The Trace Element Selenium Is Important for Redox Signaling in Phorbol Ester-Differentiated THP-1 Macrophages.

Physiological selenium (Se) levels counteract excessive inflammation, with selenoproteins shaping the immunoregulatory cytokine and lipid mediator profile. How exactly differentiation of monocytes into macrophages influences the expression of the selenoproteome in concert with the Se supply remains obscure. THP-1 monocytes were differentiated with phorbol 12-myristate 13-acetate (PMA) into macrophages and (i) the expression of selenoproteins, (ii) differentiation markers, (iii) the activity of NF-κB and NRF2, as well as (iv) lipid mediator profiles were analyzed. Se and differentiation affected the expression of selenoproteins in a heterogeneous manner. GPX4 expression was substantially decreased during differentiation, whereas GPX1 was not affected. Moreover, Se increased the expression of selenoproteins H and F, which was further enhanced by differentiation for selenoprotein F and diminished for selenoprotein H. Notably, LPS-induced expression of NF-κB target genes was facilitated by Se, as was the release of COX- and LOX-derived lipid mediators and substrates required for lipid mediator biosynthesis. This included TXB2, TXB3, 15-HETE, and 12-HEPE, as well as arachidonic acid (AA), eicosapentaenoic acid (EPA), and docosahexaenoic acid (DHA). Our results indicate that Se enables macrophages to accurately adjust redox-dependent signaling and thereby modulate downstream lipid mediator profiles.

The Synergistic Effect of Ginkgo biloba Extract 50 and Aspirin Against Platelet Aggregation.

PURPOSE: We aimed to investigate potential synergistic antiplatelet effects of Ginkgo biloba extract (GBE50) in combination with aspirin using in vitro models. METHODS: Arachidonic acid (AA), platelet activating factor (PAF), adenosine 5'-diphosphate (ADP) and collagen were used as inducers. The antiplatelet effects of GBE50, aspirin and 1:1 combination of GBE50 and aspirin were detected by microplate method using rabbit platelets. Synergy finder 2.0 was used to analyze the synergistic antiplatelet effect. The compounds in GBE50 were identified by UPLC-Q/TOF-MS analysis and the candidate compounds were screened by TCMSP database. The targets of candidate compounds and aspirin were obtained in TCMSP, CCGs, Swiss target prediction database and drugbank. Targets involving platelet aggregation were obtained from GenCLiP database. Compound-target network was constructed and GO and KEGG enrichment analyses were performed to identify the critical biological processes and signaling pathways. The levels of thromboxane B2 (TXB2), cyclic adenosine monophosphate (cAMP) and PAF receptor (PAFR) were detected by ELISA to determine the effects of GBE50, aspirin and their combination on these pathways. RESULTS: GBE50 combined with aspirin inhibited platelet aggregation more effectively. The combination displayed synergistic antiplatelet effects in AA-induced platelet aggregation, and additive antiplatelet effects occurred in PAF, ADP and collagen induced platelet aggregation. Seven compounds were identified as candidate compounds in GBE50. Enrichment analyses revealed that GBE50 could interfere with platelet aggregation via cAMP pathway, AA metabolism and calcium signaling pathway, and aspirin could regulate platelet aggregation through AA metabolism and platelet activation. ELISA experiments showed that GBE50 combined with aspirin could increase cAMP levels in resting platelets, and decreased the levels of TXB2 and PAFR. CONCLUSION: Our study indicated that GBE50 combined with aspirin could enhance the antiplatelet effects. They exerted both synergistic and additive effects in restraining platelet aggregation. The study highlighted the potential application of GBE50 as a supplementary therapy to treat thrombosis-related diseases.

Changes in PUFA and eicosanoid metabolism during/after apnea diving: a prospective single-center study.

Background: The popularity of apneic diving is continually growing. As apnea diving substantially burdens the cardiovascular system, special focus is warranted. Regarding inflammation processes and associated inflammatory-related diseases (e.g., cardiovascular diseases), eicosanoids play an important role. This study aims to investigate polyunsaturated fatty acids (PUFAs) and eicosanoids in voluntary apnea divers, and so to further improve understanding of pathophysiological processes focusing on proinflammatory effects of temporarily hypercapnic hypoxia.. Methods: The concentration of PUFAs and eicosanoids were investigated in EDTA plasma in apnea divers (n=10) before and immediately after apnea, 0.5 hour and four hours later, applying liquid chromatography-tandem mass spectrometry (LC-MS/MS). Results: Mean age was 41±10 years, and divers performed a mean breath-hold time of 317±111 seconds. PUFAs, eicosanoids and related lipids could be classified in four different kinetical reaction groups following apnea. The first group (e.g., Ω-6 and Ω-3-PUFAs) showed an immediate concentration increase followed by a decrease below baseline four hours after apnea. The second group (e.g., thromboxane B2) showed a slower increase, with its maximum concentration 0.5 hour post-apnea followed by a decrease four hours post-apnea. Group 3 (9- and 13-hydroxyoctadecadienoic acid) is characterized by two concentration increase peaks directly after apnea and four hours afterward compared to baseline. Group 4 (e.g., prostaglandin D2) shows no clear response. Conclusion: Changes in the PUFA metabolism after even a single apnea revealed different kinetics of pro- and anti-inflammatory regulations and changes for oxidative stress levels. Due to the importance of these mediators, apnea diving should be evaluated carefully and be performed only with great caution against the background of cardiovascular diseases and inflammation processes.

