Procoagulant Research Articles

2970 Non-Cirrhotic Portal Hypertension in Myeloproliferative Disease: Left and Right-Sided Component of Portal Hypertension

INTRODUCTION: NCPH is the presence of portal hypertension (PH) in the absence of cirrhosis. While Schistosomiasis is the commonest cause in West, myeloproliferative diseases (MPD) characterized by cellular proliferation of one or more hematologic cell lines resulting in extra-medullary hematopoiesis predominantly in the spleen and occasionally in liver may cause PH. Pathogenesis of NCPH in MPD revolves around the concept of left sided portal hypertension. This study presents a cohort of Indian patients of NCPH with MPD. METHODS: 260 patients with NCPH were evaluated and 9 MPD was diagnosed based on 2008 WHO-criteria. Cirrhosis was ruled out by fibroscan and liver biopsy. All patients underwent a pro coagulant screen which included protein-C, protein-S, factor-V leiden, antithrombin D, anti phospholipid antibody, prothrombin gene deficiency and Jak-2 mutation study. Routine PH evaluation and Fibroscan and liver biopsy was done in all cases. RESULTS: Out of 9 patients 4 (44.4%) each had polycythemia-rubra-vera (PRV) and myelofibrosis while 1 had essential thrombocytosis. Median age was 50 ± 7 yrs with 8 male and 1 female patient. 5 (55.5%) presented with symptomatic splenomegaly, 1 (11.1%) with ascites and 3 (33%) with GI bleed and comprised of 64.3% in PRV. Splenomegaly was universally present and hepatomegaly in 44.4%. Isolated gastric varices was present in 3 and gastroesophageal varices in 4 and isolated esophageal varices in 2 patients. Except for Jak-2 mutation found positive in 100% cases of PRV and 66.7% cases of MF, none of the other markers were positive. None, except 1 patient on liver biopsy showed sinusoidal deposits. 8/9 patients underwent endoscopic therapy for varices and were given prophylaxis with carvedilol. Mean follow up was 40 months and none had recurrent bleed or worsening of liver function due to PH. CONCLUSION: True incidence MPD causing PH is under-recognised. Our results are similar to other studies from Asia. Unlike other studies, we did not find portal vein thrombosis. MPD patients had huge splenomegaly leading to increased blood flow through the splenic vein leading to left sided portal hypertension. Myeloid metaplasia in liver can also leading to right-sided PH. Whether beta blockers would help as primary or secondary prophylaxis without this component is inconclusive. However, in our cohort of patient they were helpful. Rescue therapy like splenectomy, JAK 2 inhibitors, TIPS etc may be reserved for the refractory cases.

RERATA VOLUME TROMBOSIT DI DIABETES MELITUS (Mean Platelet Volume in Diabetes Mellitus)

Diabetes Mellitus (DM) is a state of chronically hyperglycemia, which is caused by impaired insulin secretion and function. Thealteration in coagulation system in DM patients served as the basic etiology of macro and micro vascular complication. Platelets are oneof the factors that have a role in the coagulation system and undergo pathological changes in DM patients. Mean Platelet Volume (MPV)is an indicator of platelet function and activation. Larger platelets have more dense and more reactive pro coagulant granules. The aimof this study was to know whether there were differences in MPV between DM patients compared to medical check-up participants atthe Dr. Hasan Sadikin Hospital, Bandung.. The research used analytical descriptive, retrospective data. The inclusion criteria of thisresearch was DM patients and medical check up participants whose serum fasting glucose and two (2) hours post prandial serum glucosewere within the normal range. The data were taken from medical records of DM patients and medical check up participants. The datawere further analyzed with SPSS ver. 15 program using unpaired T-test. In this study, no significant difference between DM patientsand medical check up participants (p>0.05) was found. It can be concluded that Mean Platelet Volume if used as single, can not predictvascular complication in DM patients.

