This study first optimized the processing technology for Zhangbang vinegar-processed Olibanum and investigated its in vitro anticoagulant activity. A multi-index-response surface methodology was used, with yield, powder yield, and the relative percentage of the content of six non-volatile components [11-keto-boswellic acid(KBA), 3-acetyl-11-keto-boswellic acid(AKBA), β-elemonic acid, α-boswellic acid(α-BA), β-boswellic acid(β-BA), and α-acetyl-boswellic acid(α-BA)] and three volatile components(octyl acetate, incensole, and incensole acetate) as evaluation indicators. Analytical hierarchy process(AHP) combined with coefficient of variation method was used to calculate the weight of each indicator and calculate the comprehensive score(OD). Furthermore, response surface methodology was used to investigate the effects of frying temperature(A), burning time(B), rice vinegar dosage(C), and steaming time(D) on the processing technology of vinegar-processed Olibanum. Vinegar-steamed Olibanum was prepared according to the optimal processing technology for in vitro anticoagulant experiments. The results showed that the weights of octyl acetate, incensole, incensole acetate, KBA, AKBA, β-elemonic acid, α-BA, β-BA, α-ABA, yield, and powder yield were 0.358 2, 0.104 5, 0.146 4, 0.032 9, 0.123 7, 0.044 4, 0.022 1, 0.042 2, 0.110 1, 0.012 2, and 0.0032, respectively. The optimal processing technology for Zhangbang vinegar-processed Olibanum was as follows. Olibanum(50 g) with a particle size of 1-5 mm was continuously stir-fried at a low heat of 150-180 ℃ until in a gel-like state, ignited for burning for 15 s, sprayed with 7.5 g of rice vinegar(15%), and steamed for 3 min without fire. Subsequently, the cover was removed, and the product was continuously stir-fried at 150-180 ℃ until in a soft lump shape, removed, cooled, and crushed. The results of the in vitro anticoagulant experiments showed that compared with the blank group, both Olibanum and vinegar-processed Olibanum significantly prolonged the activated partial thromboplastin time(APTT), thrombin time(TT), and prothrombin time(PT) of rat platelet-poor plasma(PPP), and the effect of vinegar-processed Olibanum was significantly better than that of Olibanum(P<0.05). The optimized processing technology for Zhangbang vinegar-processed Olibanum is stable, feasible, and beneficial for the further development and utilization of Olibanum slices. At the same time, using the content of volatile and non-volatile components, yield, and powder yield as indicators, and verifying through pharmacological experiments, the obtained results are more reasonable and credible, and have positive guiding significance for the clinical application of characteristic processed Olibanum products.
Rats ; Animals ; Frankincense ; Acetic Acid ; Powders ; Triterpenes ; Anticoagulants/pharmacology* ; Technology

This study explored the anticoagulant material basis and mechanism of Trichosanthis Semen and its shell and kernel based on spectrum-effect relationship-integrated molecular docking. High performance liquid chromatography(HPLC) fingerprints of Trichosanthis Semen and its shell and kernel were established. Prothrombin time(PT) and activated partial thromboplastin time(APTT) in mice in the low-and high-dose(5, 30 g·kg~(-1), respectively) Trichosanthis Semen, the shell, and kernel groups were determined as the coagulation markers. The spectrum-effect relationship and anticoagulant material basis of Trichosanthis Semen and its shell and kernel were analyzed with mean value calculation method of Deng's correlation degree(MATLAB) and the common effective component cluster was obtained. Then the common targets of the component cluster and coagulation were retrieved from TCMSP, Swiss-TargetPrediction, GenCLiP3, GeneCards, and DAVID, followed by Gene Ontology(GO) term enrichment and Kyoto Encyclopedia of Genes and Genomes(KEGG) pathway enrichment of the targets. The main anticoagulant molecular mechanism of the component cluster was verified by SYBYL-X 2.1.1. The spectrum-effect relationship of Trichosanthis Semen and its shell and kernel was in positive correlation with the dosage. The contribution of each component to anticoagulation was not the same, suggesting that the material basis for anticoagulation was different, but they have common effective components(i.e. common material basis), such as adenine(peak 3), uracil(peak 4), hypoxanthine(peak 6), xanthine(peak 9), and adenosine(peak 11). Network pharmacology showed that these components can act on multiple target proteins such as NOS3, KDR, and PTGS2, and exert anticoagulant effect through multiple pathways such as VEGF signaling pathway. They involved the biological functions such as proteolysis, cell component such as cytosol, and molecular functions. The results of molecular docking showed that the binding free energy of these components with NOS3(PDB ID: 1 D0 C), KDR(PDB ID: 5 AMN), and PTGS2(PDB ID: 4 COX) was ≤-5 kJ·mol~(-1), and the docking conformations were stable. Spectrum-effect relationship-integrated molecular docking can be used for the optimization, virtual screening, and verification of complex chemical and biological information of Chinese medicine. Trichosanthis Semen and its shell and kernel have the common material basis for anticoagulation and they exert the anticoagulant through multiple targets and pathways.
Animals ; Anticoagulants/pharmacology* ; Drugs, Chinese Herbal/pharmacology* ; Gene Ontology ; Mice ; Molecular Docking Simulation ; Semen

