Tetramethylpyrazine-loaded electroconductive hydrogels promote tissue repair after spinal cord injury by protecting the blood-spinal cord barrier and neurons.

Spinal cord injury (SCI) usually induces profound microvascular dysfunction. It disrupts the integrity of the blood-spinal cord barrier (BSCB), which could trigger a cascade of secondary pathological events that manifest as neuronal apoptosis and axonal demyelination. These events can further lead to irreversible neurological impairments. Thus, reducing the permeability of the BSCB and maintaining its substructural integrity are essential to promote neuronal survival following SCI. Tetramethylpyrazine (TMP) has emerged as a potential protective agent for treating the BSCB after SCI. However, its therapeutic potential is hindered by challenges in the administration route and suboptimal bioavailability, leading to attenuated clinical outcomes. To address this challenge, traditional Chinese medicine, TMP, was used in this study to construct a drug-loaded electroconductive hydrogel for synergistic treatment of SCI. A conductive hydrogel combined with TMP demonstrates good electrical and mechanical properties as well as superior biocompatibility. Furthermore, it also facilitates sustained local release of TMP at the implantation site. Furthermore, the TMP-loaded electroconductive hydrogel could suppress oxidative stress responses, thereby diminishing endothelial cell apoptosis and the breakdown of tight junction proteins. This concerted action repairs BSCB integrity. Concurrently, myelin-associated axons and neurons are protected against death, which meaningfully restore neurological functions post spinal cord injury. Hence, these findings indicate that combining the electroconductive hydrogel with TMP presents a promising avenue for potentiating drug efficacy and synergistic repair following SCI.

Independent regulation of Piezo1 activity by principal and intercalated cells of the collecting duct.

The renal collecting duct is continuously exposed to a wide spectrum of fluid flow rates and osmotic gradients. Expression of a mechanoactivated Piezo1 channel is the most prominent in the collecting duct. However, the status and regulation of Piezo1 in functionally distinct principal and intercalated cells (PCs and ICs) of the collecting duct remain to be determined. We used pharmacological Piezo1 activation to quantify Piezo1-mediated [Ca2+]i influx and single-channel activity separately in PCs and ICs of freshly isolated collecting ducts with fluorescence imaging and electrophysiological tools. We also employed a variety of systemic treatments to examine their consequences on Piezo1 function in PCs and ICs. Piezo1 selective agonists, Yoda-1 or Jedi-2, induced a significantly greater Ca2+ influx in PCs than in ICs. Using patch clamp analysis, we recorded a Yoda-1-activated nonselective channel with 18.6 ± 0.7 pS conductance on both apical and basolateral membranes. Piezo1 activity in PCs but not ICs was stimulated by short-term diuresis (injections of furosemide) and reduced by antidiuresis (water restriction for 24 h). However, prolonged stimulation of flow by high K+ diet decreased Yoda-1-dependent Ca2+ influx without changes in Piezo1 levels. Water supplementation with NH4Cl to induce metabolic acidosis stimulated Piezo1 activity in ICs but not in PCs. Overall, our results demonstrate functional Piezo1 expression in collecting duct PCs (more) and ICs (less) on both apical and basolateral sides. We also show that acute changes in fluid flow regulate Piezo1-mediated [Ca2+]i influx in PCs, whereas channel activity in ICs responds to systemic acid-base stimuli.

Dual Delivery of Tetramethylpyrazine and miR-194-5p Using Soft Mesoporous Organosilica Nanoparticles for Acute Lung Injury Therapy.

