Enhanced Blood Coagulation Research Articles

Amphiphilic Janus cotton gauze with enhanced moisture management and blood coagulation for rapid hemostasis and wound healing

Cotton gauze is commonly used in initial emergency care. However, its high hydrophilicity and limited clotting capacity can lead to the excessive absorption of blood, resulting in unnecessary blood loss. Herein, an amphiphilic Janus cotton gauze with excellent moisture management and enhanced blood coagulation has been developed via in situ generating bioactive glass (BG) onto the cotton gauze (CG), and then attaching cardanol (CA) onto one side of the BG-loaded CG (CG@BG) via click reaction. The Janus gauze (CA-CG@BG) has asymmetric wetting properties with a hydrophilic side (CA-CG@BGHL) and a hydrophobic side (HBCA-CG@BG). When applied to hemostatic, the porous and active BG on CA-CG@BGHL can rapidly initiate coagulation cascade to form a robust thrombus. CA on HBCA-CG@BG can entangled with each other, creating a hydrophobic barrier that prevents blood from flowing out. The hemostatic performance of CA-CG@BG is superior to that of CG in both rats and pigs. Interestingly, CA-CG@BG possesses unidirectional exudate removal. When applied to wound healing, the exudate can penetrate the hydrophobic HBCA-CG@BG to the hydrophilic CA-CG@BGHL, resulting in faster wound healing than CG. CA-CG@BG exhibits excellent cytocompatibility and hemocompatibility. This unique Janus dressing shows promise as a potential material for clinical applications in the future.

Facile Synthesis of Multifunctional Mesoporous Starch-Based Microparticle for Effective Hemostasis and Wound Healing.

Uncontrolled hemorrhage and infection are the principal causes of mortality associated with trauma in both military and civilian medical settings. Modified starch granules have emerged as a safe hemostatic agent for irregular and noncompressible wounds, but their performance is constrained by limited hemostasis efficiency and modest antibacterial activity. This study reported a directed self-assembly approach for a multifunctional mesoporous starch-based microparticle loaded with chitosan and calcium ions (Ca@MSMP) used for rapid hemostasis and wound healing. Directed self-assembly of uniform Ca@MSMP with a hierarchical hollow structure in the presence of chitosan was confirmed by scanning electron microscopy (SEM) analysis and pore structure analysis. The resulting Ca@MSMP exhibited a well-defined spherical shape and uniform size of 1 μm and demonstrated excellent antibacterial activity (>95%) without hemolytic activity. Importantly, Ca@MSMP enhanced blood coagulation and platelet aggregation via the synergistic effect of rapid calcium release and chitosan-mediated electrostatic interactions, leading to a significant decrease in blood loss and reduction in hemostasis time in rat tail amputation and liver injury models. In comparative analyses, Ca@MSMP significantly outperformed the commercial hemostatic agent Quickclean, notably enhancing the healing of full-thickness skin wounds in vivo by effectively preventing infection. These results underscore the potential of this innovative hemostatic material in diverse clinical scenarios, offering effective solutions for the management of bleeding in wounds that are irregularly shaped and noncompressible.

Shape memory and hemostatic silk-laponite scaffold for alveolar bone regeneration after tooth extraction trauma

Persistent bleeding and the absence of alveolar bone stress following tooth loss can hinder socket healing, complicating future dental implant procedures, and potentially leading to neighboring tooth instability. Therefore, developing materials that promote alveolar bone regeneration and possess both hemostatic and osteogenic properties is crucial for preserving the extraction sites. This study introduces a silk-based laponite composite scaffold material with proficient hemostatic and osteogenic functions, and excellent shape-memory properties for efficient extraction- site filling. In vitro studies research demonstrated that the scaffold's inherent negative charge of the scaffold significantly enhanced blood coagulation and thrombin generation. Moreover, its porous structure and slightly rough inner surface promoted blood cell adhesion and, improved the hemostatic performance. Furthermore, the scaffold facilitated stem cell osteogenic differentiation by activating the TRPM7 channel through the released of magnesium ions. In vivo tests using rat models confirmed its effectiveness in promoting coagulation and mandibular regeneration. Thus, this study proposes a promising approach for post-extraction alveolar bone regenerative repair. The composite scaffold material, with its hemostatic and osteogenic capabilities and shape-memory features, can potentially enhance dental implant success and overall oral health.

