronary artery left anterior descending branch ligation to induce a model of coronary heart disease with heart blood stasis syndrome. Three days after modeling, drug intervention was performed, and samples were taken after 14 days of intervention. General conditions were observed, and electrocardiogram and cardiac ultrasound indices were measured. Hematoxylin-eosin(HE) staining and Masson staining were used to observe tissue pathological morphology. The enzyme linked immunosorbent assay(ELISA) was used to measure the levels of triglyceride(TG) and total cholesterol(TC) in the serum. Ultra high performance liquid chromatography-quantitative exactive-mass spectrometry(UHPLC-QE-MS) technology was used to screen differential metabolites in myocardial tissue and conduct metabolic pathway enrichment analysis. The results showed that Xuefu Zhuyu Decoction significantly improved the general condition of the model rats, reduced heart rate and ST segment elevation in the electrocardiogram, increased left ventricular ejection fraction(LVEF) and left ventricular fractional shortening(LVFS), and decreased left ventricular internal diameter in diastole(LVIDd) and left ventricular internal diameter in systole(LVIDs). HE staining and Masson staining showed that Xuefu Zhuyu Decoction effectively alleviated myocardial tissue structural disorders, inflammatory cell infiltration, and collagen fiber deposition in the model rats. ELISA results showed that Xuefu Zhuyu Decoction effectively regulated serum TG and TC levels in the model rats. There were significant differences in the metabolic phenotypes of myocardial samples in each group. Fourteen differential metabolites were identified in the Xuefu Zhuyu Decoction group, involving five metabolic pathways, including arginine and proline metabolism, glycerophospholipid metabolism, aminoacyl-tRNA biosynthesis, ether lipid metabolism, and alanine, aspartate, and glutamate metabolism. Xuefu Zhuyu Decoction improved cardiac function and myocardial structural damage in the rat model of coronary heart disease with heart blood stasis syndrome, and its biological mechanism involved the regulation of lipid metabolism, choline metabolism, amino acid metabolism, energy metabolism, and protein synthesis pathways.

Xuefu Zhuyu decoction improves asthma-induced asthenozoospermia based on network pharmacology and in vivo experiment.

This study aimed to verify that Xuefu Zhuyu decoction (XFZYD) can improve asthenozoospermia caused by asthma, and explore its potential mechanism. Ovalbumin solution is used to induce asthma rat models. Sperm concentration and motility are used to evaluate semen quality. Immunohistochemistry (IHC), Western blotting and real-time reverse transcription-quantitative polymerase chain reaction (RT-qPCR) are used to detect proteins and mRNA related to rat testis tissue. Haematoxylin and eosin (H&E) staining was used to observe changes in testicular tissues. Through network pharmacology, eriodictyol, 18-ß-glycyrrhetinic acid, naringenin, chrysin and Hispidulin were prominent active ingredients of XFZYD. We found that XFZYD regulates the expression levels of albumin (ALB), vascular endothelial growth factor A (VEGFA), interleukin 6 (IL-6) protein and mRNA, thereby improving the histopathological morphology of the testis, increasing the concentration and motility of spermatozoa. We suggest that future research can increase the detection of hormones and oxidative stress and other related indicators, so as to conduct more in-depth exploration.

[Identification of active components in Xuefu Zhuyu Decoction based on targets of blood-activating function].

As a classic prescription for promoting blood circulation to remove blood stasis, Xuefu Zhuyu Decoction(XFZYD) is widely used in clinical practice and has notable curative effect. Based on the key targets of activating blood circulation, this study identified the active components of XFZYD to reveal the material basis. The components of XFZYD were collected from Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform(TCMSP). The molecular docking models were built for the blood-activating targets obtained from the previous study with the components of XFZYD. The top five active components with measurability for each target were selected as the potential blood-activating components in the prescription. The efficacy of the prescription can embody key pharmacological and high-content components. In this study, anti-platelet aggregation activity was used to characterize the effect of activating blood, and the in vivo experiments were conducted to verify the accuracy of the active components. A total of 210 chemical components of XFZYD were screened out from TCMSP and docked with the key targets with the function of activating blood. Ligustrazine, acteoside, naringin, etc. were selected as the potential active components for activating blood in XFZYD. The anti-platelet aggregation activity of the combination of Chuanxiong Rhizoma, Rehmanniae Radix, Aurantii Fructus, Glycyrrhizae Radix et Rhizoma, and Carthami Flos was 9.82%±5.11%. Compared with that in the control group, the platelet aggregation induced by adenosine diphosphate(ADP) was significantly inhibited in the test group(P<0.01), which verified the accuracy of the active components. This study can guide the research on the material basis of XFZYD and provide insights into the development and utilization of the classical prescription.

[Systematic evaluation and sequential Meta-analysis of modified Xuefu Zhuyu Decoction combined with Western medicine in treatment of leiomyoma].