Angelicae Sinensis Radix excels in activating blood, but the scientific mechanism has not been systematically analyzed, thus limiting the development of the medicinal. This study employed the computer-aided drug design methods, such as structural similarity-based target reverse prediction, complex network analysis, molecular docking, binding free energy calculation, cluster analysis, and ADMET(absorption, distribution, metabolism, excretion, toxicity) calculation, and enzyme activity assay in vitro, to explore the components and mechanism of Angelicae Sinensis Radix in activating blood. Target reverse prediction and complex network analysis yielded 40 potential anticoagulant targets of the medicinal. Gene Ontology(GO) term enrichment and Kyoto Encyclopedia of Genes and Genomes(KEGG) pathway enrichment analysis indicated that the targets mainly acted on the complement and coagulation cascade signaling pathway to exert the anticoagulant function. Among them, the key enzymes thrombin(THR) and coagulation factor Xa(FXa) in coagulation cascade and thrombosis were the drug targets for thromboembolic diseases. At the same time, molecular docking and cluster analysis showed that the medicinal had high selectivity for FXa. According to binding free energy score, 8 potential active components were selected for enzyme activity assay in vitro. The results demonstrated that 8 components inhibited THR and FXa, and the inhibition was stronger on FXa than on THR. The pharmacophore model of 8 active compounds was constructed, which suggested that the components had the common pharmacophore AAHH. The ADMET calculation result indicated that they had good pharmacokinetic properties and were safe. Based on target reverse prediction, complex network analysis, molecular docking and binding free energy calculation, anticoagulant activity in vitro, spatial binding conformation of molecules and targets, pharmacophore model construction, and ADMET calculation, this study preliminarily clarified the material basis and molecular mechanism of Angelicae Sinensis Radix in activating blood from the perspective of big data, and calculated the pharmacology and toxicology parameters of the active components. Our study, for the first time, revealed that the medicinal had obvious selectivity and pertinence for different coagulation proteins, reflecting the unique effect of different Chinese medicinals and the biological basis. Therefore, this study can provide clues for precision application of Angelicae Sinensis Radix and the development of the blood-activating components with modern technology.
Anticoagulants/pharmacology* ; Blood Coagulation ; Drug Design ; Drugs, Chinese Herbal/pharmacology* ; Molecular Docking Simulation

