The effect of fulminant hepatic failure on protein C antigen and activity.

In eighteen patients with fulminant hepatic failure (FHF), in grade III or IV coma, both protein C antigen and activity were significantly decreased (0.35 +/- 0.03 u/ml and 0.35 +/- 0.03 u/ml respectively). There was a significant correlation between protein C antigen and activity (r = 0.61, p less than 0.01). Protein C antigen levels were inversely correlated with prothrombin time (r = -0.57, p less than 0.05) as were protein C activity levels (r = -0.57, p less than 0.05). There was also significant correlations between fibrinogen and protein C antigen (r = 0.69, p less than 0.01) and protein C activity (r = 0.61, p less than 0.01). These results demonstrate that the naturally occurring inhibitor of coagulation, protein C, is present at low levels in FHF and this is probably due to the lack of synthesis of the protein in the damaged liver. The low levels of protein C may make these patients more susceptible to the disseminated intravascular coagulation which is known to occur in FHF and this in turn will lead to a further reduction in protein C levels.

Increased factor VIII complex in fulminant hepatic failure.

In 30 patients with fulminant hepatic failure (FHF) all three components of the factor VIII complex were significantly increased (VIIIC = 6.43 +/- 1.12 u/ml; VIIIRAg = 3.91 +/- 0.25 u/ml; VIIIvWF = 3.89 +/- 0.29 u/ml). There was good correlation between all three parameters in control subjects, but only between VIIIRAg and VIIIvWF (r = 0.67; p less than 0.001) in patients with FHF. VIIIC was significantly higher than VIIIRAg and VIIIvWF. These results suggest that VIIIC and VIIIRAg are increased by different mechanisms in FHF. These processes may include endothelial cell damage, reduced reticuloendothelial system function and lack of production of inactivating substances by the damaged liver. Platelet adhesion to glass beads was increased in FHF (36.4 +/- 5.9% compared to 16.6 +/- 2.1%, p less than 0.01). There was no significant correlation between platelet adhesion and any of the parameters of the factor VIII complex. Thus the increase in platelet adhesion cannot be due to the increase in VIIIvWF in FHF.

Heparin response and clearance in acute and chronic liver disease.

Patients with liver disease are at risk of bleeding due to abnormalities of the clotting system although they must be anticoagulated if they require haemodialysis or haemoperfusion. The anticoagulant of choice is heparin. In this study we have investigated heparin kinetics in patients with fulminant hepatic failure (FHF) after a single intravenous dose of heparin (2,500 units) and found there was an increased clearance of heparin whether measured by its anti-Xa effect (t 1/2 = 27.8 +/- 2.9 min compared to t 1/2 = 50.2 +/- 2.7 min in normal controls p less than 0.001) or by the whole blood activated clotting time (t 1/2 = 23.7 +/- 2.2 min compared to t 1/2 = 37.0 +/- 2.0 min p less than 0.001). There was a decreased peak level of heparin measured by anti-Xa effect (peak level in FHF = 0.48 +/- 0.05 u/ml and in controls = 0.69 +/- 0.04 u/ml, p less than 0.02), but an increased sensitivity to heparin (sensitivity in FHF = 0.072 +/- 0.011 sec/unit, in controls 0.033 +/- 0.003 sec/unit, p less than 0.001). Patients with FHF had very low levels of antithrombin III (AT III), but there was no correlation between this and any parameters of heparin effect or clearance. In a group of patients with chronic liver disease heparin kinetics did not differ from controls despite low levels of AT III. The changes in heparin kinetics in FHF are likely to be complex with the balance between the proteins that act as cofactors, (e.g. AT III) and the proteins that have heparin neutralising activity, controlling the response of added heparin.

Increased elastase-alpha 1-antitrypsin complex in fulminant hepatic failure: relationship to bacterial infection and activation of coagulation.

