[Zhongfeng Xingnao Decoction alleviates intracerebral haemorrhage-induced brain injury via inhibiting neuronal ferroptosis].

Zhongfeng Xingnao Decoction(ZFXN) has been utilized for treating intracerebral hemorrhage(ICH) in China, while the pharmacological mechanism of ZFXN remains unclear. Exploring the pharmacological roles of ZFXN is critical for guiding the treatment of cerebrovascular diseases. In this study, a rat model of ICH was constructed by injection of Ⅶ collagenase in the right caudate nucleus. SD rats were randomly assigned into five groups, and the neurological function of rats was evaluated based on the Bederson score. Magnetic resonance imaging(MRI) was used to assess the volume of ICH. Hematoxylin-eosin(HE) staining and transmission electron microscopy(TEM) were employed to observe the pathological and ultrastructural changes in the brain tissue. The levels of reactive oxygen species(ROS) were measured by flow cytometry. The immunofluorescence assay was employed to detect the expression of glutathione peroxidase 4(GPX4) in neurons surrounding the hematoma. Finally, Western blot was employed to determine the expression of ferroptosis-related proteins upstream frameshift 1(UPF1), ferroportin(FPN), acyl-CoA ligase 4(ACSL4), cyclooxyge-nase-2(COX-2), GPX4, NADPH oxidase 1(NOX1), and solute carrier family 7 member 11(SLC7A11) after ICH. Compared with the model(ICH) group, ZFXN treatment for 5 days attenuated neurological dysfunction, reduced the hematoma volume, and alleviated the pathological changes induced by ICH. Meanwhile, ZFXN lowered the levels of Fe~(2+) and oxidative stress and up-regulated the expression of proteins inhibiting ferroptosis. ZFXN improved the prognosis of ICH in rats by inhibiting neuronal ferroptosis, which provided a valuable guide for the clinical application of ZFXN. ZFXN may inhibit ferroptosis by promoting the expression of SLC7A11 and FPN.

Amygdalin prevents multidrug-resistant Staphylococcus aureus-induced lung epithelial cell injury by regulating inflammation and oxidative stress.

Methicillin-resistant Staphylococcus aureus (MRSA) is an opportunistic pathogen that can cause severe bacterial pneumonia. Amygdalin is the main active pharmaceutical ingredient of bitter almond, which has broad-spectrum antibacterial, anti-inflammatory, anti-oxidation and immunomodulatory effects. It is also the main ingredient of Yinhua Pinggan granule, which is commonly used to moisten the lung and relieve cough. However, little is known about the effects of amygdalin on MRSA. In this study, we found that amygdalin exhibited good antimicrobial activity in vitro against MRSA. Amygdalin has a protective effect on MRSA infected cells, and the effect is better when combined with levofloxacin. It also can reduce the adhesion and invasion of MRSA to cells. Amygdalin has anti-inflammatory and antioxidant effects, which can significantly reduce the increase of inflammatory factors and the production of ROS caused by infection. The protective mechanism of amygdalin on cells may be related to inhibiting the expression of NLRP3, ASC and IL-1ß pyroptosis pathways. Taken together, our study suggests that amygdalin exerts antibacterial effects by affecting biofilm formation, the expression of virulence factors, and drug resistance genes. Amygdalin combined with levofloxacin has a protective effect on A549 cells infected with MRSA, and the mechanism may be related to the inhibition of inflammatory response, oxidative damage and pyroptosis.

A Combination of Astragaloside IV and Hydroxysafflor Yellow A Attenuates Cerebral Ischemia-Reperfusion Injury via NF-κB/NLRP3/Caspase-1/GSDMD Pathway.