Direct Oral Anticoagulants in Patients with Cirrhosis Appear Safe and Effective

▪Introduction: Patients with cirrhosis have historically been excluded from clinical trials of anticoagulant therapies due to concerns about safety. While erroneously thought to be "auto-anticoagulated," patients with cirrhosis may actually be at increased risk for venous thromboemboli (VTE) due to an imbalance in pro coagulant and anticoagulant factors and do require therapeutic anticoagulation when thrombotic events occur. Traditionally low molecular weight heparins, and less commonly warfarin, have been the anticoagulants of choice in cirrhotic patients, though there are challenges with their use due to difficulty with access, administration and blood monitoring. Direct Oral Anticoagulants (DOACs) are increasingly replacing warfarin for the treatment of VTE and for stroke prevention in non-valvar atrial fibrillation. The administration and standardized dosing make DOACs an attractive alternative for safe and effective therapeutic anticoagulation but very little evidence exists for their use in patients with cirrhosis.Methods: We designed a retrospective analysis of all patients with cirrhosis at the University of Virginia who were treated with a DOAC between January 2012 and May 2016 for all indications. All patients were required to be treated with a DOAC with documented follow up for assessment of adverse events. Medical charts were reviewed for age, gender, weight, cirrhosis etiology, DOAC, dose, indication and length of treatment, Model for End-Stage Liver Disease (MELD) and Child-Turcotte-Pugh (CTP) scores, bleeding events and appearance of thrombus on repeat imaging. All bleeding events were recorded and graded according the Common Toxicity Criteria for Adverse Events, v4.0. Major bleeding events were reviewed by all investigators.Results: Eighty patients were identified, with 11 patients excluded after not starting a DOAC or being mislabeled as cirrhosis, leaving 69 patients for analysis. Median age was 73 years old (range 44-92) with 77% male. The most common etiologies of cirrhosis were NASH (66%) and HCV (22%). Thirty seven patients (54%) were treated with apixaban, 25 (36%) were treated with rivaroxaban and 7 (10%) were treated with dabigatran. Median length of time on anticoagulation was 6 months. Sixty-eight percent were treated for non-tumoral PVT or DVT while 32% were treated for stroke prevention in atrial fibrillation. Most patients had well compensated cirrhosis, but several had advanced cirrhosis (CTP class A/B/C: 33/26/10). Fourteen percent had an underlying malignancy, most commonly hepatocellular carcinoma, and 52% had esophageal varices. Of those treated for atrial fibrillation, none developed a clot while on a DOAC. Of those treated for VTE, 81% had resolution, 13% unchanged and 6% progression of their clot on repeat imaging. No patient developed drug induced liver injury or discontinued anticoagulation due to liver injury. Sixteen patients (23%) suffered a bleeding event, mostly epistaxis and easy bruising. Major bleeding, defined as grade 3 or higher, occurred in 12% of patients. Most major bleeding was gastrointestinal bleeding; with most but not all requiring endoscopy and transfusions. One subdural hematoma occurred. No variceal bleeding occurred, despite that majority of patients having esophageal varices identified on endoscopy. Of those with major bleeding, all were CTP class B or C with an average MELD of 22, compared to an average MELD of 14 in patients without major bleeding. All patients with major bleeding were treated with either apixaban or rivaroxaban.Conclusion: Similar to other anticoagulants, DOACs should be used with caution in patients with cirrhosis, particularly CTP class B or C. In this case series, the largest to our knowledge, direct oral anticoagulants in patients with cirrhosis compare favorably to historical controls (Delgado, Clin Gastroenterol Hepatol. 2012 Jul;10(7):776-83) with lower incidence of major bleeding and increased clot resolution. Further prospective studies are needed in this at risk and under studied patient population. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

RERATA VOLUME TROMBOSIT DI DIABETES MELITUS

Diabetes Mellitus (DM) is a state of chronically hyperglycemia, which is caused by impaired insulin secretion and function. The alteration in coagulation system in DM patients served as the basic etiology of macro and micro vascular complication. Platelets are one of the factors that have a role in the coagulation system and undergo pathological changes in DM patients. Mean Platelet Volume (MPV) is an indicator of platelet function and activation. Larger platelets have more dense and more reactive pro coagulant granules. The aim of this study was to know whether there were differences in MPV between DM patients compared to medical check-up participants at the Dr. Hasan Sadikin Hospital, Bandung.. The research used analytical descriptive, retrospective data. The inclusion criteria of this research was DM patients and medical check up participants whose serum fasting glucose and two (2) hours post prandial serum glucose were within the normal range. The data were taken from medical records of DM patients and medical check up participants. The data were further analyzed with SPSS ver. 15 program using unpaired T-test. In this study, no significant difference between DM patients and medical check up participants (p>0.05) was found. It can be concluded that Mean Platelet Volume if used as single, can not predict vascular complication in DM patients.

Successful treatment of coronary artery perforation using fat tissue embolization.