To study the effect of infection, a frequent complication of fulminant hepatic failure (FHF), on the release of elastase from polymorphonuclear leucocytes and its inhibition in circulation we have measured the concentrations of alpha 1-antitrypsin, which binds and inhibits elastase in the circulation, and of elastase-alpha 1-antitrypsin complex, in 30 patients with FHF. Elastase-alpha 1-antitrypsin complex was significantly increased in FHF as compared to controls (303 +/- 51 micrograms/l compared to 37 +/- 5 micrograms/l; n = 10; P less than 0.001) demonstrating activation of leucocytes in FHF. Infection caused greater release of leucocyte elastase, complex levels were significantly greater in patients who were infected when compared to those who were not (463 +/- 84 micrograms/l; n = 13 compared to 180 +/- 46 micrograms/l; n = 17; P less than 0.01). Also patients who survived had significantly lower complex levels than those who did not (212 +/- 49 micrograms/l; n = 18 compared to 440 +/- 94 micrograms/l; n = 12; P less than 0.02). alpha 1-Antitrypsin activity was not significantly different from control subjects (0.99 +/- 0.06 U/ml compared to 0.97 +/- 0.05 U/ml). However alpha 1-antitrypsin activity was significantly higher in patients who survived (1.17 +/- 0.05 U/ml; n = 18) compared to those who did not (0.71 +/- 0.03 U/ml; n = 12; P less than 0.001) and patients who died had significantly lower levels than control subjects (P less than 0.01) indicating the importance of maintenance of normal inhibitor levels in patients with FHF. The leucocyte activation and release of elastase in FHF was linked to activation of the coagulation system; elastase--alpha 1-antitrypsin complex levels correlated significantly with thrombin-antithrombin III complex levels (r = 0.68; P less than 0.001) and inversely with fibrinogen (r = -0.71; P less than 0.001).

Fibrinolytic abnormalities following liver transplantation in patients with fulminant hepatic failure.

OBJECTIVE: To evaluate the fibrinolytic system after liver transplantation in patients with fulminant hepatic failure. DESIGN: Seven patients were studied prior to, and for 4 days after, liver transplantation. METHODS: Both activators and inhibitors of the fibrinolytic system were investigated in seven patients with fulminant hepatic failure who underwent liver transplantation. RESULTS: alpha 2-antiplasmin and C1-inhibitor levels increased rapidly after transplantation (81 and 53% of normal on day 1; 106 and 99% on day 2, respectively). Plasminogen levels remained low throughout the 4-day study period. Plasminogen activator inhibitor-1 was higher than normal before transplantation (21.0 compared with 7.4 U/ml) and increased further on the first day after operation (37.5 U/ml; P < 0.05 versus pre-transplantation). Tissue plasminogen activator levels remained normal (pre-operative, 7.0 IU/ml; Day 4, 0.2 IU/ml). D-dimer remained elevated during the postoperative period showing increased fibrinolytic activity. Thrombin-antithrombin III complex was also elevated during the study period. Antithrombin III was greatly reduced prior to transplantation (13.7% of normal) and plasma levels were less than 50% of normal values during the study. CONCLUSIONS: Measures of fibrinolytic activity are raised after liver transplantation in patients with fulminant hepatic failure. This is probably due to increased fibrin formation caused by a coexisting hypercoagulable state.

Physiological inhibitors of coagulation in fulminant hepatic failure.

To study the effect of the severe loss of hepatic synthetic function on the inhibitors of coagulation we have measured protein S (total and free), protein C, heparin cofactor II and antithrombin III in 30 patients with fulminant hepatic failure. The results showed severe reduction in all inhibitor levels with mean (+/- SE) values of: protein S, 0.26 +/- 0.03 U/ml; protein C, 0.26 +/- 0.03 U/ml; heparin cofactor II, 0.12 +/- 0.02 U/ml and antithrombin III, 0.21 +/- 0.02 U/ml. Heparin cofactor II was significantly lower than the other inhibitors (P less than 0.01). Although the reduction in free protein S was significant in fulminant hepatic failure as compared to normal subjects (0.40 +/- 0.05 U/ml compared to 1.02 +/- 0.08 U/ml, P less than 0.001), the ratio of free to total protein S was significantly increased (0.67 +/- 0.02 compared to 0.40 +/- 0.04, P less than 0.01). Prothrombin time (INR) was significantly inversely correlated with total protein S (r = -0.56, P less than 0.001) and free protein S (r = -0.48, P less than 0.01), but not with the ratio of free to total protein S. No significant correlation between the different coagulation inhibitors and other measures of hepatic function could be detected. Although the loss of hepatic synthetic function appears to be the major cause of the loss of coagulation inhibitors, other effects such as increased consumption and rate of clearance may play a role. The balance of these will be reflected in the circulating levels of the coagulation inhibitors.