Cerebral ischemia-reperfusion injury (IRI), occurring after blood supply restoration, contributes significantly to stroke-related deaths. This study explored the combined impact and mechanisms of astragaloside IV (AS-IV), hydroxysafflor yellow A (HSYA), and their combination in mitigating IRI. Male Sprague-Dawley (SD) rats were randomized to the Sham, MCAO, MCAO+AS-IV, MCAO+HSYA, and MCAO+AS-IV+HSYA groups. Neurological deficits and cerebral infarction were examined after restoring the blood supply to the brain. Pathomorphological changes in the cerebral cortex were observed via HE staining. IL-1ß and IL-18 were quantified using ELISA. The expression of NF-κB and GSDMD in the ischemic cerebrum was analyzed using immunohistochemistry. The expression levels of NLRP3, ASC, IL-1ß, Caspase-1, and GSDMD in the ischemic cerebrum were evaluated using Western blot. The MCAO+AS-IV, MCAO+HSYA, and MCAO+AS-IV+HSYA groups exhibited notably better neurological function and cerebral infarction compared with the MCAO group. The combined treatment demonstrated superior brain tissue injury alleviation. Reductions in NF-κB, GSDMD positive cells, and NLRP3/ASC/IL-1ß/Caspase-1/GSDMD protein expression in the ischemic brain were significantly more pronounced with the combined therapy, indicating a synergistic effect in countering cerebral IRI via the NF-κB/NLRP3/Caspase-1/GSDMD pathway inhibition of cell pyroptosis-induced injury.

Anaerobic Glycolysis and Ischemic Stroke: From Mechanisms and Signaling Pathways to Natural Product Therapy.

Ischemic stroke is a serious condition that results in high rates of illness and death. Anaerobic glycolysis becomes the primary means of providing energy to the brain during periods of low oxygen levels, such as in the aftermath of an ischemic stroke. This process is essential for maintaining vital brain functions and has significant implications for recovery following a stroke. Energy supply by anaerobic glycolysis and acidosis caused by lactic acid accumulation are important pathological processes after ischemic stroke. Numerous natural products regulate glucose and lactate, which in turn modulate anaerobic glycolysis. This article focuses on the relationship between anaerobic glycolysis and ischemic stroke, as well as the associated signaling pathways and natural products that play a therapeutic role. These natural products, which can regulate anaerobic glycolysis, will provide new avenues and perspectives for the treatment of ischemic stroke in the future.

Catalpol reduced LPS induced BV2 immunoreactivity through NF-κB/NLRP3 pathways: an in Vitro and in silico study.

Background: Ischemic Stroke (IS) stands as one of the primary cerebrovascular diseases profoundly linked with inflammation. In the context of neuroinflammation, an excessive activation of microglia has been observed. Consequently, regulating microglial activation emerges as a vital target for neuroinflammation treatment. Catalpol (CAT), a natural compound known for its anti-inflammatory properties, holds promise in this regard. However, its potential to modulate neuroinflammatory responses in the brain, especially on microglial cells, requires comprehensive exploration. Methods: In our study, we investigated into the potential anti-inflammatory effects of catalpol using lipopolysaccharide (LPS)-stimulated BV2 microglial cells as an experimental model. The production of nitric oxide (NO) by LPS-activated BV2 cells was quantified using the Griess reaction. Immunofluorescence was employed to measure glial cell activation markers. RT-qPCR was utilized to assess mRNA levels of various inflammatory markers. Western blot analysis examined protein expression in LPS-activated BV2 cells. NF-κB nuclear localization was detected by immunofluorescent staining. Additionally, molecular docking and molecular dynamics simulations (MDs) were conducted to explore the binding affinity of catalpol with key targets. Results: Catalpol effectively suppressed the production of nitric oxide (NO) induced by LPS and reduced the expression of microglial cell activation markers, including Iba-1. Furthermore, we observed that catalpol downregulated the mRNA expression of proinflammatory cytokines such as IL-6, TNF-α, and IL-1ß, as well as key molecules involved in the NLRP3 inflammasome and NF-κB pathway, including NLRP3, NF-κB, caspase-1, and ASC. Our mechanistic investigations shed light on how catalpol operates against neuroinflammation. It was evident that catalpol significantly inhibited the phosphorylation of NF-κB and NLRP3 inflammasome activation, both of which serve as upstream regulators of the inflammatory cascade. Molecular docking and MDs showed strong binding interactions between catalpol and key targets such as NF-κB, NLRP3, and IL-1ß. Conclusion: Our findings support the idea that catalpol holds the potential to alleviate neuroinflammation, and it is achieved by inhibiting the activation of NLRP3 inflammasome and NF-κB, ultimately leading to the downregulation of pro-inflammatory cytokines. Catalpol emerges as a promising candidate for the treatment of neuroinflammatory conditions.

Post-Ischemic Stroke Cardiovascular Risk Prevention and Management.