Coronary artery rupture is a rare complication of percutaneous coronary intervention (PCI) that may lead to pericardial effusion frequently accompanied by tamponade. It is one of the most serious complications with a relatively high mortality rate ranging from 7 to 17%. 1-3 We report a case of a 63-year-old patient with acute inferoposterolateral ST-segment elevation myocardial infarction presented 4 hours from the beginning of chest pain. An urgent coronarography with left radial access was done and a 3-vessel coronary artery disease was found with occlusion of strong first marginal branch (OM1). Following stenting of the OM1 with 2nd bare metal stent (BMS), a type III distal guidewire-induced coronary perforation was recognized with extravasation of contrast into the pericardial space. Hemostasis was immediately performed inflating the balloon from the stent and a conversion of heparin with 10.000 I.U. protamine was done. As prolonged balloon inflations were unsuccessful, we decided to do a peripheral embolization with subcutaneous fat. The first guiding catheter, wire and balloon were left in place to maintain hemostasis and we placed another working guiding catheter through the right femoral artery. Prior to placing the introducer, 5 cc of subcutaneous fat was harvested from the patient’s upper right thigh. A mixture of fat and saline was injected through the over-the-wire balloon distally of the balloon for hemostasis resulting in the immediate cessation of leakage at the site of perforation. However, immediately after, most likely due to leakage of the pro coagulant fat tissue from circumflex artery (Cx) to left anterior descending artery (LAD), occlusion of the LAD at the previously diagnosed significant stenosis occurred. The PCI of the LAD was immedi ately made with BMS implantation and clinical stabilization of the patient. At the end of the procedure Cx was sealed and there was no need for the pericardiocentesis or cardiac surgery. Echocardiography reveled a moderate posterolateral hypokinesia with left ventricular ejection fraction of 45% and a mild mitral regurgitation. After 21 days PCI of the right coronary artery with BMS was done. Subcutaneous fat tissue embolization is a simple technique available to all for the treatment of potentially fatal complications such as rupture of the coronary artery. Even though potential complications must be considered.

Overview of the coagulation system.

Coagulation is a dynamic process and the understanding of the blood coagulation system has evolved over the recent years in anaesthetic practice. Although the traditional classification of the coagulation system into extrinsic and intrinsic pathway is still valid, the newer insights into coagulation provide more authentic description of the same. Normal coagulation pathway represents a balance between the pro coagulant pathway that is responsible for clot formation and the mechanisms that inhibit the same beyond the injury site. Imbalance of the coagulation system may occur in the perioperative period or during critical illness, which may be secondary to numerous factors leading to a tendency of either thrombosis or bleeding. A systematic search of literature on PubMed with MeSH terms ‘coagulation system, haemostasis and anaesthesia revealed twenty eight related clinical trials and review articles in last 10 years. Since the balance of the coagulation system may tilt towards bleeding and thrombosis in many situations, it is mandatory for the clinicians to understand physiologic basis of haemostasis in order to diagnose and manage the abnormalities of the coagulation process and to interpret the diagnostic tests done for the same.

Clinical and hemostatic responses to treatment in ventilator-associated pneumonia: Role of bacterial pathogens*

To determine pathogen-specific kinetic changes in the alveolar procoagulant (PC) activity, tissue factor (TF), and tissue factor pathway inhibitor (TFPI) expression during the course of ventilator-associated pneumonia (VAP) and to assess the relationship between clinical resolution, intra-alveolar bacterial eradication, and restoration of hemostatic balance. Prospective, multiple-center study in a cohort of VAP patients. Two university-affiliated intensive care units. Thirty-five patients with microbiologically documented VAP who received adequate antimicrobial coverage and 13 controls. Nonbronchoscopic bronchoalveolar lavage was performed at the onset of VAP and on days 4 and 8 after initiation of antibiotic therapy. Samples were assayed for PC, TF, TFPI, and thrombin-antithrombin complex (TATc). The corresponding Clinical Pulmonary Infection Score (CPIS) was collected simultaneously. Isolated pathogens included Pseudomonas aeruginosa (n=13), methicillin-resistant Staphylococcus aureus (MRSA) (n=8), methicillin-sensitive S. aureus (MSSA) (n=7), and Escherichia coli (n=7). Although PC activity and TF were increased among the various pathogens at the onset of VAP, the levels of those with P. aeruginosa remained elevated at the end of treatment compared with controls and other etiological agents. TFPI levels were elevated for the duration of the study for all pathogens. A universal increase in TATc was noted at the onset of VAP, but the difference among the group of pathogens was significant at days 4 and 8 posttherapy. Despite the persisting hemostatic imbalance and incomplete intra-alveolar eradication of P. aeruginosa at end of therapy, the CPIS fell comparably at each time point irrespective of the etiological agents. Alveolar activation of the TF-dependent pathway may be species-specific in VAP and may not be adequately balanced by TFPI. The disparity between clinical response and eradication of P. aeruginosa from the intra-alveolar space suggests the need for biological markers to guide response to therapy.