Background: The respiratory system is intensely damaged by acute lung injury (ALI). The anti-inflammatory effects of tetramethylpyrazine (TMP) against ALI have been confirmed, but it exhibits a short half-life. miR-194-5p could directly target Rac1, but the internalization rate of miRNA cells was low. Purpose: To explore the potential of the soft mesoporous organic silica nanoplatform (NPs) as carriers for delivery of TMP and miR-194-5p through the tail vein. Methods: NPs@TMP and NPs@PEI@miR-194-5p were added to the HUVEC cell-lines, in vitro, to observe the cell uptake and cytotoxic effects. In vivo experiments were conducted by injecting fluorescently labeled NPs through the tail vein and tracking distribution. Therapeutic and toxic side-effects were analyzed systemically. Results: In vitro study exhibited that NPs have no toxic effect on HUVECs within the experimental parameters and have excellent cellular uptake. The IVIS Spectrum Imaging System shows that NPs accumulate mainly in the lungs. NPs@TMP treatment can improved oxidative stress and inflammation levels in ALI mice and inhibited the TLR4/NLRP3/caspase 1 pathway. NPs@PEI@miR-194-5p can inhibit the Rac1/ZO-1/occludin pathway and improved endothelial cell permeability in ALI mice. The co-treatment of NPs@TMP and NPs@PEI@miR-194-5p can significantly improved the survival rates of the mice, reduced pulmonary capillary permeability and improved pathological injury in ALI mice. Innovation: This study combined traditional Chinese medicine, bioinformatics, cellular molecular biology and nanobiomedicine to study the pathogenesis and treatment of ALI. The rate of cellular internalization was improved by changing the shape and hardness of nanoparticles. NPs@TMP and NPs@PEI@miR-194-5p combined application can significantly improve the survival condition and pathological injury of mice. Conclusion: NPs loaded with TMP and miR-194-5p showed a greater therapeutic effect in ALI mice.

Dietary Flavonoids with Catechol Moiety Inhibit Anticancer Action of Bortezomib: What about the other Boronic Acid-based Drugs?

Approval of the first boronic acid group-containing drug, bortezomib, in 2003 for the treatment of multiple myeloma sparked an increased interest of medicinal chemists in boronic acidbased therapeutics. As a result, another boronic acid moiety-harboring medication, ixazomib, was approved in 2015 as a second-generation proteasome inhibitor for multiple myeloma; and dutogliptin is under clinical investigation in combination therapy against myocardial infarction. Moreover, a large number of novel agents with boronic acid elements in their structure are currently in intensive preclinical studies, allowing us to suppose that at least some of them will enter clinical trials in the near future. On the other hand, only some years after bortezomib approval, direct interactions between its boronic acid group and catechol moiety of green tea catechins as well as some other common dietary flavonoids like quercetin and myricetin were discovered, leading to the formation of stable cyclic boronate esters and abolishing the anticancer activities. Although highly relevant, to date, no reports on possible co-effects of catechol group-containing flavonoids with new-generation boronic acidbased drugs can be found. However, this issue cannot be ignored, especially considering the abundance of catechol moiety-harboring flavonoids in both plant-derived food items as well as over-thecounter dietary supplements and herbal products. Therefore, in parallel with the intensified development of boronic acid-based drugs, their possible interactions with catechol groups of plant-derived flavonoids must also be clarified to provide dietary recommendations to patients for maximizing therapeutic benefits. If concurrently consumed flavonoids can indeed antagonize drug efficacy, it may pose a real risk to clinical outcomes.

Tetramethylpyrazine: A review on its mechanisms and functions.

Ligusticum chuanxiong Hort (known as Chuanxiong in China, CX) is one of the most widely used and long-standing medicinal herbs in China. Tetramethylpyrazine (TMP) is an alkaloid and one of the active components of CX. Over the past few decades, TMP has been proven to possess several pharmacological properties. It has been used to treat a variety of diseases with excellent therapeutic effects. Here, the pharmacological characteristics and molecular mechanism of TMP in recent years are reviewed, with an emphasis on the signal-regulation mechanism of TMP. This review shows that TMP has many physiological functions, including anti-oxidant, anti-inflammatory, and anti-apoptosis properties; autophagy regulation; vasodilation; angiogenesis regulation; mitochondrial damage suppression; endothelial protection; reduction of proliferation and migration of vascular smooth muscle cells; and neuroprotection. At present, TMP is used in treating cardiovascular, nervous, and digestive system conditions, cancer, and other conditions and has achieved good curative effects. The therapeutic mechanism of TMP involves multiple targets, multiple pathways, and bidirectional regulation. TMP is, thus, a promising drug with great research potential.

Tetramethylpyrazine induces the release of BDNF from BM-MSCs through activation of the PI3K/AKT/CREB pathway.

