In silico study of combination thrombolytic therapy with alteplase and mutant pro-urokinase for fibrinolysis in ischemic stroke.

The synergistic advantage of combining tissue plasminogen activator (tPA) with pro-urokinase (proUK) for thrombolysis has been demonstrated in several in vitro experiments, and a single site proUK mutant (m-proUK) has been developed for better stability in plasma. Based on these studies, combination thrombolytic therapy with intravenous tPA and m-proUK has been suggested as a promising treatment for patients with ischemic stroke. This paper evaluates the efficacy and safety of the dual therapy by computational simulations of pharmacokinetics and pharmacodynamics coupled with a local fibrinolysis model. Seven dose regimens are simulated and compared with the standard intravenous tPA monotherapy. Our simulation results provide more insights into the complementary reaction mechanisms of tPA and m-proUK during clot lysis and demonstrate that the dual therapy can achieve a similar recanalization time (about 50 min) to tPA monotherapy, while keeping the circulating fibrinogen level within a normal range. Specifically, our results show that for all dual therapies with a 5 mg tPA bolus, the plasma concentration of fibrinogen remains stable at around 7.5 µM after a slow depletion over 50 min, whereas a rapid depletion of circulating fibrinogen (to 5 µM) is observed with the standard tPA therapy, indicating the potential advantage of dual therapy in reducing the risk of intracranial hemorrhage. Through simulations of varying dose combinations, it has been found that increasing tPA bolus can significantly affect fibrinogen level but only moderately improves recanalization time. Conversely, m-proUK doses and infusion duration exhibit a mild impact on fibrinogen level but significantly affect recanalization time. Therefore, future optimization of dose regimen should focus on limiting the tPA bolus while adjusting m-proUK dosage and infusion rate. Such adjustments could potentially maximize the therapeutic advantages of this combination therapy for ischemic stroke treatment.

Stimulation of platelet P2Y1 receptors by different endogenous nucleotides leads to functional selectivity via biased signalling.

BACKGROUND AND PURPOSE: Platelet function during inflammation is dependent on activation by endogenous nucleotides. Non-canonical signalling via the P2Y1 receptor is important for these non-thrombotic functions of platelets. However, apart from ADP, the role of other endogenous nucleotides acting as agonists at P2Y1 receptors is unknown. This study compared the effects of ADP, Ap3A, NAD+ , ADP-ribose, and Up4A on platelet functions contributing to inflammation or haemostasis. EXPERIMENTAL APPROACH: Platelets obtained from healthy human volunteers were incubated with ADP, Ap3A, NAD+ , ADP-ribose, or Up4A, with aggregation and fibrinogen binding measured (examples of function during haemostasis) or before exposure to fMLP to measure platelet chemotaxis (an inflammatory function). In silico molecular docking of these nucleotides to the binding pocket of P2Y1 receptors was then assessed. KEY RESULTS: Platelet aggregation and binding to fibrinogen induced by ADP was not mimicked by NAD+ , ADP-ribose, and Up4A. However, these endogenous nucleotides induced P2Y1 -dependent platelet chemotaxis, an effect that required RhoA and Rac-1 activity, but not canonical PLC activity. Analysis of molecular docking of the P2Y1 receptor revealed distinct differences of amino acid interactions and depth of fit within the binding pocket for Ap3A, NAD+ , ADP-ribose, or Up4A compared with ADP. CONCLUSION AND IMPLICATIONS: Platelet function (aggregation vs motility) can be differentially modulated by biased-agonist activation of P2Y1 receptors. This may be due to the character of the ligand-binding pocket interaction. This has implications for future therapeutic strategies aimed to suppress platelet activation during inflammation without affecting haemostasis as is the requirement of current ant-platelet drugs. LINKED ARTICLES: This article is part of a themed issue on Platelet purinergic receptor and non-thrombotic disease. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v181.4/issuetoc.

Characterization and Usefulness of Clot-Fibrinolysis Waveform Analysis in Critical Care Patients with Enhanced or Suppressed Fibrinolysis.

