Purification of a fibrinolysis inhibitor in serum from post-traumatic patients.

A fibrinolysis inhibitor was purified in serum from post-traumatic patients by the use of flat bed electrofocusing of serum desalted by gel chromatography followed by affinity chromatography on a column of matrix-linked plasminogen. Disc gel electrophoresis yielded one protein band. The inhibitor protein was also found in normal serum, but in a lower concentration.

Heparin-coated circuits reduce activation of granulocytes during cardiopulmonary bypass. A clinical study.

Activated granulocytes release highly active enzymes such as myeloperoxidase and lactoferrin, which can be involved in tissue destruction mediated by oxygen free radicals. Cardiopulmonary bypass has been reported to activate granulocytes. Bypass circuits coated with heparin have been shown to reduce release of granulocyte factors in experimental studies. In the present study, heparin-coated circuits were compared with noncoated circuits. In seven patients undergoing coronary bypass, heparin-coated circuits were used (group HC), and seven served as control patients (group C). In group HC the heparin dose was reduced to 75% (225 IU/kg). Group C had the standard dose of 300 IU/kg. No preoperative differences in myeloperoxidase and lactoferrin were observed between the groups. At the end of bypass in both groups, there was a significant increase of these enzymes (p less than 0.001) followed by a later decrease. In group HC, however, the release of myeloperoxidase was significantly lower than in group C (215 +/- 24 versus 573 +/- 133 micrograms/L, mean +/- standard error of the mean). The release of lactoferrin was significantly lower in group HC than in group C both at the end of cardiopulmonary bypass (659 +/- 79 versus 1448 +/- 121 micrograms/L) and 3 hours after bypass (224 +/- 37 versus 536 +/- 82 micrograms/L). Granulocytes as well as total number of leukocytes continued to increase until 1 hour after bypass (p less than 0.001) and then manifested a slow decrease. It was concluded that the use of heparin-coated circuits reduced the release of granulocyte factors because of lower activation of leukocytes.

Primary haemostasis, plasmatic coagulation and fibrinolysis in renal transplantation.

The disturbed coagulation state seen in patients with uremia has been suggested to contribute to thrombotic events in kidney grafts following transplantation. Primary haemostasis, plasmatic coagulation and fibrinolysis were investigated in eight patients before and during four weeks after kidney transplantation. In spite of an improved renal function there was postoperatively still a depressed platelet aggregation and the prostacyclin concentrations in plasma were low. The plasma coagulation seemed to be activated according to short activated partial thromboplastin time, high levels of FVIII:C and prothrombin complex. The fibrinolysis was increased and the PAI-1 levels were decreased. It is concluded that the overall haemostatic balance is characterized by a high degree of activation in uremic patients and that this activation persisted four weeks after transplantation.

Specific coagulation and fibrinolysis tests as biochemical markers in traumatic-induced adult respiratory distress syndrome.

There is a great demand for more specific methods for assaying individual components of coagulation and fibrinolysis, with the chief aim being to use them as biochemical markers of the Adult Respiratory Distress Syndrome (ARDS) induced by severe trauma. This prospective study was undertaken on 18 severely traumatized patients in various stages of shock admitted to the Intensive Care Unit of the University Hospital in Uppsala, Sweden. After haemodynamic restitution, during which surgery was often required as an intervening procedure, the patients were carefully studied regarding pulmonary function, coagulation and fibrinolysis. Eight patients developed ARDS according to our criteria, and one patient died from this condition. It was found that patients who developed ARDS had significantly lower levels of antithrombin-III, higher levels of von Willebrand factor levels, higher levels of tissue plasminogen activator inhibitors and lower levels of plasminogen as compared with those who did not develop this condition. We believe that these coagulation and fibrinolysis variables can be used along with appropriate pulmonary function tests as specific biochemical markers to disclose the development of traumatic-induced ARDS.