Cardiac death is the second most common cause of death among patients with acute ischemic stroke (IS), following neurological death resulting directly from acute IS. Risk prediction models and screening tools including electrocardiograms can assess the risk of adverse cardiovascular events after IS. Prolonged heart rate monitoring and early anticoagulation therapy benefit patients with a higher risk of adverse events, especially stroke patients with atrial fibrillation. IS and cardiovascular diseases have similar risk factors which, if optimally managed, may reduce the incidence of recurrent stroke and other major cardiovascular adverse events. Comprehensive risk management emphasizes a healthy lifestyle and medication therapy, especially lipid-lowering, glucose-lowering, and blood pressure-lowering drugs. Although antiplatelet and anticoagulation therapy are preferred to prevent cardiovascular events after IS, a balance between preventing recurrent stroke and secondary bleeding should be maintained. Optimization of early rehabilitation care comprises continuous care across environments thus improving the prognosis of stroke survivors.

Efficacy of Astragalus membranaceus-Carthamus tinctorius in cerebral ischemia/reperfusion injury: Insights from metabolomics and mass spectrometry imaging.

BACKGROUND: The combination of Astragalus membranaceus and Carthamus tinctorius (AC) exhibits significant therapeutic effects in cerebral ischemia/reperfusion injury (CIRI). Understanding the metabolic characteristics of brain microregions and disturbances in tissues and systemic circulation is crucial for elucidating the mechanisms of CIRI and the therapeutic benefits of AC. However, in situ metabolic regulation of the complex brain structure has not been adequately studied, and the therapeutic mechanism of AC requires immediate clarification. PURPOSE: The present study aimed to unveil the specific metabolic reprogramming of CIRI at systemic and microregional levels, identify key metabolic pathways and metabolites, and elucidate the therapeutic mechanisms of AC. METHODS: Air flow-assisted desorption electrospray ionization mass spectrometry imaging (AFADESI-MSI), a newly developed technique, was used to investigate metabolites in brain microregions. Hematoxylin-eosin, Nissl, and immunofluorescence staining were performed to visualize the microscopic changes associated with spatial metabolism. A comprehensive metabolomics study was conducted on serum, brain tissue, and microregions, along with neurological assessments, cerebral infarction measurements, and Evans blue experiments, to assess the systemic and local metabolic effects of AC treatment for CIRI. RESULTS: AC significantly reduced neurological damage, minimized infarct size, and repaired blood-brain barrier damage in CIRI rats. AFADESI-MSI demonstrated that the metabolic imbalance caused by CIRI primarily occurs in the cerebral cortex, hippocampus, caudate putamen, thalamus, cerebellar cortex, and fiber tract regions. Significant changes in 16 metabolites were observed in these regions, corresponding to neuron damage, glial cell activation, and neural repair. 20 metabolites from serum and 4 from brain tissue varied significantly with the sham group. Comprehensive metabolomics analysis indicated a close relationship among serum, tissue, and microregional metabolism. CIRI-induced systemic and localized metabolic disorders involve 14 metabolic pathways. AC conferred therapeutic benefits in CIRI by reversing various metabolic imbalances. CONCLUSION: AFADESI-MSI efficiently visualized brain microregion metabolism. Comprehensive metabolomics analysis revealed detailed insights into the specific metabolic reprogramming in CIRI and the therapeutic impacts of AC. AC demonstrated significant clinical potential as an adjunct therapy to existing CIRI treatments.

Mechanisms of cuproptosis and its relevance to distinct diseases.

Copper is a trace element required by the organism, but once the level of copper exceeds the threshold, it becomes toxic and even causes death. The underlying mechanisms of copper-induced death are inconclusive, with different studies showing different opinions on the mechanism of copper-induced death. Multiple investigations have shown that copper induces oxidative stress, endoplasmic reticulum stress, nucleolar stress, and proteasome inhibition, all of which can result in cell death. The latest research elucidates a copper-dependent death and denominates it as cuproptosis. Cuproptosis takes place through the combination of copper and lipoylated proteins of the tricarboxylic acid cycle, triggering agglomeration of lipoylated proteins and loss of iron-sulfur cluster proteins, leading to proteotoxic stress and ultimately death. Given the toxicity and necessity of copper, abnormal levels of copper lead to diseases such as neurological diseases and cancer. The development of cancer has a high demand for copper, neurological diseases involve the change of copper contents and the binding of copper to proteins. There is a close relationship between these two kinds of diseases and copper. Here, we summarize the mechanisms of copper-related death, and the association between copper and diseases, to better figure out the influence of copper in cell death and diseases, thus advancing the clinical remedy of these diseases.

