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Dissolution of the fibrin blood clot is regulated in large part by plasminogen activator inhibitor-1 (PAI-1). Elevated levels of plasma PAI-1 may be an important risk factor for atherosclerotic vascular disease and are associated with premature myocardial infarction. The role of the endogenous plasminogen activation system in limiting thrombus formation following atherosclerotic plaque disruption is unknown. This study found that genetic deficiency for PAI-1, the primary physiologic regulator of tissue-type plasminogen activator (tPA), prolonged the time to occlusive thrombosis following photochemical injury to carotid atherosclerotic plaque in apolipoprotein E-deficient (apoE−/−) mice. However, anatomic analysis revealed a striking difference in the extent of atherosclerosis at the carotid artery bifurcation between apoE−/− mice and mice doubly deficient for apoE and PAI-1 (PAI-1−/−/apoE−/−). Consistent with a previous report, PAI-1+/+/apoE−/−and PAI-1−/−/apoE−/− mice developed similar atherosclerosis in the aortic arch. The marked protection from atherosclerosis progression at the carotid bifurcation conferred by PAI-1 deficiency suggests a critical role for PAI-1 in the pathogenesis of atherosclerosis at sites of turbulent flow, potentially through the inhibition of fibrin clearance. Consistent with this hypothesis, intense fibrinogen/fibrin staining was observed in atherosclerotic lesions at the carotid bifurcation compared to the aortic arch. These observations identify significant differences in the pathogenesis of atherosclerosis at varying sites in the vascular tree and suggest a previously unappreciated role for the plasminogen activation system in atherosclerosis progression at sites of turbulent flow.

Recombinant tissue plasminogen activator (rTPA) is commonly used in patients with myocardial infarction. Recently, it has also been applied intraocularly to dissolve postoperative fibrin with no serious complications being reported so far. In this study we describe our own experience with rTPA in 25 patients with persisting fibrinous membranes in the anterior segment. rTPA (Actilyse, Dr. Karl Thomae GmbH) was given in a single dose of 25 micrograms and injected into the anterior chamber via a paracentesis. We did not encounter any complications during the injection of rTPA. In 21 eyes fibrin could be reduced significantly, albeit sometimes only slowly. In 13 patients, the membrane had dissolved almost completely by the following day. In contrast, no success was observed after glaucoma surgery (2 eyes) and in chronic iritis (1 eye), or when fibrin mixed with blood was treated (1 eye). There were two (controllable) post-operative hemorrhages (rTPA after vitrectomy, and for fibrin/blood after cataract surgery). In addition, we noted 2 cases of irreversible superficial corneal clouding (rTPA after cataract surgery). We conclude that injection of rTPA can be a useful addition to steroid treatment in selective cases of persisting fibrin in the anterior segment. Long-standing membranes, however, are unlikely to be dissolved. Care should also be taken and rTPA be avoided when there is evidence of recent bleeding. Most worrying to us were the corneal complications that we cannot explain to date. With regard to the definite time correlation we feel that rTPA or one of the solution components might be the cause of this unusual feature.

Coagulation factor V is a critical cofactor for the activation of prothrombin to thrombin, the penultimate step in the generation of a fibrin blood clot. Genetic deficiency of factor V results in a congenital bleeding disorder (parahaemophilia), whereas inheritance of a mutation rendering factor V resistant to inactivation is an important risk factor for thrombosis. We report here that approximately half of homozygous embryos deficient in factor V (Fv-/-), which have been generated by gene targeting, die at embryonic day (E) 9-10, possibly as a result of an abnormality in the yolk-sac vasculature. The remaining Fv-/- mice progress normally to term, but die from massive haemorrhage within 2 hours of birth. Considered together with the milder phenotypes generally associated with deficiencies of other clotting factors, our findings demonstrate the primary role of the common coagulation pathway and the absolute requirement for functional factor V for prothrombinase activity. They also provide direct evidence for the existence of other critical haemostatic functions for thrombin in addition to fibrin clot formation, and identify a previously unrecognized role for the coagulation system in early mammalian development.