Fragmin (LMWH) vs heparin for anticoagulation during in vitro recycling of human blood in cardiopulmonary bypass circuits: dose-dependence and mechanisms of clotting.

Low-molecular-weight heparin (LMWH) (Fragmin) vs heparin was studied in vitro in order to investigate its antithrombotic efficacy in the isolated thrombogenic link of cardiopulmonary bypass (CPB). Fresh human blood (400 ml) with various dosages of the anticoagulant was recycled in a CPB circuit for 120 min. The standard dosage of heparin (1,500 IU, n = 6) was compared with a lower dosage (1,000 IU, n = 3) and several dosages of Fragmin (IU anti-FXa): 750 (n = 1), 1,500 (n = 3), 2,100 (n = 4) and 2,500 (n = 3). Clotting occurred in three Fragmin experiments at dosages of 750, 1,500 and 2,100 IU. This was associated with short activated clotting time (ACT) and activated partial thromboplastin time (aPTT) but was independent of the levels of anti-FXa, FVIII, von Willebrand factor and prothrombin complex. It was concluded that at least twice the dose of Fragmin (anti-FXa), compared with heparin, was required, suggesting that thrombin inhibition is crucial for the antithrombotic efficacy of heparin in CPB circuits. Absence of fibrinolytic markers suggests that the well known enhancement of fibrinolysis often seen during CPB, is not due to heparin interaction with normally circulating blood components, but rather to interaction with the vessel walls or to the surgical trauma itself.

Low-molecular-weight heparin (Fragmin) versus heparin for anticoagulation during cardiopulmonary bypass in open heart surgery, using a pig model.

Fragmin and heparin were studied in pigs during 120 min of cardiopulmonary bypass (CPB) and up to 240 min postoperatively, with respect to clotting, bleeding and the effects of protamine. Thirty-three pigs received bolus injections of 300 IU/kg with or without additional dosage during CPB and with or without subsequent protamine sulphate. Doses of Fragmin 60% higher were necessary to prevent clotting. These had 100% higher anti-FXa levels but about 50% shorter activated coagulation time (ACT) compared with heparin. Anti-FXa increased with cumulative doses of heparin and Fragmin but ACT and activated partial thromboplastin time (aPTT) did not, indicating a larger loss of thrombin inhibition compared with anti-FXa in both drugs during CPB. Thrombin inhibition was crucial for prevention of clotting. Protamine efficiently normalized ACT in the Fragmin group but left a residual 20% anti-FXa, which did not increase the bleeding tendency. Fragmin could adequately be monitored with ACT and would be a safe alternative to heparin in CPB.

Heparin-coated cardiopulmonary bypass circuits reduce blood cell trauma. Experiments in the pig.

Blood cell trauma and postoperative bleeding remain important problems in cardiopulmonary bypass (CPB). We compared heparin-coated with non-coated circuits in the pig. Twenty animals were perfused for 2 h at normothermia using membrane oxygenators (Bentley Bos 50). Two groups were studied. In the non-coated group (NC, n = 11) the CPB circuits used were without a heparin coating. This group had systemic heparinization of 400 IU/kg to maintain an ACT (activated clotting time) of over 400 s during CPB. In the coated group (C, n = 9), all surfaces exposed to blood in the CPB circuits were heparin-coated. This group had the heparin dose reduced to 25% (100 IU/kg) without further administration regardless of ACT. During CPB, group C displayed shorter ACT (per definition), higher platelet count, platelet adhesion and lower fibrinolysis and haemolysis (P less than 0.05) as compared to group NC. No thromboembolic events were detected during CPB. Three animals in group NC and 4 animals in group C were weaned from CPB and protaminized. Four hours postoperatively, the leucocyte consumption was two-fold greater and blood loss about four-fold greater in group NC as compared with group C (P less than 0.05). Perfusion with heparin-coated surfaces reduces blood cell trauma. The decreased postoperative blood loss observed in group C is probably explained by the reduced dosages of heparin and protamine.