Naoxintong capsule remodels gut microbiota and ameliorates early-stage atherosclerosis in apolipoprotein E-deficient mice.

BACKGROUND: Naoxintong capsule (NXT) is a compound traditional Chinese medicine prescription with demonstrated effect for the treatment of cardiovascular and cerebrovascular diseases including atherosclerosis (AS). However, the pharmacological mechanisms of NXT in ameliorating early-stage AS are still unclear, especially regarding the role of gut microbiota. PURPOSE: This study is aiming to evaluate the therapeutic effect of NXT against early-stage AS, and further illustrate the potential correlations among AS, gut microbiota, and NXT. METHODS: Thirty-two male ApoE knockout mice (C57BL/6 background) were fed with a high cholesterol diet (HCD) for 4 weeks to establish an early-stage AS model. NXT in two different dosages and simvastatin (Simv) were than administrated for another 8 weeks. Lipid metabolism indicators and inflammation levels were measured with corresponding assay kits. Changes in blood vessels, liver lesions, and intestinal barrier proteins were evaluated with different staining methods. Furthermore, the gut microbiota structure was analyzed using 16S rRNA sequencing technology, while GC-MS was utilized to determine the fecal contents of short-chain fatty acids (SCFAs). RESULTS: Administration of NXT significantly ameliorated obesity, hyperlipidemia, systemic inflammation, vasculopathy, liver injury, and intestinal barrier disorder in AS mice. Administration of NXT also significantly regulated the gut microbiota disturbance and increased the total contents of fecal SCFAs in AS mice. Furthermore, acetic acid content and the relative abundance of Faecalibacterium in feces were proposed as potential therapeutic biomarkers of NXT for AS treatment as indicated via the correlation analysis. CONCLUSION: This study demonstrated that NXT could effectively treat early-stage AS induced by HCD in mice. NXT regulated the gut microbiota and metabolites, maintained intestinal homeostasis, and improved the systemic inflammatory response. These findings may provide robust experimental support for the clinical use of NXT for AS treatment.

Synergistic promotion of angiogenesis after intracerebral hemorrhage by ginsenoside Rh2 and chrysophanol in rats.

BACKGROUND: Intracerebral hemorrhage (ICH) is a debilitating condition characterized by the rupture of cerebral blood vessels, resulting in profound neurological deficits. A significant challenge in the treatment of ICH lies in the brain's limited capacity to regenerate damaged blood vessels. This study explores the potential synergistic effects of Ginsenoside Rh2 and Chrysophanol in promoting angiogenesis following ICH in a rat model. METHODS: Network pharmacology was employed to predict the potential targets and pathways of Ginsenoside Rh2 and Chrysophanol for ICH treatment. Molecular docking was utilized to assess the binding affinity between these compounds and their respective targets. Experimental ICH was induced in male Sprague-Dawley rats through stereotactic injection of type VII collagenase into the right caudate putamen (CPu). The study encompassed various methodologies, including administration protocols, assessments of neurological function, magnetic resonance imaging, histological examination, observation of brain tissue ultrastructure, reverse transcription-quantitative polymerase chain reaction (RT-qPCR), immunofluorescence staining, Western blot analysis, and statistical analyses. RESULTS: Network pharmacology analysis indicated that Ginsenoside Rh2 and Chrysophanol may exert their therapeutic effects in ICH by promoting angiogenesis. Results from animal experiments revealed that rats treated with Ginsenoside Rh2 and Chrysophanol exhibited significantly improved neurological function, reduced hematoma volume, and diminished pathological injury compared to the Model group. Immunofluorescence analysis demonstrated enhanced expression of vascular endothelial growth factor receptor 2 (VEGFR2) and CD31, signifying augmented angiogenesis in the peri-hematomal region following combination therapy. Importantly, the addition of a VEGFR2 inhibitor reversed the increased expression of VEGFR2 and CD31. Furthermore, Western blot analysis revealed upregulated expression of angiogenesis-related factors, including VEGFR2, SRC, AKT1, MAPK1, and MAPK14, in the combination therapy group, but this effect was abrogated upon VEGFR2 inhibitor administration. CONCLUSION: The synergistic effect of Ginsenoside Rh2 and Chrysophanol demonstrated a notable protective impact on ICH injury in rats, specifically attributed to their facilitation of angiogenesis. Consequently, this research offers a foundation for the utilization of Ginsenosides Rh2 and Chrysophanol in medical settings and offers direction for the advancement of novel pharmaceuticals for the clinical management of ICH.