Compelling evidences suggest that transplantation of bone marrow-derived mesenchymal stem cells (BM-MSCs) can be therapeutically effective for central nervous system (CNS) injuries and neurodegenerative diseases. The therapeutic effect of BM-MSCs mainly attributes to their differentiation into neuron-like cells which replace injured and degenerative neurons. Importantly, the neurotrophic factors released from BM-MSCs can also rescue injured and degenerative neurons, which plays a biologically pivotal role in enhancing neuroregeneration and neurological functional recovery. Tetramethylpyrazine (TMP), the main bioactive ingredient extracted from the traditional Chinese medicinal herb Chuanxiong, has been reported to promote the neuronal differentiation of BM-MSCs. This study aimed to investigate whether TMP regulates the release of neurotrophic factors from BM-MSCs. We examined the effect of TMP on brain-derived neurotrophic factor (BDNF) released from BM-MSCs and elucidated the underlying molecular mechanism. Our results demonstrated that TMP at concentrations of lower than 200 µM increased the release of BDNF in a dose-dependent manner. Furthermore, the effect of TMP on increasing the release of BDNF from BM-MSCs was blocked by inhibiting the phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K)/protein kinase B (AKT)/cAMP-response element binding protein (CREB) pathway. Therefore, we concluded that TMP could induce the release of BDNF from BM-MSCs through activation of the PI3K/AKT/CREB pathway, leading to the formation of neuroprotective and proneurogenic microenvironment. These findings suggest that TMP possesses novel therapeutic potential to promote neuroprotection and neurogenesis through improving the neurotrophic ability of BM-MSCs, which provides a promising nutritional prevention and treatment strategy for CNS injuries and neurodegenerative diseases via the transplantation of TMP-treated BM-MSCs.

A systems-level study reveals host-targeted repurposable drugs against SARS-CoV-2 infection.

Understanding the mechanism of SARS-CoV-2 infection and identifying potential therapeutics are global imperatives. Using a quantitative systems pharmacology approach, we identified a set of repurposable and investigational drugs as potential therapeutics against COVID-19. These were deduced from the gene expression signature of SARS-CoV-2-infected A549 cells screened against Connectivity Map and prioritized by network proximity analysis with respect to disease modules in the viral-host interactome. We also identified immuno-modulating compounds aiming at suppressing hyperinflammatory responses in severe COVID-19 patients, based on the transcriptome of ACE2-overexpressing A549 cells. Experiments with Vero-E6 cells infected by SARS-CoV-2, as well as independent syncytia formation assays for probing ACE2/SARS-CoV-2 spike protein-mediated cell fusion using HEK293T and Calu-3 cells, showed that several predicted compounds had inhibitory activities. Among them, salmeterol, rottlerin, and mTOR inhibitors exhibited antiviral activities in Vero-E6 cells; imipramine, linsitinib, hexylresorcinol, ezetimibe, and brompheniramine impaired viral entry. These novel findings provide new paths for broadening the repertoire of compounds pursued as therapeutics against COVID-19.

The antibacterial activity of Libanstin from Ilex paraguariensis (Yerba Mate).

Infectious diseases are reported to be one of the major causes of death in the world. The World Health Organization (WHO) warns of an increase in the deaths number because of antibacterial resistance. Lately, a trend towards searching for new active antibacterial compounds in plants has been observed. Ilex paraguariensis, known as Yerba Mate, is a plant known to be rich in numerous bioactive compounds that have an important role in human health. In this study, Yerba Mate was extracted with acetone: water (1:1) and further fractionated with hexane, chloroform and ethyl acetate. The obtained fractions were tested for antibacterial activity against Staphylococcus aureus and Salmonella species. The minimum inhibitory concentration (MIC) values on S. aureus ranged from 1.56 to 3.12 mg/mL for both the chloroform and ethyl acetate fractions. Whereas for the water fraction, the MIC values ranged from 0.78 to 3.12 mg/mL on S. aureus and ranged from 1.56 mg/mL to 3.12 mg/mL on Salmonella species. The aqueous fraction was further treated with different enzymes to mimic in vivo digestion and the fractions obtained were then tested for antibacterial activity. Furthermore, the Yerba Mate aqueous fraction was run on High Performance Liquid Chromatography (HPLC) and collected fractions were tested for antibacterial activity, to identify the active metabolite. Fraction 3 was tested on different strains of S. aureus and the MIC values ranged from 0.19 to 1.56 µg/mL. A novel pyrazinone, Libanstin, from Ilex paraguariensis was identified using NMR spectroscopy.