Stroke in Young at High Altitude: A Case-Control Study from India.

There is limited knowledge about stroke in young individuals in high altitude (HA). We present the clinico-hematological profile of 21 cases of stroke in young seen between July 2000 to March 2002, at a hospital at a height of 3250 meters above mean sea level. All of them were investigated at the time of occurrence of the event with detailed hematological, neurological, biochemical, radiological, carotid doppler, neuroimaging and vasculitic workup. The thrombophilia work up was done at least 12 weeks after the occurrence of stroke, consequent to stopping anticoagulant therapy. Their mean age was 32.6 years (SD 7.9) and they were low landers staying at HA for 3 to 24 months (mean 8 months). 13 (62 percent) presented at heights between 3000–4000 meters and rest (38 percent) were evacuated from heights greater then 4000 meters. 14 (66.7 percent) had ischemic stroke, 2 (9.5 percent) had intracranial hemorrhage, 3 (14.3 percent) had TIA (transient ischemic attack)/RIND (reversible ischemic neurological deficiency) and 2 (9.5 percent) cerebro venous thrombosis. CT scan showed multiple cerebral infarcts in 10 (48 percent) cases. Initial presentation was considered to be high altitude cerebral edema (HACO) in all the cases. 2 (9.5 percent) presented with seizures and 1 (5 percent) had additionally left axillary artery thrombosis. 3 (14.3 percent) had primary hypertension, 2 were smokers, 2 had loose motions and 2-forced inactivity prior to event. All cases recovered adequate activities of daily living (ADL). Hyper homocysteinemia was found in 3 patients, protein C and S deficiency in 1 each respectively. Pro coagulant factors were identified in 5 cases (24 percent) as compared to stroke in young in low altitude where we earlier found these factors in 30%. The increased incidence of cerebral thrombosis at HA thus shows that long-term stay at HA is associated with a higher risk of stroke. Table shows the results of comparison of values of Hemoglobin, platelet count, BTG, PF4, fibrinogen and PAI-1, between patients at the time of stroke and controls matched for age, sex and duration of stay at HA. Platelet counts showed a significant fall in patients probably due to utilization in the acute thrombus. Platelet activation factors, BTG, PF4, fibrinogen and PAI-1 were significantly higher in the patients as compared to controls. Thereby showing that in patients who develop thrombosis at HA the milieu is pro thrombotic. Asymptomatic individuals who have protein C and S deficiency and hyper homocysteinemia may get cerebral thrombosis when inducted to HA.Comparison Of Hematological Tests Between Cases Of Stroke And Matched ControlsTests doneCases Mean (SD)Controls Mean (SD)P ValueHemoglobin (gm/dl)16.0 (1.09)16.5 (0.59)0.071 *Platelet Count (x10 ³/mm³)255 (53.89)350.75 (27.91)0.000¤Beta Thromboglobulin (IU/ml)59.45 (11.23)47.25 (5.64)0.000¤Platelet Factor 4 (IU/ml)18.64 (4.16)13.71 (3.39)0.000¤Fibrinogen (mg/dl)456.19 (83.65)395.25 (50.75)0.013¤PAI-1 (ng/ml)50.42 (7.38)39.32 (6.14)0.000¤* Not Statistically Significant; ¤ Statistically Significant

Calcium and calmodulin regulate lipopolysaccharide-induced alveolar macrophage production of tumor necrosis factor and procoagulant activity.