CNKI, PubMed and other databases were retrieved to extract eligible randomized controlled trial(RCT) about modified Xuefu Zhuyu Decoction(MXZD) combined with Western medicine(trial group) versus Western medicine alone(control group) in the treatment of leiomyoma. Therefore, a total of 25 RCTs were included, involving 2 328 patients. Bias risk evaluation tool in Cochrane Handbook 5.1.0 was used for evaluating the quality of these RCTs. Meta-analysis was performed for the reported indicators, including total efficiency, serum hormone level [progesterone(P), luteinizing hormone(LH), estradiol(E_2), follicle stimulating hormone(FSH)], uterine size, fibroids size and adverse reactions by using Stata 14.0 software. Meta-analysis showed that the total efficiency(RR=1.21,95%CI[1.17,1.25],P<0.05) of trial group was better than that of control group. Serum hormone level(WMD_P=-3.86,95%CI[-4.31,-3.41],P<0.05; WMD_(LH)=-3.64,95%CI[-4.47,-2.82],P<0.05; WMD_(E_2)=-39.99,95%CI[-53.45,-26.52],P<0.05; WMD_(FSH)=-3.79,95%CI[-4.86,-2.72],P<0.05), uterine size(WMD=-50.02,95%CI[-55.98,-44.06],P<0.05), fibroids size(WMD=-15.79,95%CI[-18.11,-13.46],P<0.05) and adverse reactions(RR=0.65,95%CI[0.48,0.88],P<0.05) of trial group were all lower than those of control group, with statistical significances. Trial sequential analysis(TSA) was performed by using TSA 0.9 software, and showed a reliable therapeutic effect of the experimental group. In short, our study indicated that modified Xuefu Zhuyu Decoction combined with Western medicine had a better therapeutic effect on leiomyoma than Western medicine alone, but more high-quality studies are needed to verify this conclusion in the future.

[Mechanism of Xuefu Zhuyu Decoction in treatment of myocardial infarction based on network pharmacology and molecular docking].

To explore the action mechanism of Xuefu Zhuyu Decoction in treating myocardial infarction based on network pharmaco-logy and molecular docking. Active components and corresponding targets of Xuefu Zhuyu Decoction were obtained through Traditional Chinese Medicine Systems Pharmacology Database(TCMSP), and related targets of myocardial infarction were obtained through GeneCards, DisGeNET, and OMIM databases. Then the intersection targets were obtained by integrating the drug targets and disease targets. The "active component-target" network was constructed by Cytoscape software, and protein-protein interaction(PPI) network was drawn using STRING platform. Protein cluster analysis was carried out using MCODE. GO enrichment analysis and KEGG pathway analysis were carried out using DAVID database and ClueGO, and molecular docking was carried out using Autodock Vina and Pymol. Finally, 226 active components of Xuefu Zhuyu Decoction were obtained, 257 corresponding targets, 1 340 targets of myocardial infarction, and 109 drug and disease intersection targets were obtained. From GO enrichment analysis, 208 biological process terms, 38 molecular function terms, and 33 cellular component terms were obtained. From KEGG pathway analysis, NF-κB signaling pathway, IL-17 signaling pathway, HIF-1 signaling pathway, and other related pathways were obtained. The molecular docking results showed that the main active components(quercetin, kaempferol, ß-sitosterol, luteolin, stigmasterol and baicalein) of Xuefu Zhuyu Decoction in the treatment of myocardial infarction had good binding properties with the core proteins IL6, ALB, VEGFA, TNF, MAPK3 and CASP3. The results suggested that Xuefu Zhuyu Decoction may play a role in the treatment of myocardial infarction by reducing the inflammatory response, reducing oxidative stress, inhibiting cell apoptosis, and promoting angiogenesis.

1H-NMR-based metabolomics study on coronary heart disease with blood-stasis syndrome and phlegm syndrome. / 基于1H-NMRçš„å† å¿ƒç— è¡€ç˜€è¯ã€ç—°æµŠè¯çš„ä»£è°¢ç»„å­¦ç ”ç©¶.