Electrospun Food Polysaccharides Loaded with Bioactive Compounds: Fabrication, Release, and Applications.

Food polysaccharides are well acclaimed in the field of delivery systems due to their natural safety, biocompatibility with the human body, and capability of incorporating/releasing various bioactive compounds. Electrospinning, a straightforward atomization technique that has been attracting researchers worldwide, is also versatile for coupling food polysaccharides and bioactive compounds. In this review, several popular food polysaccharides including starch, cyclodextrin, chitosan, alginate, and hyaluronic acid are selected to discuss their basic characteristics, electrospinning conditions, bioactive compound release characteristics, and more. Data revealed that the selected polysaccharides are capable of releasing bioactive compounds from as rapidly as 5 s to as prolonged as 15 days. In addition, a series of frequently studied physical/chemical/biomedical applications utilizing electrospun food polysaccharides with bioactive compounds are also selected and discussed. These promising applications include but are not limited to active packaging with 4-log reduction against E. coli, L. innocua, and S. aureus; removal of 95% of particulate matter (PM) 2.5 and volatile organic compounds (VOCs); heavy metal ion removal; increasing enzyme heat/pH stability; wound healing acceleration and enhanced blood coagulation, etc. The broad potentials of electrospun food polysaccharides loaded with bioactive compounds are demonstrated in this review.

Physiological and Biochemical Characteristics of Rainbow Trout with Severe, Moderate and Asymptomatic Course of Vibrio anguillarum Infection.

Simple SummaryDuring the past decades, bacterial infections have been a serious problem in aquaculture that causes very large economic losses. Currently, antibiotics are the most common method of disease prevention and control. A combination of water quality monitoring, early detection of fish infections, and other preventive biosecurity measures in fish farms can help prevent the spread of infection. We investigated the natural bacterial infection in fish farms and characterized the parameters of the health status of rainbow trout Oncorhynchus mykiss (Walbaum, 1792) during disease. Depending on the course of the disease (severity of the pathology, leukocyte profile, and expression of immune-related genes), three subpopulations of fish with severe damage, a moderate course of the infectious process, and asymptomatic fish were characterized. An unexpected result was a small metabolic difference between fish with moderate symptoms and fish with weak signs of pathology. Thus, we have described the characteristics of a trout subpopulation with a mild course of infection which has potential for recovery after infection.This article describes the clinical manifestation of natural Vibrio anguillarum infection in rainbow trout (Oncorhynchus mykiss) during an outbreak on a fish farm. (i) Using an integrated approach, we characterized the pathogenesis of vibriosis from the morphological, hematological, and biochemical points of view. The molecular mechanisms associated with the host immune response were investigated using mass spectrometric analysis of trout plasma proteins. (ii) According to the severity of infection (the extent of tissue damage, the level of expression of pro-inflammatory genes, and changes in the leukocyte profile) three fish populations were identified among infected trout: fish with severe lesions (SL), fish with the moderate infectious process (IP) and asymptomatic fish (AS). (iii) Lymphopenia, granulocytosis, and splenomegaly were strong trends during the progression of infection and informative indicators of severe manifestation of disease, associated with hemorrhagic shock, metabolic acidosis, and massive tissue damage. (iv) As expected, pro-inflammatory interleukins, complement components, acute phase proteins, and antimicrobial peptides were implicated in the acute pathogenesis. Systemic coagulopathy was accompanied by increased antithrombotic reactions. (v) Reconstruction of metabolic pathways also revealed a high energy requirement for the immune response in severely affected fish. (vi) An unexpected result was a small difference between fish with moderate symptoms and fish with no or minor external signs of pathology (putatively resistant to infection). Increased production of antiproteases and enhanced blood coagulation cascade were observed in healthier fish, which may underlie the mechanisms of a controlled, non-self-damaging immune response to infection. (vii) Depending on the progression of the disease and the presence of the pathogen, a stepwise or linear change in the abundance of some plasma proteins was revealed. These proteins could be proposed as molecular markers for diagnosing the health and immune status of trout when cultured in fish farms.