Invasion of renal tumor into retroperitoneal major vessels with thrombosis should be characterized as local spread of renal carcinoma and a serious complication. Extensive interventions were conducted in 30 subjects out of 196 nephrectomy cases. Nephrectomy was attended by colectomy (3 cases), pancreatic resection and adrenalectomy (3 cases), resection of the liver (2 cases), one-stage lobectomy (2 cases), adrenalectomy (9 cases), resection of the uterine appendages (1 case), resection of the colon, splenectomy, opening of an intraorganic abscess. 12 patients underwent thrombectomy from the major vein via the thoracophrenoabdominal approach. Cavathrombectomy was carried out in 7 (3.6%) patients, in 3 of which vena cava inferior was resected. Removal of the thrombus from the renal vein with resection of the opening and suturing of the vena cava inferior was performed in 5 patients. The thrombus originated from the right kidney in 9, while from the left one in 3 patients treated surgically. The thrombi occupied 4-10 cm along the renal vein from its opening. The removed kidney weighted from 400 to 3200 g. One death occurred due to pulmonary embolism during the operation, one on day 5 due to cardiopulmonary insufficiency. Histological examinations of the thrombi showed them to consist of fibrin, blood elements and tumor cells within the thrombus. The thrombi grow slowly, undergo organization and vascularization. Tumor cells multiply in the thrombus. Fibrin coating restricts cancer cell free dissemination via the venous system. Cavathrombectomy is considered the only way to prolong survival for the above patients.

Exposure of type III collagen coats on plastic cover slips in parallel-plate perfusion chambers to flowing nonanticoagulated human blood resulted in deposition of platelets and fibrin. Blood was drawn directly from an antecubital vein by an occlusive roller pump over the collagen coats in chambers having flow slits of different dimensions, so that wall shear rates of 100, 650, and 2600 s-1 were obtained at 10 ml/min. Coagulation was minimally activated during the passage of blood from the vein to the chamber as shown by fibrinopeptide A levels of 3.7 ng/ml after 5-minute perfusions. The surface coverage with platelets increased from 18% at 100 s-1 to 59% at 2600 s-1, and the corresponding thrombus volumes increased from 2 to 22 microns 3/microns 2, respectively. This contrasted with the coverage with fibrin on collagen, which decreased from 28% at 100 s-1 to 9% at 2600 s-1. Fibrin deposits on the thrombi covered 6% of the surface irrespective of the shear rate, indicating that some of the deposited platelets accelerated the deposition of fibrin. The type III collagen preparation did not activate factor XII and did not possess tissue factor activity, indicating that the surface itself was not procoagulant. However, a correlation between deposited leukocytes and surface coverage with fibrin was observed (r = 0.78, p less than 0.01), suggesting a role for these cells in the deposition of fibrin. The data demonstrate that thrombogenesis is triggered by pure type III collagen, although the deposition of fibrin is not initiated by the collagen itself but presumably by deposited leukocytes.(ABSTRACT TRUNCATED AT 250 WORDS)

Glanzmann thrombasthenia (GT) is a rare autosomal-recessive bleeding disorder that is characterized by genetic defects of the platelet-specific integrin, αIIbβ3. Molecular defects in either the αIIb- or β3-subunit can disrupt receptor synthesis, assembly, and/or function thereby preventing platelets from binding the receptors major adhesive ligands (von Willebrand factor and fibrinogen) to form platelet aggregates as a primary response to vascular injury. To develop methods for gene therapy of inherited bleeding disorders, we targeted expression of the human integrin αIIb-subunit in megakaryocytes of dogs affected with GT due to a molecular defect in αIIb. cDNA encoding the human αIIb-subunit was subcloned into an HIV type-1 lentivirus-derived vector under the transcriptional control of the lineage-specific human αIIb gene promoter (nucleotides −889 to +35). The vector also contained a cassette encoding the P140K form of methylguanine methyltransferase (MGMT) DNA repair protein under the transcriptional control of the murine stem cell virus promoter to allow in vivo chemo-selection of transduced stem cells. CD34+ cells were isolated from the apheresis product of G-CSF/SCF mobilized peripheral blood cells (PBC), transduced with virions, and autologously transplanted into a sublethally-irradiated animal. One dog received 3 x 106 transduced CD34+ PBC/kg of body weight following 300 cGy of total body irradiation (TBI). Platelet counts decreased below 10,000/ul at two weeks following PBC transplant and the animal succumbed to an intracranial hemorrhage in the third week. Another animal received an autologous transplant of 0.5 x 106 transduced CD34+ PBC/kg and 3 x 108 CD34(−) PBC fraction/kg of body weight following 100 cGy of TBI. The platelet count decreased to 35,000/ul at three weeks post-transplant and recovered to 100,000/ul at week four without incident. Flow cytometric analysis using αIIbβ3 specific antibodies detected integrin on the surface of 3% of circulating platelets at 3–6 weeks post-transplant. Since the drug-resistance marker, P140K MGMT, was detectable within cultured PBC, the dog was treated with chemocytotoxic reagents (O6BG, BCNU) three times at six-week intervals post-transplant for in vivo enrichment of transduced-PBC. Flow cytometric analysis detected peak expression of the hybrid human/canine αIIbβ3 integrin complex on the surface of 34% circulating platelets following the third drug treatment at 6 months post-transplant. Similar to platelets from normal dogs, αIIb-transduced platelets could be induced to form aggregates and retract a fibrin blood clot upon activation with physiological agonists of platelet activation. Bruising was also reduced significantly on the animal’s legs, thus further indicating correction of the GT phenotype. These ongoing studies reveal the potential feasibility for utilization of a lentivirus vector encoding a drug-selectable gene in conjunction with a therapeutic gene for enabling platelets to deliver therapy at the site of a vascular injury in patients with inherited bleeding disorders.

