Chuanxiong rhizoma (CX) has been utilized for centuries as a traditional herb to treat blood stasis syndromes. However, the pharmacological mechanisms are still not completely revealed. This research was aimed at exploring the molecular mechanisms of CX treatment for thrombosis. Network pharmacology was used to predict the potential anti-thrombosis mechanism after correlating the targets of active components with targets of thrombosis. Furthermore, we verified the mechanism of using CX to treat thrombosis via molecular docking and in vitro experiments. Network pharmacology results showed that a total of 18 active ingredients and 65 targets of CX treatment for thrombosis were collected, including 8 core compounds and 6 core targets. We revealed for the first time that tissue factor (TF) had a close relationship with most core targets of CX in the treatment of thrombosis. TF is a primary coagulation factor in physiological hemostasis and pathological thrombosis. Furthermore, core components of CX have strong affinity for core targets and TF according to molecular docking analysis. The in vitro experiments indicated that Ligustilide (LIG), the representative component of CX, could inhibit TF procoagulant activity, TF mRNA and protein over-expression in a dose-dependent manner in EA.hy926 cells through the PI3K/Akt/NF-κB signaling pathway. This work demonstrated that hemostasis or blood coagulation was one of the important biological processes in the treatment of thrombosis with CX, and TF also might be a central target of CX when used for treating thrombosis. The inhibition of TF might be a novel mechanism of CX in the treatment of thrombosis.

Cardiovascular metal stents have shown potential in the treatment of coronary artery disease using percutaneous coronary intervention. However, thrombosis, endothelialization, and new atherosclerosis after stent implantation remain unsolved problems. Herein, a multifunctional coating material based on phase-transited lysozyme was developed to promote stent endothelialization and simultaneously reduce thrombus events by embedding moieties of heparin and co-immobilized copper ions for in-situ catalyzing nitric oxide (NO) generation. The lysozyme-based biomimetic coating is compatible with blood and enables facile loading and sustainable release of copper ions to produce NO with donors via catalytic reaction. The novel coating strategy displayed several bio-effects of anti-thrombosis; it synergistically promoted endothelial cell growth and inhibited smooth muscle cell growth. Thus, this systemic in vitro study will provide a foundation for developing multifunctional cardiovascular stents in clinical settings.

Stenting is the primary treatment for vascular obstruction-related cardiovascular diseases, but it inevitably causes endothelial injury which may lead to severe thrombosis and restenosis. Maintaining nitric oxide (NO, a vasoactive mediator) production and grafting endothelial glycocalyx such as heparin (Hep) onto the surface of cardiovascular stents could effectively reconstruct the damaged endothelium. However, insufficient endogenous NO donors may impede NO catalytic generation and fail to sustain cardiovascular homeostasis. Here, a dopamine-copper (DA-Cu) network-based coating armed with NO precursor L-arginine (Arg) and Hep (DA-Cu-Arg-Hep) is prepared using an organic solvent-free dipping technique to form a nanometer-thin coating onto the cardiovascular stents. The DA-Cu network adheres tightly to the surface of stents and confers excellent NO catalytic activity in the presence of endogenous NO donors. The immobilized Arg functions as a NO fuel to generate NO via endothelial nitric oxide synthase (eNOS), while Hep works as eNOS booster to increase the level of eNOS to decompose Arg into NO, ensuring a sufficient supply of NO even when endogenous donors are insufficient. The synergistic interaction between Cu and Arg is analogous to a gas station to fuel NO production to compensate for the insufficient endogenous NO donor in vivo. Consequently, it promotes the reconstruction of natural endothelium, inhibits smooth muscle cell (SMC) migration, and suppresses cascading platelet adhesion, preventing stent thrombosis and restenosis. We anticipate that our DA-Cu-Arg-Hep coating will improve the quality of life of cardiovascular patients through improved surgical follow-up, increased safety, and decreased medication, as well as revitalize the stenting industry through durable designs.

Background: The florets of Carthamus tinctorius L. (Safflower) is an important traditional medicine for promoting blood circulation and removing blood stasis. However, its bioactive compounds and mechanism of action need further clarification. Objective: This study aims to investigate the effect and possible mechanism of 6-hydroxykaempferol 3,6-di-O-glucoside-7-O-glucuronide (HGG) from Safflower on endothelial injury in vitro, and to verify its anti-thrombotic activity in vivo. Methods: The endothelial injury on human umbilical vein endothelial cells (HUVECs) was induced by oxygen-glucose deprivation followed by reoxygenation (OGD/R). The effect of HGG on the proliferation of HUVECs under OGD/R was evaluated by MTT, LDH release, Hoechst-33342 staining, and Annexin V-FITC apoptosis assay. RNA-seq, RT-qPCR, Enzyme-linked immunosorbent assay and Western blot experiments were performed to uncover the molecular mechanism. The anti-thrombotic effect of HGG in vivo was evaluated using phenylhydrazine (PHZ)-induced zebrafish thrombosis model. Results: HGG significantly protected OGD/R induced endothelial injury, and decreased HUVECs apoptosis by regulating expressions of hypoxia inducible factor-1 alpha (HIF-1α) and nuclear factor kappa B (NF-κB) at both transcriptome and protein levels. Moreover, HGG reversed the mRNA expression of pro-inflammatory cytokines including IL-1β, IL-6, and TNF-α, and reduced the release of IL-6 after OGD/R. In addition, HGG exhibited protective effects against PHZ-induced zebrafish thrombosis and improved blood circulation. Conclusion: HGG regulates the expression of HIF-1α and NF-κB, protects OGD/R induced endothelial dysfunction in vitro and has anti-thrombotic activity in PHZ-induced thrombosis in vivo.