Particularity of environmental biotic factors impact on mammal’s organism

This article explores ecological significance of animal venom as a stressful biotic factor and its role in disruption of homeostatic processes in mammal’s organism. The variety of venomous species of animals and their widely distribution in the world forces other species of animals to adapt to the coexistence with them. Thus, venom toxins have an important ecological significance, because they act as stressors and appreciably impact on the biotic interaction between organisms. According to scientists’ estimates, 220.000 species of animals, or 15% of the world’s biodiversity, are venomous, and there are 3.709 species of venomous snakes. The increase of snakebites cases is not only related to geographical distribution of snakes, but also to climate changes (high humidity and an increase of average annual temperature), which contributes to the reproduction of snakes and a shift of their natural habitats. Every year, around 5.4 million people are bitten by snakes worldwide. This leads to heavy social and economic costs in many countries of the world and it is the reason for constant attention to this problem. Among the variety of venomous animals, snakes have attracted the greatest attention of scientists due to peculiarities of the composition of their venom, its biochemical properties and the possibility of use in pharmacology and medicine. However, despite annual large-scale studies of the composition of snake toxins, the specifics of their effects on animals and human is not fully indentified. It has been established that among venomous snakes only vipers are common in Ukraine. Among them there are two subspecies of the common adder (Vipera berus Linnaeus, 1758): Vipera b. berus, which can be found in Polissia, Forest Steppe and the Carpathians, and Nikolsky’s viper (Vipera berus nikolskii Vedmederja, Grubant et Rudaeva, 1986), which occurs mainly in the Left Forest Steppe. Their bites can be very painful, but are rarely fatal for adults. Vipera berus venom has hemolytic, proteolytic and cytotoxic properties due to its composition: phospholipase A2, serine proteases, metallo proteases, amino acid L-oxidase, hemorrhagic factors and blood coagulation inhibitors, vasoactive peptides, cysteine-rich secretory protein. The composition of Vipera b. berus and Vipera b. nikolskii venom and mechanism of its action in victim’s organism were analyzed too. It is known that among the components of Nikolsky viper’s venom (Vipera b. nikolskii) the most part belongs to various enzymes. Of all enzymes, phospholipase A2 reached about 65% of the dry weight, serine proteases — 19%. Experiments on mammals (mice) showed that Nikol­sky’s viper (Vipera b. nikolskii) venom cause both local symptoms — swelling, pain, lymphangitis, and systemic symptoms — hypotension, mild neurotoxic effect. In addition, there are reports of damage to the cranial nerves and progressive paralysis of the limbs in mice. The components of Vipera b. berus viper venom have mainly a hemotoxic effect due to variety of enzymes from the metalloproteinase family, serine proteases, L-amino acid oxidases and C-type lectinlike proteins and cause enhanced blood coagulation. It is known that L-amino acid oxidases block the neuromuscular transmission of impulses and cause the destruction of cell membranes. The venom of this viper also contains several proteins that have a neurotoxic effect. A comprehensive and thorough analysis of experimental research data testify that the composition of toxins of various animal species, including vipers, is extremely diverse. Plenty of studies state about the hemovasotoxic and neurotoxic effects of these venoms. There are also reports of damage to the cardiovascular, urinary, respiratory, and digestive systems. It is noted that not all targets of viper toxins in mammal’s organism have been identified yet, and not all mechanisms of its action are understood, so further research is relevant.