Fluorescent basic drugs have been shown to selectively accumulate in granules of blood platelets. The transformation kinetics of these organelles was studied in the course of clot formation. Simultaneous recording of fluorescence intensity and light transmission demonstrated that processes of fibrin filaments sedimentation and fibrin-blood platelet interaction were followed by the “collapse” of cells and liberation of the fluorescent marker. The platelet degranulation takes place due to contractile properties of platelets. This process was found to be inhibited by cytochalasin B and colchicine. Redistribution of platelets into aggregated, caused by the addition of ADP and stirring, led to the inhibition of the liberation. These experiments provide the evidence that secretion by platelets is realized by different mechanisms. Fluorescent basic drugs are a useful tool for investigating the function of the granules of blood platelets.

A 3-mm-diameter synthetic polytetrafluoroethylene (PTFE) cervical carotid bypass graft 20 cm in length was implanted in 30 dogs for the evaluation of blood flow, tissue response, and patency at intervals of 1 to 120 days. Although 4 of 5 grafts removed after 5 to 8 days were patent (80%), long term patency was observed in only 1 graft (10%). Aspirin treatment did not influence patency. Scanning electron microscopy demonstrated the lack of a neoendothelial layer upon the luminal surfaces of patent grafts, which were covered with a fibrin-blood cell lining. Subintimal fibrosis resulted in stenosis at sites of anastomosis in thrombosed grafts. The graft length, its small caliber, and a 40% decrease in blood flow after implantation may have contributed to thrombosis of the bypass graft in this model. Synthetic PTFE microvascular grafts may not be suitable for clinical use in extracranial-intracranial arterial bypass surgery.

Summary Subacute bacterial endocarditis is a distinet pathologic and clinical entity. The vegetative endocarditis that characterizes this disease, with its exudation of fibrin, blood platelets, and enmeshed bacteria, represents a state of high local and general tissue immunity to bacterial invasion, rather than lowered resistance; the immunity resides in the local endothelial structures, as well as in the general reticuloendothelial system. Since the reaction to this bacterial invasion is largely endothelial, the disease can rightfully be termed infectious endotheliosis . Subacute bacterial endocarditis occurs typically in patients who have valvular disease, usually rheumatic, but who have acquired a high degree of immunity, so that reinfection, recurrent endocarditis or pancarditis, and congestive heart failure do not occur. This increased immunity is responsible for the fact that the patient is not heart-conscious even during the attack of acute valvular infection. Then a transient bacteremia, caused by an upper respiratory infection, tonsillectomy, the extraction of an infected tooth, or pyelitis, permits secondary invasion and implantation of bacteria (usually the ubiquitous Streptococcus viridans ) on the damaged valve. A careful history will reveal that some infection which lowered the general body resistance is usually the beginning of an illness that later develops into subacute bacterial endocarditis. Although the bacteria are able to implant themselves on the damaged valves, they find in the valve an altered tissue reactivity which does not permit much local damage, such as ulceration or extension into the myocardium or pericardium. A more favorable outcome would be likely if the bacteria were not localized in a focus that communicates directly with the blood stream. If embolism does not cut the disease short, death occurs when the local and systemic endothelial systems become exhausted. Subacute bacterial endocarditis is a true endocardial sepsis, for the valve acts as the primary distributing focus of the infection. As such, if the focus can be removed surgically (as in some cases of patency of the ductus arteriosus) or chemotherapeutically before complications set in, there is some hope for a cure of this dreaded disease.