Ligustrazine Attenuates Hyperhomocysteinemia-induced Alzheimer-like Pathologies in Rats.

Ligustrazine, an alkaloid extracted from the traditional Chinese herbal medicine Ligusticum Chuanxiong Hort, has been clinically applied to treat the cerebrovascular diseases. Hyperhomocysteinemia (Hhcy) is an independent risk factor for Alzheimer's disease (AD). Memory deficits can be caused by Hhcy via pathologies of AD-like tau and amyloid-ß (Aß) in the hippocampus. Here, we investigated whether homocysteine (Hcy) can induce AD-like pathologies and the effects of ligustrazine on these pathologies. The Hcy rat model was constructed by 14-day Hcy injection via vena caudalis, and rats were treated with daily intragastric administration of ligustrazine at the same time. We found that the pathologies of tau and Aß were induced by Hcy in the hippocampus, while the Hcy-induced tau hyperphosphorylation and Aß accumulation could be markedly attenuated by simultaneous ligustrazine treatment. Our data demonstrate that ligustrazine may be used as a promising neuroprotective agent to treat the Hcy-induced AD-like pathologies.

Vinegar/Tetramethylpyrazine Induces Nutritional Preconditioning Protecting the Myocardium Mediated by VDAC1.

Vinegar is good for health. Tetramethylpyrazine (TMP) is the main component of its flavor, quality, and function. We hypothesized that vinegar/TMP pretreatment could induce myocardial protection of "nutritional preconditioning (NPC)" by low-dose, long-term supplementation and alleviate the myocardial injury caused by anoxia/reoxygenation (A/R). To test this hypothesis, TMP content in vinegar was detected by HPLC; A/R injury model was prepared by an isolated mouse heart and rat cardiomyocyte to evaluate the myocardial protection and mechanism of vinegar/TMP pretreatment by many enzymatic or functional, or cellular and molecular biological indexes. Our results showed that vinegar contained TMP, and its content was in direct proportion to storage time. Vinegar/TMP pretreatment could improve hemodynamic parameters, decrease lactate dehydrogenase (LDH) and creatine phosphokinase activities, and reduce infarct size and apoptosis in the isolated hearts of mice with A/R injury. Similarly, vinegar/TMP pretreatment could increase cell viability, decrease LDH activity, and decrease apoptosis against A/R injury of cardiomyocytes. Vinegar/TMP pretreatment could also maintain the mitochondrial function of A/R-injured cardiomyocytes, including improving oxygen consumption rate and extracellular acidification rate, reducing reactive oxygen species generation, mitochondrial membrane potential loss, mitochondrial permeability transition pore openness, and cytochrome c releasing. However, the protective effects of vinegar/TMP pretreatment were accompanied by the downregulation of VDAC1 expression in the myocardium and reversed by pAD/VDAC1, an adenovirus that upregulates VDAC1 expression. In conclusion, this study is the first to demonstrate that vinegar/TMP pretreatment could induce myocardial protection of NPC due to downregulating VDAC1 expression, inhibiting oxidative stress, and preventing mitochondrial dysfunction; that is, VDAC1 is their target, and the mitochondria are their target organelles. TMP is one of the most important myocardial protective substances in vinegar.

Antiviral drug screen identifies DNA-damage response inhibitor as potent blocker of SARS-CoV-2 replication.

SARS-CoV-2 has currently precipitated the COVID-19 global health crisis. We developed a medium-throughput drug-screening system and identified a small-molecule library of 34 of 430 protein kinase inhibitors that were capable of inhibiting the SARS-CoV-2 cytopathic effect in human epithelial cells. These drug inhibitors are in various stages of clinical trials. We detected key proteins involved in cellular signaling pathways mTOR-PI3K-AKT, ABL-BCR/MAPK, and DNA-damage response that are critical for SARS-CoV-2 infection. A drug-protein interaction-based secondary screen confirmed compounds, such as the ATR kinase inhibitor berzosertib and torin2 with anti-SARS-CoV-2 activity. Berzosertib exhibited potent antiviral activity against SARS-CoV-2 in multiple cell types and blocked replication at the post-entry step. Berzosertib inhibited replication of SARS-CoV-1 and the Middle East respiratory syndrome coronavirus (MERS-CoV) as well. Our study highlights key promising kinase inhibitors to constrain coronavirus replication as a host-directed therapy in the treatment of COVID-19 and beyond as well as provides an important mechanism of host-pathogen interactions.