Thiol Isomerase Activity of β3 Integrin Psi Domain and L33P Polymorphism: Implications in Blood Coagulation and Anti-Thrombotic Therapy

Background: Integrins are α and β subunit heterodimeric receptors expressed ubiquitously on metazoan cells that play key roles in cell adhesion, movement, and signaling. The αIIbβ3 integrin on platelets is essential for thrombosis and hemostasis through its binding of fibrinogen and other ligands to mediate platelet adhesion and aggregation. The plexin-semaphorin-integrin (PSI) domain is an approximately 54 amino acid sequence at the N-terminus of the β3 subunit located in close proximity to the “knee” region. Our lab recently discovered that two “CXXC” motifs within PSI domains in the integrin family exert endogenous thiol isomerase activity, and blockade of this activity in β3 integrin with our novel anti-PSI domain monoclonal antibodies (mAbs) significantly attenuated platelet adhesion and aggregation without impairing hemostasis (Blood, 2017). Interestingly, intravital microscopy studies demonstrate that our anti-PSI antibodies inhibited thrombosis in vivo 10-30 fold more potent than their effects in vitro under anti-coagulant conditions, suggesting a possible inhibitory effect on blood coagulation. Notably, A L33P polymorphism (HPA-1b) within the β3 integrin PSI domain is linked to an approximately two-fold increased risk for cardiovascular disease (CVD), however, the underlying mechanism for this remains unclear.Methods/Results: To investigate the role of PSI domain in blood coagulation, we first employed thromboelastography (TEG) to compare our anti-PSI domain mAbs with other anti-β3 mAbs that do not directly bind to β3 PSI domain. Results show that anti-PSI domain mAbs inhibited blood coagulation in human and murine whole blood or platelet-rich plasma (PRP) significantly more than anti-αIIbβ3 antibodies JAN-D1, M1, and Abciximab precursor 7E3. To address whether the L33P polymorphism affects PSI domains thiol isomerase activity, we generated L33P PSI domain via site-directed mutagenesis in E. coli. Using a scrambled RNase assay, we found that L33P polymorphism enhanced thiol-isomerase activity relative to WT PSI domain. We further corroborated these findings through an insulin β chain reduction assay, and a MPB (N-Maleimidopropionyl-biocytin) western blot assay, which quantifies thiol isomerase activity through MPB binding to free thiols that have not been oxidized into RNase. Interestingly, TEG results show that recombinant human PSI domain enhanced blood coagulation in platelet-microparticle (PMP) free plasma, which was generated through high-speed centrifugation (17,000 x g) of platelet poor plasma (PPP)for 15 minutes that removed residual platelets and microparticles .Conclusion: We have discovered a novel role of integrin β3 PSI domain in blood coagulation, which is enhanced by the L33P polymorphism (HPA-1b). These data highlight the β3 PSI domain as a suitable therapeutic target for its roles in both platelet adhesion/aggregation, and blood coagulation. Furthermore, these data may explain the increased risk of CVD such as myocardial infarction and deep vein thrombosis for individuals with the L33P polymorphism. DisclosuresNo relevant conflicts of interest to declare.

Fabrication of thrombin loaded TEMPO-oxidized cellulose nanofiber-gelatin sponges and their hemostatic behavior in rat liver hemorrhage model

Excessive blood loss due to trauma or major surgical intervention can be life threatening which necessitates rapid hemorrhage management for the prevention of such bleeding related sufferings. Broad interest in developing new hemostatic technologies have been paid for bleeding control but none of them found completely satisfactory especially in terms of rapid clotting, absorbability, porosity, cost effectiveness and safety. To address these issues, a combination of active and passive hemostatic materials from biological sources could be a wise choice. Therefore, plant-derived TEMPO-oxidized nanocellulose (TOCN)/biopolymer gelatin (G) sponge was successfully prepared in co-operation with intrinsic blood coagulation enzyme thrombin (Th) via freeze drying method and their application as rapid hemostatic dressing was investigated. Morphological and in vitro characteristics of the samples were evaluated where uniformity, porosity, swelling, degradation behavior had direct relationship with the percent gelatin incorporation. In vitro hemocompatibility and cyto-compatibility of these sponges were confirmed as well. Among the samples, TOCN 2.5G-Th sponge exhibited excellent hemostatic effect, rapid absorbability, minimum clotting time (1.37 ± 0.152 min) and reduction of blood loss was ensured through rat liver punch biopsy model. The results demonstrated that, Th enhanced blood coagulation, platelet and red blood cell aggregation following application of biopolymer TOCN 2.5G-Th sponge compared with samples devoid of Th. In short, the functional, cost effective and nontoxic sponge developed via facile preparation could potentially be used as an absorbable biomaterial to achieve immediate hemostasis. Highlights Plant-derived TEMPO-oxidized nanocellulose (TOCN) and biopolymer gelatin (G) was successfully used to prepare a hemostatic sponge in combination with intrinsic blood coagulation enzyme thrombin (Th). The TG sponge combines the advantages of TOCN and gelatin, exhibiting biocompatibility, biodegradability and superior blood-absorption performance. The TOCN 2.5G-Th sponge improves plasma absorption, red blood cell adhesion, aggregation, platelet adhesion and activation leading to enhanced hemostasis effect and shorter hemostasis time in vitro and in vivo