INTRODUCTION: Recently, clot-fibrinolysis waveform analysis (CFWA), which is a coagulation and fibrinolysis global assay based on assessing the activated partial thromboplastin time with tissue-type plasminogen activator, was developed. This study aimed to investigate the characteristics of CFWA using plasma samples from patients in the critical care unit. MATERIALS AND METHODS: The fibrinolysis times using CFWA were measured in 298 plasma samples. These samples were divided into three groups based on the reference interval (RI) of fibrinolysis time using CFWA: shortened group, less than RI; within group, within RI; prolonged group, more than RI. The coagulation and fibrinolysis markers, including D-dimer, plasmin-α2 plasmin inhibitor complex (PIC), fibrin monomer complex (FMC), plasmin-α2 plasmin inhibitor (α2-PI), plasminogen (Plg), and fibrinogen (Fbg) were analyzed and compared among the three groups. RESULTS: The FMC level decreased in the order of shortened, within, and prolonged groups, and the decrease was statistically significant among all three group pairs. The opposite tendency was observed for Fbg and fibrinolysis-related markers of α2-PI and Plg, and significant differences were recognized in all pair comparisons except for between within and prolonged groups in Plg. The mean values of the fibrinolysis markers D-dimer and PIC in all three groups were higher than the cut-off values, and the PIC value differed significantly between the within and prolonged groups. CONCLUSION: The fibrinolysis reaction was detected in all three groups, but the status differed. CFWA has the potential to reflect the fibrinolysis status in one global assay.

Advances in tumor immunomodulation based on nanodrug delivery systems.

Immunotherapy is a therapeutic approach that employs immunological principles and techniques to enhance and amplify the body's immune response, thereby eradicating tumor cells. Immunotherapy has demonstrated effective antitumor effects on a variety of malignant tumors. However, when applied to humans, many immunotherapy drugs fail to target lesions with precision, leading to an array of adverse immune-related reactions that profoundly limit the clinical application of immunotherapy. Nanodrug delivery systems enable the precise delivery of immunotherapeutic drugs to targeted tissues or specific immune cells, enhancing the immune antitumor effect while reducing the number of adverse reactions. A nanodrug delivery system provides a feasible strategy for activating the antitumor immune response by the following mechanisms: 1) increased targeting and uptake of vaccines by DCs, which enhances the efficacy of the immune response; 2) increased tumor cell immunogenicity; 3) regulation of TAMs and other cells by, for example, regulating the polarization of TAMs and interfering with TAN formation, and ECM remodeling by CAFs; and 4) interference with tumor immune escape signaling pathways, namely, the PD-1/PD-L1, FGL1/LAG-3 and IDO signaling pathways. This paper reviews the progress of nanodrug delivery system research with respect to tumor immunotherapy based on tumor immunomodulation over the last few years, discussing the promising future of these delivery systems under this domain.

The mechanism of oleic acid inhibiting platelet activation stimulated by collagen.

BACKGROUND: Abnormal platelet activation is a key factor in the occurrence and development of thrombotic diseases. However, the physiological mechanisms that underlie platelet homeostasis remain unclear. Oleic acid, one of the most abundant lipids in the human diet, has potential antithrombotic effects. This study aimed to investigate the effects of oleic acid on platelet activation and thrombosis. METHODS: Platelet aggregation, ATP release, and fibrinogen spread were evaluated to determine the role of oleic acid in platelet activation. A ferric chloride-induced carotid injury model was used to establish the effect of oleic acid on thrombus formation in vivo. Western blotting analysis and transfection experiments were performed to determine the mechanisms involved in this process. RESULTS: Oleic acid inhibited platelet aggregation, granule release, and calcium mobilization. Furthermore, it inhibited the spread of platelets on fibrinogen. We also found that oleic acid delayed arterial thrombosis in mice, as demonstrated in a murine model of ferric chloride-induced carotid artery thrombosis. The molecular mechanism of its inhibition of platelet activity may be through the Syk-PLCγ2 and CaMKKß/AMPKα/VASP pathways. In addition, we demonstrated that the phosphorylation of AMPK at Ser496 was an important mechanism of platelet activation. CONCLUSIONS: Our study showed that oleic acid inhibits platelet activation and reduces thrombogenesis by inhibiting the phosphorylation of multiple signaling molecules, offering new insights into the research and development of antiplatelet drugs. Video Abstract.