Deep venous thrombosis (DVT) poses a major challenge to public health worldwide. Endothelial cell injury evokes inflammatory and oxidative responses that contribute to thrombus formation. Tea polyphenol (TP) in the form of epigallocatechin-3-gallate (EGCG) has anti-inflammatory and oxidative effect that may ameliorate DVT. However, the precise mechanism remains incompletely understood. The current study was designed to investigate the anti-DVT mechanism of EGCG in combination with warfarin (an oral anticoagulant). Rabbits were randomly divided into five groups. A DVT model of rats was established through ligation of the inferior vena cava (IVC) and left common iliac vein, and the animals were orally administered with EGCG, warfarin, or vehicle for seven days. In vitro studies included pretreatment of human umbilical vein endothelial cells (HUVECs) with different concentrations of EGCG for 2 h before exposure to hydrogen peroxide. Thrombus weight and length were examined. Histopathological changes were observed by hematoxylin-eosin staining. Blood samples were collected for detecting coagulation function, including thrombin and prothrombin times, activated partial thromboplastin time, and fibrinogen levels. Protein expression in thrombosed IVCs and HUVECs was evaluated by Western blot, immunohistochemical analysis, and/or immunofluorescence staining. RT-qPCR was used to determine the levels of AGTR-1 and VEGF mRNA in IVCs and HUVECs. The viability of HUVECs was examined by CCK-8 assay. Flow cytometry was performed to detect cell apoptosis and ROS generation was assessed by 2',7'-dichlorofluorescein diacetate reagent. In vitro and invivo studies showed that EGCG combined with warfarin significantly reduced thrombus weight and length, and apoptosis in HUVECs. Our findings indicated that the combination of EGCG and warfarin protects HUVECs from oxidative stress and prevents apoptosis. However, HIF-1α silencing weakened these effects, which indicated that HIF-1α may participate in DVT. Furthermore, HIF-1α silencing significantly up-regulated cell apoptosis and ROS generation, and enhanced VEGF expression and the activation of the PI3K/AKT and ERK1/2 signaling pathways. In conclusion, our results indicate that EGCG combined with warfarin modifies HIF-1α and VEGF to prevent DVT in rabbits through anti-inflammation via the PI3K/AKT and ERK1/2 signaling pathways.
Animals ; Anticoagulants/pharmacology* ; Catechin/analogs & derivatives* ; Eosine Yellowish-(YS)/pharmacology* ; Fibrinogen/pharmacology* ; Hematoxylin/pharmacology* ; Human Umbilical Vein Endothelial Cells ; Humans ; Hydrogen Peroxide/pharmacology* ; Hypoxia-Inducible Factor 1, alpha Subunit/metabolism* ; MAP Kinase Signaling System ; Phosphatidylinositol 3-Kinases/metabolism* ; Polyphenols/pharmacology* ; Proto-Oncogene Proteins c-akt/metabolism* ; RNA, Messenger ; Rabbits ; Rats ; Reactive Oxygen Species/metabolism* ; Signal Transduction ; Sincalide/pharmacology* ; Tea ; Thrombin/pharmacology* ; Vascular Endothelial Growth Factor A/metabolism* ; Venous Thrombosis/pathology* ; Warfarin/pharmacology*

Whitmania pigra is the most widely distributed species of leeches in the market. In this study, the effect of heavy metal lead pollution on the anticoagulant activity of Wh. pigra was studied and the potential mechanism was explored. Pb(NO_3)_2 was used to contaminate the breeding soil which was then used to rear Wh. pigra for 50 days(lead-contaminated group, LC group), and meanwhile the blank control group(CG group) was set. Proteins were extracted from the obtained leech samples, and the differentially expressed proteins between LC and CG groups were analyzed by label-free proteomics technology. In this study, a total of 152 differentially expressed proteins were screened out, of which 93 proteins were up-regulated and 59 proteins were down-regulated in LC group. Bioinformatics analysis showed that the biological processes enriched with the differentially expressed proteins were mainly vesicle-mediated transport and transport positive regulation; the enriched cell components were mainly endocytosis vesicles and apical plasma membrane; the enriched molecular functions mainly included carbohydrate binding. The differentially expressed proteins were enriched in 76 KEGG pathways, which mainly involved metabolic pathways, biosynthesis of secondary metabolites, and bacterial invasion of epithelial cells. In this study, two differentially expressed proteins with Antistasin domain were presumed, which provides reference for further exploring the regulatory mechanism and signal transduction underlying the effect of lead pollution on the anticoagulant activity of leech.
Animals ; Anticoagulants/pharmacology* ; Environmental Pollution ; Leeches ; Metals, Heavy ; Proteomics