Delayed incubation as an alternative method to sample storage for enumeration of E. coli and culturable bacteria in water.

The effects of sample storage on enumeration of Escherichia coli in marine bathing water and culturable bacteria in drinking water were evaluated. Results showed that overnight storage at 0-5 degrees C significantly reduced the counts of E. coli in bathing water (p = 0.0001) with a mean reduction of 25%. A similar effect of sample storage was observed for the enumeration of culturable bacteria in drinking water at 22 +/- 2 degrees C for 66 +/- 4 h (p = 0.0074; mean reduction = 25%) or at 36 +/- 2 degrees C for 44 +/- 4h (p = 0.0353; mean reduction = 6%). The use of a delayed incubation method, i.e. overnight storage at 0-5 degrees C of inoculated agar plates prior to incubation, did not significantly affect the counts of culturable bacteria when plates were incubated at 22 + 2 degrees C for 66 +/- 4 h, whereas it resulted in a significant increase of the bacterial numbers when plates were incubated at 36 +/- 2 degrees C for 44 +/- 4 h (p = 0.0002; mean increase = 32%). Based on these results, it is suggested to avoid the use of overnight or longer sample storage for the enumeration of E. coli in bathing water samples, as well as for the enumeration of culturable bacteria in drinking water. The delayed incubation method appears to be a reliable procedure for the enumeration of culturable bacteria and could represent a valid alternative to sample storage in order to overcome problems associated with the performance of bacteriological counts during weekends or statutory holidays. However, a multi-laboratory study is needed to evaluate the reproducibility of the delayed incubation method for the enumeration of culturable bacteria and its possible use for the enumeration of E. coli by membrane filtration.

Effect of nicotinic acid on the posttraumatic increase in free fatty acids and fibrinolysis inhibition activity in the rat.

Nicotinic acid effectively inhibited the posttraumatic increase in both free fatty acids (FFA) and fibrinolysis inhibition activity (FIA) in the blood in rats, indicating that FFA might be involved in the posttraumatic increase of FIA. The FIA in the liver was greater than that in other organs studied and was increased in the posttraumatic phase. The possible role of the liver in the posttraumatic increase of FIA is discussed.

Fibrinolysis inhibition after a major standardized trauma.

The present investigation on 20 patients after total hip replacement surgery has confirmed that the posttraumatic increase of the fibrinolysis inhibition activity (FIA) in serum and plasminogen-depleted serum is due to the primary fibrinolysis inhibitor (PFI, alpha 2-antiplasmin). This protein exists in at least two forms and it was indicated that PFI alpha with affinity to immobilized plasminogen, is mainly responsible for the posttraumatic variations of the FIA in plasminogen-depleted serum. PFI beta, the major part of the PFI-related antigen, which has none or low such affinity, displayed weak FIA and relatively small increase after the surgical trauma. It was established that the posttraumatic increase of the FIA was not derived from the low molecular weight fraction in serum of those patients.

Fibrinolysis inhibition and fibronectin in the blood in patients with the delayed microembolism syndrome.

Various parameters of fibrinolysis inhibition and the plasma concentration of fibronectin (alpha 2-surface binding glycoprotein, cold insoluble globulin) were measured in patients at risk of developing acute progressive respiratory sufficiency following trauma or sepsis - the delayed microembolism syndrome (DMS). Most parameters measuring fibrinolysis inhibition were significantly higher in the five patients with DMS than in five patients who did not develop the syndrome. Thus, the primary fibrinolysis inhibitor (alpha 2-antiplasmin) was enhanced and the alpha-form of this inhibitor, with affinity to plasminogen, showed the greatest increment and might be of major importance for the delayed elimination of fibrin from the lungs occurring in these patients. The fibronectin concentrations were not lower in patients with DMS than in those who did not develop the syndrome.

Heparin-coated cardiopulmonary bypass circuits and 25% reduction of heparin dose in coronary artery surgery--a clinical study.