Pathogenic Role of Fibrinogen in the Neuropathology of Multiple Sclerosis: A Tale of Sorrows and Fears.

Multiple sclerosis (MS) is an autoimmune demyelinating neurodegenerative disease of the central nervous system (CNS) due to injury of the myelin sheath by immune cells. The clotting factor fibrinogen is involved in the pathogenesis of MS by triggering microglia and the progress of neuroinflammation. Fibrinogen level is correlated with MS severity; consequently, inhibition of the fibrinogen cascade may reduce MS neuropathology. Thus, this review aimed to clarify the potential role of fibrinogen in the pathogenesis of MS and how targeting of fibrinogen affects MS neuropathology. Accumulation of fibrinogen in the CNS may occur independently or due to disruption of blood-brain barrier (BBB) integrity in MS. Fibrinogen acts as transduction and increases microglia activation which induces the progression of inflammation, oxidative stress, and neuronal injury. Besides, brain fibrinogen impairs the remyelination process by inhibiting the differentiation of oligodendrocyte precursor cells. These findings proposed that fibrinogen is associated with MS neuropathology through interruption of BBB integrity, induction of neuroinflammation, and demyelination with inhibition of the remyelination process by suppressing oligodendrocytes. Therefore, targeting of fibrinogen and/or CD11b/CD18 receptors by metformin and statins might decrease MS neuropathology. In conclusion, inhibiting the expression of CD11b/CD18 receptors by metformin and statins may decrease the pro-inflammatory effect of fibrinogen on microglia which is involved in the progression of MS.

Slow Intravenous Infusion of a Novel Damage Control Cocktail Decreases Blood Loss in a Pig Polytrauma Model.

BACKGROUND: Our objective was to optimize a novel damage control resuscitation (DCR) cocktail composed of hydroxyethyl starch, vasopressin, and fibrinogen concentrate for the polytraumatized casualty. We hypothesized that slow intravenous infusion of the DCR cocktail in a pig polytrauma model would decrease internal hemorrhage and improve survival compared with bolus administration. METHODS: We induced polytrauma, including traumatic brain injury (TBI), femoral fracture, hemorrhagic shock, and free bleeding from aortic tear injury, in 18 farm pigs. The DCR cocktail consisted of 6% hydroxyethyl starch in Ringer's lactate solution (14mL/kg), vasopressin (0.8U/kg), and fibrinogen concentrate (100mg/kg) in a total fluid volume of 20mL/kg that was either divided in half and given as two boluses separated by 30 minutes as control or given as a continuous slow infusion over 60 minutes. Nine animals were studied per group and monitored for up to 3 hours. Outcomes included internal blood loss, survival, hemodynamics, lactate concentration, and organ blood flow obtained by colored microsphere injection. RESULTS: Mean internal blood loss was significantly decreased by 11.1mL/kg with infusion compared with the bolus group (p = .038). Survival to 3 hours was 80% with infusion and 40% with bolus, which was not statistically different (Kaplan Meier log-rank test, p = .17). Overall blood pressure was increased (p < .001), and blood lactate concentration was decreased (p < .001) with infusion compared with bolus. There were no differences in organ blood flow (p > .09). CONCLUSION: Controlled infusion of a novel DCR cocktail decreased hemorrhage and improved resuscitation in this polytrauma model compared with bolus. The rate of infusion of intravenous fluids should be considered as an important aspect of DCR.

Study on the Mechanism of Allergic Rhinitis Based on the Expression of FIB, PCT, hs-CRP, and Th17/Treg-IL10/IL-17 Axis Balance.