Therapeutic effect of Yinhuapinggan granules mediated through the intestinal flora in mice infected with the H1N1 influenza virus.

Objective: In this study, we examined the therapeutic effects of Yinhuapinggan granules (YHPGs) in influenza-infected mice. We also examined how YHPGs affect the composition of the intestinal flora and associated metabolites. Methods: We used the nasal drip method to administer the influenza A virus (IAV) H1N1 to ICR mice. Following successful model construction, the mice were injected with 0.9% sterile saline and low (5.5 g/kg), medium (11 g/kg), and high (22 g/kg) doses of YHPGs. The pathological changes in the lungs and intestines were evaluated by gavage for 5 consecutive days. Detection of sIgA, IL-6, TNF-α, INF-γ, and TGF-ß cytokine levels in serum by enzyme-linked immunosorbent assay. Real-time fluorescence quantitative polymerase chain reaction and Western blot were used to measure the mRNA and protein expression of the tight junction proteins claudin-1, occludin, and zonula occludens-1 (ZO-1) in the colon. To assess the influence of YHPGs on the intestinal microbiota, feces were obtained from the mice for 16s rRNA sequencing, and short-chain fatty acids (SCFAs) were measured in the feces. Results: By reducing the production of pro-inflammatory cytokines and increasing the relative expression of claudin-1, occludin, and ZO-1 in colon tissues, YHPGs had a protective effect in tissues from the lungs and colon. When YHPGs were administered to mice with IAV infection, the relative abundance of Lactobacillus, Coprobacillus, Akkermansia, Prevotella, Oscillospira, and Ruminococcus increased, whereas the relative abundance of Desulfovibrio decreased. Conclusion: The therapeutic mechanism of YHPGs against IAV infection in mice may be underpinned by modulation of the structural composition of colonic bacteria and regulation of SCFA production.

Research progress on immune-related therapeutic targets of brain injury caused by cerebral ischemia.

Stroke is the second leading cause of death worldwide and a leading cause of disability. The innate immune response occurs immediately after cerebral ischemia, resulting in adaptive immunity. More and more experimental evidence has proved that the immune response caused by cerebral ischemia plays an important role in early brain injury and later the recovery of brain injury. Innate immune cells and adaptive cells promote the occurrence of cerebral ischemic injury but also protect brain cells. A large number of studies have shown that cytokines and immune-related substances also have dual functions of promoting injury, reducing injury, or promoting injury recovery in the later stage of cerebral ischemia. They can be an important target for treating cerebral ischemic recovery. Therefore, this study discussed the immune cells, cytokines, and immune-related substances with dual roles in cerebral ischemia and summarized the therapeutic targets of cerebral ischemia. To explore more effective methods to treat cerebral ischemia, promote the recovery of brain function, and improve the prognosis of patients.

Development and Application of Traditional Chinese Medicine Using AI Machine Learning and Deep Learning Strategies.

Traditional Chinese medicine (TCM) has been used for thousands of years and has been proven to be effective at treating many complicated illnesses with minimal side effects. The application and advancement of TCM are, however, constrained by the absence of objective measuring standards due to its relatively abstract diagnostic methods and syndrome differentiation theories. Ongoing developments in machine learning (ML) and deep learning (DL), specifically in computer vision (CV) and natural language processing (NLP), offer novel opportunities to modernize TCM by exploring the profound connotations of its theory. This review begins with an overview of the ML and DL methods employed in TCM; this is followed by practical instances of these applications. Furthermore, extensive discussions emphasize the mature integration of ML and DL in TCM, such as tongue diagnosis, pulse diagnosis, and syndrome differentiation treatment, highlighting their early successful application in the TCM field. Finally, this study validates the accomplishments and addresses the problems and challenges posed by the application and development of TCM powered by ML and DL. As ML and DL techniques continue to evolve, modern technology will spark new advances in TCM.

Qualitative and Quantitative Analysis of Chemical Components in Yinhua Pinggan Granule with High-Performance Liquid Chromatography Coupled with Q-Exactive Mass Spectrometry.