The Effects of Tocotrienol-Rich Vitamin E (Tocovid) on Diabetic Neuropathy: A Phase II Randomized Controlled Trial.

Chronic hyperglycemia increases oxidative stress, activates inflammatory pathways and reduces nerve growth factor (NGF) among diabetic patients, which contribute to development of diabetic peripheral neuropathy (DPN). Tocotrienol-Rich Vitamin E (Tocovid) possesses potent antioxidant and anti-inflammatory properties which are postulated to target these pathogeneses in order to ameliorate DPN. This study aims to evaluate the effects of Tocovid on nerve conduction parameters and serum biomarkers among diabetic patients. This multicenter, prospective, randomized, double-blind, placebo-controlled clinical trial was conducted on 80 eligible participants. The intervention group (n = 39) was randomly allocated to receive 200 mg of Tocovid twice a day, and the control group (n = 41) received placebo twice a day. At the end of eight weeks, the nerve conduction parameters, as assessed by nerve conduction study, as well as serum biomarkers (NGF, malondialdehyde, vascular cell adhesion molecule 1, tumor necrosis factor receptor 1 and thromboxane B2) were compared between the two groups. Compared to placebo, Tocovid significantly improves the nerve conduction velocities of all nerves (+1.25 m/s, interquartile range [IQR] 3.35, p < 0.001, median nerve; +1.60 m/s, IQR 1.80, p < 0.001, sural nerve; +0.75 m/s, IQR 2.25, p < 0.001, tibial nerve). Meanwhile, the levels of serum NGF were significantly higher in the Tocovid group as compared to placebo at eight weeks post-intervention. Participants receiving Tocovid illustrated highly significant improvement in terms of nerve conduction velocities for all nerves tested after eight weeks of supplementation. In addition, Tocovid supplementation elevated the levels of serum NGF, in which its increase is postulated to reflect enhanced neuronal functions. This novel finding suggests that Tocovid could be a disease-modifying agent targeting serum NGF to improve nerve conduction velocities.

Extract of Danggui-Shaoyao-San ameliorates cognition deficits by regulating DHA metabolism in APP/PS1 mice.

ETHNOPHARMACOLOGICAL RELEVANCE: The traditional Chinese medicine formula Danggui-Shaoyao-San (DSS) has been reported to show therapeutic effect on alleviating the symptoms of Alzheimer's disease (AD). AIM OF THE STUDY: The present study aims to investigate the relation between DSS treatment of AD and DHA metabolism and evaluates its neuroprotective effect on cognitive in APP/PS1 mice. MATERIAL AND METHODS: DSS (1.6, 3.2, 6.4 g/kg/day) or Aricept (3 mg/kg/day) was orally administered (i.g.) to APP/PS1 mice, and saline was orally administered to Wild-type (WT) male mice as control group. Then, the Morris water maze (MWM) test, Y-maze spontaneous alternation test, open filed test and fear conditioning test were conducted for evaluation of learning and memory abilities. The DHA content was assessed by HPLC-MS/MS. Physiological indices were determined, including triglyceride (TG), total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C), ROS level, activity of superoxide dismutase (SOD), glutathione (GSH), malondialdehyde (MDA), PEG2, TXB2 and LTB4. The expressions of COX-1, COX-2, cPLA2, iPLA2, 15-LOX, and were assessed by Western blot. RESULTS: APP/PS1 mice showed serious cognitive impairment in behavioral tests. However, treatment of DSS extract significantly ameliorated the cognitive deficits of APP/PS1 mice. Biochemical measurements showed the increases in TG, TC, LDL-c and the decrease in HDL-c in APP/PS1 mice compared with WT mice, and DSS extract significantly retarded these changes. Low content of DHA, low expression of iPLA2 and 15-LOX were observed both in hippocampus and cortex of APP/PS1 mice, while DSS extract significantly restored these changes. Additionally, the abnormal activity of SOD and ROS level, the decreased levels of MDA and GSH were observed in APP/PS1 mice, while DSS extract prominently lessened these changes. Moreover, DSS extract decreased the level of PEG2, TXB2 and LTB4 and also attenuated the expression of cPLA2, COX-1 and COX-2 in hippocampus as well as cortex of APP/PS1 mice. CONCLUSIONS: Based on these results, we suggest that DSS play a positive effective role in increasing DHA content by up-regulating iPLA2 and 15-LOX, resulting in ameliorating oxidative stress and inflammation and finally ameliorating cognition deficits in APP/PS1 mice.