OBJECTIVES: Coronary heart disease (CHD) is a serious threat to human health because of its high morbidity. It is very urgent to study the pathogenesis of CHD and the effective drug target. The purpose of this paper is using the 1H-nuclear magnetic resonance spectroscopy (1H-NMR) metabolomics technology to establish the metabolic fingerprint and find the potential biomarker metabolites of CHD with blood-stasis syndrome and phlegm syndrome, and to reveal the metabolic mechanism of Xuefu Zhuyu Decoction for the treatment of CHD with blood stasis syndrome. METHODS: The plasma samples of 69 patients with CHD blood-stasis syndrome, 60 patients with CHD phlegm syndrome, and 40 healthy volunteers were collected in this study. Based on the 1H-NMR metabolomics technology, the metabolic fingerprint of CHD with blood-stasis syndrome and phlegm syndrome was established. Multivariate statistical analysis methods including principal component analysis (PCA) and orthogonal partial least squares-discriminant analysis (OPLS-DA) were used to find the potential biomarker metabolites of CHD with blood-stasis syndrome and phlegm syndrome. Xuefu Zhuyu Decoction was used to randomly selected blood-stasis syndrome patient. The plasma samples of pre-treatment and post-treatment were collected. 1H-NMR and multivariate statistical analysis were used to analyze the changes of metabolites in patients with CHD blood-stasis syndrome before and after Xuefu Zhuyu Decoction treatment. RESULTS: A total of 15 potential biomarkers were identified in the plasma of patients with CHD blood-stasis syndrome, including 3-hydroxybutyrate (3-HB), lactate, alanine, glutamate, glutamine, pyruvate,phosphatidylcholine (PC), glycerylphosphorylcholine (GPC), glycine, glucose, phenylalanine, citrate,tyrosine, formate,very low density lipoprotein (VLDL). The levels of glucose, 3-HB, and VLDL increased, while the levels of other 12 metabolites decreased. A total of 16 potential biomarkers were identified in the plasma of patients with CHD phlegm syndrome, including valine, lactate, alanine, N-acetyl-ß-glucosaminidase (NAG), glutamate, glutamine, pyruvate, creatine, choline, glycine, glucose, phenylalanine, citrate, histidine, tyrosine, and formate. The levels of glucose and choline increased, while the levels of other 12 metabolites decreased. After treatment with Xuefu Zhuyu Decoction, the levels of choline, phospholipids/glycerolipids, creatine, lipids, and citrate increased, while the level of lactate decreased in patients with CHD blood-stasis syndrome. CONCLUSIONS: 1H-NMR combined with multivariate statistical method could effectively establish the diagnostic model for CHD blood-stasis syndrome and CHD phlegm syndrome, and find the metabolites related to the syndrome type. The metabolic mechanism of Xuefu Zhuyu Decoction on CHD blood-stasis syndrome may be associated with regulation of lipid metabolism and energy metabolism.

A comparative pharmacogenomic analysis of three classic TCM prescriptions for coronary heart disease based on molecular network modeling.

Traditional Chinese medicine (TCM) has evolved over several thousands of years, which has been shown to be efficacious in the treatment of ischemic heart disease. Three classical TCM prescriptions, namely Xuefu Zhuyu Decoction, Zhishi Xiebai Guizhi Decoction, and Gualou Xiebai Banxia Decoction, have been extensively used in the treatment of coronary heart disease (CHD). Based on molecular network modeling, we performed a comparative pharmacogenomic analysis to systematically determine the drug-targeting spectrum of the three prescriptions at molecular level. Wide-area target molecules of CHD were covered, which was a common feature of the three decoctions, demonstrating their therapeutic functions. Meanwhile, collective signaling involved metabolic/pro-metabolic pathways, driving and transferring pathways, neuropsychiatric pathways, and exocrine or endocrine pathways. These organized pharmacological disturbance was mainly focused on almost all stages of CHD intervention, such as anti-atherosclerosis, lipid metabolism, inflammation, vascular wall function, foam cells formation, platelets aggregation, thrombosis, arrhythmia, and ischemia-reperfusion injury. In addition, heterogeneity analysis of the global pharmacological molecular spectrum revealed that signaling crosstalk, cascade convergence, and key targets were tendentious among the three decoctions. After all, it is unadvisable to rank the findings on targeting advantages of the three decoctions. Comparative pharmacological evidence may provide an appropriate decoction scheme for individualized intervention of CHD.

Development of a sensitive and rapid UHPLC-MS/MS method for simultaneous quantification of nine compounds in rat plasma and application in a comparative pharmacokinetic study after oral administration of Xuefu Zhuyu Decoction and nimodipine.

Xuefu Zhuyu Decoction (XFZYD) is a traditional Chinese medicine prescription used for the clinical treatment of traumatic brain injury (TBI). The purpose of this work was to develop a sensitive and rapid UHPLC-MS/MS method to simultaneously study the pharmacokinetics of nimodipine and eight components of XFZYD, namely, amygdalin, hydroxysafflor yellow A, rutin, liquiritin, narirutin, naringin, neohesperidin and saikosaponin A, in rats with and without TBI. Multiple reaction monitoring was highly selective in the detection of nine analytes and the internal standard without obvious interference. The calibration curves displayed good linearity (r > 0.99) over a wide concentration range. The mean absolute recoveries of the nine analytes were 85-106%, and all matrix effects were in the range 80-120%. The intra- and inter-day precision and accuracy were acceptable (RSD, <15%; RE%, ±20%). The validated method was successfully applied to compare the pharmacokinetics in four experimental groups, including control rats orally administered XFZYD and TBI model rats orally administered XFZYD, XFZYD and nimodipine, or nimodipine alone. The results showed that herb-drug interactions occurred between XFZYD and nimodipine in the treatment of TBI, nimodipine affected the pharmacokinetics of XFZYD, and XFZYD affected the absorption, distribution and excretion of nimodipine in vivo.

Transcriptomic analysis reveals the critical role of chemokine signaling in the anti-atherosclerosis effect of Xuefu Zhuyu decoction.