Yinhua Pinggan Granule (YPG) is an approved compounded traditional Chinese medicine (TCM) prescription for the treatment of cold, cough, viral pneumonia, and related diseases. Due to its complicated chemical composition, the material basis of YPG has not been systematically investigated. In this study, an analytical method based on high-performance liquid chromatography (HPLC) coupled with Q-Exactive mass spectrometry was established. Together with the help of a self-built compound database and Compound Discoverer software 3.1, the chemical components in YPG were tentatively identified. Subsequently, six main components in YPG were quantitatively characterized with a high-performance liquid chromatography-diode array detector (HPLC-DAD) method. As a result, 380 components were annotated, including 19 alkaloids, 8 organic acids, 36 phenolic acids, 27 other phenols, 114 flavonoids, 75 flavonoid glycoside, 72 terpenes, 11 anthraquinones, and 18 other compounds. Six main components, namely, chlorogenic acid, puerarin, 3'-methoxypuerarin, polydatin, glycyrrhizic acid, and emodin, were quantified simultaneously. The calibration curves of all six analytes showed good linearity (R2 > 0.9990) within the test ranges. The precision, repeatability, stability, and recovery values were all in acceptable ranges. In addition, the total phenol content and DPPH scavenging activity of YPG were also determined. The systematic elucidation of the chemical components in YPG in this study may provide clear chemical information for the quality control and pharmacological research of YPG and related TCM compounded prescriptions.

Injectable Hydrogels Based on Hyaluronic Acid and Gelatin Combined with Salvianolic Acid B and Vascular Endothelial Growth Factor for Treatment of Traumatic Brain Injury in Mice.

Traumatic brain injury (TBI) leads to structural damage in the brain, and is one of the major causes of disability and death in the world. Herein, we developed a composite injectable hydrogel (HA/Gel) composed of hyaluronic acid (HA) and gelatin (Gel), loaded with vascular endothelial growth factor (VEGF) and salvianolic acid B (SAB) for treatment of TBI. The HA/Gel hydrogels were formed by the coupling of phenol-rich tyramine-modified HA (HA-TA) and tyramine-modified Gel (Gel-TA) catalyzed by horseradish peroxidase (HRP) in the presence of hydrogen peroxide (H2O2). SEM results showed that HA/Gel hydrogel had a porous structure. Rheological test results showed that the hydrogel possessed appropriate rheological properties, and UV spectrophotometry results showed that the hydrogel exhibited excellent SAB release performance. The results of LIVE/DEAD staining, CCK-8 and Phalloidin/DAPI fluorescence staining showed that the HA/Gel hydrogel possessed good cell biocompatibility. Moreover, the hydrogels loaded with SAB and VEGF (HA/Gel/SAB/VEGF) could effectively promote the proliferation of bone marrow mesenchymal stem cells (BMSCs). In addition, the results of H&E staining, CD31 and α-SMA immunofluorescence staining showed that the HA/Gel/SAB/VEGF hydrogel possessed good in vivo biocompatibility and pro-angiogenic ability. Furthermore, immunohistochemical results showed that the injection of HA/Gel/SAB/VEGF hydrogel to the injury site could effectively reduce the volume of defective tissues in traumatic brain injured mice. Our results suggest that the injection of HA/Gel hydrogel loaded with SAB and VEGF might provide a new approach for therapeutic brain tissue repair after traumatic brain injury.

Exploring the Potential Mechanisms of Guanxinshutong Capsules in Treating Pathological Cardiac Hypertrophy based on Network Pharmacology, Computer-Aided Drug Design, and Animal Experiments.

Cardiovascular diseases (CVDs) are significant causes of morbidity and mortality worldwide, and pathological cardiac hypertrophy (PCH) is an essential predictor of many heart diseases. Guanxinshutong capsule (GXST) is a Chinese patent medicine widely used in the clinical treatment of CVD, In our previous research, we identified 111 compounds of GXST. In order to reveal the potential molecular mechanisms by which GXST treats PCH, this study employed network pharmacology methods to screen for the active ingredients of GXST in treating PCH and predicted the potential targets. The results identified 26 active ingredients of GXST and 110 potential targets for PCH. Through a protein-protein interaction (PPI) network, gene ontology (GO), and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis, we confirmed AKT1, MAPK1, and MAPK3 as the core proteins in GXST treatment of PCH, thus establishing the PI3K/AKT and MAPK signaling pathways as the significant mechanisms of GXST in treating PCH. The results of molecular docking (MD) demonstrate that flavonoid naringenin and diterpenoid tanshinone iia have the highest binding affinity with the core protein. Before performing molecular dynamics simulations (MDSs), the geometric structure of naringenin and tanshinone iia was optimized using density functional theory (DFT) at the B97-3c level, and RESP2 atomic charge calculations were carried out at the B3LYP-D3(BJ)/def2-TZVP level. Further MDS results demonstrated that in the human body environment, the complex of naringenin and tanshinone iii with core proteins exhibited high stability, flexibility, and low binding free energy. Additionally, naringenin and tanshinone iia showed favorable absorption, distribution, metabolism, excretion, and toxicity (ADMET) characteristics and passed the drug similarity (DS) assessment. Ultrasound cardiograms and cardiac morphometric measurements in animal experiments demonstrate that GXST can improve the PCH induced by isoproterenol (ISO). Protein immunoblotting results indicate that GXST increases the expression of P-eNOS and eNOS by activating the PI3K/AKT signaling pathway and the MAPK signaling pathway, further elucidating the mechanism of action of GXST in treating PCH. This study contributes to the elucidation of the key ingredients and molecular mechanisms of GXST in treating PCH.