Evaluation Procoagulant Activity and Mechanism of Astragalin.

Astragalin, isolated from flowers of Rosa chinensis Jacq., is a kind of flavonoid, with anti-inflammatory, antioxidant, antiviral, analgesic, antibacterial, antiallergic, and antihepatotoxic effects. However, no studieson the procoagulant effect of astragalin have been reported. This study aimed to investigate the procoagulant activity of astragalin and its mechanism. Its procoagulant effect was investigated by activated partial thromboplastin time (APTT), thrombin time (TT), prothrombin time (PT), and fibrinogen (FIB) in vitro, and a rat model established by heparin sodium was used to evaluate the mechanism for the procoagulant effect in vivo. The results showed that astragalin had good procoagulant effects compared with the control group in vitro. Compared with the model group in vivo, astragalin could shorten the coagulation time and significantly increase the number of platelets. Meanwhile, astragalin could significantly reduce the effectual time of PT and APTT and increase the content of FIB. The contents of 6-keto-PGF1α and eNOS significantly decreased. Astragalin could increase whole blood viscosity (WBV), plasma viscosity (PV), erythrocyte sedimentation rate (ESR) and packedcell volume (PCV). All of the above revealed that astragalin had good procoagulant effects by promoting the intrinsic and extrinsic coagulation system.

Untargeted metabolomics reveals alterations in metabolites of lipid metabolism and immune pathways in the serum of rats after long-term oral administration of Amalaki rasayana.

Amalaki rasayana, a traditional preparation, is widely used by Ayurvedic physicians for the treatment of inflammatory conditions, cardiovascular diseases, and cancer. Metabolic alterations induced by Amalaki rasayana intervention are unknown. We investigated the modulations in serum metabolomic profiles in Wistar rats following long-term oral administration of Amalaki rasayana. Global metabolic profiling was performed of the serum of rats administered with either Amalaki rasayana (AR) or ghee + honey (GH) for 18 months and control animals which were left untreated. Amalaki rasayana components were confirmed from AR extract using HR-LCMS analysis. Significant reductions in prostaglandin J2, 11-dehydrothromboxane B2, and higher levels of reduced glutathione and glycitein metabolites were observed in the serum of AR administered rats compared to the control groups. Eleven different metabolites classified as phospholipids, glycerophospholipids, glucoside derivatives, organic acids, and glycosphingolipid were exclusively observed in the AR administered rats. Pathway analysis suggests that altered metabolites in AR administered rats are those associated with different biochemical pathways of arachidonic acid metabolism, fatty acid metabolism, leukotriene metabolism, G-protein mediated events, phospholipid metabolism, and the immune system. Targeted metabolomics confirmed the presence of gallic acid, ellagic acid, and arachidonic acid components in the AR extract. The known activities of these components can be correlated with the altered metabolic profile following long-term AR administration. AR also activates IGF1R-Akt-Foxo3 signaling axis in heart tissues of rats administered with AR. Our study identifies AR components that induce alterations in lipid metabolism and immune pathways in animals which consume AR for an extended period.

[Anti-platelet aggregation and anti-thrombotic mechanism of Trichosanthis Fructus combined with aspirin based on network pharmacology].

To explore the anti-platelet aggregation and anti-thrombotic mechanisms of Trichosanthis Fructus combined with aspirin based on network pharmacology and the validation of arteriovenous by pass model in rats. The databases of Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform(TCMSP),Drug Repositioning and Adverse Drug Reaction Chemical-Protein Interactome(DRAR-CPI),Universal Protein Resource(Uniprot) and the Database for Annotation,Visualization,and Integrated Discovery(DAVID) were used to predict protein targets and analyze biological pathway and signal pathway in the combination of Trichosanthis Fructus with aspirin. The effects of pretreatment with Trichosanthis Fructus pellets,aspirin pellets and their combination on thromboxane B2(TXB2),6-keto prostaglandin F1α(6-keto-PGF1α) and cyclic adenosine monophosphate(c AMP) in rat thrombotic model were studied. Through the study of network pharmacology,12 components of aspirin and Trichosanthis Fructus,including hydroxygenkwanin,quercetin and adenosine,were found to show the anti-platelet aggregation and anti-thrombosis mechanisms through9 common protein targets,such as SRC,RAC1,MAPK14,MAPK1,AKT1,and 14 common signaling pathways,such as VEGF signaling pathway. After the intervention with Trichosanthis Fructus pellets combined with aspirin pellets,the vascular endothslia growth factor(VEGF) signaling pathway can be activated to inhibit platelet aggregation and improve vascular endothelial function,and show the anti-platelet aggregation and anti-thrombosis mechanisms,which verify the results of the network pharmacology,and explain the anti-platelet aggregation and anti-thrombotic mechanisms of the combination of Trichosanthis Fructus pellets with aspirin pellets.