Cardiopulmonary bypass with systemic heparinization causes trauma to blood cells and coagulation defects. Artificial surfaces could be coated by end-linkage binding of heparin (Carmeda Bioactive Surface, CBAS). Use of such surfaces during cardiopulmonary bypass in animals resulted in less postoperative blood loss and better preservation of blood cells. In this study heparin-coated circuits were employed during coronary artery grafting in 7 patients (Group HC). Concomitantly, the heparin dose was reduced by 25% and an activated clotting time (ACT) of 300 sec was accepted. Additional 7 patients were operated with standard circuits (Group C), requiring ACT above 400 sec with normal doses of heparin. There were no thromboembolic complications in Group HC. The postoperative bleeding was generally low and without significant intergroup differences. Coagulation parameters displayed significantly lower ACT and anti-Factor Xa during bypass in Group HC. A tendency towards less blood cell trauma was observed with heparin-coated circuits. The protamine dose could be reduced by 50%, which significantly reduced the protamine/heparin quotient. This study indicates that routine cardiopulmonary bypass could be performed safely with heparin-coated circuits and reduced intravenous doses of heparin and protamine. It is suggested that the use of heparin-coated circuits may lead to less blood cell trauma.

[Legionella pneumophila in pool water]. / Legionella pneumophila i bassinvand.

We investigated 87 samples of pool water for the presence of Legionella. The samples were from cold water pools (< 28 degrees C) and from hot water pools (> 32 degrees C). Sampling was furthermore done on normal water samples, on water from the bottom of pools and of water from departure from the activated carbon filters. Legionella was not detected in any of the samples from cold water pools, whereas in the hot water pools 10% of the pool water samples and 80% of the water from the filters were infected with Legionella pneumophila. The highest concentrations of Legionella were found in the filter samples, whereas the concentrations in the pool water (10-100 cfu/liter) were not alarming. This investigation demonstrates the potential risk of presence of Legionella in hot water pools with activated carbon filters being the site with best growth potential. It should be stressed that a high level of disinfection (at least 1.0 mg free chlorine/liter and pH 7.2) is essential for the prevention of Legionella in pool water at higher temperatures.

Induction of endogenous fibrinolysis inhibition in the dog. Effect of intravascular coagulation and release of free fatty acids.

Possible mechanisms underlying the development of fibrinolysis inhibition following trauma were studied. In order to investigate the role of intravascular coagulation dogs were subjected to infusions of thrombin or endotoxin, which both caused an increase in urokinase inhibitor activity in serum after 24 and 48 hours. The inhibitor increase following thrombin infusion was not, however, prevented by previous defibrinogenation with Defibrase or by induction of thrombocytopenia with antiplatelet serum, suggesting that neither platelets nor fibrinogen are necessary for the post-traumatic occurrence of fibrinolysis inhibition. In all groups subjected to infusion of thrombin an increase in plasma free fatty acids (FFA) was observed. The role of this increase for the development of fibrinolysis inhibition was tested by infusion of norepinephrine alone and in combination with nicotinic acid. Norepinephrine caused an increase of FFA after 2 hours and in urokinase inhibitor activity after 24-48 hours. Both of these were diminished by high doses of nicotinic acid, indicating that the release of FFA rather than intravascular coagulation might be the principal mechanism underlying the occurrence of fibrinolysis inhibition following trauma.

Coagulation, fibrinolysis and bleeding after open-heart surgery.

To investigate the disputed pathogenesis of excessive bleeding after open-heart surgery, variables representing different hemostatic systems were correlated to postoperative blood loss in 29 patients. The general bleeding tendency in the early postoperative phase was probably attributable to depletion of hemostatic agents due to hemodilution, decreased antiplasmin activity, instantaneous but reversible platelet dysfunction following protaminization, and the natural interval to development of complete hemostasis. Heavy bleeding (greater than 800 ml/16 h) occurred in ten patients, who had significantly reduced levels of von Willebrand factor and lower active platelet count than in eight patients with minor bleeding. Defective primary hemostasis thus seemed to be the main cause of increased postoperative bleeding in these patients. Determination of platelet function by glass retention test showed good clinical relevance and gave considerably more reliable diagnosis than conventional platelet count alone. The patient with the greatest blood loss also showed drastic decrease in the plasminogen-binding form of alpha 2-antiplasmin, suggesting that additionally impaired fibrinolysis inhibition may contribute to development of severe hemorrhagic complications.