ETHNOPHARMACOLOGICAL RELEVANCE: The process of atherosclerosis (AS) is complicated. Transcriptomics technology can assist in discovering the underlying mechanisms and exploring the key targets of Traditional Chinese Medicine (TCM) against atherosclerosis. AIM: This study aimed to investigate targets and signaling pathways significantly related to AS and the potential intervention targets of Xuefu Zhuyu decoction by transcriptomics. MATERIALS AND METHODS: AS models were established by subjecting ApoE-/-mice to an 8-week high-fat diet. Structural changes and plaque formation in the aortic root were observed using hematoxylin-eosin staining (HE staining), while Oil Red O staining was employed to visualize lipid deposition within the aortic root plaque. Movat staining and immunohistochemical staining were conducted to examine the components present in the aortic root plaque. Macrophage content within the plaque was observed through immunofluorescence. Additionally, mRNA sequencing was performed on aortic tissues to identify differentially expressed genes. Enrichment analysis was performed using GO and KEGG analysis. Visualization of the protein-protein interaction (PPI) network was achieved using Cytoscape 3.7.1 and STRING. Western blotting (WB) was employed to assess the protein expression of major differentially expressed genes in the aortic tissue. The drug freeze-dried powder of Xuefu Zhuyu decoction was prepared and the RAW264.7 cells were induced by lipopolysaccharide (LPS) to build an in vitro model. Real-time quantitative PCR was employed to measure the mRNA expression of major differential genes. RESULTS: After ApoE-/- mice were fed with an 8-week high-fat diet, observable changes included the thinning of the aortic root wall, the accumulation of foam cells within the plaque, and the formation of cholesterol crystals in the model group. Treatment with Xuefu Zhuyu (XFZY) decoction for 12 weeks significantly reduced the lipid deposition and the number of macrophages (P < 0.05) and significantly increased the collagen content within the plaque (P < 0.01). Enrichment analysis revealed a high enrichment of the Cytokine-cytokine receptor interaction pathway and Chemokine signaling pathway. Noteworthy genes involved in this response included Ccl12, Ccl22, Cx3cr1, Ccr7, Ccr2, Tnfrsf25, and Gdf5. Xuefu Zhuyu decoction significantly downregulated the expression of CX3CL1 and CX3CR1 (P < 0.05) and upregulated the expression of GDF5 (P < 0.01). Compared with control group, in cell models, the mRNA expressions of Ccl12, Ccl22, and Ccr2 were significantly upregulated (P < 0.05 or P < 0.01). Xuefu Zhuyu decoction significantly downregulated the expression of Ccl12, Ccl22, Cx3cr1, Ccr7 and Ccr2 (P < 0.05 or P < 0.01). CONCLUSION: Xuefu Zhuyu decoction demonstrates effective regulation of plaque components, retarding plaque progression and preserving plaque stability by modulating lipid metabolism and inflammatory responses. Subsequent transcriptome analysis identified the Cytokine-cytokine receptor interaction and Chemokine signaling pathway as potential key pathways for the therapeutic effects of Xuefu Zhuyu decoction. This insight not only provides crucial avenues for further exploration into the mechanisms underlying Xuefu Zhuyu decoction but also offers valuable perspectives and hypotheses for enhancing disease prevention and treatment strategies.

Spatial amine metabolomics and histopathology reveal localized brain alterations in subacute traumatic brain injury and the underlying mechanism of herbal treatment.

INTRODUCTION: Spatial changes of amine metabolites and histopathology of the whole brain help to reveal the mechanism of traumatic brain injury (TBI) and treatment. METHODS: A newly developed liquid microjunction surface sampling-tandem mass tag-ultra performance liquid chromatography-mass spectrometry technique is applied to profile brain amine metabolites in five brain regions after impact-induced TBI at the subacute stage. H&E, Nissl, and immunofluorescence staining are performed to spatially correlate microscopical changes to metabolic alterations. Then, bioinformatics, molecular docking, ELISA, western blot, and immunofluorescence are integrated to uncover the mechanism of Xuefu Zhuyu decoction (XFZYD) against TBI. RESULTS: Besides the hippocampus and cortex, the thalamus, caudate-putamen, and fiber tracts also show differentiated metabolic changes between the Sham and TBI groups. Fourteen amine metabolites (including isomers such as L-leucine and L-isoleucine) are significantly altered in specific regions. The metabolic changes are well matched with the degree of neuronal damage, glia activation, and neurorestoration. XFZYD reverses the dysregulation of several amine metabolites, such as hippocampal Lys-Phe/Phe-Lys and dopamine. Also, XFZYD enhances post-TBI angiogenesis in the hippocampus and the thalamus. CONCLUSION: This study reveals the local amine-metabolite and histological changes in the subacute stage of TBI. XFZYD may promote TBI recovery by normalizing amine metabolites and spatially promoting dopamine production and angiogenesis.

A Chinese classical prescription Xuefu Zhuyu decoction in the treatment of coronary heart disease: An overview.

