Cellular and Plasma Protein Crosstalk in Arterial and Venous Thrombosis

Coagulation is an enzymatic cascade culminating in the formation of a clot. Intravascular coagulation is termed thrombosis. Studies using platelet-rich plasma, whole blood, and animal models reveal the complex crosstalk between clotting factors and the cellular environment that promote thrombosis. Three distinct, yet related, studies included in this dissertation exemplify this crosstalk. First, the vascular bed-dependent prothrombotic effects of elevated prothrombin were examined. We found that elevated prothrombin increased venous, but not arterial, thrombosis in mice. The prothrombotic effects of elevated prothrombin were manifested by increased fibrin deposition but no significant increase in platelet accumulation. These data show that elevated prothrombin would not be expected to contribute to platelet-dominated arterial thrombosis. Second, the procoagulant properties of microparticles from different cell types were investigated. We found that monocyte-derived microparticles contributed to initiation and propagation of clotting via the extrinsic coagulation pathway, while platelet-derived microparticles contributed to primarily to propagation of clotting via the intrinsic coagulation pathway. These data suggest monocyte-derived microparticles may contribute to the initiation of a thrombotic event, while platelet-derived microparticles may propagate an existing thrombotic event. Third, the role of fibrinogen and factor XIII (FXIII) in venous thrombosis was studied. We discovered the binding site of FXIII on fibrinogen, the necessity for FXIII activity for red blood cell retention in clots, and that reduction or deficiency of FXIII activity reduces venous thrombus size. These data indicate a novel role for FXIII in the pathogenesis of thrombosis and implicate FXIII as a novel therapeutic target. Together, these studies emphasize the importance of complementary in vitro and in vivo studies of coagulation and thrombosis to properly elucidate the pathophysiological relevance of clotting factor and cell function.Doctor of Philosoph

Influence of Cellular and Plasma Procoagulant Activity on the Fibrin Network

At the nexus of cellular and plasma procoagulant activities lies fibrin, which is necessary to provide a clot's structural support. Abnormalities in fibrin network formation or function can result in either bleeding or thrombotic complications. Understanding relationships between procoagulant activity and normal fibrin formation, as well as pathophysiologic mechanisms leading to abnormal fibrin deposition, is essential for the continued development of hemostatic and antithrombotic therapies

Interaction of gonadal steroids and growth factors in brain sex differentiation

Sex hormones have developmental trophic actions on neurons and glial cells and activational effects in the adult brain. It has been proposed that sex steroids may interact with peptide trophic factors to induce part of their biological effects in the nervous system. The first evidence of such an interaction was provided by Toran-Allerand et al (Brain Research 1980; 184: 517-524), showing that in explant cultures of fetal rodent hypothalamus, estrogen and insulin have synergistic effects on neurite growth, an effect probably mediated by insulin-like growth factor-1 receptors. Recent data indicate that estrogen and insulin-like growth factor-1 signaling pathways interact on hypothalamic neurons to regulate survival and differentiation and that sex steroids interact with a variety of different trophic signals in vivo to regulate neuroendocrine events. These findings suggest that trophic factors may be involved in the genesis of sex differences in the developing brain and in the maintenance of a sexually differentiated brain function in the adult.Biomedical Reviews 1997; 7: 67-74