The binding of autotaxin to integrins mediates hyperhomocysteinemia-potentiated platelet activation and thrombosis in mice and humans

AbstractHyperhomocysteinemia (HHcy) is associated with an exaggerated platelet thrombotic response at sites of vascular injury. In this study, human medical examination showed that elevated human plasma Hcy levels correlated positively with enhanced blood coagulation and platelet activity, suggesting that humans with HHcy are more prone to thrombus formation at the sites of vascular injury. Accordingly, we observed accelerated platelet activation, primary hemostasis, and thrombus formation in apolipoprotein E-deficient (ApoE−/−) mice with acute or chronic HHcy. Upon homocysteine (Hcy) administration in C57BL/6J mice, platelet aggregation, spreading and clot retraction were markedly induced. More important, Hcy increased the affinity of platelet integrin αIIbβ3 with ligands and enhanced integrin outside-in signaling by promoting membrane phosphatidylserine exposure in vitro. Mechanistically, lipidomics analysis showed that lysophosphatidylcholines were the primary metabolites leading to clustering of HHcy-stimulated platelets. Cytosolic phospholipase A2 (cPLA2) activity and autotaxin (ATX, a secreted lysophospholipase D) secretion were upregulated by Hcy, leading to membrane phospholipid hydrolysis and PS exposure. Moreover, secreted ATX directly interacted with integrin β3. Inhibitors of cPLA2 and ATX activity blocked integrin αIIbβ3 outside-in signaling and thrombosis in HHcy ApoE−/− mice. In this study, we identified a novel mechanism by which HHcy promotes platelet membrane phospholipid catabolism and extracellular ATX secretion to activate integrin outside-in signaling, consequently exacerbating thrombosis and the results revealed an innovative approach to treating HHcy-related thrombotic diseases.

Enhanced blood coagulation and antibacterial activities of carboxymethyl-kappa-carrageenan-containing nanofibers

Ideal wound dressings should be biocompatible, exhibit high antibacterial activity, and promote blood coagulation. To impart these imperative functions, carboxymethyl-kappa-carrageenan was incorporated into poly(vinyl alcohol) nanofibers (PVA-CMKC). The antibacterial activity of the nanofibers was evaluated. Adsorption of two important blood proteins, fibrinogen and albumin, was also assessed. The adhesion and activation of platelets, and the clotting of whole blood were evaluated to characterize the ability of the nanofibers to promote hemostasis. Adhesion and morphology of both Staphylococcus aureus and Pseudomonas aeruginosa were evaluated using fluorescence microscopy and scanning electron microscopy. CMKC-containing nanofibers demonstrated significant increases in platelet adhesion and activation, percentage of coagulation in whole blood clotting test and fibrinogen adsorption, compared to PVA nanofibers, showing blood coagulation activity. Incorporating CMKC also reduces adhesion and viability of S. aureus and P. aeruginosa bacteria after 24 h of incubation. PVA-CMKC nanofibers show potential application as dressings for wound healing applications.