Ligustrazine Suppresses Platelet-Derived Growth Factor-BB-Induced Pulmonary Artery Smooth Muscle Cell Proliferation and Inflammation by Regulating the PI3K/AKT Signaling Pathway.

Pulmonary arterial hypertension (PAH) is a serious pulmonary vascular disease. Excessive proliferation of pulmonary artery smooth muscle cells (PASMCs) plays an important role in the course of this disease. Ligustrazine is an alkaloid monomer extracted from the rhizome of the herb Ligusticum chuanxiong. It is often used to treat cardiovascular diseases, but its effect on PAH has rarely been reported. This study aims to explore the protective effect and mechanism of ligustrazine on PAH. In the in vivo experiment, monocrotaline (MCT) was used to induce PAH in rats, and then ligustrazine (40, 80, 160 mg/kg/day) or sildenafil (25 mg/kg/day) was administered. Four weeks later, hemodynamic changes, right ventricular hypertrophy index, lung morphological characteristics, inflammatory factors, phosphoinositide 3-kinase (PI3K), and AKT expression were evaluated. In addition, primary rat PASMCs were extracted by the tissue adhesion method, a proliferation model was established with platelet-derived growth factor-BB (PDGF-BB), and the cells were treated with ligustrazine to investigate its effects on cell proliferation, inflammation, and cell cycle distribution. The results indicate that ligustrazine can markedly alleviate right ventricular systolic pressure, right ventricular hypertrophy, pulmonary vascular remodeling, and inflammation caused by MCT, and that it decreased PI3K and AKT phosphorylation expression. Moreover, ligustrazine can inhibit the proliferation and inflammation of PASMCs and arrest the progression of G0/G1 to S phase through the PI3K/AKT signaling pathway. Therefore, we conclude that ligustrazine may inhibit the proliferation and inflammation of PASMCs by regulating the activation of the PI3K/AKT signaling pathway, thereby attenuating MCT-induced PAH in rats. Collectively, these findings suggest that ligustrazine may be a promising therapeutic for PAH.

Salviae miltiorrhizae and ligustrazine hydrochloride injection combined with mecobalamin for treating diabetic peripheral neuropathy: A protocol for systematic review and meta-analysis.

OBJECTIVE: Currently, it is unclear whether the salviae miltiorrhizae (Danshen Salvia) and ligustrazine hydrochloride (Chuanxiong Chuanxiong) (SMLH) injection combined with mecobalamin can improve diabetic peripheral neuropathy (DPN). We conducted a systematic analysis to evaluate the clinical effects of SMLH injection combined with mecobalamin for treating DPN. METHODS: Seven databases, including PubMed, Embase, Cochrane Library, China National Knowledge Infrastructure (CNKI), Wan Fang Database (Wang Fang), Chinese Biomedical Literature Database (CBM), and VIP Database for Chinese Technical Periodicals (VIP) were searched for systematic literature retrieval. Each database was searched up to 2020 to identify randomized controlled trials on DPN treated with SMLH injection combined with mecobalamin. We used the RevMan 5.3 and Stata 14.0 software to assess the risk of bias in the included trials. RESULTS: A total of 15 publications, including 1349 samples, were reviewed. The total effective rate of SMLH injection combined with mecobalamin was 31% higher than that of mecobalamin alone (95% confidence interval [CI] = 1.23-1.38; P < .00001). The experimental group showed a significant increase in the motor conduction velocity (MCV) of the peroneal nerve (weighted mean difference [WMD] = 4.81, 95% CI 3.53-6.09; P < .00001). In addition, SMLH injection combined with mecobalamin showed a statistical significant effect on the sensory conduction velocity (SCV) of the peroneal nerve (WMD = 5.03, 95% CI = 4.16-5.90; P < .00001), and MCV of the median nerve (WMD = 5.38, 95% CI = 4.05-6.72; P < .00001). The WMD for the change in SCV in the median nerve was 4.89 m/s (95% CI = 3.88-5.89; P < .00001). The P-values of the Egger and Begg tests were 0.967 and 0.961, respectively, indicating no publication bias. Subgroup and sensitivity analyses indicated that the results for MCV and SCV of the peroneal nerve and the median nerve were stable. CONCLUSION: SMLH injection combined with mecobalamin can improve DPN, compared with mecobalamin alone.