Anti-platelet aggregation and anti-thrombotic mechanism of Trichosanthis Fructus combined with aspirin based on network pharmacology / 中国中药杂志

To explore the anti-platelet aggregation and anti-thrombotic mechanisms of Trichosanthis Fructus combined with aspirin based on network pharmacology and the validation of arteriovenous by pass model in rats. The databases of Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform(TCMSP),Drug Repositioning and Adverse Drug Reaction Chemical-Protein Interactome(DRAR-CPI),Universal Protein Resource(Uniprot) and the Database for Annotation,Visualization,and Integrated Discovery(DAVID) were used to predict protein targets and analyze biological pathway and signal pathway in the combination of Trichosanthis Fructus with aspirin. The effects of pretreatment with Trichosanthis Fructus pellets,aspirin pellets and their combination on thromboxane B2(TXB2),6-keto prostaglandin F1α(6-keto-PGF1α) and cyclic adenosine monophosphate(c AMP) in rat thrombotic model were studied. Through the study of network pharmacology,12 components of aspirin and Trichosanthis Fructus,including hydroxygenkwanin,quercetin and adenosine,were found to show the anti-platelet aggregation and anti-thrombosis mechanisms through9 common protein targets,such as SRC,RAC1,MAPK14,MAPK1,AKT1,and 14 common signaling pathways,such as VEGF signaling pathway. After the intervention with Trichosanthis Fructus pellets combined with aspirin pellets,the vascular endothslia growth factor(VEGF) signaling pathway can be activated to inhibit platelet aggregation and improve vascular endothelial function,and show the anti-platelet aggregation and anti-thrombosis mechanisms,which verify the results of the network pharmacology,and explain the anti-platelet aggregation and anti-thrombotic mechanisms of the combination of Trichosanthis Fructus pellets with aspirin pellets.

Aspirin intake in the morning is associated with suboptimal platelet inhibition, as measured by serum Thromboxane B2, during infarct-prone early-morning hours.

Aspirin is traditionally taken once daily in the morning and considered to be effective throughout the 24h interval. Cardiovascular events occur most frequently in the early morning, suggesting that these hours are critical in terms of adequate platelet inhibition. This study therefore assed platelet function in the early morning-8.00 AM-in healthy volunteers, during a once-daily (OD) 80 mg morning in comparison with an OD evening regimen and a twice-daily (BID) 40 mg regimen. It was an open-label randomized cross-over study, comprising 12 healthy subjects. Subjects were allocated to three sequential dosage regimens: 80 mg OD at 8.00 AM, 80mg OD at 8.00 PM, and 40 mg BID at 8.00 AM and PM. Platelet function 12 and 24 hours after aspirin intake was measured by means of serum thromboxane B2 (sTxB2) levels, the collagen/epinephrine closure time (Platelet Function Analyzer(PFA)-200®) and the Aspirin Reaction Units (ARU, VerifyNow®). The results demonstrated that early morning sTxB2 concentrations were 5843pg in the morning regimen, 2877pg in the evening OD regimen, and 3343pg in the BID regimen (morning- vs evening regimen p = < 0.01; morning- vs BID regimen p = < 0.01). Early morning PFA-closure time (p = 0.12)) as well as VerifyNow ARU (p = 0.17) mean values were similar for all three regimens. In conclusion, the OD-morning regimen seems to acquire the lowest level of platelet inhibition during the critical early morning window. Switching to an OD-evening or BID intake seems prudent, although further research on clinical cardiovascular outcome in patients with stable cardiovascular disease is needed.

Antithrombotic effects and related mechanisms of Salvia deserta Schang root EtOAc extracts.

Salvia deserta Schang (SDS) belongs to the same family as Salvia miltiorrhiza bunge, one of the antithrombotic Chinese herbal medicines. In our study, EtOAc root extracts were analyzed for their effects on adenosine diphosphate (ADP)-induced platelet aggregation in rabbits and FeCl3-induced rat common carotid artery thrombosis as well as on rat blood plasma concentrations of thromboxane B2 (TXB2), 6-keto-prostaglandin F1 alpha (6-keto-PGF1α), antithrombin-III (AT-III), protein C (PC), plasminogen (PLG), plasminogen activator inhibitor (PAI-1), von Willebrand factor (vWF) and tissue-type plasminogen activator (t-PA). EtOAc extracts from SDS roots had significant inhibitory effects on ADP-induced maximum platelet aggregation rate (10.2 ± 2.6 vs control 35.7 ± 5.2; P < 0.05), reduced the FeCl3-induced rat common carotid artery thrombus weight and thrombus area ratio (P < 0.05), significantly decreased plasma TXB2, vWF and PAI-1 levels and increased 6-keto-PGF1α and t-PA levels in a dose dependent manner (all P < 0.05). Thus, the ratio of TXB2/6-keto-PGF1α was significantly decreased (P < 0.05), while the ratio of t-PA/PAI-1 was significantly increased (P < 0.05). In addition, enhanced AT-III and PC activities indicated coagulation inactivation effects of EtOAc SDS root extracts. EtOAc extraction from SDS showed antithrombotic effects, which are likely due to platelet adhesion and aggregation inhibition as well as anticoagulant activities.