Hepatocyte growth factor and plasminogen activation in fulminant hepatic failure.

Human hepatocyte growth factor (hHGF) has considerable sequence homology with plasminogen and both proteins can be activated by plasminogen activators. The aim of this study was to investigate the relationship between plasma hHGF and fibrinolysis in patients with fulminant hepatic failure (FHF), in whom proteases of coagulation are known to be activated and hHGF levels have been shown to be raised as a consequence of hepatic regeneration. Serum hHGF measured by ELISA was increased in FHF (median 6.67 ng/ml, range 1.2-62 ng/ml), but the values did not correlate with the decreased plasminogen level (median 9%., range 0.7-35.5%) or the level of t-PA which was normal. There was a significant correlation between serum hHGF and increased plasma D-dimer (median 2,163 microgram/l, range 39-7 311 microgram/l), produced by the action of plasmin on fibrin and increased plasma thrombin-antithrombin III complexes (TAT, median 31.7 microgram/l, range 3.7-105 microgram/l). These relationship could be indicative of an involvement of blood coagulation, possibly a specific serine protease, in hHGF activity. After liver transplantation, plasma hHGF was rapidly cleared to almost normal levels, whereas D-dimer and TAT continued to be at elevated levels.

9 beta-methylcarbacyclin, a stable prostacyclin analogue, in prevention of platelet losses during charcoal haemoperfusion.

The effect of 9 beta-methylcarbacyclin, a stable analogue of prostacyclin, as a platelet protective agent, has been investigated during in-vitro charcoal haemoperfusion with fresh heparinized human blood, a model of progressive platelet damage. There were no platelet losses over 3 h perfusion after an initial bolus (1 microgram ml-1) of 9 beta-methylcarbacyclin (101.6 +/- s.e. 1.9% of initial value) whereas in the paired control circuit there was a significant reduction in the platelet level (73.5 +/- 4.1%, P less than 0.05). Prevention of rises in plasma thromboxane B2 by 9 beta-methylcarbacyclin confirmed the lack of platelet damage and a significant, but less marked, effect on white cell losses was observed. In a second series of experiments a bolus of 9 beta-methylcarbacyclin in one circuit was compared with prostacyclin infusion in the other which demonstrated that this prostaglandin analogue was as effective as prostacyclin and on the basis of these initial results it would merit clinical evaluation.

Determination of the ketone body ratio in fulminant hepatic failure.

The ratio of acetoacetate to beta-hydroxybutyrate, the ketone body ratio, was measured in arterial blood from 28 patients with fulminant hepatic failure as an index of the hepatic energy charge. The ketone body ratio was significantly reduced in the total group of patients with fulminant hepatic failure as compared with control subjects (0.27 +/- 0.03 SE as compared with 0.48 +/- 0.03; p less than 0.001). Patients who survived had significantly less reduction of the ketone body ratio on admission than those who died (0.39 +/- 0.06, n = 10 as compared with 0.20 +/- 0.02, n = 19; p less than 0.02). In seven patients who died, in whom ketone body ratio was measured less than 12 hours before death there was a significant decrease in ketone body ratio as compared with that on admission (0.24 +/- 0.05 to 0.15 +/- 0.04; p less than 0.05). In contrast, in seven patients who survived there was no significant change in ketone body ratio when measured within 12 hours of regaining consciousness as compared with the figures on admission. Measurement of arterial ketone body ratio may give an indication of prognosis, and may be of use in testing the efficacy of treatments which aim to enhance hepatic regeneration or to remove toxic substances that may reduce the hepatic energy charge.

In-vitro blood compatibility studies on a new polymer-coated charcoal for haemoperfusion.

The blood compatibility of new polyhema-coated charcoal (Biotec) has been studied in an in-vitro haemoperfusion circuit. The mean inlet platelet levels after 4 hours of perfusion were 99.5 +/- SE 4.2% of the initial value and mean white cell levels were 81.9 +/- SE 5.8% of initial. No rises in Swank screen filtration pressure were detected. Initial evaluation of this charcoal in patients with fulminant hepatic failure should be possible without platelet-protective agents.

Platelet function during haemoperfusion in acute liver failure.