The Tetramethylpyrazine Analogue T-006 Alleviates Cognitive Deficits by Inhibition of Tau Expression and Phosphorylation in Transgenic Mice Modeling Alzheimer's Disease.

T-006, a small-molecule compound derived from tetramethylpyrazine (TMP), has potential for the treatment of neurological diseases. In order to investigate the effect of T-006 prophylactic treatment on an Alzheimer's disease (AD) model and identify the target of T-006, we intragastrically administered T-006 (3 mg/kg) to Alzheimer's disease (AD) transgenic mice (APP/PS1-2xTg and APP/PS1/Tau-3xTg) for 6 and 8 months, respectively. T-006 improved cognitive ability after long-term administration in two AD mouse models and targeted mitochondrial-related protein alpha-F1-ATP synthase (ATP5A). T-006 significantly reduced the expression of phosphorylated-tau, total tau, and APP while increasing the expression of synapse-associated proteins in 3xTg mice. In addition, T-006 modulated the JNK and mTOR-ULK1 pathways to reduce both p-tau and total tau levels. Our data suggested that T-006 mitigated cognitive decline primarily by reducing the p-tau and total tau levels in 3xTg mice, supporting further investigation into its development as a candidate drug for AD treatment.

2,3,5,6-Tetramethylpyrazine protects retinal photoreceptors against endoplasmic reticulum stress by modulating ATF4-mediated inhibition of PRP aggregation.

Endoplasmic reticulum (ER) stress is a common threat to photoreceptors during the pathogenesis of chronic retinopathies and often results in irreversible visual impairment. 2,3,5,6-Tetramethylpyrazine (TMP), which possesses many beneficial pharmacological activities, is a potential drug that could be used to protect photoreceptors. In the present study, we found that the cellular growth rate of 661 W cells cultured under low glucose conditions was lower than that of control cells, while the G2/M phase of the cell cycle was longer. We further found that the mitochondrial membrane potential (ΔΨm) was lower and that ER stress factor expression was increased in 661 W cells cultured under low glucose conditions. TMP reversed these trends. Visual function and cell counts in the outer nuclear layer (ONL) were low and the TUNEL-positive rate in the ONL was high in a C3H mouse model of spontaneous retinal degeneration. Similarly, visual function was decreased, and the TUNEL-positive rate in the ONL was increased in fasted C57/BL6j mice compared with control mice. On the other hand, ER stress factor expression was found to be increased in the retinas of both mouse models, as shown by reverse transcription real-time PCR (RT-qPCR) and western blotting. TMP reversed the physiological and molecular biological variations observed in both mouse models, and ATF4 expression was enhanced again. Further investigation by using western blotting illustrated that the proportion of insoluble prion protein (PRP) versus soluble PRP was reduced both in vitro and in vivo. Taken together, these results suggest that TMP increased the functions of photoreceptors by alleviating ER stress in vitro and in vivo, and the intrinsic mechanism was the ATF4-mediated inhibition of PRP aggregation. TMP may potentially be used clinically as a therapeutic agent to attenuate the functional loss of photoreceptors during the pathogenesis of chronic retinopathies. KEY MESSAGES: • Already known: TMP is a beneficial drug mainly used in clinic to enhance organ functions, and the intrinsic mechanism is still worthy of exploring. • New in the study: We discovered that TMP ameliorated retinal photoreceptors function via ER stress alleviation, which was promoted by ATF4-mediated inhibition of PRP aggregation. • Application prospect: In prospective clinical practices, TMP may potentially be used in the clinic as a therapeutic agent to attenuate the photoreceptors functional reduction in chronic retinopathies.