Pharmacokinetic-pharmacodynamic modeling to study the anti-dysmenorrhea effect of Guizhi Fuling capsule on primary dysmenorrhea rats.

BACKGROUND: Primary dysmenorrhea (PDM) is one of the most common gynaecological disorders among women, which seriously affects women's life quality due to its high incidence rate. Guizhi Fuling capsule (GZFLC), a well-known traditional Chinese medical prescription, has been widely used to treat gynecological blood stasis syndromes such as PDM. However, its mechanisms of action and combination were still unknown. PURPOSE: The aim of this study was to develop a pharmacokinetic-pharmacodynamic (PK-PD) model to assess time-concentration-effect relationships for anti-dysmenorrhea effect of GZFLC and provide better understanding for mechanisms of action and combination of GZFLC. STUDY DESIGN AND METHODS: The PDM rats model was induced by oxytocin exposure following estradiol benzoate pretreatment. Gallic acid (GA), amygdalin (AMY), albiflorin (ALB), prunasin (PA) and cinnamic acid (CA) were evaluated as bioactive ingredients for investigating PK processes. GA, AMY, ALB and PA exhibited appropriate PK parameters and were selected as the PK markers to map the anti-dysmenorrhea effect of GZFLC. A PK-PD model was established on the basis of GA, AMY, ALB and PA plasma concentrations vs. the values of two ratios (PGE2/PGF2α and 6-Keto-PGF1α/TXB2), by a two-compartment PK model with a simple Emax model to explain the time delay between the drug plasma concentrations of PK markers and the anti-dysmenorrhea effect. RESULTS: The PDM rat model has been successfully established. Compared with the normal treated group, the bioactive ingredients in PDM treated group exhibited significant changing trends of PK behaviors, such as better absorption and distribution, slower elimination and delays in reaching the maximum concentration (Tmax). The analysis of PK-PD parameters indicated that the active metabolites and prototypes of bioactive ingredients in GZFLC were inclined to regulate the activity of prostacyclin synthetase and thromboxane synthetase to control the production of TXA2 and PGI2 so as to treat PDM. As the main effective medicinal materials for the treatment of PDM in GZFLC prescription Persicae Semen, Moutan Cortex and Paeonia lactiflora Pall, Persicae Semen played the most important role, while the role of Paeonia lactiflora Pall was the weakest. CONCLUSION: The PK-PD model results provided scientific basis for clarifying compatibility mechanisms of GZFLC prescription and a better understanding for biosynthetic mechanisms of four prostaglandins (PGE2, PGF2α, 6-Keto-PGF1α and TXB2) in the treatment of PDM by GZFLC. Investigations on the relationship between the effects and the bioactive ingredients are of benefit to explore the mechanisms of action and combination for traditional Chinese medical prescriptions (TCP) and facilitate the development of future clinical applications of TCP.

[Effects of Gui Zhi Fu Ling Jiao Nang on Hemorheology and Expression of Phenotypic Proteins in Uterine Smooth Muscle Cells in Rats with An Intrauterine Device].

Objective To observe the effect of Gui Zhi Fu Ling Jiao Nang (GZFLJN) on the expressions of alpha smooth muscle actin (α-SMA) and proliferating cell nuclear antigen (PCNA) in uterine vascular smooth muscle cells (VSMC) of rat models with an intrauterine device (IUD) and to determine the thromboxane B2 (TXB2) and 6-keto-prostaglandin F1α (6-keto-PGF1α) levels in peripheral blood. Methods Female Wistar rats were randomly divided into four groups:normal group (n=16,with normal breed without treatment),model group (n=18,drenching 0.9% normal saline after modeling of IUD),GZFLJN group (n=18),and aminocaproic acid tablets group (n=17). Immunohistochemical SP method was used to detect the expressions of α-SMA and PCNA in uterine VSMC.ELISA was served to detect the levels of TXB2 and 6-keto-PGF1α in peripheral blood. Results The positive rate of α-SMA were (50.89±9.41)%,(26.93±6.80)%,(48.92±6.80)%,and (34.63±7.26)%,respectively,in normal group,model group,GZFLJN group,and aminocaproic acid tablets group;obviously,it was significantly higher in normal group (t=14.43,P=0.00) and GZFLJN group (t=11.37,P=0.00) than that in model group and it was significantly lower in aminocaproic acid tablets group than in normal group (t=9.96,P=0.00) and GZFLJN group (t=8.23,P=0.00). The positive rate of PCNA were (25.66±7.24)%,(61.26±9.98)%,(28.36±9.17)%,and (50.23±8.71)%,respectively,in these four groups;obviously,it was significantly lower in the normal group (t=20.86,P=0.00) and GZFLJN group (t=19.12,P=0.00) than in model group and it was significantly higher in aminocaproic acid tablets group than in normal group (t=17.82,P=0.00) and GZFLJN group (t=16.05,P=0.00). Serum TXB2 level in these four groups were (445.86±24.43),(508.78±12.42),(448.11±9.63),and (498.11±13.63)ng/L;obviously,it was significantly higher in model group than in normal group (t=16.55,P=0.00) and aminocaproic acid tablets group (t=-4.12,P=0.00) and it was significantly lower in GZFLJN group than in model group (t=-15.23,P=0.00) and aminocaproic acid tablets group (t=-12.08,P=0.00). Serum 6-keto-PGF1α level in these four groups were (23.17±1.93),(18.09±0.93),(22.70±1.61),and (20.70±1.41)ng/L,respectively;obviously,it was significantly lower in model group than in normal group (t=-13.98,P=0.00) and aminocaproic acid tablets group (t=5.26,P=0.00) and it was significantly higher in GZFLJN group than in model group (t=11.43,P=0.00) and aminocaproic acid tablets group (t=8.76,P=0.00). Conclusion GZFLJN can regulate the expressions of α-SMA and PCNA of VSMC in the endometrium of IUD rats and the concentrations of TXB2 and 6-keto-PGF1α in the serum.