The changes in platelet-related haemostatic parameters have been studied during haemoperfusion of eleven patients with acute liver failure. Five patients were treated by haemoperfusion with an albumin-coated resin column and six with a polymer-coated charcoal column. The platelet and white cell losses over four hours' haemoperfusion were small in both groups. Significant increases in beta-thromboglobulin (mean 341 +/- SE 145 ng/ml) were seen after one hour in the patients treated by charcoal haemoperfusion. One patient in the charcoal group with the greatest rises in beta-thromboglobulin (860 ng/ml) and screen filtration pressure (205 mmHg) developed severe hypotension and haemoperfusion was terminated after 1 hour. One patient in the resin group showed rapid consumption of heparin after 2 hours. Measurement of beta-thromboglobulin is a sensitive assay of platelet activation during haemoperfusion. Albumin-coated resin haemoperfusion appears to be a more blood-compatible procedure with respect to platelets than charcoal haemoperfusion.

Evaluation of the BioLogic-DT sorbent-suspension dialyser in patients with fulminant hepatic failure.

The BioLogic-DT sorbent suspension dialyser was developed to remove toxic substances from the blood of patients with liver failure. In the present study a randomised controlled trial was carried out in 10 patients with fulminant hepatic failure who had developed grade 4 encephalopathy to evaluate the safety and biocompatibility of the dialyser in such severely ill patients. A total of 18 treatments were performed in 5 patients. Haemodynamic stability was maintained throughout. There was a significant loss of platelets (163 +/- 34 to 101 +/- 13 x 10(9)/l) and decrease in plasma fibrinogen (0.53 +/- 0.09 to 0.31 +/- 0.08 g/l) with a rise in blood activated clotting time (190 +/- 17 to 223 +/- 22 sec)--not seen in the controls--, which was a result of the dialysis being carried out without the use of heparin as anticoagulant. Removal of metabolites by treatment was limited, with no significant effect on blood ammonia level and further developments of the system will be needed for this very sick group of patients.

Effect of ELAD liver support on plasma HGF and TGF-beta 1 in acute liver failure.

The aim of this study was to investigate the effects of treatment with the extracorporeal liver assist device (ELAD) in patients with acute liver failure (ALF) on plasma hepatocyte growth factor (HGF), the most potent growth factor, and transforming growth factor-beta 1 (TGF-beta 1), an inhibitory factor for liver regeneration. Initial plasma HGF, measured by ELISA, was significantly increased in the ALF patients (7.86 +/- SEM 1.76 ng/ml) compared with normal subjects (0.10 +/- 0.02 ng/ml, p < 0.001). After 6 hours of ELAD haemoperfusion, plasma HGF increased further (30.5 +/- 6.19 ng/ml, p < 0.001), with a subsequent decrease towards the initial value by 48 hours. Initial plasma levels of TGF-beta 1 determined by ELISA were significantly increased in the ALF patients (43.4 +/- 5.9 ng/ml) compared with normal subjects (25.1 +/- 2.3 ng/ml, p < 0.01), but there was no change in plasma TGF-beta 1 during the study period in either the ELAD or control ALF group. As HGF is a heparin-binding growth factor and similar changes in HGF were observed during CVVHD, one possible explanation is that heparin administered as anticoagulant for extracorporeal circulation is involved in the effects observed on HGF.

The use of prostacyclin, PGI2, in the prevention of platelet aggregation during charcoal haemoperfusion.

In previous studies severe side effects, including irreversible hypotension, platelet losses and rises in Swank screen filtration pressure, have been recorded during the treatment by charcoal haemoperfusion of patients with fulminant hepatic failure. It has also been demonstrated that rises in Swank screen filtration pressure in an in vitro test circuit are due to the presence of platelet aggregates. In this study Prostacyclin was infused at a constant rate, to give a blood concentration of 5 ng/ml, into the in vitro circuit just prior to the charcoal column. Rises in Swank screen filtration pressure were prevented, and it was also shown that the giving of a bolus dose of Prostacyclin (400 ng/ml blood concentration) reversed rises in Swank screen filtration pressure that have already occurred. On the basis of these findings, Prostacyclin would be expected to be of considerable clinical value in the prevention of adverse platelet reactions to charcoal haemoperfusion when carried out in patients with fulminant hepatic failure.