Alterations in macrophage (M phi) function are responsible, in part, for adult respiratory distress syndrome and multiple organ failure developing in patients with sepsis. Elucidation and control of these M phi mechanisms during sepsis are crucial to our understanding of this disease and, ultimately, to improving survival of these patients. To investigate the involvement of calcium flux in endotoxin-induced alveolar M phi production of tumor necrosis factor (TNF) and procoagulant (PC) activity. Rabbit alveolar M phi obtained by bronchoalveolar lavage were exposed to endotoxin in the form of lipopolysaccharide (LPS) extracted from Escherichia coli 0111:B4 in the presence of different specific calcium agonists and antagonists. The TNF expression was measured in the supernatant by L929 bioassays. The PC activity was determined in cell lysates by a one-step coagulation assay. Macrophages activated by LPS produce enormous levels of TNF and PC. Either W7 (20 mumol/L), a calmodulin antagonist, or TMB-8 (50 mumol/L), which prevents calcium release from the endoplasmic reticulum, inhibited production of both TNF and PC activity. Verapamil (50 mumol/L) alone or combined with TMB-8 significantly inhibited both TNF and PC production by LPS-stimulated M phi. Elevating intracellular calcium ([Ca2+]i), using the calcium ionophore, A23187, or thapsigargin alone, did not induce M phi production of TNF but significantly augmented LPS-stimulated TNF production. Our results indicate that increased intracellular calcium causing signal transduction activation through the calmodulin pathway is a necessary, but insufficient, component of the LPS signaling in M phi.

Disordered pathways of fibrin turnover in lung lavage of premature infants with respiratory distress syndrome.

Premature infants who have self-limited respiratory distress syndrome (RDS) rapidly improve, whereas infants with a complicated respiratory course are more likely to develop bronchopulmonary dysplasia (BPD), a chronic lung disorder that is the result of prolonged lung injury and impaired healing. The balance of competing activities of coagulation and fibrinolysis may contribute to the premature lung's response to acute injury and determine, in part, whether there is early resolution or protracted alveolar inflammation. To determine the relative activities of the coagulation and fibrinolytic pathways in neonatal lung injury, procoagulant (PC) and plasminogen activator (PA) activities were measured in undiluted cell-free lung lavage samples obtained serially over the first 28 days of life from 11 infants with self-limited RDS, 11 infants with evolving BPD, and 5 mechanically ventilated control infants without lung disease. Lung lavage from all three groups contained readily detectable procoagulant activity due mainly to the tissue factor-Factor VII complex. Plasminogen activator activity was relatively high in control lavage samples but depressed on the first day of life in the two groups of infants with lung disease: median, 0.3814 IU/ml (control); 0.0541 IU/ml (RDS); and 0.0454 IU/ml (BPD), p < 0.05 in each case compared with control. Two infants with severe lung disease had no detectable plasminogen activator activity in lung lavage on the first day of life. Depressed fibrinolytic activity correlated with severity of lung disease assessed radiographically and by pulmonary function measurements. Plasminogen activator activity was due to both tissue plasminogen activator and urokinase.(ABSTRACT TRUNCATED AT 250 WORDS)

Changes of Procoagulant and Fibrinolytic Activities in the Alveoli of Rats Exposed to Ozone.

The purpose of this study was to evaluate the role of ozone, a reactive product of environmental photochemical oxidation, in the development of pulmonary fibrosis. Male Wistar rats were exposed continuously to 0.5 ppm ozone for 1,4,7 and 14 days, and alveolar macrophages and lavage fluid obtained by bronchoalveolar lavage were examined. The results were as follows: 1) The total protein content in the lavage fluid was significantly increased compared to the control at 1 to 7 days by ozone exposure. Both alveolar macrophage and neutrophil counts increased in response to ozone exposure. However, approximately 90% of the free cells recovered were alveolar macrophages throughout the exposure period. 2) The plasminogen activator (PA) activity released from alveolar macrophages did not change in the group exposed for 1 day. But the activities were significantly high in the groups exposed for 4 to 14 days. 3) The PA activity of the lavage fluid showed a marked increase on the 1st day of ozone exposure, and subsequently decreased rapidly. However, the significantly increased activity was maintained throughout the exposure period. 4) In contrast, the procoagulant (PC) activity was unchanged on the 1st day of ozone exposure but the activity increased significantly on the 4th day, and was maintained at a high level until the 14th day. 5) The elastase inhibitory capacity (EIC) of the lavage fluid was significantly increased compared to the control by ozone exposure, but this difference was not seen throughout the exposure period when the EIC was corrected for the total protein content in the lavage fluid. These results revealed that both PA and PC activities increased in the alveolar fluid of rats exposed to 0.5 ppm ozone. The transition in the respective activities suggested that the fibrinolytic pathway in the alveoli was enhanced early in the exposure to ozone, while the coagulation pathway was enhanced later. This imbalance in coagulation homeostasis may be important in the regulation of fibrotic responses in the lungs of rats exposed to ozone. These findings are in agreement with morphological reports indicating that ozone exposure initially damaged the alveoli and later caused pulmonary fibrosis.