Thromboembolic complications in critically ill COVID-19 patients are associated with impaired fibrinolysis

BackgroundThere is emerging evidence for enhanced blood coagulation in coronavirus 2019 (COVID-19) patients, with thromboembolic complications contributing to morbidity and mortality. The mechanisms underlying this prothrombotic state remain enigmatic. Further data to guide anticoagulation strategies are urgently required.MethodsWe used viscoelastic rotational thromboelastometry (ROTEM) in a single-center cohort of 40 critically ill COVID-19 patients.ResultsClear signs of a hypercoagulable state due to severe hypofibrinolysis were found. Maximum lysis, especially following stimulation of the extrinsic coagulation system, was inversely associated with an enhanced risk of thromboembolic complications. Combining values for maximum lysis with D-dimer concentrations revealed high sensitivity and specificity of thromboembolic risk prediction.ConclusionsThe study identifies a reduction in fibrinolysis as an important mechanism in COVID-19-associated coagulopathy. The combination of ROTEM and D-dimer concentrations may prove valuable in identifying patients requiring higher intensity anticoagulation.

Age-related gene expression and DNA methylation changes in rhesus macaque

Aging is a very complicated biological process that can change gene expressions. The Chinese rhesus macaque (Macaca mulatta lasiota; CR) is closely related to humans. We explored gene expression with increasing age and DNA methylation changes in young and old CRs. Results showed blood transcriptome and DNA methylome significantly changed from young to old CRs. The age-associated differentially expressed genes (DEGs) and differentially methylated regions (DMRs) were associated with age-related biological features, such as immunity, blood coagulation, and biosynthetic process. The measurements of coagulation indicators confirmed old CRs had shorter coagulation time than young CRs, and the activities of coagulation factor II (FII) and factor VIII (FVIII) were enhanced in old CRs. Humans and CRs exhibited the same enhanced blood coagulation with age phenotype. Our study found aging is a critical factor affecting gene expression in CRs, and also provided new insights into the blood coagulation changes in non-human primates.

An enzymatically self-assembled DNA patch for enhanced blood coagulation.

DNA has been widely investigated as a carrier for drug delivery. Here, we describe a macroscopic DNA film that has been generated enzymatically. This DNA film was subsequently coated with thrombin (TB) using an aptamer-protein interaction, to expedite hemostasis over a large area. The DNA film coated with TB (DNA patch) significantly improved plasma hemostasis by acting as a coagulative scaffold for TB, as well as carrying localized TB. This study elucidates the benefits of using enzymatic amplification-based DNA structures in the context of topical drug treatment.

Increased Neutrophil Activation and Plasma DNA Levels in Patients with Pre-Eclampsia.

Pre-eclampsia (PE) is a chronic inflammatory disease in pregnancy, which is associated with enhanced blood coagulation and high thrombotic risk. To date, the mechanisms underlying such an association are not fully understood. Previous studies reported high levels of plasma deoxyribonucleic acid (DNA) in PE women, but the cellular source of the circulation DNA remains unknown. In this study, we tested the hypothesis that activated neutrophils undergoing cell death, also called NETosis, may be responsible for the elevated plasma DNA levels in PE women. We analysed plasma samples from non-pregnant, normal pregnant and PE women and found high levels of double-stranded DNA, myeloperoxidase (an abundant neutrophil granular enzyme) and histones (the major nucleosome proteins) in PE-derived samples, indicating increased NETosis in the maternal circulation. The high plasma DNA levels positively correlated with enhanced blood coagulation in PE women. When isolated neutrophils from normal individuals were incubated with PE-derived plasma, an elevated NETosis-stimulating activity was detected. Further experiments showed that endothelial micro-particles, but not soluble proteins, in the plasma were primarily responsible for the NETosis-stimulating activity in PE women. These results indicate that circulating micro-particles from damaged maternal endothelium are a potent stimulator for neutrophil activation and NETosis in PE women. Given the pro-coagulant and pro-thrombotic nature of granular and nuclear contents from neutrophils, enhanced systemic NETosis may represent an important mechanism underlying the hyper-coagulability and increased thrombotic risk in PE.

Synthesis of Phosphatidylserine and Its Stereoisomers: Their Role in Activation of Blood Coagulation.