BACKGROUND: The pathogenesis of allergic rhinitis (AR) is ambiguous, while it is clear that various immune cells and cytokines play crucial roles in its occurrence and development. AIM: To investigate the effect of exogenous interleukin-10 (IL-10) on the expression of fibrinogen (FIB), procalcitonin (PCT), hypersensitive C-reactive protein (hs-CRP), and Th17/Treg-IL10/IL-17 axis balance in the nasal mucosa of rats with AR. METHOD: In this study, 48 female-specific pathogen-free Sprague-Dawley rats were randomly divided into 3 groups: blank control group, AR group, and IL-10 intervention group. The AR model was established in the AR group and IL-10 group. The rats in the control group were treated with normal saline; the rats in the AR group were given 20 µL of saline containing 50 µg of ovalbumin (OVA) every day. The rats in the IL-10 intervention group were intraperitoneally injected with 1 mL of 40 pg/kg IL-10 and provided with OVA. The IL-10 intervention group was composed of mice with AR that received IL-10. The behavior of nasal allergic symptoms (such as nasal itching, sneezing, and runny nose) and the hematoxylin and eosin staining of nasal mucosa were observed. The levels of FIB, PCT, hs-CRP, IgE, and OVA sIgE in serum were determined by enzyme-linked immunosorbent assay. The levels of Treg and Th17 cells in serum were detected by flow cytometry. The protein levels of TGF-ß, IL-10, and IL-17 in nasal mucosa were detected by the Western-blot method. RESULTS: The scores of snots, nasal itching, and sneezing in the AR group were significantly higher than those in the control group, while the scores of the above symptoms in the IL-10 intervention group were lower than those in the AR group. The levels of FIB, PCT, hs-CRP, IgE, and OVA sIgE in serum and the protein levels of IL-10 and IL-17 in the nasal mucosa in the AR group were higher than those in the blank control group. Meanwhile, the levels of FIB, PCT, hs-CRP, IgE, and OVA sIgE in serum and IL-10 and IL-17 protein in the nasal mucosa in the IL-10 group were lower than those in the AR group. CONCLUSION: IL-10 can relieve the allergy of AR rats by affecting the expression of FIB, PCT, and hs-CRP, as well as the balance of the Th17/Treg-IL10/IL-17 axis in the nasal mucosa of AR rats.

In vitro study on the effect of fibrinogen γ-chain peptide-coated ADP-encapsulated liposomes on postcardiopulmonary bypass coagulopathy using patient blood.

BACKGROUND: Fibrinogen γ-chain peptide-coated, adenosine 5'-diphosphate (ADP)-encapsulated liposomes (H12-ADP-liposomes) are potent hemostatic adjuvants that promote platelet thrombi formation at bleeding sites. Although we have reported the efficacy of these liposomes in a rabbit model of cardiopulmonary bypass coagulopathy, we are yet to address the possibility of their hypercoagulative potential, especially in human beings. OBJECTIVES: Considering its future clinical applications, we herein investigated the safety of using H12-ADP-liposomes in vitro using blood samples from patients who had received platelet transfusion after cardiopulmonary bypass surgeries. METHODS: Ten patients receiving platelet transfusions after cardiopulmonary bypass surgery were enrolled. Blood samples were collected at the following 3 points: at the time of incision, at the end of the cardiopulmonary bypass, and immediately after platelet transfusion. After incubating the samples with H12-ADP-liposomes or phosphate-buffered saline (PBS, as a control), blood coagulation, platelet activation, and platelet-leukocyte aggregate formation were evaluated. RESULTS: Patients' blood incubated with H12-ADP-liposomes did not differ from that incubated with PBS in coagulation ability, degree of platelet activation, and platelet-leukocyte aggregation at any of the time points. CONCLUSION: H12-ADP-liposomes did not cause abnormal coagulation, platelet activation, or platelet-leukocyte aggregation in the blood of patients who received platelet transfusion after a cardiopulmonary bypass. These results suggest that H12-ADP-liposomes could likely be safely used in these patients, providing hemostasis at the bleeding sites without causing considerable adverse reactions. Future studies are needed to ensure robust safety in human beings.

In Vitro Staphylococcal Aggregate Morphology and Protection from Antibiotics Are Dependent on Distinct Mechanisms Arising from Postsurgical Joint Components and Fluid Motion.