As a result of thrombosis or intimal hyperplasia, synthetic artificial vascular grafts had a low success rate when they were used to replace small-diameter arteries (inner diameter < 6 mm). C-type natriuretic peptides (CNP) have anti-thrombotic effects, and can promote endothelial cell (EC) proliferation and inhibit vascular smooth muscle cell (SMC) over-growth. In this study, poly(ε-caprolactone) (PCL) vascular grafts loaded with CNP (PCL-CNP) were constructed by electrospinning. The PCL-CNP grafts were able to continuously release CNP at least 25 days in vitro. The results of scanning electron microscopy (SEM) and mechanical testing showed that the loading of CNP did not change the microstructure and mechanical properties of the PCL grafts. In vitro blood compatibility analysis displayed that PCL-CNP grafts could inhibit thrombin activity and reduce platelet adhesion and activation. In vitro cell experiments demonstrated that PCL-CNP grafts activated ERK1/2 and Akt signaling in human umbilical vein endothelial cells (HUVECs), as well as increased cyclin D1 expression, enhanced proliferation and migration, and increased vascular endothelial growth factor (VEGF) secretion and nitric oxide (NO) production. The rabbit arteriovenous (AV)-shunt ex vitro indicated that CNP loading significantly improved the antithrombogenicity of PCL grafts. The assessment of vascular grafts in rat abdominal aorta implantation model displayed that PCL-CNP grafts promoted the regeneration of ECs and contractile SMCs, modulated macrophage polarization toward M2 phenotype, and enhanced extracellular matrix remodeling. These findings confirmed for the first time that loading CNP is an effective approach to improve the hemocompatibility and vascular regeneration of synthetic vascular grafts. STATEMENT OF SIGNIFICANCE: Small-diameter (< 6 mm) vascular grafts (SDVGs) have not been made clinically available due to their prevalence of thrombosis, limited endothelial regeneration and intimal hyperplasia. The incorporation of bioactive molecules into SDVGs serves as an effective solution to improve hemocompatibility and endothelialization. In this study, for the first time, we loaded C-type natriuretic peptides (CNP) into PCL grafts by electrospunning and confirmed the effectiveness of loading CNP on improving the hemocompatibility and vascular regeneration of artificial vascular grafts. Regenerative advantages included enhancement of endothelialization, modulation of macrophage polarization toward M2 phenotypes, and improved contractile smooth muscle cell regeneration. Our investigation brings attention to CNP as a valuable bioactive molecule for modifying cardiovascular biomaterial.

UNASSIGNED: Prolonged uncontrolled hyperglycaemia has shown to cause oxidative stress, inflammation, thrombosis and upregulation of angiogenesis in diabetics, which all contributes to diabetic retinopathy development and progression. Vitamin E is found to have anti-inflammatory, anti-oxidative, anti-thrombogenic and anti-angiogenesis which could play an important role in early treatment of diabetic retinopathy. This study aims to investigate the effect of Tocotrienol-rich vitamin E (Tocovid) on the progression of retinal microhaemorrhages and diabetic macular oedema in patients with diabetic retinopathy.
UNASSIGNED: This is a multi-centred, randomized, double-blinded, placebo-controlled trial which involved 55 eligible participants. The participants in the treatment group (n = 22) received Tocovid 200 mg twice daily while those in the placebo group (n = 23) would receive placebo twice daily. Both groups will be on the treatment for a total duration of 12 months. Both retinal signs will be assessed at baseline, 2 months, 6 months and 12 months of treatment to determine the progression of diabetic retinopathy. Serum vascular endothelial growth factor which reflects on the angiogenesis process in the eye was analysed as well at similar time points as the retinal findings.
UNASSIGNED: After 12 months of treatment, the placebo group had a significant increase of 23.42% in retinal microhaemorrhages (p < 0.05), but the Tocovid group had no significant changes. Moreover, the Tocovid group showed a significant decrease of 48.38% in area of diabetic macular oedema over the 12 months period (p < 0.05), but the placebo group had no significant changes. Meanwhile, there was no significant difference in serum vascular endothelial growth factor level when comparing between both groups.
UNASSIGNED: These findings could indicate that Tocovid has an important role in preventing early diabetic retinopathy progression.