Procoagulant Activity in Hemostasis and Thrombosis: Virchowʼs Triad Revisited

Virchow’s triad is traditionally invoked to explain pathophysiologic mechanisms leading to thrombosis, alleging concerted roles for abnormalities in blood composition, vessel wall components, and blood flow in the development of arterial and venous thrombosis. Given the tissue-specific bleeding observed in hemophilia patients, it may be instructive to consider the principles of Virchow’s triad when investigating mechanisms operant in hemostatic disorders as well. Blood composition (the function of circulating blood cells and plasma proteins) is the most well-studied component of the triad. For example, increased levels of plasma procoagulant proteins such as prothrombin and fibrinogen are established risk factors for thrombosis, whereas deficiencies in plasma factors VIII and IX result in bleeding (hemophilia A and B, respectively). Vessel wall (cellular) components contribute adhesion molecules that recruit circulating leukocytes and platelets to sites of vascular damage, tissue factor, which provides a procoagulant signal of vascular breach, and a surface upon which coagulation complexes are assembled. Blood flow is often characterized by two key variables: shear rate and shear stress. Shear rate affects several aspects of coagulation, including transport rates of platelets and plasma proteins to and from the injury site, platelet activation, and the kinetics of fibrin monomer formation and polymerization. Shear stress modulates adhesion rates of platelets and expression of adhesion molecules and procoagulant activity on endothelial cells lining the blood vessels. That no one abnormality in any component of Virchow’s triad fully predicts coagulopathy a priori suggests coagulopathies are complex, multifactorial and interactive. In this review, we focus on contributions of blood composition, vascular cells, and blood flow to hemostasis and thrombosis, and suggests cross-talk among the three components of Virchow’s triad is necessary for hemostasis and determines propensity for thrombosis or bleeding. Investigative models that permit interplay among these components are necessary to understand the operant pathophysiology, and effectively treat and prevent thrombotic and bleeding disorders

Fibrinogen and red blood cells in venous thrombosis

Deep vein thrombosis and pulmonary embolism, collectively termed venous thromboembolism (VTE), affect over 1 million Americans each year. VTE is triggered by inflammation and blood stasis leading to the formation of thrombi rich in fibrin and red blood cells (RBCs). However, little is known about mechanisms regulating fibrin and RBC incorporation into venous thrombi, or how these components mediate thrombus size or resolution. Both elevated circulating fibrinogen (hyperfibrinogenemia) and abnormal fibrin(ogen) structure and function, including increased fibrin network density and resistance to fibrinolysis, have been observed in plasmas from patients with VTE. Abnormalities in RBC number and/or function have also been associated with VTE risk. RBC contributions to VTE are thought to stem from their effects on blood viscosity and margination of platelets to the vessel wall. More recent studies suggest RBCs also express phosphatidylserine, support thrombin generation, and decrease fibrinolysis. RBC interactions with fibrin(ogen) and cells, including platelets and endothelial cells, may also promote thrombus formation. The contributions of fibrin(ogen) and RBCs to the pathophysiology of VTE warrants further investigation

Cystamine Preparations Exhibit Anticoagulant Activity

Transglutaminases are a superfamily of isoenzymes found in cells and plasma. These enzymes catalyze the formation of ε-N-(γ-glutamyl)-lysyl crosslinks between proteins. Cystamine blocks transglutaminase activity and is used in vitro in human samples and in vivo in mice and rats in studies of coagulation, immune dysfunction, and inflammatory disease. These studies have suggested cystamine blocks fibrin crosslinking and has anti-inflammatory effects, implicating transglutaminase activity in the pathogenesis of several diseases. We measured the effects of cystamine on fibrin crosslinking, tissue factor-triggered plasma clot formation and thrombin generation, and coagulation factor enzymatic activity. At concentrations that blocked fibrin crosslinking, cystamine also inhibited plasma clot formation and reduced thrombin generation. Cystamine inhibited the amidolytic activity of coagulation factor XI and thrombin towards chromogenic substrates. These findings demonstrate that cystamine exhibits anticoagulant activity during coagulation. Given the close relationship between coagulation and inflammation, these findings suggest prior studies that used cystamine to implicate transglutaminase activity in disease pathogenesis warrant re-examination

Diagnosis of solid waste management in the petrochemical-plastic sector of Cartagena de indias, Northern Colombia