Considerable progress has been made toward elucidating the mechanism of Staphylococcus aureus aggregation in synovial fluid. In this study, aggregate morphology was assessed following incubation under several simulated postsurgical joint conditions. Using fluorescently labeled synovial fluid polymers, we show that aggregation occurs through two distinct mechanisms: (i) direct bridging between S. aureus cells and host fibrinogen and (ii) an entropy-driven depletion mechanism facilitated by hyaluronic acid and albumin. By screening surface adhesin-deficient mutants (clfA, clfB, fnbB, and fnbA), we identified the primary genetic determinant of aggregation in synovial fluid to be clumping factor A. To characterize this bridging interaction, we employed an atomic force microscopy-based approach to quantify the binding affinity of either wild-type S. aureus or the adhesin mutant to immobilized fibrinogen. Surprisingly, we found there to be cell-to-cell variability in the binding strength of the bacteria for immobilized fibrinogen. Superhigh-resolution microscopy imaging revealed that fibrinogen binding to the cell wall is heterogeneously distributed at both the single cell and population levels. Finally, we assessed the antibiotic tolerance of various aggregate morphologies arising from newly deciphered mechanisms of polymer-mediated synovial fluid-induced aggregation. The formation of macroscopic aggregates under shear was highly tolerant of gentamicin, while smaller aggregates, formed under static conditions, were susceptible. We hypothesize that aggregate formation in the joint cavity, in combination with shear, is mediated by both polymer-mediated aggregation mechanisms, with depletion forces enhancing the stability of essential bridging interactions. IMPORTANCE The formation of a bacterial biofilm in the postsurgical joint environment significantly complicates the resolution of an infection. To form a resilient biofilm, incoming bacteria must first survive the initial invasion of the joint space. We previously found that synovial fluid induces the formation of Staphylococcus aureus aggregates, which may provide rapid protection during the early stages of infection. The state of the host joint environment, including the presence of fluid flow and fluctuating abundance of synovial fluid polymers, determines the rate and size of aggregate formation. By expanding on our knowledge of the mechanism and pathogenic implications of synovial fluid-induced aggregation, we hope to contribute insights for the development of novel methods of prevention and therapeutic intervention.

Physiological correction of hereditary mild hypofibrinogenemia during pregnancy.

INTRODUCTION: Hereditary hypofibrinogenemia is a rare fibrinogen disorder characterised by decreased levels of fibrinogen. Pregnant women with hypofibrinogenemia are at risk of adverse obstetrical outcomes, depending on the fibrinogen level. AIM: We investigated how the physiological changes of hemostasis throughout the pregnancy impact the hemostatic balance in a woman with hereditary mild hypofibrinogenemia. METHODS: Fibrin clot properties were analyzed by turbidimetry and scanning electron microscopy, clot weight and red blood cells retention were measured by whole clot contraction, and in vitro thrombin generation was assessed by calibrated automated thrombogram and ex vivo by TAT. RESULTS: Throughout the pregnancy, the fibrinogen levels increased reaching normal values in the third trimester (activity 3.1 g/L, antigen 3.2 g/L). In parallel, the fibrin polymerisation increased, the fibrinolysis decreased, the fibrin clot network became denser with thicker fibrin fibers, and the fibrin clot weight and red blood cells retention increased, reaching control's value at the third trimester. Similarly, in vitro and ex vitro thrombin generation increased, reaching maximum values at the delivery. CONCLUSION: In this case of hereditary mild hypofibrinogenemia we observed a physiological increase of fibrinogen and thrombin generation. Future studies should focus on moderate and severe hypofibrinogenemia, to assess fibrinogen variation and the overall impact of increased TG on the hemostasis balance.

The impact of high on-treatment platelet reactivity and fibrinogen levels on ischemic events in patients with ST elevation myocardial infarction: a prospective observational study.