Background: Coronary heart disease (CHD) is the leading cause of morbidity and mortality worldwide and is a hot topic in cardiovascular disease research. Western medicine treats CHD with stent implantation, anti-angina pectoris, anti-platelet aggregation and other operations or drugs. According to the whole concept and the characteristics of syndrome differentiation, traditional Chinese medicine (TCM) treats CHD according to different syndromes and points out that qi deficiency and blood stasis are the basic pathogenesis of CHD. Xuefu Zhuyu Decoction (XFZYD), as a classic prescription of TCM, has certain value in the treatment of CHD, with the effects of promoting qi, activating blood circulation, dredging collaterals and relieving pain. In addition, it also exhibits advantages in high efficiency, low toxicity, high cost performance, few side effects, and high patient acceptance. Objective: The therapeutic effect and mechanism of XFZYD in the treatment of CHD were searched by literature search, and the components and targets of XFZYD in the treatment of CHD were analyzed by computer simulation technology for molecular docking, providing theoretical basis for clinical treatment of CHD. Method: This study comprehensively searched CNKI, Wanfang, VIP, CBM, Pubmed, Embase, Web of science and other databases, included clinical studies with efficacy evaluation indicators in hospitals according to randomization, and excluded literatures with low quality and no efficacy evaluation indicators. Clinical cases and studies, molecular mechanisms and pharmacological effects of XFZYD in the treatment of CHD were searched, and the effective ingredients and core targets of XFZYD in the treatment of CHD were docked through molecular docking, providing theoretical support for clinical treatment of CHD. Results and Conclusion: Through this study, we found that XFZYD has a significant therapeutic effect in the clinical treatment of coronary heart disease, which can play a role in the treatment of CHD by inhibiting atherosclerosis, inhibiting cardiovascular remodeling, improving oxidative stress damage, improving hemorheology, improving myocardial fibrosis and other mechanisms. Through computer simulation, it was found that the main effective components of XFZYD treatment for CHD were quercetin, kaempferol and luteolin, and the key core targets were IL6, VEGFA and P53, and each component had a high VEGFA libdock score. It is speculated that VEGFA is the key target of XFZYD in the treatment of CHD. Kaempferol and VEGFA had the highest libdock score. kaempferol and IL6 have the highest number of hydrogen bonds, kaempferol and IL6 have the highest number of hydrogen bonds, which indicates that they are most stable, indicating that kaempferol is the key component of XFZYD in the treatment of CHD, which provides a theoretical basis for follow-up experimental research.

Clinical intervention effect of Xuefu Zhuyu decoction on chronic heart failure complicated with depression.

BACKGROUND: The diagnosis and treatment of depression in patients with chronic heart failure (CHF) is challenging, with no ideal treatment at present. AIM: To analyze the clinical intervention effect of Xuefu Zhuyu decoction (XFZYD) on CHF complicated with depression. METHODS: The study cohort comprised 116 patients with CHF complicated with depression who received treatment from July 2020 to July 2023, of which 55 received Western medicine (control group) and 61 received XFZYD (research group). Data on clinical effectiveness, traditional Chinese medicine (TCM) syndrome score, cardiac function, negative emotions, and serum inflammatory factors, were collected for comparative analyses. RESULTS: Compared with the control group, the research group had an evidently higher total effective rate. Furthermore, there were marked reductions in TCM symptom score, left ventricular end-diastolic diameter, left ventricular end-systolic diameter, Self-Rating Depression Scale, Hamilton Depression Scale, high-sensitivity C-reactive protein, monocyte chemoattractant protein-1, and matrix metalloproteinase-9 in the research group after treatment, and these were lower than the corresponding values in the control group. Left ventricular ejection fraction was increased and higher in the research group compared with the control group after treatment. CONCLUSION: Our findings conclusively proved that XFZYD was considerably superior to Western medicine for treating CHF complicated with depression because it significantly alleviated patients' symptoms, improved cardiac function, relieved negative emotions, and reduced the levels of serum inflammatory factors.

Xuefu Zhuyu decoction alleviates deep vein thrombosis through inhibiting the activation of platelets and neutrophils via sirtuin 1/nuclear factor kappa-B pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Xuefu Zhuyu Decoction (XZD), a renowned traditional Chinese medicine prescription, is widely employed for the management of conditions characterized by qi-stagnation and blood stasis. Although its anti-thrombotic effect on deep vein thrombosis (DVT) patients has been clinically observed, the underlying mechanism remains largely unexplored. AIM OF THE STUDY: Our aim was to investigate the mechanisms by which XZD exerted its effect on DVT. MATERIALS AND METHODS: The ultra performance liquid chromatography (UPLC) technique was employed to evaluate quality of XZD. To examine the effect of XZD on DVT, a DVT rat model with inferior vena cava (IVC) stenosis was established. The 4D-label-free proteomics approach was then utilized to uncover the possible mechanisms of XZD against DVT. Based on proteomics, citrullinated histone H3 (CitH3), along with serum levels of tumor necrosis factor-alpha (TNF-α) and interleukin-1 beta (IL-1ß) were observed the inhibitory activity of XZD on neutrophil activation. Subsequently, the marker of platelet activation, specifically glycoprotein IIb (CD41) and glycoprotein IIIa (CD61), were assessed along with the secretion of von Willebrand factor (vWF) to investigate the inhibitory activity of XZD on platelet activation. Finally, we explored the impact of XZD on the sirtuin 1 (SIRT1)/nuclear factor kappa-B (NF-κB) pathway, which was associated with the activation of platelets and neutrophils. RESULTS: Eight distinct components were identified for the quality control of XZD. XZD effectively reduced thrombus weight and length in DVT rats, without affecting the coagulation function or hematological parameters in the systemic circulation. Proteomics analysis revealed that XZD alleviated DVT by inhibiting the activation of platelets and neutrophils. The protein expression of CitH3, along with serum levels of TNF-α and IL-1ß, were reduced in XZD-treated DVT rats. Similarly, protein expressions of CD41 and CD61, along with the release of vWF, were markedly down-regulated in XZD-treated DVT rats. Finally, treatment with XZD resulted in an up-regulation of SIRT1 protein expression and a down-regulation of both acetylated NF-κB/p65 and phosphorylated NF-κB/p65 protein expressions in endothelium. CONCLUSIONS: XZD alleviates DVT by inhibiting the activation of platelets and neutrophils at the injured endothelium via the regulation of SIRT1/NF-κB pathway.