Mahuang decoction mitigates airway inflammation and regulates IL-21/STAT3 signaling pathway in rat asthma model.

ETHNOPHARMACOLOGICAL RELEVANCE: Nowadays, bronchial asthma is still a severe disease threatening human health, and it is incumbent upon us to seek effective therapeutic drugs. Mahuang decoction (MHD), a classic famous Chinese prescription, has been used for thousands of years to prevent phlegm from forming, stop coughing and relieve asthma, but the relevant mechanism has not been thoroughly clarified. This study aims to investigate the anti-airway inflammation effect of MHD and the possible molecular mechanism underlying IL21/STAT3 signaling pathway, so as to provide guidance for the treatment of MHD on bronchial asthma. MATERIALS AND METHODS: Specific pathogen free SD rats were randomly divided into 6 groups: normal control group, model group, positive group (Compound methoxyphenamine), MHD-treated groups at doses of 10 ml/kg, 5 ml/kg and 2.5 ml/kg, 10 rats in each group. Except for the normal control group, rats in other groups were sensitized with ovalbumin via introperitoneal injection and challenged with ovalbumin inhalation to trigger asthma model. At 24 h after the last excitation, bronchoalveolar lavage fluid (BALF) of every rat was drawn and the number of inflammatory cells was analyzed using cell counting method. ELISA method was performed to determine the concentrations of TXB2, 6-keto-PGF1α, MMP-9, TIMP-1, IL-2, IL-4, IL-5 and TNF-α in rat serum. The protein expressions of IL-21, IL-21R, STAT3 and p-STAT3 in murine pulmonary tissues were assessed with western blotting analysis. RESULTS: Compared with the control group, the airway wall and airway smooth muscle of murine pulmonary tissues significantly thickened and massive inflammatory cells infiltration occurred around the bronchus in the model group, and the cell counts of WBC and EOS in BALF were also apparently increased, which indicated the rat asthma model was successfully established. MHD or Compound methoxyphenamine not only alleviated the pulmonary inflammatory pathological damages, but also down- regulated the numbers of WBC and EOS in BALF. What's more, the levels of TXB2, MMP-9, TIMP-1, ILs-(2, 4, 5) and TNF-α in rat serum were lessened by the treatment of MHD. In western blotting analysis, treatment with 10 ml/kg or 5 ml/kg MHD markedly declined the increased protein expressions of IL-21, IL-21R, STAT3 and p-STAT3 in lung tissues of asthmatic rats to normal level. CONCLUSION: MHD intervention demonstrated a strong inhibitory action on the secretion of inflammatory mediators as well as the inflammatory cell infiltration in pulmonary tissues of asthmatic rats, and also depressed the protein expressions of IL-21, IL-21R, STAT3 and p-STAT3 in pulmonary tissues. MHD effectively mitigates airway inflammation and regulates the IL-21/STAT3 signaling pathway in rat asthma model.

[Comparison of integration processing technology of origin and traditional cutting processing technology of Moslae Herba for lung-Yang deficiency rats].