Natural phosphatidylserine (PS), which contains two chiral centers, enhances blood coagulation. However, the process by which PS enhanced blood coagulation is not completely understood. An efficient and flexible synthetic route has been developed to synthesize all of the possible stereoisomers of PS. In this study, we examined the role of PS chiral centers in modulating the activity of the tissue factor (TF)-factor VIIa coagulation initiation complex. Full length TF was relipidated with phosphatidylcholine, and the synthesized PS isomers were individually used to estimate the procoagulant activity of the TF-FVIIa complex via a FXa generation assay. The results revealed that the initiation complex activity was stereoselective and had increased sensitivity to the configuration of the PS glycerol backbone due to optimal protein-lipid interactions.

ラットに対するカノーラ油および水素添加大豆油の有害作用と油中の微量因子―ジヒドロビタミンＫ１様物質の関与の可能性―

Since the survival-shortening activity of some vegetable oils, including canola and hydrogenated soybean oil, was reported in stroke-prone spontaneously hypertensive rats (SHRSP), several studies have been carried out to identify the causatives, but without success. We evaluated changes in pathophysiological parameters in several strains of rats caused by canola and/or hydrogenated soybean oil to help identify the causatives. No single target organ was found for canola oil, but several changes were observed in the parameters. That is, a diet containing 10% canola oil, which shortens the life of SHRSP and similar survival-shortening is also caused by hydrogenated soybean oil, increased blood pressure; decreased platelet counts; enhanced blood coagulation; increased hepatic glucose-6-phosphate dehydrogenase activity, with an elevation of plasma lipids; altered Nas, Ks-ATPase activities; decreased antioxidant enzyme activities; and increased plasma aldosterone levels, with a decrease in plasma and/or testicular testosterone levels in several strains of rats. Moreover, histochemical examinations revealed increased expression of cyclooxygenase 2 in macula densa cells in the kidney. In this review, the physiological and pathological changes are summarized in relation to canola oil-induced exacerbation of the hypertension-related conditions in SHR and SHRSP. In addition, possible involvement of dihydro-vitamin K1-like substance is discussed in relation to the observed changes.

Trivalent methylated arsenical-induced phosphatidylserine exposure and apoptosis in platelets may lead to increased thrombus formation

Trivalent methylated metabolites of arsenic, monomethylarsonous acid (MMAIII) and dimethylarsinous acid (DMAIII), have been found highly reactive and toxic in various cells and in vivo animal models, suggesting their roles in the arsenic-associated toxicity. However, their effects on cardiovascular system including blood cells, one of the most important targets for arsenic toxicity, remain poorly understood. Here we found that MMAIII and DMAIII could induce procoagulant activity and apoptosis in platelets, which play key roles in the development of various cardiovascular diseases (CVDs) through excessive thrombus formation. In freshly isolated human platelets, treatment of MMAIII resulted in phosphatidylserine (PS) exposure, a hallmark of procoagulant activation, accompanied by distinctive apoptotic features including mitochondrial membrane potential disruption, cytochrome c release, and caspase-3 activation. These procoagulant activation and apoptotic features were found to be mediated by the depletion of protein thiol and intracellular ATP, and flippase inhibition by MMAIII, while the intracellular calcium increase or reactive oxygen species generation was not involved. Importantly, increased platelet procoagulant activity by MMAIII resulted in enhanced blood coagulation and excessive thrombus formation in a rat in vivo venous thrombosis model. DMAIII also induced PS-exposure with apoptotic features mediated by protein thiol depletion, which resulted in enhanced thrombin generation. In summary, we believe that this study provides an important evidence for the role of trivalent methylated arsenic metabolites in arsenic-associated CVDs, giving a novel insight into the role of platelet apoptosis in toxicant-induced cardiovascular toxicity.