Concurrent expression of procoagulant and plasminogen activator activities by rabbit alveolar macrophages in vitro: Opposite modulating effects of prostaglandin E 2

We examined the effects of arachidonic acid metabolites on the simultaneous expression of procoagulant (PC) and plasminogen activator (PA) activities by rabbit alveolar macrophages. Incubation with lymphocyte-conditioned medium (LCM) caused a significant increase in cell-associated PC activity. Co-treatment with indomethacin (1 μM) reduced this augmentation in PC activity by 33% (p < 0.05). In contrast, indomethacin caused a 42% increase in PA activity released into incubation medium (p < .05). Both effects of indomethacin were reversed by the addition of PGE 2 in concentrations as low as 1 nM. Addition of 100 nM PGE 2 to these cells caused an increase in PC activity 2.7-fold greater than that achieved by LCM alone, while PGE 2 suppressed released PA activity by 62%. PGE 2 and indomethacin had similar but less pronounced effects on phorbol myristate acetate-treated cells. These effects of PGE 2 could be duplicated by PGE 1, but not by any other arachidonic acid metabolite (PGF 2α, PGI 2, PGD 2, ddPGF 2α, LTB 4, or LTC 4). While PGE 2 increases intracellular levels of cAMP, the observed effects on PC and PA activities could not be reproduced fully by treatment with dibutyryl cAMP. We conclude that PGE 2 amplifies the augmentation of PC activity by stimulated alveolar macrophages while concurrently inhibiting expression of plasminogen activator. This suggests that PGE 2 may be a significant mediator in regulating the highly interactive processes of inflammation and coagulation/fibrinolysis.

Procoagulant and plasminogen activator activities of bronchoalveolar fluid in patients with pulmonary sarcoidosis.

Tissue fibrin deposition may be an important component of inflammatory reactions. Current evidence suggests that intraalveolar procoagulant (PC) and plasminogen activator (PA) activities may be important determinants of local fibrin turnover in lung injury. In this study, we measured the PC and PA activities in cell-free bronchoalveolar lavage fluid (BALF) obtained from 17 patients with pulmonary sarcoidosis and 12 normal volunteers. Procoagulant activity was assayed by timing clot formation in a one-stage coagulation assay, and plasminogen activator activity was determined by measuring plasminogen-dependent lysis of [125I]fibrin. Mean PC activity in the sarcoidosis group was significantly elevated (102 +/- 25 versus 31.5 +/- 8.1 tissue thromboplastin units/ml; p less than 0.002), with 6 of 17 patient values beyond the 95% confidence limits of normals. These differences were not seen when PC activity was corrected for total protein in BAL. In contrast, PA activity tended to be lower in the sarcoidosis group (0.54 +/- 0.094 versus 0.643 +/- 0.106 Plough units/ml, p less than 0.3), and this difference became significant when PA was normalized to total protein (p less than 0.001). The ratio of procoagulant activity compared to plasminogen activator (PC/PA) was greater in the patients with sarcoidosis than normals (258 +/- 54 versus 40.3 +/- 6.4; p less than 0.001). The PC/PA ratios in 14 of 17 patients exceeded the 95% confidence limits of normals. In the sarcoidosis group, the PC/PA ratio correlated weakly with the number and percentage of lymphocytes retrieved by BAL. The plasminogen activator was a urokinase by molecular weight (53 kDa) and by comparing neutralization of PA activity by antibodies against urokinase and tissue plasminogen activator. The procoagulant was particulate and functioned as a factor X activator comprised of tissue thromboplastin and factor VII. We conclude that in pulmonary sarcoidosis, abnormal expression of procoagulant and plasminogen activator activities in alveolar fluid may favor accumulation of fibrin matrix at inflammatory foci.