Combination of Danshen and ligustrazine has dual anti-inflammatory effect on macrophages and endothelial cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Salvia Miltiorrhiza Radix et Rhizoma (Danshen) and Chuanxiong Rhizoma (Chuanxiong) are both traditional Chinese medicines with vascular protective effects, and their combination is widely used in China to treat occlusive or ischemic diseases of the cerebrovascular or cardiovascular system. Although it is widely accepted that these diseases have high relevance to inflammation, little is known about the anti-inflammatory effect of Danshen, Chuanxiong, and their combination. AIM OF STUDY: We aimed to investigate the complex mode of action of Danshen, Chuanxiong, and their combination and the molecular mechanisms underlying their anti-inflammatory activity. Specifically, toll-like receptor (TLR1/2, 3, and 4)-triggered macrophages and endothelial cells, the two major cell players in atherosclerosis as well as in related cardiovascular and cerebrovascular injuries, were emphasized. METHODS: TLR1/2-, TLR3-, and TLR4-induced bone marrow macrophages (BMMs) and human umbilical vein endothelial cells (HUVECs) were treated with Danshen extract (S. miltiorrhiza extract, SME), ligustrazine (2, 3, 5, 6-tetramethylpyrazine, TMP), and their combination (S. miltiorrhiza and TMP injection, SLI), respectively. The proinflammatory cytokines interleukin 6 (IL-6), IL-12, and tumor necrosis factor alpha (TNF-α) were detected as the preliminary indicators of inflammation. In addition, RNA sequencing (RNA-seq)-based transcriptional profiling analyses were conducted for TLR2-activated BMMs to determine the molecular mode of action of SLI as well as the contribution of SME to SLI activity. RESULTS: SLI mitigated inflammation in both BMMs and HUVECs. Refer to the combination, SME had pronounced anti-inflammatory effect on BMMs but had only a slight effect on HUVECs. In contrast, TMP had considerable anti-inflammatory effect on HUVECs but not on BMMs. Bioinformatic analysis identified a broad spectrum of regulatory genes, in addition to IL-6 gene, and C-X-C motif chemokine ligand 10 (CXCL10) appeared to be another key molecule involved in the mechanism underlying SLI and SME effects. At the molecular level, SME was a major contributor of the anti-inflammatory activity of SLI. CONCLUSIONS: In TLR-activated inflammation, SLI exhibits a "multiple ingredient-multiple target" effect, with SME primarily affecting macrophages and TMP affecting HUVECs. Our study provides evidence for the clinical application of SLI in treating complex diseases involving inflammation-induced injury of both macrophages and epithelial cells. Further bioinformatics studies are required to reveal the entire molecular network involved in TMP, SME, and SLI activity.

Protective Effects of Astragalus Membranaceus and Ligustrazine on Rat Brain Microvascular Endothelial Cell Injury after Oxygen-Glucose Deprivation/ Reoxygenation by Suppressing the PKCδ/MARCKS Pathway.

AIM AND OBJECTIVE: Cell death is a main pathological change in brain ischemia. Astragalus membranaceus (Ast) and ligustrazine (Lig), as traditional Chinese herbs, have a protective effect against ischemia-reperfusion injury. We aim to find whether the underlying protective mechanism of Astragalus membranaceus and ligustrazine against Oxygen-glucose deprivation/reoxygenation (OGD/R) -induced injury in RBMECs is related to PKCδ/MARCKS pathway. MATERIALS AND METHODS: OGD/R preconditioning was instituted in rat brain microvascular endothelial cells (RBMECs). The survival and apoptosis of RBMECs were detected by a Cell Counting Kit-8 and TUNEL staining; PKCδ/MARCKS and MMP9 expression were examined by immunofluorescence, western blot and quantitative real-time PCR. RESULTS: OGD/R stimulation significantly increased RBMEC apoptosis, whereas Ast+Lig, Rottlerin or Ast+Lig+Rottlerin treatment evidently reduced cellular apoptosis and increased cell viability (P <0.05). Furthermore, Ast+Lig, Rottlerin or Ast+Lig+Rottlerin treatment significantly reduced mRNA expression levels of PKCδ/MARCKS and MMP9 (P <0.05), compared to OGD/R control group. Moreover, Ast+Lig, Rottlerin or Ast+Lig+Rottlerin treatment evidently reduced protein expression levels of PKCδ, MMP9, and MARCKS (P <0.05), compared to OGD/R control group, detected by western blotting or immunofluorescence. CONCLUSION: The administration of Astragalus membranaceus and ligustrazine protected RBMECs against OGD/R-induced apoptosis. PKCδ/MARCKS and MMP9 expression were significantly increased after OGD/R stimulation, while Astragalus membranaceus and ligustrazine treatment evidently suppressed. Collectively, Astragalus membranaceus and ligustrazine play protective effects against OGD/R-induced injury in RBMECs through regulating PKCδ/MARCKS pathway to inhibit MMP9 activation.