Xuefu Zhuyu decoction promotes synaptic plasticity by targeting miR-191a-5p/BDNF-TrkB axis in severe traumatic brain injury.

BACKGROUND: Xuefu Zhuyu decoction (XFZYD) is a traditional Chinese herbal formula known for its ability to eliminate blood stasis and improve blood circulation, providing neuroprotection against severe traumatic brain injury (sTBI). However, the underlying mechanism is still unclear. PURPOSE: We aim to investigate the neuroprotective effects of XFZYD in sTBI from a novel mechanistic perspective of miRNA-mRNA. Additionally, we sought to elucidate a potential specific mechanism by integrating transcriptomics, bioinformatics, and conducting both in vitro and in vivo experiments. METHODS: The sTBI rat model was established, and the rats were treated with XFZYD for 14 days. The neuroprotective effects of XFZYD were evaluated using a modified neurological severity score, hematoxylin and eosin staining, as well as Nissl staining. The anti-inflammatory effects of XFZYD were explored using quantitative real-time PCR (qRT-PCR), Western blot analysis, and immunofluorescence. Next, miRNA sequencing of the hippocampus was performed to determine which miRNAs were differentially expressed. Subsequently, qRT-PCR was used to validate the differentially expressed miRNAs. Target core mRNAs were determined using various methods, including miRNA prediction targets, mRNA sequencing, miRNA-mRNA network, and protein-protein interaction (PPI) analysis. The miRNA/mRNA regulatory axis were verified through qRT-PCR or Western blot analysis. Finally, morphological changes in the neural synapses were observed using transmission electron microscopy and immunofluorescence. RESULTS: XFZYD exhibited significant neuroprotective and anti-inflammatory effects on subacute sTBI rats' hippocampus. The analyses of miRNA/mRNA sequences combined with the PPI network revealed that the therapeutic effects of XFZYD on sTBI were associated with the regulation of the rno-miR-191a-5p/BDNF axis. Subsequently, qRT-PCR and Western blot analysis confirmed XFZYD reversed the decrease of BDNF and TrkB in the hippocampus caused by sTBI. Additionally, XFZYD treatment potentially increased the number of synaptic connections, and the expression of the synapse-related protein PSD95, axon-related protein GAP43 and neuron-specific protein TUBB3. CONCLUSIONS: XFZYD exerts neuroprotective effects by promoting hippocampal synaptic remodeling and improving cognition during the subacute phase of sTBI through downregulating of rno-miR-191a-5p/BDNF axis, further activating BDNF-TrkB signaling.

Mechanisms of Xuefu Zhuyu Decoction in the treatment of coronary heart disease based on integrated metabolomics and network pharmacology approach.

Coronary heart disease (CHD) has become the leading cause of mortality, morbidity, and disability worldwide. Though the therapeutic effect of Xuefu Zhuyu Decoction (XFZY) on CHD has been demonstrated in China, the active ingredients and molecular mechanisms of XFZY have not been elucidated. The purpose of the current study is to explore the molecular mechanisms of XFZY in the treatment of CHD via network pharmacology, metabolomics, and experimental validation. First, we established a CHD rat model by permanently ligating the left anterior descending coronary artery (LAD), and evaluated the therapeutic effect of XFZY by hemorheology and histopathology. Second, network pharmacology was employed to screen the active ingredients and potential targets of XFZY for the treatment of CHD. Metabolomic was applied to identify the molecules present in the serum after XFZY treatment. Third, the results of network pharmacology and metabolomics were further analyzed by Cytoscape to elucidate the core ingredients and pathways. Finally, the obtained key pathways were verified by transmission electron microscopy and immunofluorescence assay. The results showed that XFZY was effective in the treatment of CHD in the rat model, and the highest dose exerted the best effect. Network pharmacology analysis revealed 215 active ingredients and 129 key targets associated with XFZY treatment of CHD. These targets were enriched in pathways of cancer, lipid and atherosclerosis, fluid shear stress and atherosclerosis, proteoglycans in cancer, chemical carcinogenesis - receptor activation, HIF-1 signaling, et al. Serum metabolomic identified 1081 metabolites involved in the therapeutic effect of XFZY on CHD. These metabolites were enriched in taurine and hypotaurine metabolism, histidine metabolism, retrograde endocannabinoid signaling pathways, et al. Cytoscape analysis combining the data from serum metabolomic and network pharmacology revealed that energy metabolism as the core pathway for XFZY treatment of CHD. Electron microscope observation identified changes in the level of autophagy in the mitochondrial structure of cardiomyocytes. Immunofluorescence assay showed that the expression levels of autophagy-related proteins LC3-B and P62/SQSTM1 were consistent with the levels of autophagy observed in mitochondria. In conclusion, our findings suggest that the possible mechanisms of XFZY in the treatment of CHD are reducing the level of autophagy, improving energy metabolism, and maintaining mitochondrial homeostasis in cardiomyocytes. Our study also shows that the combined strategies of network pharmacology, metabolomics, and experimental validation may provide a powerful approach for TCM pharmacology study.