Leech has a good anticoagulant activity and is one of the raw materials for treatment of many cardiovascular and cerebrovascular diseases. This study was based on in vitro anticoagulant experiments( APTT and PT) to investigate the effects of lead contamination on the anticoagulant effect of leech. At present,the Hirudo circulating in the market are dominated by Whitmania pigra,therefore Wh. pigra were cultivated under a different lead pollution for 50 days. Then,the effects of Wh. pigra extract,extracting from different cultivating environment,on activated partial thrombin time( APTT) and prothrombin time( PT) were determined by automatic coagulation instrument. The results showed that the Wh. pigra extract significantly prolonged the APTT compared with the saline group.The APTT of the lead-high residual Wh. pigra was shorter than that of the blank Wh. pigra. The Wh. pigra extracts from different treatment groups had little effect on PT. The results showed that the lead residue in the Wh. pigra increased with the increase of lead in the cultured soil,the lead residual of the Pb-H group was( 10. 66±2. 79) mg·kg~(-1),which exceeded the lead limit specified in the 2015 edition of the Chinese Pharmacopoeia. The results indicated that growth environment pollution is one of the important factors causing excessive lead in Wh. pigra. Lead pollution will reduce the anticoagulant effect of Wh. pigra and affect its clinical efficacy.
Animals ; Anticoagulants ; Biological Products/pharmacology* ; Blood Coagulation ; Environmental Pollution ; Lead/toxicity* ; Leeches/drug effects* ; Prothrombin Time ; Thrombin Time

Lung cancer is the first leading cause of morbidity and mortality in the world. Venous thromboembolism (VTE) is a recognized complication in patients with lung cancer, which is one of the leading cause of death in lung cancer patients. The cancer-related, patient-related and treatment-related factors are the main causes of VTE in lung cancer patients. Malignant cells can directly activate blood coagulation by producing tissue factor (TF), cancer procoagulance (CP), inflammatory factors and cytokines; And the one of predominant mechanisms in cancer-related thrombosis is the overexpression of TF. The 10th edition of the antithrombotic therapy guidelines for VTE with cancer patients (AT-10) published in 2016 by American College of Chest Physicians (APCC) recommended that anticoagulant therapy is the basic treatment for patients with lung cancer complicated with VTE; And low molecular-weight-heparin (LMWH) is preferred as an anticoagulant drug, but can be use with caution due to increasing risk of bleeding.
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Anticoagulants ; pharmacology ; therapeutic use ; Humans ; Lung Neoplasms ; complications ; Risk Factors ; Venous Thromboembolism ; complications ; drug therapy

BACKGROUND: The incidence of pulmonary embolism (PE) in lung cancer patients who underwent surgery increased during the perioperative period, and prophylactic anticoagulation therapy was important part of enhanced recovery after surgery (ERAS). However, the timing of preventive anticoagulation in patients with lung cancer remained controversial. This study was designed to investigate the safety and efficacy of perioperative prophylactic anticoagulation therapy for lung cancer patients. METHODS: Retrospective research was conducted on 562 lung cancer patients who underwent video-assisted thoracoscopic (VATS) anatomic pulmonary resections in West China Hospital from June 2016 to December 2016. 56 patients were treated with low molecular weight heparin (LMWH) anticoagulation 12 hours before operation until discharge, while the other 506 patients were treated with LMWH 24 hours after operation until discharge. The postoperative chest drainage volume, postoperative bleeding, pulmonary embolism incidence, and respiratory complications were analyzed. RESULTS: (1) There were no significant differences in prothrombin time (PT), activated partial thromboplastin time (APTT) and international normalized ratio (INR) between the pre-operation prophylactic anticoagulation group (PRE group) [(11.5±3.9) s, (27.8±3.5) s, (0.96±0.06) s] and the post-operation prophylactic anticoagulation group (POST group) [(11.4±1.4) s, (28.3±4.0) s, (0.98±0.07) s] (P=0.796, P=0.250, P=0.137), and there was no significant difference in Caprini score between the PRE group (3.1±1.8) and the POST group (3.3±1.5) (P=0.104). (2) There were no significant differences in anesthesia time and intraoperative bleeding between PRE group [(130.2±53.9) min, (76.8±49.3) mL] and POST group [(142.2±56.5) min, (73.7±41.6) mL] (P=0.067, P=0.201). (3) The total drainage volume in 72 hours after operation in PRE group [(728.1±505.7) mL] was significantly higher than that of POST group [(596.4±373.5) mL] (P=0.005), while there were no significant differences between the two groups in total postoperative drainage volume [(1,066.8±1,314.6) mL vs (907.8±999.8) mL, P=0.203]. (4) There were no significant differences between the two groups in pulmonary embolism incidence (1.785% vs 0.019%, P=0.525) and postoperative bleeding rates (1.785% vs 0.039%, P=0.300). (5) There were no significant differences between PRE group and POST group in subcutaneous emphysema incidence (1.785% vs 1.581%, P=0.989) and pulmonary infection rates (14.285% vs 6.324%, P=0.085). CONCLUSIONS The clinical value of preoperative prophylactic anticoagulation is equal to postoperative prophylactic anticoagulation for lung cancer patients.
Adult ; Aged ; Anticoagulants ; pharmacology ; Female ; Heparin, Low-Molecular-Weight ; pharmacology ; Humans ; Lung Neoplasms ; surgery ; Male ; Middle Aged ; Perioperative Period ; Postoperative Complications ; etiology ; prevention & control ; Retrospective Studies ; Safety ; Young Adult