Investigating the Mechanistic of Danhong Injection in Brain Damage Caused by Cardiac I/R Injury via Bioinformatics, Computer Simulation, and Experimental Validation.

OBJECTIVE: Cardiac ischemia-reperfusion (I/R) injury has negative effects on the brain and can even lead to the occurrence of ischemic stroke. Clinical evidence shows that Danhong injection (DHI) protects the heart and brain following ischemic events. This study investigated the mechanisms and key active compounds underlying the therapeutic effect of DHI against brain damage induced by cardiac I/R injury. METHODS: The gene expression omnibus database provided GSE66360 and GSE22255 data sets. The R programming language was used to identify the common differentially expressed genes (cDEGs). Gene ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analysis were performed, and protein-protein interaction network was constructed. Active compounds of DHI were collected from the Traditional Chinese Medicine Systems Pharmacology database. Molecular docking and molecular dynamics simulations were performed. The MMPBSA method was used to calculate the binding-free energy. The pkCSM server and DruLiTo software were used for Absorption, Distribution, metabolism, excretion, and toxicity (ADMET) analysis and drug-likeness analysis. Finally, in vitro experiments were conducted to validate the results. RESULTS: A total of 27 cDEGs had been identified. The PPI and enrichment results indicated that TNF-α was considered to be the core target. A total of 80 active compounds were retrieved. The molecular docking results indicated that tanshinone I (TSI), tanshinone IIA (TSIIA), and hydroxyl safflower yellow A (HSYA) were selected as core active compounds. Molecular dynamics verification revealed that the conformations were relatively stable without significant fluctuations. MMPBSA analysis revealed that the binding energies of TSI, TSIIA, and HSYA with TNF-α were -36.01, -21.71, and -14.80 kcal/mol, respectively. LEU57 residue of TNF-α has the highest contribution. TSI and TSIIA passed both the ADMET analysis and drug-likeness screening, whereas HSYA did not. Experimental verification confirmed that DHI and TSIIA reduced the expression of TNF-α, NLRP3, and IL-1ß in the injured H9C2 and rat brain microvascular endothelial cells. CONCLUSION: TNF-α can be considered to be a key target for BD-CI/R. TSIIA in DHI exerts a significant inhibitory effect on the inflammatory damage of BD-CI/R, providing new insights for future drug development.

Exploring the Mechanism of Salvianolic Acid B against Myocardial Ischemia-Reperfusion Injury Based on Network Pharmacology.

This study aimed to explore the mechanisms through which salvianolic acid B (Sal-B) exerts its effects during myocardial ischemia-reperfusion injury (MI/RI), aiming to demonstrate the potential pharmacological characteristics of Sal-B in the management of coronary heart disease. First, Sal-B-related targets and MI/RI-related genes were compiled from public databases. Subsequent functional enrichment analyses using the protein-protein interaction (PPI) network, gene ontology (GO), and the Kyoto Encyclopedia of Genes and Genomes (KEGG) predicted the core targets and approaches by which Sal-B counters MI/RI. Second, a Sal-B-treated MI/RI mouse model and oxygen-glucose deprivation/reoxygenation (OGD/R) H9C2 cell model were selected to verify the main targets of the network pharmacological prediction. An intersectional analysis between Sal-B and MI/RI targets identified 69 common targets, with a PPI network analysis highlighting caspase-3, c-Jun N-terminal kinase (JNK), and p38 mitogen-activated protein kinase (p38) as central targets. GO and KEGG enrichment analyses indicated remarkable enrichment of the apoptosis pathway among these targets, suggesting their utility in experimental studies in vivo. Experimental results demonstrated that Sal-B treatment not only mitigated myocardial infarction size following MI/RI injury in mice but also modulated the expression of key apoptotic regulators, including Bcl-2-Associated X (Bax), caspase-3, JNK, and p38, alongside enhancing the B-cell lymphoma-2 (Bcl-2) expression, thereby inhibiting myocardial tissue apoptosis. This study leveraged an integrative network pharmacology approach to predict Sal-B's potential targets in MI/RI treatment and verified the involvement of key target proteins within the predicted signaling pathways through both in vivo and in vitro experiments, offering a comprehensive insight into Sal-B's pharmacological mechanism in MI/RI management.