Identification of prototype compounds and their metabolites in rats' serum from Xuefu Zhuyu Decoction by UPLC-Q-TOF/MS.

Objective: As a classic prescription in traditional Chinese medicine, Xuefu Zhuyu Decoction (XFZYD) has been widely used in the clinical treatment of cardiovascular and cerebrovascular diseases. In order to unveil the potentially effective compounds, a rapid ultra-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS) method was established to identify prototype compounds and their metabolites from XFZYD in rats' serum. Methods: The serum from rats after intragastric administration of XFZYD aqueous extract was analyzed by UPLC-Q-TOF/MS method. The prototype compounds and their metabolites were identified by comparison with the reference standards and tentatively characterized by comprehensively analyzing the retention time, MS data, characteristic MS fragmentation pattern and retrieving literatures. Results: A total of 175 compounds (24 prototype compounds and 151 metabolites) were identified and tentatively characterized. The metabolic pathways of prototype compounds in vivo were also summarized, including glucuronidation, hydrolyzation, sulfation, demethylation, and hydroxylation, and so on. Conclusion: In this study, a UPLC-Q-TOF/MS technique was developed to analyze prototype compounds and their metabolites from XFZYD in serum, which would provide the evidence for further studying the effective compounds of XFZYD.

N6-methyladenosine profiling reveals that Xuefu Zhuyu decoction upregulates METTL14 and BDNF in a rat model of traumatic brain injury.

ETHNOPHARMACOLOGICAL RELEVANCE: The traditional Chinese herbal formula Xuefu Zhuyu decoction (XFZYD) is a classic formula in the category of invigorating blood circulation and resolving blood stasis. It has been proven to improve the neurological and ethological prognosis of traumatic brain injury. XFZYD promotes synaptic and axonal regeneration after traumatic brain injury, which is functionally modulated by the N6-methyladenosine (m6A) modification of RNA. However, the epigenetic effects of XFZYD on m6A modification remain unknown. AIM OF THE STUDY: To explore how XFZYD protects against traumatic brain injury induced by controlled cortical impact (CCI) injury by altering RNA m6A modification. MATERIALS AND METHODS: The modified neurological severity scoring and Morris water maze were performed to evaluate the neuroprotective effects of XFZYD for 14 days and screen the dose. Then, dot blot, western blotting, and methylated RNA immunoprecipitation sequencing (MeRIP-Seq) were used to explore changes in RNA m6A modification in the perilesional cortex. The Metascape platform was used to analyze the Gene Ontology, Kyoto Encyclopedia of Genes and Genomes (KEGG), and Reactome pathway of the differential m6A-tagged genes. Furthermore, MeRIP-qPCR was conducted to quantify differences in the hub differential m6A modification gene brain-derived neurotrophic factor (Bdnf). RESULTS: XFZYD significantly ameliorated the neurological deficits, spatial learning, and memory impairments in rats post-CCI on day 14. XFZYD enhanced the m6A level, and the expression of METTL14 and YTHDC2 in the perilesional cortex of CCI rats. In all three groups, the 3'-untranslated regions and coding sequence were primarily enriched for m6A peaks. XFZYD reversed the increased proportion of 3'-untranslated regions, and the decreased proportion of coding sequence and 5'-untranslated regions post-CCI. Moreover, XFZYD markedly downregulated 41 elevated m6A-tagged transcripts and upregulated 119 decreased m6A-tagged transcripts following CCI. Gene ontology and KEGG pathway analysis revealed that XFZYD-regulated m6A-tagged transcripts were predominantly enriched in synapse assembly, synaptic plasticity, learning or memory, and MAPK signaling pathway. Then, the hub-regulated m6A-tagged gene BDNF was identified. Both the m6A methylation level and the protein level of BDNF were ascended by XFZYD treatment. CONCLUSION: XFZYD improves neurological deficits, spatial learning and memory impairments in rats post-TBI probably through increasing the expression of METTL14 and BDNF in the cortex. Our study highlights a novel post-transcriptional regulation mechanism mediated by herbal medicine for traumatic brain injury treatment.

Network pharmacological analysis of Xuefu Zhuyu decoction in the treatment of atherosclerosis.