To evaluate the safety and efficiency of citrate anticoagulant-based continuous blood purification in patients at high risk of bleeding. 
 Methods: One hundred and fifty-two patients at high risk of bleeding were divided into local citrate group (group A, n=68) and heparin group (group B, n=84). Clotting function, change of pH, ionized sodium, bicarbonate ion, ionized calcium, activated clotting time (ACT) and complications were monitored before and during treatment. 
 Results: Compared to the group A, the incidence of clotting in filter and chamber, the degree of bleeding or fresh bleeding were significantly reduced in the group B (P<0.05). ACT of post-filter at 4, 8 and 12 h during the treatment in the group A was significantly extended compared with that without treatment (P<0.05), while there was no significant change in group B (P>0.05). The pH value, the levels of ionized sodium, bicarbonate ion and ionized calcium during the treatment were maintained in normal range in both group A and group B.
 Conclusion: Local citrate-based continuous blood purification can achieve effective anticoagulation and decrease the incidence of bleeding. It is an ideal choice for patients at high risk of bleeding.
Anticoagulants ; pharmacology ; Bicarbonates ; blood ; Blood Coagulation ; drug effects ; Blood Coagulation Tests ; Calcium ; blood ; Citrates ; Citric Acid ; therapeutic use ; Female ; Hemodiafiltration ; adverse effects ; methods ; Hemofiltration ; Hemorrhage ; etiology ; prevention & control ; Heparin ; therapeutic use ; Humans ; Intensive Care Units ; Male ; Reference Values ; Renal Dialysis ; Sodium ; blood ; Treatment Outcome

BACKGROUND: The identification of in vitro hemolysis (IVH) using a hematology analyzer is challenging because centrifugation of the specimens cannot be performed for cell counts. In the present study, we aimed to develop a scoring system to help identify the presence of hemolysis in anticoagulated blood specimens. METHODS: Thirty-seven potassium EDTA anticoagulated blood specimens were obtained, and each specimen was divided into 3 aliquots (A, B, and C). Aliquots B and C were mechanically hemolyzed by aspirating 2 and 5 times, respectively, using a 27-gauge needle and then tested; aliquot A was analyzed immediately without any hemolysis. After the cells were counted, aliquots B and C were centrifuged and the supernatants were tested for the hemolytic index and lactate dehydrogenase levels. RESULTS: The 4 hematologic parameters were selected and scored from 0 to 3 as follows:< 34.0, 34.0-36.2, 36.3-38.4, and > or =38.5 for mean cell hemoglobin concentration (MCHC, g/dL); <0.02, 0.02, 0.03, and > or =0.04 for red blood cell ghosts (10(12)/L); <0.13, 0.13-0.38, 0.39-1.30, and > or =1.31 for difference value (g/dL) of measured hemoglobin and calculated hemoglobin; and <0.26, 0.26-0.95, 0.96-3.34, and > or =3.35 for difference value (g/dL) of MCHC and cell hemoglobin concentration mean. The hemolysis score was calculated by adding all the scores from the 4 parameters. At the cutoff hemolysis score of 3, the IVH of aliquots B and C were detected as 64.9% and 91.9%, respectively. CONCLUSIONS: The scoring system might provide effective screening for detecting spurious IVH.
Anticoagulants/*pharmacology ; *Blood Specimen Collection ; Edetic Acid/pharmacology ; Hemoglobins/analysis ; Hemolysis/drug effects ; Humans