Haemostasis at low heparin dosage during cardiopulmonary bypass with heparin-coated circuits in pigs.

Cardiopulmonary bypass (CPB) causes activation of cascade systems. Although heparin coating of CPB circuits improves biocompatibility, the effects on coagulation remain controversial. Theoretically, heparin coating should permit the reduction of systemic anticoagulation during CPB. We investigated influences on haemostatic variables in animal CPB, comparing heparin-coated circuits and reduced systemic heparinization (group HC) with uncoated circuits and full heparinization (group C). Twenty pigs underwent 2-h CPB. Seven (HC, n = 4; C, n = 3) were weaned from CPB and studied for up to 4 h. Total administered heparin was 470 +/- 6 IU/kg (mean +/- SEM) in group C and 100 +/- 0 IU/kg in group HC. Protamine dosage was significantly reduced in group HC. In group C, levels of prothrombin complex, factor VIII and adhesive platelets were reduced significantly during CPB, and postoperatively there were significantly lower values of prothrombin complex, fibrinogen, antithrombin III, factor VIII and adhesive platelets but a significantly increased concentration of von Willebrand factor and cumulative bleeding after 4 h. In conclusion, heparin-coated CPB circuits combined with lowered heparin dosage reduced coagulation factor consumption and preserved platelet function, possibly contributing to improved postoperative haemostasis.

Haemostasis at low heparin dosage during cardiopulmonary bypass with heparin-coated circuits in pigs.

Cardiopulmonary bypass (CPB) causes activation of cascade systems. Although heparin coating of CPB circuits improves biocompatibility, the effects on coagulation remain controversial. Theoretically, heparin coating should permit the reduction of systemic anticoagulation during CPB. We investigated influences on haemostatic variables in animal CPB, comparing heparin-coated circuits and reduced systemic heparinization (group HC) with uncoated circuits and full heparinization (group C). Twenty pigs underwent 2-h CPB. Seven (HC, n = 4; C, n = 3) were weaned from CPB and studied for up to 4 h. Total administered heparin was 470 +/- 6 IU/kg (mean +/- SEM) in group C and 100 +/- 0 IU/kg in group HC. Protamine dosage was significantly reduced in group HC. In group C, levels of prothrombin complex, factor VIII and adhesive platelets were reduced significantly during CPB, and postoperatively there were significantly lower values of prothrombin complex, fibrinogen antithrombin III, factor VIII and adhesive platelets but a significantly increased concentration of von Willebrand factor and cumulative bleeding after 4 h. In conclusion, heparin-coated CPB circuits combined with lowered heparin dosage reduced coagulation factor consumption and preserved platelet function, possibly contributing to improved postoperative haemostasis.

Role of extradural and of general anaesthesia in fibrinolysis and coagulation after total hip replacement.

Thirty patients undergoing total hip replacement were randomly allocated to one of two groups. One group (n = 14) received extradural anaesthesia with 0.5% bupivacaine with adrenaline continued into the postoperative period (24 h) for pain relief. The other group (n = 16) received general anaesthesia with controlled ventilation, using nitrous oxide in oxygen and fentanyl i.v. Following surgery they received a narcotic analgesic i.m. on demand. Analysis of fibrinolysis inhibition activity and plasminogen activators revealed a significantly better fibrinolytic function in patients given continuous extradural anaesthesia than in those who received general anaesthesia followed by narcotics in the period after operation. Furthermore, the capacity for activation of factor VIII was significantly lower after operation in the former group.