The Value of Factor VIII Levels in Acquired Hypofibrinogenemia

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1 The Value of Factor VIII Levels in Acquired Hypofibrinogenemia JAMES J. CORRIGAN, JR., M.D., BOBBIE B. BENNETT, B.S., A BERNARD BUEFFEL, M.D. Section of Pediatric Hematology, Department of Pediatrics, University of Arizona Medical Center, Tucson, Arizona ABSTRACT Corrigan, James J., Jr., Bennett, Bobbie B., and Bueffel, Bernard: The value of factor VIII levels in acquired hypofibrinogenemia. Am. J. Clin. Pathol. 60: , Acquired hypofibrinogenemia can result from either severe hepatic disease or disseminated intravascular coagulation (DIC). Differentiation between these two entities in the laboratory can be difficult because of similar reductions in certain coagulation factors. In addition, both pathologic events may be present in the same patient. The purpose of this investigation was to study the coagulation mechanisms in 35 patients with acquired hypofibrinogenemia (< 150 mg. per 100 ml.). Of these, 14 had DIC, 18 had hepatic disorders, and three had combinations of hepatic disease and DIC. Similar reductions in Factors II and V were found in the DIC and hepatic-disease cases. Although the platelet counts and fibrin split products values (FSP's) tended to be normal with hepatic disease, and abnormal with DIC, many patients had thrombocytopenia and/or elevated FSP's for reasons other than DIC. Factor VIII levels were 404 ± 18 per cent with hepatic disease and 31 ± 14 per cent with DIC (normal range %), clearly different in the two gnoups. Since fibrinogen, Factor II and Factor V are synthesized by the liver and are consumed with DIC, the similar results are explainable. Factor VIII has been reported to be produced by the reticuloendothelial system, but more recent evidence suggests that the liver may play a major role in its synthesis. The data suggest that if the liver is involved in Factor VIII production it probably is not totally from the hepatocyte. The data show that Factor VIII levels can be used to distinguish DIC from hepatic disease in acquired hypofibrinogenemic states. Received March 5, 1973; received revised manuscript April 25, 1973; accepted for publication April 27, Address reprint requests to: Dr. Corrigan, Arizona Medical Center, Tucson, Arizona Presented in part at the joint meetings of the Western Society for Pediatric Research, Western Society for Clinical Research, and Western Section American Federation of Clinical Research, Carmel, California, February 4, 1972, and published in abstract form. 6 This work was supported in part by research grants AI and AI from the National Institutes of Health, Bethesda, Maryland, U.S.A. and general institutional research support funds. 897 THE CONCENTRATION of fibrinogen is reduced below normal hemostatic levels in a number of acquired pathologic states in man. The mechanisms which produce hypofibrinogenemia are inadequate production by the liver and excessive utilization or destruction by either intravascular coagulation or systemic hyperfibrinolysis ' 20 ' 24 In the course of our studies of disseminated intravascular coagulation (DIC) in

2 898 CORRIGAN T/fZ.. AJ.C.P. Vol. 60 Table 1. Coagulation and Fibrinolytic Data for Patients with Hepatic Disease and DIC Test Hepatic Disease (18 Patients) DIC (14 Patients) Normal Range Fibrinogen (mg. per 100 ml.) Factor II (%) Factor V (%) Platelets (x 10 3 per cu. mm.) Fibrin split products (% positive) Euglobulin lysis time and fibrin plate (% abnormal) * Mean. t Range. man, it was noted that patients with hepatocellular disease frequently had changes in their coagulation mechanism that could be confused with DIC. Successful therapy directed toward correcting the hemostatic defects depends upon determining which mechanism (or both) may be responsible for the coagulation defects. The purpose of this investigation was to analyze the coagulation mechanisms in 35 patients who had acquired hypofibrinogenemia, 18 of whom had severe hepatocellular disease, 14, classic DIC, and three, combinations of the two. The results show that the level of plasma coagulation factor VIII can be used to differentiate the hypofibrinogenemia due to hepatic disease from that resulting from DIC. Materials and Methods Clinical Material For this study, patients were selected on the basis of a fibrinogen concentration of less than 150 mg./100 ml. plasma. Eighteen cases of hepatic disease were studied. These consisted of six cases of acute hepatitis, one acute yellow atrophy, one Reyes syndrome, one subacute necrosis, three chronic active hepatitis, five cirrhosis, and one with 90% hepatectomy. 90* (0-145)t 29 (10-62) 23 (<l-78) 155 (26-400) (0-126) 28 (4-72) 14 (<l-39) 55 (4-100) The patients' ages ranged from 3 months to 52 years; 75% were less than 20 years old.:the female-to-male ratio was 1.8:1.0. Diagnoses were histologically confirmed by biopsy or at autopsy in all but one case. Three patients showed various severe derangements in other liver function studies in addition to the fibrinogen levels. No attempt was made to correlate the abnormal chemistries with the coagulation data, since this has been reported by other investigators ,25 No patient had received blood or blood components before the studies were obtained. Fourteen patients with laboratory and pathologic evidence of DIC were studied. Eleven cases were due to bacterial septicemia, one hemorrhagic shock secondary to a gunshot wound of the abdomen, one massive pulmonary embolus, and one postopen-heart surgery for cyanotic congenital heart disease. The septicemic patients were children and the other three were adult. Three patients with hepatic disease and DIC were also studied. These were children aged 3, 7, and 7!4 years with acute leukemia and fulminating hemorrhagic varicella. All three died; varicella hepatitis was found at necropsy in each case, and diffuse fibrin thrombi in one. Laboratory evidence of DIC was recorded in all three cases. 0