To compare the effect of integration processing technology of origin (IPTO) and traditional cutting processing technology (TCPT) of Moslae Herba for lung-Yang deficiency rats caused by complex factors, analyze the mechanism, and provide the modern pharmacology basis for the implementation of IPTO of Moslae Herba. The rat models of lung-Yang deficiency were established by smoking + swimming in ice water + drinking ice water. The model rats were randomly divided into different groups, and were treated with intragastric administration for 30 d. Then the general signs, anal temperature and autonomic activity of the rats were observed. The pathological morphology of lung tissues was observed, and the positive expression of tumor necrosis factor (TNF-α) was observed by immunohistochemical method, and the hematological indexes were determined. Enzyme linked immunosorbent assay (ELISA) method was used to detect serum nitric oxide (NO), immunoglobulin G (IGG), malondialdehyde (MDA), thromboxane B2 (TXB2) and interleukin-8 (IL-8) level, and the organ coefficients of heart, liver, spleen, lung, kidney and other organs were calculated. According to the results, Moslae Herba volatile oil and decoction could improve the general signs and autonomic activities of lung-Yang deficiency rats, improve the body weight, rectal temperature, and the content of IGG in serum of lung-Yang deficiency rats, reduce organ coefficients of heart, liver, spleen, lung and kidney, serum NO, MDA, TXB2, IL-8 contents, white blood cell and TNF-α mean optical density in the lung tissues of rats. witg statistically significant difference (P<0.01 or P<0.05). The effects of IPTO volatile oil and water decoction were slightly higher. Therefore, Moslae Herba has therapeutic effect on lung-Yang deficiency rats, and ICPT has better effect, whose mechanism may be related to the intervention of TNF-α expression, improving the level of IGG, and inhibiting NO, MDA, IL-8, and TXB2 levels.

Evaluation of the antithrombotic activity of Zhi-Xiong Capsules, a Traditional Chinese Medicinal formula, via the pathway of anti-coagulation, anti-platelet activation and anti-fibrinolysis.

Zhi-Xiong Capsules (ZXC) involving Hirudo, Ligusticum chuanxiong, Salvia miltiorrhiza, Leonurus artemisia, and Pueraria lobata, is an empirical prescription used in Chinese clinics applied for treating cerebral arteriosclerosis and blood-stasis in clinic. However, the mechanism of its antithrombotic activity has not been investigated until now. The present study was designed to investigate its antithrombotic effects, the mechanism of ZXC on anti-thrombus action and to identify the main chemical composition of ZXC using HPLC-DAD-ESI-IT-TOF-MS. Two animal models were used to evaluate the antithrombotic effect of ZXC, the arterial thrombosis model and a venous thrombosis model. ZXC prolonged the plasma recalcification time (PRT), the activated partial thromboplastin time (APTT), the thrombin time (TT) and the prothrombin time (PT) and clearly reduced the content of fibrinogen (FIB) obviously in the arterial thrombosis model. Furthermore, it markedly suppressed the level of TXB2 and up-regulated the level of 6-keto-PGF1a. In addition, it significantly up-regulated the level of t-PA and down-regulated the level of PAI-1 (p < 0.05). These results revealed that ZXC played a vital role in the prevention of thrombosis through interacting with multiple targets, including inhibition of coagulation and platelet aggregation and increasing thrombolysis. A total of 23 compounds were identified as the main components of ZXC by HPLC-DAD-ESI-IT TOF-MS.

Mirabegron, a ß3-adrenoceptor agonist reduced platelet aggregation through cyclic adenosine monophosphate accumulation.

Mirabegron is a ß3-adrenoceptor agonist and released on the marked for the treatment of overactive bladder. Because mirabegron is the only ß3-adrenoceptor agonist available and substances that increase the levels of cyclic adenosine monophosphate (cAMP) inhibit platelet activity, we tested the hypothesis that mirabegron could have antiplatelet activity. Collagen- and thrombin induced platelet aggregation, thromboxane B2 (TXB2) and cyclic nucleotides quantification and calcium (Ca2+) mobilization were determined in the absence and presence of mirabegron in human washed platelets. Our results revealed that mirabegron (10-300 µM) produced significant inhibitions on platelet aggregation induced by collagen- or thrombin, accompanied by greater intracellular levels of cAMP. The ß3-adrenoceptor antagonist L 748,337 (1 µM) and the adenylate cyclase inhibitor, SQ 22,536 (100 µM) reversed the inhibition induced by mirabegron in thrombin-stimulated platelets. The selective antagonists for ß1-and ß2-adrenoceptors, atenolol and ICI 117,551 (3 µM), respectively did not interfere on the inhibition induced by mirabegron. In Fluo-4 loaded platelets, mirabegron reduced the total and intracellular Ca2+ levels. Pre-incubation with mirabegron almost abolished the levels of TXB2. Mirabegron did not augment the intracellular levels of cyclic guanosine monophosphate. In conclusion, mirabegron inhibited human platelet aggregation through cAMP accumulation, thus suggesting that substances that activate ß3-adrenoceptor could be beneficial as adjuvant antiplatelet therapy.

Panax quinquefolius saponins combined with dual antiplatelet drug therapy alleviate gastric mucosal injury and thrombogenesis through the COX/PG pathway in a rat model of acute myocardial infarction.