BACKGROUND: After treatment, high residual platelet reactivity (HRPR) is considered as an essential risk factor for recurrent ischemic events. AIM: To evaluate the impact of fibrinogen on HRPR after implantation of emergency drug-eluting stents (DES) in patients treated with aspirin and clopidogrel or ticagrelor due to ST-elevation myocardial infarction (STEMI) and to explore the predictive values of HRPR and fibrinogen for adverse ischemic events at 12 months. METHOD: This single-center prospective observational study analyzed patients with STEMI who underwent primary percutaneous coronary intervention (PCI) with second-generation DES implantation from January 2017 to December 2018. Platelet reactivity was measured by thromboelastography (TEG) at 60-72 h after primary PCI. HRPR was defined as the adenosine diphosphate-induced maximum amplitude (MAADP) > 47 mm. RESULTS: A total of 919 patients were analyzed, of which 512 (55.8%) received aspirin and clopidogrel and 406 (44.2%) received aspirin and ticagrelor. Elevated fibrinogen levels were associated with an increased prevalence of HRPR (P < 0.001). High fibrinogen (quartile IV, ≥ 410 mg/dL) was an independent risk factor for HRPR after multivariate regression (odds ratio 6.556, 95% confidence interval [CI]: 3.200-13.431, P < 0.001). When analyzed by Kaplan-Meier survival curves, the combination of high fibrinogen and HRPR was strongly predictive for ischemic major adverse cardiac events at 12 months compared to the group without HRPR and with low fibrinogen (hazard ratio 9.681, 95% CI: 4.467-20.98, log-rank P < 0.001). Similar results were confirmed in subgroups according to different dual antiplatelet therapies. CONCLUSION: A combination of high fibrinogen and HRPR may identify recurrent adverse ischemic events over 12 months. Ticagrelor exhibited more potent platelet inhibition and a better prognosis than clopidogrel.

A comparative high-resolution physicochemical analysis of commercially available fibrin sealants: Impact of sealant osmolality on biological performance.

Fibrin sealants are well-established components of the surgical toolbox, especially in procedures that harbor a high risk of perioperative bleeding. Their widespread use as hemostats, sealants or tissue-adhesives in various surgical settings has shown that the choice of the appropriate sealant system affects the clinical outcome. While many studies have compared the hemostatic efficiency of fibrin sealants to that of other natural or synthetic sealants, there is still limited data on how subtle differences in fibrin sealant formulations relate to their biological performance. Here, we performed an in-depth physicochemical and biological characterization of the two most commonly used fibrin sealants in the US and Europe: TISSEEL™ ("FS") and VISTASEAL™/VERASEAL™ ("FS+Osm"). Our chemical analyses demonstrated differences between the two sealants, with lower fibrinogen concentrations and supraphysiological osmolality in the FS+Osm formulation. Rheological testing revealed FS clots have greater clot stiffness, which strongly correlated with network density. Ultrastructural analysis by scanning electron microscopy revealed differences between FS and FS+Osm fibrin networks, the latter characterized by a largely amorphous hydrogel structure in contrast to the physiological fibrillar network of FS. Cytocompatibility experiments with human fibroblasts seeded on FS and FS+Osm fibrin networks, or cultured in presence of sealant extracts, revealed that FS+Osm induced apoptosis, which was not observed with FS. Although differential sealant osmolality and amounts of fibrinogen, as well as the presence of Factor XIII or additives such as antifibrinolytics, may explain the mechanical and structural differences observed between the two fibrin sealants, none of these substances are known to cause apoptosis at the respective concentrations in the sealant formulation. We thus conclude that hyper osmolality in the FS+Osm formulation is the primary trigger of apoptosis-a mechanism that should be evaluated in more detail, as it may affect the cellular wound healing response in situ.

Beyond uterine atony: characterizing postpartum hemorrhage coagulopathy.

BACKGROUND: Postpartum hemorrhage is a leading cause of morbidity and mortality worldwide, yet the associated early coagulopathy is not well defined. OBJECTIVE: We hypothesized that women who develop postpartum hemorrhage have a distinct derangement of thrombin generation and coagulation factors compared with postpartum women without postpartum hemorrhage. STUDY DESIGN: This prospective study of pregnant patients with postpartum hemorrhage was completed at a single urban hospital. Blood was drawn on postpartum hemorrhage diagnosis and 2 and 4 hours later. Assays of patients with postpartum hemorrhage included thrombelastography, whole blood thrombin generation, coagulation factor activity, tissue factor levels and activity, and tissue factor pathway inhibitor levels, which were compared with that of patients without postpartum hemorrhage. RESULTS: A total of 81 patients were included in this study. Of those patients, 66 had postpartum hemorrhage, and 15 served as controls. Compared with patients without PPH, patients with postpartum hemorrhage had lower fibrinogen levels (469.0 mg/dL vs 411.0 mg/dL; P=.02), increased tissue plasminogen activator resistance (fibrinolysis 30 minutes after maximal clot strength: 8.7% vs 4.2%; P=.02), decreased peak thrombin concentration (150.2 nM vs 40.7 nM; P=.01), and decreased maximal rate of thrombin generation (60.1 nM/minute vs 2.8 nM/minute; P=.02). Furthermore, compared with patients without postpartum hemorrhage, patients with postpartum hemorrhage had decreased tissue factor levels (444.3 pg/mL vs 267.1 pg/mL; P=.02) and increased tissue factor pathway inhibitor levels (0.6 U/mL vs 0.8 U/mL; P=.04), with decreased tissue factor pathway inhibitor ratios (624 vs 299; P=.01). CONCLUSION: PPH is not only an issue of uterine tone and mechanical bleeding but also a distinct coagulopathy that is characterized by decreased fibrinogen level, clot breakdown resistance, and markedly low thrombin generation. This pathology seemed to be driven by low tissue factor and high tissue factor pathway inhibitor levels.