Lymphocyte collaboration is not required for the induction of murine macrophage procoagulant by endotoxin

The putative requirement for lymphocytes as instructor cells in the induction of macrophage procoagulant (PCA) by endotoxin (LPS) was tested on elicited mouse peritoneal macrophages and on bone marrow-derived macrophages. Percoll purification of thioglycollate macrophages to at least 99.8 percent failed to diminish PCA induction by LPS. Bone marrow macrophages synthesized most PCA in response to LPS when they constituted more than 95 percent of the cells. In addition, PCA synthesis by these cells was not enhanced by the addition of splenic lymphocytes in a ratio of four to one. Exudate macrophages from endotoxin unresponsive C3H/HeJ mice failed to increase PCA synthesis in the presence of LPS. The addition of responsive C3H/HeN splenic lymphocytes to non-responsive HeJ macrophages did not permit LPS to induce the synthesis of PCA, suggesting the absence of unidirectional lymphocyte-instructed pathway. These data provide no evidence for lymphocyte collaboration in the LPS induction of murine macrophage PCA. LPS appears to induce PCA by acting directly on the macrophage.

Tissue fibrin deposition during acute lung injury in rabbits and its relationship to local expression of procoagulant and fibrinolytic activities.

Parenchymal fibrin deposition is well recognized in many forms of acute lung injury. Proteins derived from the actions of the coagulation and fibrinolytic systems may potentiate these inflammatory reactions as well as influence the subsequent repair process. However, the factors regulating fibrin formation and dissolution in acute pneumonitis have not been defined. In this study, we characterized the procoagulant (PC) and fibrinolytic activities simultaneously present in the alveolar space during the course of acute lung injury induced in rabbits by an intravenous injection of phorbol myristate acetate (PMA). Within 6 h of PMA injection, this injury was characterized histologically by extensive intra-alveolar fibrin formation and marked accumulation in pulmonary parenchyma of intravenously administered 125I-fibrinogen. Clearance of fibrin ensued over the remainder of the 72-h study period. Normal BAL fluid contained high levels of procoagulant activity which did not vary after the onset of inflammation. The procoagulant activity was attributed to particle-bound tissue thromboplastin as well as other factors of the extrinsic coagulation pathway. There were low levels of plasminogen activator (PA) activity in normal BAL fluid, but the mean activity increased 9.3-fold over control values by 12 h after PMA injection (p less than 0.01). When plasminogen activator activity in BAL fluid was referenced to the concomitant procoagulant activity, this ratio (PA/PC) increased 17.8-fold over controls, peaking 24 h after PMA injection (p less than 0.01). The levels of both procoagulant and plasminogen activator activities associated with alveolar macrophages were stable during the study period. Compared to alveolar macrophages, granulocytes expressed similar levels of plasminogen activator but negligible procoagulant activity.(ABSTRACT TRUNCATED AT 250 WORDS)

Procoagulant and fibrinolytic activity of human ovarian carcinoma cells in culture

We have investigated here the coordinate expression of both procoagulant (PCA) and fibrinolytic (FA) activity of cells from 16 human ovarian carcinoma cases. To avoid interference of contaminating host cells, we used cells isolated in primary culture from ascitic fluid or from solid tumor. The FA was determined in cellular extracts by an amidolytic assay in the presence of fibrin monomers. FA, which was plasminogen dependent in almost all of the cases, showed a wide range of activity (from < 0.001 to 2.30 UK units/mg protein). The molecular analysis of plasminogen activator (by SDS-PAGE and fibrin autography) showed a single molecular form of 52,000 daltons, inhibited by an antibody against human urokinase. PCA, studied with a one stage clotting assay in disrupted cells, was of tissue thromboplastin type in all instances and varied from 12.0 to 1300 thromboplastin units/ 10 4 cells. No simple correlation was found between FA and PCA in the cellular samples studied; moreover, for neither parameter was it possible to find any changes with the staging of the disease.

An innate host response to the neoplastic cell: syngeneic rat tumor cells can elicit a rapid de novo lymphoid procoagulant response.