Urine white from the suspension of a quantity of fatty matter in it, has been called chylous urine, and the albumen and fibrin which escape from the blood with the fat have been considered to belong to the chyle and not to the blood. An opportunity of observing a case of this disease having occurred to me, I was led to make the following experiments. The conclusions therefrom are,— 1st. That the fat on which the white colour of the urine depends does indeed ap­pear in the urine after the chyle is absorbed, but the albumen, fibrin, blood-globules and alkaline salts, may be found in the urine previous to any food being taken, and these substances can be made to appear in, or disappear from the urine according as the circulation is hurried by motion or quieted by rest.

Objective Bacterial endocarditis is a frequent infectious cardiac disease, especially in patients with congenital or acquired heart defects. It is characterized by bacterial colonization of the heart valves and the appearance of vegetations consisting of fibrin, blood cells, and bacteria. The glycoprotein Deleted in Malignant Brain Tumors 1 is a scavenger receptor cysteine-rich protein with functions in innate immunity and epithelial differentiation. Because of the aggregating capacity of Deleted in Malignant Brain Tumors 1, we hypothesized that an up-regulation in bacterial endocarditis may be linked to the development of vegetations. Methods Heart tissue of 19 patients with bacterial endocarditis and 10 controls without bacterial endocarditis was analyzed by immunohistochemistry. The effect of human recombinant Deleted in Malignant Brain Tumors 1 on erythrocyte aggregation was measured using an automated red blood cell aggregometer MA1. Binding of human recombinant Deleted in Malignant Brain Tumors 1 to erythrocyte membranes, platelets, fibrin, and fibrinogen was analyzed by Western blotting and enzyme-linked immunosorbent assay. Results Deleted in Malignant Brain Tumors 1 expression was up-regulated in affected heart valves with bacterial endocarditis and limited to the colonizing bacteria on the heart valves and granulocyte-depleted fibrin/fibrinogen formations, and around localized atheromatosis. Patients with aggressive bacteria showed higher DMBT1 levels than patients with less aggressive bacteria. Human recombinant Deleted in Malignant Brain Tumors 1 aggregates erythrocytes and binds to erythrocyte membranes, platelets, and fibrin/fibrinogen. Conclusion Deleted in Malignant Brain Tumors 1 up-regulation at sites of bacterial endocarditis, its association with platelets and fibrin/fibrinogen, and its ability to aggregate erythrocytes through binding to their membranes indicate a potential role in the development of vegetations and thrombosis.

The fluorescent marker method and measurements of aggregation were utilized to study the significance of the acidic intragranular space in blood platelets. Acridine orange is liberated from 5-HT granules after alkalinization by the ammonia ion which inhibits the induction of platelet aggregation by ADP and causes the platelet disaggregation. At low doses, ammonia ion enhances ADP-induced aggregation. The transformation kinetics of 5-HT granules was investigated in the course of clot formation with a new instrument that simultaneously monitors the light transmittance (aggregation, clot formation) and fluorescence intensity (accumulation and liberation of fluorescent marker). These experiments provide the evidence that processes of fibrin filament sedimentation and fibrin-blood platelet interaction were followed by a liberation of fluorescent marker from 5-HT granules.

A 3-mm-diameter synthetic polytetrafluoroethylene (PTFE) cervical carotid bypass graft 20 cm in length was implanted in 30 dogs for the evaluation of blood flow, tissue response, and patency at intervals of 1 to 120 days. Although 4 of 5 grafts removed after 5 to 8 days were patent (80%), long term patency was observed in only 1 graft (10%). Aspirin treatment did not influence patency. Scanning electron microscopy demonstrated the lack of a neoendothelial layer upon the luminal surfaces of patent grafts, which were covered with a fibrin-blood cell lining. Subintimal fibrosis resulted in stenosis at sites of anastomosis in thrombosed grafts. The graft length, its small caliber, and a 40% decrease in blood flow after implantation may have contributed to thrombosis of the bypass graft in this model. Synthetic PTFE microvascular grafts may not be suitable for clinical use in extracranial-intracranial arterial bypass surgery.