Objective: Using a network pharmacological approach, this study will evaluate the effect of Xuefu Zhuyu Decoction in the treatment of atherosclerosis. Methods: The data were imported into the STRING database to construct a protein-protein interaction network, and the network topology was analysed with the Bisogenet plug-in by Cytoscape 3.7.2. Using the R language Bioconductor platform, Gene Ontology (GO) enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis for potential targets of Xuefu Zhuyu Decoction in the treatment of atherosclerosis were performed, and import the results were imported into Cytoscape 3.7.2. To map the results and create a KEGG network diagram, we used Cytoscape 3.7.2 for analysis. Results: A total of 91 chemical components and 1320 disease targets were obtained, including 138 cross-targets. TNF, AKT1 and ALB were identified as important targets, and Gene Ontology functional analysis indicated that biological process was the primary cause of oxidative stress. The primary action of molecular function is binding. KEGG has explored and enriched 149 signalling pathways, including the AGE-RAGE signalling system and the TNF signalling network. According to a study involving molecular docking, quercetin and ß-carotene have a strong binding affinity for AKT1 and ALB. Conclusion: The potential of Xuefu Zhuyu Decoction to treat atherosclerosis through multiple components and targets provides a way to further study its mechanism.

Bioinformatics Analysis of miRNAs and mRNAs Network-Xuefu Zhuyu Decoction Exerts Neuroprotection of Traumatic Brain Injury Mice in the Subacute Phase.

Xuefu Zhuyu decoction (XFZYD) is used to treat traumatic brain injury (TBI). XFZYD-based therapies have achieved good clinical outcomes in TBI. However, the underlying mechanisms of XFZYD in TBI remedy remains unclear. The study aimed to identify critical miRNAs and putative mechanisms associated with XFYZD through comprehensive bioinformatics analysis. We established a controlled cortical impact (CCI) mice model and treated the mice with XFZYD. The high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) confirmed the quality of XFZYD. The modified neurological severity score (mNSS) and Morris water maze (MWM) tests indicated that XFZYD improved the neurological deficit (p < 0.05) and cognitive function (p < 0.01). Histological analysis validated the establishment of the CCI model and the treatment effect of XFZYD. HE staining displayed that the pathological degree in the XFZYD-treated group was prominently reduced. The transcriptomic data was generated using microRNA sequencing (miRNA-seq) of the hippocampus. According to cluster analysis, the TBI group clustered together was distinct from the XFZYD group. Sixteen differentially expressed (5 upregulated; 11 downregulated) miRNAs were detected between TBI and XFZYD. The reliability of the sequencing data was confirmed by qRT-PCR. Three miRNAs (mmu-miR-142a-5p, mmu-miR-183-5p, mmu-miR-96-5p) were distinctively expressed in the XFZYD compared with the TBI and consisted of the sequencing results. Bioinformatics analysis suggested that the MAPK signaling pathway contributes to TBI pathophysiology and XFZYD treatment. Subsequently, the functions of miR-96-5p, miR-183-5p, and miR-142a-5p were validated in vitro. TBI significantly induces the down-expression of miR-96-5p, and up-expression of inflammatory cytokines, which were all inhibited by miR-96-5p mimics. The present research provides an adequate fundament for further knowing the pathologic and prognostic process of TBI and supplies deep insights into the therapeutic effects of XFZYD.

The mechanisms underlying the actions of Xuefu Zhuyu decoction pretreatment against neurological deficits after ischemic stroke in mice: The mediation of glymphatic function by aquaporin-4 and its anchoring proteins.

Ischemic stroke (IS) has been associated with an impairment in glymphatic function. Xuefu Zhuyu Decoction (XFZYD) is widely used in the prevention and treatment of ischemic stroke. We hypothesized that Xuefu Zhuyu decoction pretreatment could attenuate early neurological deficits after ischemic stroke by enhancing the function of the glymphatic system. To prove our hypothesis, we carried out temporary middle cerebral artery occlusion and reperfusion surgery on C57BL/6 mice and then measured neurological score, infarct size and performed hematoxylin-eosin staining to assess stroke outcomes after 24 h of reperfusion. Subsequently, we injected fluorescent tracers in to the cisterna magna and evaluated tracer distribution in coronal brain sections. The polarization of aquaporin-4 (AQP4), colocalization of aquaporin-4, α-dystroglycan, ß-dystroglycan and agrin were determined by immunofluorescence. Our research showed that pretreatment with Xuefu Zhuyu decoction significantly alleviated neurological scores, neurological deficits and pathological abnormalities in a mouse model of ischemic stroke. Importantly, Xuefu Zhuyu decoction pretreatment enhanced cerebrospinal fluid influx, protected aquaporin-4 depolarization and promoted the colocalization of aquaporin-4 with its anchoring proteins in the brain. Our findings highlight novel mechanisms underlying the neuroprotective effect of Xuefu Zhuyu decoction pretreatment on ischemic stroke-induced brain damage through the glymphatic system. Xuefu Zhuyu decoction pretreatment may offer a promising approach to slow the onset and progression of ischemic stroke.