Effects of Floium ginkgo extract and tertram ethypyrazine sodium chloride injection on expression of inflammatory cytokines and cerebral infarction.

This experiment was carried out to observe and analyze the effect of floium ginkgo extract and tertram ethypyrazine sodium chloride injection in patients with cerebral infarction. A total of 200 patients diagnosed with cerebral infarction were enrolled in the study. They were randomly divided into a research group and control group, each containing 100 patients. The control group was given routine treatment measures while the research group was given floium ginkgo extract and tertram ethypyrazine sodium chloride injection on the basis of routine treatment. The therapeutic effects of the two groups were observed and compared. After implementing different treatment schemes, the levels of MMP-9, SOD, CBV and CBF in the research group were significantly higher than those in the control group, p<0.05. The research group was lower in hs-CRP, MDA, MTT, TTP and TNF-α as compared with the control group, p<0.05. In terms of the quality of life of the two groups after six months of treatment, the scores of various indicators in the research group were all significantly superior, p<0.05. Conclusion: The treatment of cerebral infarction patients with floium ginkgo extract and tertram ethypyrazine sodium chloride injection can significantly improve the therapeutic effect, which is a relatively ideal treatment.

FLT3 inhibitors in the treatment of acute myeloid leukemia: current status and future perspectives.

Mutations in the FMS-like tyrosine kinase 3 (FLT3) gene arise in 25-30% of all acute myeloid leukemia (AML) patients. These mutations lead to constitutive activation of the protein product and are divided in two broad types: internal tandem duplication (ITD) of the juxtamembrane domain (25% of cases) and point mutations in the tyrosine kinase domain (TKD). Patients with FLT3 ITD mutations have a high relapse risk and inferior cure rates, whereas the role of FLT3 TKD mutations still remains to be clarified. Additionally, growing research indicates that FLT3 status evolves through a disease continuum (clonal evolution), where AML cases can acquire FLT3 mutations at relapse - not present in the moment of diagnosis. Several FLT3 inhibitors have been tested in patients with FLT3-mutated AML. These drugs exhibit different kinase inhibitory profiles, pharmacokinetics and adverse events. First-generation multi-kinase inhibitors (sorafenib, midostaurin, lestaurtinib) are characterized by a broad-spectrum of drug targets, whereas second-generation inhibitors (quizartinib, crenolanib, gilteritinib) show more potent and specific FLT3 inhibition, and are thereby accompanied by less toxic effects. Notwithstanding, all FLT3 inhibitors face primary and acquired mechanisms of resistance, and therefore the combinations with other drugs (standard chemotherapy, hypomethylating agents, checkpoint inhibitors) and its application in different clinical settings (upfront therapy, maintenance, relapsed or refractory disease) are under study in a myriad of clinical trials. This review focuses on the role of FLT3 mutations in AML, pharmacological features of FLT3 inhibitors, known mechanisms of drug resistance and accumulated evidence for the use of FLT3 inhibitors in different clinical settings.

Design, synthesis, and biological evaluation of ligustrazine/resveratrol hybrids as potential anti-ischemic stroke agents.

To search for potent anti-ischemic stroke agents, a series of tetramethylpyrazine (TMP)/resveratrol (RES) hybrids 6a-t were designed and synthesized. These hybrids inhibited adenosine diphosphate (ADP)- or arachidonic acid (AA)-induced platelet aggregation, among them, 6d, 6g-i, 6o and 6q were more active than TMP. The most active compound 6h exhibited more potent anti-platelet aggregation activity than TMP, RES, as well as positive control ticlopidine (Ticlid) and aspirin (ASP). Furthermore, 6h exerted strong antioxidative activity in a dose-dependent manner in rat pheochromocytoma PC12 cells which were treated with hydrogen peroxide (H2O2) or hydroxyl radical (·OH). Importantly, 6h significantly protected primary neuronal cells suffered from oxygen-glucose deprivation/reoxygenation (OGD/R) injury, comparable to an anti-ischemic drug edaravone (Eda). Together, our findings suggest that 6h may be a promising candidate warranting further investigation for the intervention of ischemic stroke.