BACKGROUND: Protease-activated receptor 4 (PAR4), belonging to a subfamily of G-protein-coupled receptors (GPCR), is expressed on the surface of Human platelets, and the activation of it can lead to platelets aggregation. Studies demonstrated that PAR4 inhibition protect mice from arterial/arteriolar thrombosis, pulmonary embolism and cerebral infarct, while do not affect the hemostatic responses integrity. Therefore, PAR4 has been a promising target for the development of anti-thrombotic agents.
METHODS: This review covers recent patents and literature on PAR4 and their application published between 2013 and 2021.
CONCLUSIONS: PAR4 is a promising anti-thrombotic target and PAR4 inhibitors are important biologically active compounds for the treatment of thrombosis. Most the recent patents and literature focus on PAR4 selective inhibitors, and BMS-986120 and BMS-986141, which were developed by BMS, have entered clinical trials. With the deep understanding of the crystal structures and biological functions of PAR4, we believe that many other novel types of molecules targeting PAR4 would enter the clinical studies or the market.

BACKGROUND: To prevent unsolved problems of medical devices, we hypothesized that combinatorial effects of zwitterionic functional group and anti-bacterial metal ions can reduce effectively the thrombosis and bacterial infection of polymeric biomaterials. In this research, we designed a novel series of zwitterionic polyurethane (zPU) additives to impart anti-thrombotic properties to a polyvinyl chloride (PVC) matrix.
METHODS: We have synthesized zPUs by combination of various components and zPUs complexed with metal ions. Zwitterion group was prepared by reaction with 1,3-propane sultone and Nmethyldiethanolamine and metal ions were incorporated into sulfobetaine chains via molecular complexation. These zPU additives were characterized using FT-IR, 1H-NMR, elemental analysis, and thermal analysis. The PVC film blended with zPU additives were prepared by utilizing a solvent casting and hot melting process.
RESULTS: Water contact angle demonstrated that the introduction of zwitterion group has improved hydrophilicity of polyurethanes dramatically. Protein adsorption test resulted in improved anti-fouling effects dependent on additive concentration and decreases in their effects by metal complexation. Platelet adhesion test revealed anti-fouling effects by additive blending but not significant as compared to protein resistance results.
CONCLUSIONS: With further studies, the synthesized zPUs and zPUs complexed with metal ions are expected to be used as good biomaterials in biomedical fields. Based on our results, we can carefully estimate that the enhanced anti-fouling effect contributed to reduced platelet adhesion. Schematic explanation of the effect of zwitterionic polyurethane additives for blood-compatible and anti-bacterial bulk modification.

Hirudin, an acidic polypeptide secreted by the salivary glands of Hirudo medicinalis (also known as \"Shuizhi\" in traditional Chinese medicine), is the strongest natural specific inhibitor of thrombin found so far. Hirudin has been demonstrated to possess potent anti-thrombotic effect in previous studies. Recently, increasing researches have focused on the anti-thrombotic activity of the derivatives of hirudin, mainly because these derivatives have stronger antithrombotic activity and lower bleeding risk. Additionally, various bioactivities of hirudin have been reported as well, including wound repair effect, anti-fibrosis effect, effect on diabetic complications, anti-tumor effect, anti-hyperuricemia effect, effect on cerebral hemorrhage, and others. Therefore, by collecting and summarizing publications from the recent two decades, the pharmacological activities, pharmacokinetics, novel preparations and derivatives, as well as toxicity of hirudin were systematically reviewed in this paper. In addition, the clinical application, the underlying mechanisms of pharmacological effects, the dose-effect relationship, and the development potential in new drug research of hirudin were discussed on the purpose of providing new ideas for application of hirudin in treating related diseases.

Danqi tablet composed of the dried roots of Salvia miltiorrhiza and Panax notoginseng is a well-known Chinese patent medicine commonly used for the treatment of cardio-cerebrovascular diseases such as coronary heart disease and myocardial ischemia. Numerous chemical constituents belonging to S. miltiorrhiza and P. notoginseng were detectable in Danqi tablet. Here, we established and validated a rapid and sensitive ultra-performance liquid chromatography coupled with triple quadrupole mass spectrometry method for simultaneous quantification of 23 components in Danqi tablet and then successfully applied to assay 12 batches of samples from ten manufacturers. Our results demonstrated that the contents of 23 components in 12 batches of Danqi tablets varied significantly and their quality indeed existed differently based on the principal component analysis. According to the quantitative data and the loading plot of principal component analysis, five abundant compounds in Danqi tablet were selected as characteristic chemical markers possibly responsible for the quality assessment. Among them, salvianolic acid B and ginsenoside Rg1 were further chosen to be combined at 2:5 ratio to evaluate the anti-thrombotic activity on phenylhydrazine-induced zebrafish heart thrombosis model. Expectedly, this component combination increased the heart red blood cells intensity compared with the model group and the median effective concentration was 123.4 µg/mL, suggestive of its well anti-thrombotic effect. This study contributed to the quantitative evaluation of Danqi tablet and indicated the combination of salvianolic acid B and ginsenoside Rg1 may be capable of reflecting the effect of Danqi tablet, thereby providing a reference for further investigations on the improvement of quality control and clinical application of Danqi tablet.