Abstract To analyze unique molecular differences between normal and neoplastic cells, we have examined host responses to tumor cells. The present study provides the first evidence for an innate rapid recognitive response of the lymphoid system to some syngeneic tumors. The lymphoid procoagulant (PCA) response, a T cell-instructed monocyte response that activates proteases of the coagulation cascade culminating in thrombin formation, is considered a component of classic delayed-type hypersensitivity responses. We have demonstrated that three syngeneic rat mammary carcinomas elicit this cellular response in vitro in lymphoid cells of the unimmunized rat. The response was rapid, reaching maximum within 6 hr. Analysis was compounded by the constitutive PCA activity of some tumors; however, the PCA product produced in the response to tumor challenge in vitro was newly biosynthesized and was of lymphoid cell origin, differing from the PCA of tumor cells. The lymphoid PCA response was prothrombinase-like and did not require vitamin K for biosynthesis, nor were other gamma-carboxylation-dependent extrinsic pathway proteases other than prothrombin required for thrombin generation. Both in vivo and in vitro derived mammary carcinoma cells elicited the response, whereas a fibrosarcoma and nontransformed syngeneic cells did not. Tumor shed substances, which were devoid of PCA and sedimentable only in part at 100,000 X G, induced this cellular response. The same stimuli shed from tumor cells did not directly elicit a PCA response from elicited peritoneal macrophages; however, in the presence of T lymphocytes a PCA response of these macrophages was produced. This study provides novel information to indicate that a T-enriched lymphocyte-dependent monocyte-macrophage response to some tumors, before effective in vivo immunization, may participate in initial local protease generation and fibrin deposition, both thought to play a significant role in the local pathobiology of tumors.

936 PROCOAGULANT (PC) AND PLASMINOGEN ACTIVATOR (PA) FUNCTION OF CHILDHOOD LEUKEMIC BLASTS

The incidence and etiology of coagulopathy in childhood acute leukemia (AL) have not been well characterized. We have studied 11 children with AL prospectively at the time of diagnosis or relapse. A DIC screen, fibrinopeptide A (FPA) assay and assays of blast cell procoagulant (PC) and plasminogen activator (PA) were performed. Patients were evaluated for clinical prognostic features, blast cell morphology and surface markers. Blast cells were separated on Ficoll-Hypaque and then used as a source of procoagulant material in normal plasma and factor VIII, X and VII deficient plasma. Fibrinolytic activity was measured using the substrate S-2251 with des-fibrin I. We found: Overt laboratory evidence of DIC was present at diagnosis and worsened after therapy in one patient with ALL. This patient's blasts produced the highest level of procoagulant material. From these preliminary data, we conclude: 1. Coagulation abnormalities are common in childhood AL. 2. FPA levels are frequently increased in childhood AL. 3. Some leukemic cells produce potent procoagulant activity which may be associated with overt DIC. 4. ALL cells also frequently produce a plasminogen activator. 5. Production of PC and/or PA by blast cells was not clearly related to morphologic or immunologic characteristics.

Induction of a Fibrin-Gel Investment: An Early Event in Line 10 Hepatocarcinoma Growth Mediated by Tumor-Secreted Products

Abstract Within a few hours of implantation in the subcutaneous space of strain 2 guinea pigs syngeneic line 10 tumors became invested in a fibrin-gel. We hypothesized that tumors might induce and modulate this fibrin-gel by secreting molecules capable of activating the host's clotting and fibrinolytic systems. To test this possibility, line 10 tumor cells, and several non-neoplastic control cells, were cultured for 2 to 4 hr in serum-free medium and the culture supernatants were assayed for their capacity to activate host inflammatory pathways. Four distinct mediator activities were recognized in line 10 tumor cell culture supernatants: a vascular permeability factor (VPF), a procoagulant (PC), a plasminogen activator (PA), and a macrophage-migration inhibitory activity (MIF). Nonmalignant control cells secreted none of these activities with the exception of rabbit kidney cells that not unexpectedly released a PA activity, presumably urokinase. Based on conditions of culture, physical and chemical properties, inhibitor studies, and kinetics of secretion, it is likely that four distinct molecules are responsible for these activities. The complexity of the culture medium was not critical but low pH was an important variable favoring PA secretion. Secretion of all mediators was inhibited by cold and that of VPF and PA by puromycin. Cytochalasin B and EDTA caused significant increases in PA secretion. The line 10 PA, like that from other sources, was irreversibly inactivated by diisopropyl fluorophosphate and is probably a serine esterase. The capacity to secrete mediators that activate important host inflammatory pathways provides tumors with a mechanism for stroma induction and may be an important factor in determining a tumor's histologic pattern and degree of malignancy.

Uroplastin, Its Nature and Its Excretion Pattern in Health and in Certain Disease States

SummaryExperimental studies with a partially purified, potent procoagulant from human urine are reported. The material having the electrophoretic property of a â-globulin appears to functionally closely resemble tissue thromboplastin. A significant difference in the excretion of this pro coagulant between normal males and females and deviations from the normal excretion pattern in patients with impaired kidney function and certain other disease states are demonstrated. Possible pathophysiological implications of these findings are discussed.