In the production of most petrochemical products there is a generation of associated solid waste. The objective is to diagnose solid waste management in a company in the petrochemical-plastic sector. The methodology initially consisted in the identification of the different operational areas and type of waste under the NTC GTC 24, in the second instance the solid waste classification was carried out taking into account the simple method of analysis of CEPIS and finally the quantification of The waste taking as a reference what Ruiz (2012)[1] did. It is evident that there is an increase (10.2%) in the generation of solid waste generated, from 545,750 kg in 2013 to 601,492 kg in 2014. The activities with the largest generation of solid waste (cardboard, plastic, wood) Are those of the reception and storage processes and the packaging process in each of the plants

IL-1 receptor antagonist reduces endotoxin-induced airway inflammation in healthy volunteers

Asthma with neutrophil predominance is challenging to treat with corticosteroids. Novel treatment options for asthma include those that target innate immune activity. Recent literature has indicated a significant role for IL-1β in both acute and chronic neutrophilic asthma

Insulin regulates neurovascular coupling through astrocytes

Mice with insulin receptor (IR)-deficient astrocytes (GFAP-IR knockout [KO] mice) show blunted responses to insulin and reduced brain glucose uptake, whereas IRdeficient astrocytes show disturbed mitochondrial responses to glucose. While exploring the functional impact of disturbed mitochondrial function in astrocytes, we observed that GFAP-IR KO mice show uncoupling of brain blood flow with glucose uptake. Since IR-deficient astrocytes show higher levels of reactive oxidant species (ROS), this leads to stimulation of hypoxia-inducible factor-1¿ and, consequently, of the vascular endothelial growth factor angiogenic pathway. Indeed, GFAP-IR KO mice show disturbed brain vascularity and blood flow that is normalized by treatment with the antioxidant N-acetylcysteine (NAC). NAC ameliorated high ROS levels, normalized angiogenic signaling and mitochondrial function in IR-deficient astrocytes, and normalized neurovascular coupling in GFAP-IR KO mice. Our results indicate that by modulating glucose uptake and angiogenesis, insulin receptors in astrocytes participate in neurovascular coupling.We are thankful to M.Garcia and R. Cañadas for technical support. This work was funded by Centro de Investigación Biomédica en Red de Enfermedades Neurodegenerativas (CIBERNED) (Instituto de Salud CarlosIII, Spain) to I.T.A., A.G., and T.I.; an Inter-CIBER project (PIE14/00061) to I.T.A.that forms part of the projects PID2019-104376RB-I00 (I.T.A.) and RTI2018-094887-B-I00 (M.N.) funded by MCIN/AEI/10.13039/501100011033; a grant from Junta de Andalucia Consejería de Economía y Conocimiento (P18-RT-2233 to A.G.) cofinanced by Programa Operativo FEDER 2014–2020; a grant from Instituto de Salud Carlos III Spain (cofinanced by FEDER funds from the European Union; PI21/00915 to A.G.); Grant PID2020-115218RB-I00 to T.I. funded by Ministerio de Ciencia e Innovación/Agencia Española de Investigación (MCIN/AEI/10.13039/501100011033); and a grant from Comunidad de Madrid through the European Social Fund (ESF)–financed programme Neurometabolismo-Comunidad de Madrid (NEUROMETAB-CM) (B2017/BMD-3700 to I.T.A.and T.I.). M.N. was also supported by the Spanish Ministry of Science and Innovation (Ramón y Cajal RYC-2016-20414). J.P.-U. was contracted by CIBERNED

A proof-of-concept clinical study examining the NRF2 activator sulforaphane against neutrophilic airway inflammation

Abstract Sulforaphane (SFN), a naturally occurring isothiocyanate found in cruciferous vegetables, is implicated as a possible therapy for airway inflammation via induction of the transcription factor NF-E2-related factor 2 (NRF2). In this proof-of-concept clinical study, we show that supplementation of SFN with broccoli sprout homogenate in healthy human subjects did not induce expression of antioxidant genes or protect against neutrophilic airway inflammation in an ozone-exposure model. Therefore, dietary sulforaphane supplementation is not a promising candidate for larger scale clinical trials targeting airway inflammation. Trial registration: NCT01625130 . Registered 19 June, 2012