Hematotoxicity of Co-Administration of Bisphenol A and Acetaminophen in Rats and its Amelioration by Melatonin.

BACKGROUND: Hematotoxicity is an underexplored endpoint of toxicity in most of the chemical exposures. An adverse effect on the hematological system arising out of xenobiotic exposure causes impaired hemostasis and coagulation leading to disease. BPA and acetaminophen are widely used synthetic chemicals worldwide and both are known and have numerous toxic effects. Since both can be simultaneously exposed to humans over a period of time, we hypothesized that their exposure can cause hematotoxicity, which may be ameliorated by melatonin. OBJECTIVE: In the current study, we aimed to find the effect of single and co-treatment of bisphenol A and acetaminophen on the RBC and coagulation factors in rats, and amelioration of impairment by melatonin. METHODS: Oxidative stress in red blood cells, bleeding time, blood clotting time, prothrombin time, partial thromboplastin time, and fibrinogen levels were assessed as indicators of hematotoxicity with single and co-exposure to bisphenol A and acetaminophen in rats. The effect of melatonin as a hemato-protective agent was assessed in the co-exposure. RESULTS: An increase in RBC oxidative stress and decrease in bleeding time, blood clotting time, prothrombin time, and partial thromboplastin time along with an increase in fibrinogen levels were observed with bisphenol A and acetaminophen treatment, which were further aggravated with cotreatment of the two. Melatonin treatment, however, was seen to decrease the increase in oxidative stress and ameliorate the impairment in coagulation factors. CONCLUSION: Bisphenol A and acetaminophen cause an increase in the oxidative stress in the red blood cells, and cause a shift toward pro-coagulation, which is alleviated by treatment with melatonin.

The role of fibrinogen in traumatic brain injury: from molecular pathological mechanisms to clinical management.

Fibrinogen is the substrate of plasma coagulation. It plays an important role in the formation of reticular network, which is crucial to the strength and stability of blood clots. In addition to directly participating in coagulation, fibrinogen also participates in the destruction of blood-brain barrier and neuroinflammation. This article reviews the pathophysiological changes of fibrinogen after traumatic brain injury. Considerable efforts have been made to understand the mechanisms by which fibrinogen damages the central nervous system. Combined with the latest research hotspots, potentially promising treatment strategies at the molecular level were discussed. We believe that understanding the role of fibrinogen-mediated damage in nerve and blood-brain barrier function will enable timely intervention in patients with nerve damage, and guide the development of novel targeted therapeutics.

AIF-1, a potential biomarker of aggressive tumor behavior in patients with non-small cell lung cancer.