An auxiliary strategy of partial least squares regression in pharmacokinetic/pharmacodynamic studies: A case of application of guhong injection in myocardial ischemia/reperfusion rats.

Guhong injection (GHI) has been applied in the therapy of cardio-cerebrovascular disease in clinic, but there is no report about the pharmacokinetic/pharmacodynamic (PK/PD) research on GHI treating myocardial ischemia/reperfusion (MI/R) injury in rats. In this study, eight compounds of GHI in plasma, including N-acetyl-L-glutamine (NAG), chlorogenic acid (CGA), hydroxysafflor yellow A (HSYA), p-coumaric acid ( pCA), rutin, hyperoside, kaempferol-3-O-rutinoside, and kaempferol-3-O-glucoside, were quantified by LC-MS/MS. We discovered that the values of t1/2ß, k12, V2, and CL2 were larger than those of t1/2α, k21, V1, and CL1 for all compounds. The levels of four biomarkers, creatine kinase-MB (CK-MB), cardiac troponin I (cTn I), ischemia-modified albumin (IMA), and alpha-hydroxybutyrate dehydrogenase (α-HBDH) in plasma were determined by ELISA. The elevated level of these biomarkers induced by MI/R was declined to different degrees via administrating GHI and verapamil hydrochloride (positive control). The weighted regression coefficients of NAG, HSYA, CGA, and pCA in PLSR equations generated from The Unscrambler X software (version 11) were mostly minus, suggesting these four ingredients were positively correlated to the diminution of the level of four biomarkers. Emax and ED50, two parameters in PK/PD equations that were obtained by adopting Drug and Statistics software (version 3.2.6), were almost enlarged with the rise of GHI dosage. Obviously, all analytes were dominantly distributed and eliminated in the peripheral compartment with features of rapid distribution and slow elimination. With the enhancement of GHI dosage, the ingredients only filled in the central compartment if the peripheral compartment was replete. Meanwhile, high-dose of GHI generated the optimum intrinsic activity, but the affinity of compounds with receptors was the worst, which may be caused by the saturation of receptors. Among the eight analytes, NAG, HSYA, CGA, and pCA exhibited superior cardioprotection, which probably served as the pharmacodynamic substance basis of GHI in treating MI/R injury.

Lipid regulation of protocatechualdehyde and hydroxysafflor yellow A via AMPK/SREBP2/PCSK9/LDLR signaling pathway in hyperlipidemic zebrafish.

The consumption of a high-cholesterol diet is known to cause hyperlipidemia, which is one of the main risk factors for cardiovascular disease. Protocatechualdehyde (PCA) and hydroxysafflor yellow A (HSYA) are the active components of Salvia miltiorrhiza and safflower, respectively. However, their exact mechanism is still unclear. The aim of this study is to investigate its effects on lipid deposition and liver damage in hyperlipidemic zebrafish and its mechanism of anti-hyperlipidemia. The results showed that the use of PCA and HSYA alone and in combination can improve lipid deposition, slow behavior, abnormal blood flow and liver tissue damage, and the combined use is more effective. Further RT-qPCR results showed that PCA + HSYA can regulate the mRNA levels of PPAR-γ, SREBP2, SREBP1, HMGCR, PCSK9, mTOR, C/EBPα, LDLR, AMPK, HNF-1α and FoxO3a. The PCA + HSYA significantly improves lipid deposition and abnormal liver function in hyperlipidemic zebrafish larvae, which may be related to the AMPK/SREBP2/PCSK9/LDLR signaling pathway.