Blood and plasma clots are frequently abnormal in patients with multiple myeloma a evidenced by the prolongation of clotting time, formation of bulky and gelatinous cloes and inhibition of clos retraction. It has been demonstrated in several multiple myeloma cases that the isolated immunoglobulin inhibited clot formation in normal plasma and decreased the rate of fibrin polymerization. In this work a hypothesis was tested whether these phenomena originate from binding of multiple myeloma immunoglobulin with fibrinogen and fibrin. Blood was obtained from a patient (J.I.) having IgAH gammopath; with prolonged clotting time and abnormal clot retraction. Washed plasma clots contaii ed largo amount of monoclonal IgAλ demonstrated by precipitation with anti-α or anti-λ antibodies and by the presence of heavy (65,000) and light (25,000) polypeptide chains. This myeloma IgA was incorporated into clots regardless of crosslinking and was extract able. Affinity chromatography of serum or heated plasma on insolubilized fibrin monoment resulted in recovery of large amount of myeloma monoclonal IgA, but not IgG, IgH or Igl. The isolated fibrin-specific IgA was taken up by clots obtained from normal plasma or purified fibrinogen, lnso]ubilized fibrinogen bound myeloma monoclonal IgA From plasma. Two-dimensional agarose/polyacrylamide slab gel electrophoresis demonstrated the presen of a fibrinogen-IgA complex in patient’s plasma. The results indicate that a myeloma monoclonal immunoglobulin binds with fibrinogen and fibrin, probably forming an antibot antigen complex, implying the Formation of autoantibodies against these proteins.

The possibility of prevention of intravascular blood coagulation in rats by DIP-alpha-thrombin devoid of proteolytic activity and capable of stimulating the reaction of anticoagulation system was studied. The injection of lethal thromboplastin dose was shown to produce a sharp increase in soluble fibrin blood content, total disappearance of fibrinolytic activity and intravascular blood coagulation. The animals died of thrombosis in 90% of cases. It was established that the injection of lethal thromboplastin dose 5 min after DIP-alpha-thrombin injection caused a 13% lethality from thrombosis. No reliable changes in fibrinolytic activity and soluble fibrin content were observed. A significant increase in thrombin and recalcification time was recorded. It is suggested that DIP-alpha-thrombin prevents intravascular blood coagulation induced by lethal thromboplastin dose due to mobilization of the reserve capacities of neuro-humoral anticoagulation system.

Postpartum haemorrhage (PPH) is responsible for about 100,000 maternal deaths every year, most of which occur in low- and middle-income countries. Tranexamic acid (TXA) reduces bleeding by inhibiting the enzymatic breakdown of fibrin blood clots. TXA decreases blood loss in surgery and reduces death due to bleeding after trauma. When given within 3 h of birth, TXA reduces deaths due to bleeding in women with PPH. However, for many women, treatment of PPH is too late to prevent death. Over one third of pregnant women in the world are anaemic and many are severely anaemic. These women have an increased risk of PPH and suffer more severe outcomes if PPH occurs. There is an urgent need to identify a safe and effective way to reduce postpartum bleeding in anaemic women. The WOMAN-2 trial is an international, multicentre, randomised, double-blind, placebo-controlled trial to quantify the effects of TXA on postpartum bleeding in women with moderate or severe anaemia. Ten thousand women with moderate or severe anaemia who have given birth vaginally will be randomised to receive 1 g of TXA or matching placebo by intravenous injection immediately (within 15 min) after the umbilical cord is cut or clamped. The primary outcome is the proportion of women with a clinical diagnosis of primary PPH. The cause of PPH will be described. Data on maternal health and wellbeing, maternal blood loss and its consequences, and other health outcomes will be collected as secondary outcomes. The main analyses will be on an ‘intention-to-treat’ basis, irrespective of whether the allocated treatment was received. Results will be presented as appropriate effect estimates with a measure of precision (95% confidence intervals). Subgroup analyses will be based on the severity of anaemia (moderate versus severe) and type of labour (induced or augmented versus spontaneous). A study with 10,000 patients will have over 90% power to detect a 25% relative reduction from 10 to 7.5% in PPH. The trial will be conducted in hospitals in Africa and Asia. The WOMAN-2 trial should provide reliable evidence for the effects of TXA for preventing postpartum bleeding in women with anaemia. ISRCTN, ISRCTN62396133 . Registered on 7 December 2017; ClincalTrials.gov, ID: NCT03475342 . Registered on 23 March 2018.

Disseminated alterative arteriolitis is a disease entity which may be best understood by an analysis of the literature which led to the concept of acute bacterial endocarditis and what is known in the United States as Libman-Sacks' disease, or verrucous endocarditis without bacteremia. The two cases to be discussed are of interest because of the cerebral symptomatology and the presence of the typical vascular lesions in the brain as well as in other organs of the body, although endocarditis was not present in either case. In acute bacterial endocarditis the endothelial lining of the heart responds with proliferative changes on the wall and especially on the valves; verrucous masses are formed by the acquisition of thrombotic material consisting of fibrin, blood platelets and leukocytes of the blood stream, and the underlying inflammatory processes are secondary. In the subacute process of Libman-Sacks' disease bacteremia cannot be proved, and the inflammatory reaction