The use of an in vitro haemoperfusion circuit to evaluate the blood compatibility of albumin-coated Amberlite XAD-7 resin.

We have developed an in vitro haemoperfusion circuit, which closely simulates the conditions encountered in a clinical haemoperfusion, for testing the blood compatibility of new adsorbent materials. Fresh heparinised human blood is perfused for 2 hours over two model columns arranged in parallel closed circuits. The present study was performed to investigate the blood compatibility of human serum albumin-coated Amberlite XAD-7 resin. Mean platelet losses over 2 hours were greater following perfusion of the uncoated resin (44 +/- SE 5.6%) than the albumin-coated resin (17 +/- SE 2.2%) (p less than 0.01). Mean white cell losses were similar (55%) for both resins. No rises in Swank screen filtration pressure, which detects the presence of cellular aggregates in blood, were observed. Coating of XAD-7 resin with HSA therefore improves its blood compatibility with respect to platelet losses and we thus intend to use this preparation in a clinical trial of resin haemoperfusion in fulminant hepatic failure.

Temporary extracorporeal liver support for severe acute alcoholic hepatitis using the BioLogic-DT.

Patients with severe acute alcoholic hepatitis develop multiple organ failure which is associated with production of inflammatory cytokines and a poor prognosis. The aim of the present pilot study was to evaluate the effects of the BioLogic-DT sorption-suspension dialyser in patients with severe acute alcoholic hepatitis. Ten patients with encephalopathy (grade II-IV) were entered into the study, 5 received treatment with the BioLogic-DT for 6 hours daily for 3 days and 5 received conventional treatment as controls. The system was biocompatible with no adverse effects on blood pressure or platelet counts, factor V, fibrinogen or antithrombin III. No bleeding episodes were observed even with the use of small doses of heparin. After 3 days, blood ammonia was lower in the BioLogic-DT treated patients than in the controls, although blood lactate was higher. There were slight increases in plasma TNF and IL-8 during treatment over and above the higher levels present initially, possibly as a result of activation of white cells in the extracorporeal circuit. The further development of the BioLogic-DT dialyser with the addition of a plasma treatment module capable of removing cytokines would be worth evaluating in acute alcoholic hepatitis.

Effects of serial resin hemoperfusion in fulminant hepatic failure.

Resin hemoperfusion using an albumin coated Amberlite XAD-7 column was performed in 19 patients in coma due to fulminant hepatic failure. The procedure was clinically well tolerated, with good blood compatibility, platelet and white cell levels being 97.3 +/- SE 3.2% and 105 +/- 3.8% of the respective initial values after four hours hemoperfusion. No significant changes were observed in beta-thromboglobulin, screen filtration pressure, plasma electrolytes, calcium, protein or albumin. The total plasma bilirubin fell by a mean of 24 mumol/l, with a reduction in 21 of the 25 perfusions studied of up to 104 mumol/l during perfusion. Mean plasma levels of total bile acids were 137 +/- 19 mumol/l and 115 +/- 16 mumol/l respectively before and after four hours hemoperfusion. The amount of bile acids recovered by elution of the resin column was over three times greater than that apparent from the change in plasma levels. Column chromatography on Sephadex G-25 of material eluted from the resin column showed various peaks to be removed, including substances in the middle molecular weight range (1000-5000 daltons). Of the patients treated, eight (42%) survived to leave hospital.

The management of abnormalities of hemostasis in acute liver failure.

The liver is the primary site of synthesis of most coagulation and fibrinolytic proteins, and also plays a role in the clearance of hemostasis factors and their degradation products. In acute liver failure, these functions are severely disturbed, and the risk of hemorrhage is increased. Following a brief summary of the physiology of hemostasis, this review describes the nature and frequency of hemostatic abnormalities in acute liver failure. These abnormalities include quantitative and qualitative platelet defects, impaired synthesis and clearance of the coagulation factors and related inhibitory proteins, and enhanced fibrinolysis. Disseminated intravascular coagulation may also play a role, although this syndrome is difficult to distinguish from changes due to the failure of hepatic synthesis and clearance alone. At present, management options are limited to support with blood products, although pharmacological manipulation of the coagulation and fibrinolytic systems represent a potential area for future study.