Enhanced blood coagulation and fibrinolysis in mice lacking histidine‐rich glycoprotein (HRG)

Histidine-rich glycoprotein (HRG) is a serum protein belonging to the cystatin superfamily. HRG may play a regulatory role in hemostasis and innate immunity. However, this role is uncertain because of a lack of rigorous testing in an animal model. We generated mice lacking the translation start point of exon 1 of the Hrg gene, effectively resulting in a null mutation (Hrg-/-). The mice were viable and fertile but had no HRG in their blood. Antithrombin activity in the plasma of Hrg-/- mice was higher than in the plasma of heterozygous Hrg+/- or wild-type Hrg+/+ mice. The prothrombin time was shorter in Hrg-/- mice than in Hrg+/- and Hrg+/+ mice. Bleeding time after tail tip amputation in Hrg-/- mice was shorter than in Hrg+/+ mice. The spontaneous fibrinolytic activity in clotted blood of Hrg-/- mice was higher than in Hrg+/+ mice. These findings suggest that HRG plays a role as both an anticoagulant and an antifibrinolytic modifier, and may regulate platelet function in vivo.

Ratio of serum vascular endothelial growth factor to platelet count correlates with disease activity in a patient with POEMS syndrome

Vascular endothelial growth factor (VEGF) is known to be involved in the pathogenesis of POEMS (polyneuropathy, organomegaly, endocrine disorder, M-protein, and skin lesion) syndrome. Because platelets have recently been recognized as transporters of VEGF, enhanced blood coagulation activity in this syndrome may accelerate vasopermeability by releasing VEGF from platelets in vivo. Here we report a case of POEMS syndrome with anasarca showing a high ratio of serum VEGF to platelet count, indicative of massive VEGF release from aggregated platelets in vivo. Changes in the ratio clearly reflected disease activity. This observation suggests that the ratio of serum VEGF to platelet count is more precise in monitoring disease activity than serum VEGF alone, and that VEGF released in vivo is critically involved in the pathogenesis of POEMS syndrome, causing hypervasopermeability.

SYSTEMIC EFFECT OF PARTICULATE AIR POLLUTION

The association in epidemiological studies between particulate air pollution (PM10) and increased mortality and morbidity from cardiovascular disease is well established. However, the mechanism(s) by which PM10 produces these effects is unknown. Wehave developed a hypothesis that ultrafine components of PM10 cause lung inflammation by creating local lung oxidative stress, which activates transcription factors such as NF-kappaB and hence the transcription of genes for inflammatory mediators. In a series of studies in vivo in the rat and using cultured airspace epithelial cells in vitro, we have tested this hypothesis using environmental PM10 and fine (CB 260 nm in diameter) and ultrafine (UfCB, 14 nm in diameter) carbon particles, since carbon is a major component of PM10. Wehave shown that compared with CB, which produces trivial effects, UFCB and PM10 produce local lung inflammation, increased epithelial permeability,and evidence of oxidative stress. Wehave also shown that PM10 also activates NF-kappaB in airspace epithelial cells. Wealso hypothesized that the local lung inflammation produced by PM10 may result in systemic effects, in particular systemic oxidative stress and enhanced blood coagulation, which may have a role in the adverse cardiovascular effects induced by PM10. To test this hypothesis we measured oxidative stress and changes in coagulation factors in plasma following the inhalation of UFCB, CB, or instillation of PM10 in the rat. Immediately after 7 h inhalation of UFCB (1000 (g/m3) there was a significant decrease in the antioxidant capacity in rat plasma, which fell further 16 and 48 h after cessation of the inhalation. A similar fall in plasma antioxidant capacity was shown 6 h after instillation of PM10 (125 mug). In contrast, there were no significant changes in antioxidant capacity in rats after 7 h of inhalation of CB at similar concentrations. The levels of factor VII in plasma, which is a key factor in the intrinsic pathway of coagulation cascade, increased at time points from 6 h to 7 days after inhalation of UFCB, but did not change after CB exposure. There were no changes in plasma fibrinogen or other coagulation factors after inhalation of particles. Thus, inhalation of ultrafine carbon black particles and instillation of PM10 in rats decreases plasma antioxidant capacity as an indication of systemic oxidative stress. Ultrafine carbon black particles also cause increased factor VII levels in the plasma, a known risk factor for adverse cardiovascular events. These studies help to explain the relationship between the levels of particulate air pollutants and cardiovascular morbidity/mortality.