OBJECTIVES: Previous studies have found that Panax quinquefolius saponins (PQS) combined with dual antiplatelet therapy (DAPT) of aspirin and clopidogrel enhances antithrombotic effects while reducing gastric mucosal injury induced by DAPT. We investigated the effects of the combined drug therapy (PQS+DAPT) through the COX/PG pathways. METHODS: Acute myocardial infarction (AMI) was induced in Wistar rats by ligation of the left anterior descending (LAD) coronary artery, and the animals were randomly divided into Model, DAPT, and PQS+DAPT groups. Rats in the sham group did not undergo artery ligation. They were intragastrically treated for 14 days. Myocardial infarct size; myocardial pathology; platelet aggregation rate, CD62p activation, concentrations of thromboxane B2 (TXB2), 6-keto-PGF1α, tissue plasminogen activator (t-PA), and plasminogen activator inhibitor (PAI), the TXB2/6-keto-PGF1α ratio were measured. The ultrastructure of the gastric mucosa was observed by scanning electron microscopy. The expression of PGE2 and 6-keto-PGF1α in gastric mucosa was measured by radioimmunoassay, and levels of COX-1, COX-2, and VEGF in gastric mucosa were assessed using immunohistochemistry. RESULTS: The addition of Panax quinquefolius saponins (PQS+DAPT) to standard DAPT therapy significantly decreased the myocardial infarct area, degree of myocardial lesions, TXB2 and PAI levels, and the TXB2/6-keto-PGF1α ratio, while increasing 6-keto-PGF1α and t-PA levels and reducing the degree of gastric mucosal injury. Expression of PGE2, 6-keto-PGF1α, COX-2, and VEGF in the gastric mucosa was upregulated in the PQS+DAPT group compared with the standard DAPT group. CONCLUSION: PQS increases the degree of DAPT inhibition of myocardial necrosis and antiplatelet effects in AMI rats, as well as reducing damage to the gastric mucosa caused by DAPT. The mechanism may be related to inhibition of TXB2 and PAI activity and elevation of 6-keto-PGF1α and t-PA levels in blood, and may be associated with upregulated expression of COX-2, PGE2, PGI2, and VEGF in gastric tissue.

Antithrombotic components of Malus halliana Koehne flowers.

Flowers of Malus halliana (M. halliana) Koehne have been used as a Chinese traditional medicine to treat metrorrhagia and in our study, its chemical composition and anticoagulant effect were investigated. Five compounds were isolated and identified from M. halliana flowers, including limocitrin-3-O-glucoside (1), baohuoside Ⅱ (2), kaempferol-3-O-α-L-furan arabinoside (3), phloretin-4'-O-glycosidase (4) and afzeloside (5). Compound 1-3 were isolated for the first time from this genus. The anticoagulant effect of the compounds and extracts of M. halliana flowers were evaluated by APTT, PT, TT and FIB on plasma of rabbit in vitro. The results indicated that several fractions of M. halliana flowers and compounds 2-5 exhibited anticoagulant activity in vitro. Subsequently, afzeloside (5), the abundant component in M. halliana flowers, was investigated further for its antithrombotic effect in vivo and its antithrombotic mechanisms were evaluated on rats acute blood-stasis model. The antithrombotic effect was evaluated by WBV, PV, HCT, ESR, APTT, PT, TT, FIB, 6-keto-PGF1α, TXB2, ET-1 and eNOS in vivo. Afzeloside demonstrated inhibitory effect of thrombus formation, and its underlying antithrombotic mechanism was found to be related to the regulation of vascular endothelium active substance, activating blood flow and anticoagulant effect. Hence, we postulate that flavonoids may be the active ingredients of the plant.

Anti- Versus Pro-Inflammatory Metabololipidome Upon Cupping Treatment.

BACKGROUND/AIMS: This study aimed to explore the metabololipidome in mice upon cupping treatment. METHODS: A nude mouse model mimicking the cupping treatment in humans was established by administrating four cupping sets on the back skin for 15 minutes. UPLC-MS/ MS was performed to determine the PUFA metabolome in mice skin and blood before and after cupping treatment. The significantly changed lipids were administered in macrophages to assess the production of pro-inflammatory cytokines IL-6 and TNF-α by ELISA. RESULTS: The anti-inflammatory lipids, e.g. PGE1, 5,6-EET, 14,15-EET, 10S,17S-DiHDoHE, 17R-RvD1, RvD5 and 14S-HDoHE were significantly increased while pro-inflammatory lipids, e.g. 12-HETE and TXB2 were deceased in the skin or plasma post cupping treatment. Cupping treatment reversed the LPS-stimulated IL-6 and TNF-α expression in mouse peritoneal exudates. Moreover, 5,6-EET, PGE1 decreased the level of TNF-α, while 5,6-EET, 5,6-DHET downregulated IL-6 production in macrophages. Importantly, 14,15-EET and 14S-HDoHE inhibited both IL-6 and TNF-α induced by lipopolysaccharide (LPS). 17-RvD1, RvD5 and PGE1 significantly reduced the LPS-initiated TNF-α, while TXB2 and 12-HETE further upregulated the LPS-enhanced IL-6 and TNF-α expression in macrophages. CONCLUSION: Our results reveal the identities of anti-inflammatory versus pro-inflammatory metabolipidome and suggest the potential therapeutic mechanism of cupping treatment.