Allogeneic inflammatory factor-1 (AIF-1) overexpression has been reported to be associated with tumorigenesis and tumor metastasis. This study aimed to investigate the role of AIF-1 in the development and progression of non-small cell lung cancer (NSCLC). AIF-1, IL-6, and VEGF expressions in human NSCLC tissue were examined by immunofluorescence staining. Bioinformatics analyses were performed to identify AIF-1-related molecules and pathways in NSCLC. Human lung cancer A549 cell proliferation was assessed by CCK-8 assay, and cell migration was evaluated with wound-healing assay. IL-6 and VEGF secretions in A549 cell culture supernatants were quantified using the Elecsys IL-6 immunoassay kit and Vascular Endothelial Growth Factor Assay Kit. RT-PCR and western blot were performed to quantify the expressions of AIF-1, IL-6, and VEGF mRNAs and proteins involved in p38-MAPK and JAK/STAT3 signaling such as p-p38 and p-STAT3. The effects of AIF-1 on A549 cell proliferation and the expressions of IL-6 and VEGF were assessed using SB203580 and ruxolitinib. The results showed that AIF-1 expression was higher in human NSCLC tissue than that in paracancer tissue. High AIF-1 expression was associated with metastasis, higher TNM stage, and poorer survival. Bioinformatics connected AIF-1 to JAK/STAT signaling in NSCLC. AIF-1 increased A549 cell proliferation, migration, IL-6 secretion and, VEGF secretion, and these effects were attenuated by inhibition of p38-MAPK or JAK/STAT3 signaling. In conclusion, AIF-1 may promote aggressive NSCLC behavior via activation of p38-MAPK and JAK/STAT signaling.

Fibrinolytic Changes in Women with Preeclampsia.

OBJECTIVES: Preeclampsia (PE) is a serious complication of pregnancy. The fibrinolytic system play crucial roles regarding placentation and evolution of PE. AIM: To study comprehensively components of the fibrinolytic system and fibrin lysability in women with PE. DESIGN AND METHODS: 117 women with PE and matched controls were included. Tissue type plasminogen activator (t-PA), plasminogen, PAI-1, plasmin inhibitor (PI), D-dimer, the fibrinolytic potential of dextran sulphate euglobulin fraction (DEF), PAI-2, polymere PAI-2, fibrin clot lysability, thrombin activatable fibrinolysis inhibitor (TAFI) and fibrinogen were assessed. RESULTS: Women with PE had significantly increased concentrations of t-PA and PAI-1, whereas the plasma concentration of PAI-2 was significantly lower compared to controls, p < 0.0001. Polymere PAI-2 was detected in both groups. DEF, TAFI and fibrinogen were not different between the groups. D-dimer was significantly increased and plasminogen/PI together with fibrin clot lysability time decreased in the PE-group, p = 0.0004 p = 0.04, p = 0.03, p < 0.0001 respectively. CONCLUSION: This study demonstrates that PE is associated with an affected t-PA/PAI-1 system, decreased PAI-2 and increased fibrin lysability. Furthermore, PAI-2 has the potential to polymerize during pregnancy.

The combination of EGCG with warfarin reduces deep vein thrombosis in rabbits through modulating HIF-1α and VEGF via the PI3K/AKT and ERK1/2 signaling pathways.

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.

Effects of berberine hydrochloride on immune response in the crab Charybdis japonica.

Berberine hydrochloride is the main effective component of Coptis spp. used in Chinese herbal medicine and its underlying molecular mechanisms, responsible for inducing effects in crustacean species, are not fully understood. In this study, the molecular response of the crab Charybdis japonica to berberine hydrochloride exposure was studied using transcriptome sequencing. The survival rate, gene expression and activities of several immune enzymes were measured after berberine hydrochloride treatments, with or without injection of the pathogenic bacterium Aeromonas hydrophila. A total of 962 differentially expressed genes (464 up-regulated and 498 down-regulated) were observed during exposure to 100 mg/L of berberine hydrochloride and in the control group after 48 h. Enrichment analysis revealed that these genes are involved in metabolism, cellular processes, signal transduction and immune functions, indicating that exposure to berberine hydrochloride activated the immune complement system. This bioactive compound simultaneously activated fibrinogen beta (FGB), fibrinogen alpha (FGA), alpha-2-macroglobulin (A2M), kininogen (KNG), fibrinogen gamma chain (FGB), alpha-2-HS-glycoprotein (AHSG), caspase-8 (CASP8), cathepsin L (CTSL), adenylate cyclase 3 (Adcy3) and MMP1. Its action could significantly increase the survival rate of the crabs injected with A. hydrophila and promote the activity of LZM, Caspas8, FGA, ACP and AKP in the hepatopancreas. When A. hydrophila was added, the neutralization of 300 mg/L berberine hydrochloride maximized the activities of Caspas8, LZM, ACP and AKP. Our results provide a new understanding of the potential effects of berberine hydrochloride on the immune system mechanisms in crustaceans.