3 December 1973 FACTOR VIII A HYPOFIBRINOGENEMIA 899 Coagulation Methods Platelet counts were performed by the direct method of Brecher and Cronkite. 3 Plasma was obtained by anticoagulating 9 volumes of whole blood with 1 volume of citric acid-sodium citrate and processed as previously described. 7 The following determinations were performed on the plasma: activated partial thromboplastin time (PTT); 23 prothrombin time (PT); 18 thrombin time; 12 one-stage assays for Factors II, 17 V, 22 IX, 23 and VIII; 23 the P and P test of Owren and Aas; 17 fibrinogen concentration; 21 euglobulin clot lysis time; 5 unheated fibrin plate lysis. 1 Serum was obtained by collecting 1-2 ml. of whole blood into epsilon-aminocaproic acid and allowing it to clot for 2 hours in a sterile glass tube at 37 C. Bovine thrombin (100 units) was added to insure complete fibrinogen utilization. After centrifugation, the serum was stored at -20 C. until used for detection of fibrinolytic split products (FSP's) by immunologic technic. 15 >500-f ) 400 I 300- o 200 o <1 Liver Disease DIC FIG. 1. Comparison of Factor VIII levels in hepatic disease and DIC. The shaded area represents the normal range, each dot a single patient, and the small horizontal bar the mean value for each group. Table 2. Effects of Partial Hepatectomy on Coagulation Factors Test Fibrinogen (mg./loo ml.) Factor II (%) Factor V (%) Factor IX (%) P and P test 17 (%) Factor VIII (%) Preoperative (Not done) Results Hepatic Disease Cases Postoperative (Week) < All 18 patients had abnormally prolonged PTT's and PT's, and twelve had prolonged thrombin times. As can be seen on Table 1, the mean fibrinogen concentration was 90 mg. per 100 ml., with a range of Sixty-six per cent of the patients had fibrinogen levels below 100 mg. per 100 ml. Although the mean platelet count was normal, seven patients had fewer than 150,000 platelets. Of these, five had splenomegaly (four patients with cirrhosis and one with chronic active hepatitis). Factors V and II were abnormally low in 17 of 18 patients and the P and P test was reduced in all. Factor IX was measured in seven and found to be reduced (< 50%) in six. Factor VIII levels (Fig. 1) ranged from 152 to 800% (normal %). All patients were above the normal range, and 14 were above 200%. The effects of 90% hepatectomy in a child with a hepatoma are shown in Table 2. During the first postoperative week, severe hypofibrinogenemia with a reduction in Factors V, II, and IX occurred. Factor VIII was above normal and remained unchanged over the next month. The mean Factor VIII value for the hepatic-disease cases was 404%, clearly different from that of patients with DIC (p < 0.001).

4 900 CORRIGAN T/lL. A.J.C.P. Vol. 60 Table 3. Coagulation and Fibrinolytic Data in Three Cases of Hemorrhagic Varicella Test Day 1 Case 1 Day 2 Day 3 Day 1 Case 2 Day 2 Day 1 Case 3 Day 2 Normal Range Platelets (xlo 3 per cu. mm.) Fibrinogen (mg. per 100 ml.) Factor II (%) Factor V (%) Factor VIII (%) Fibrin split products Oxg. per ml.) Euglobulin lysis time (hours) Results of the fibrinolytic studies showed that 23% of the patients had elevated FSP's (>4.0 fig. per ml. serum), and 20% had euglobulin lysis times <2 hours and/or greater than 100 sq. mm. zones of lysis on unheated fibrin plates. DIC Cases All patients had abnormally prolonged PTT's, prothrombin times and thrombin times. The mean fibrinogen concentration in the 14 cases was 66 mg. per 100 ml. (Table 1), with 86% less than 100 mg. per 100 ml. All patients had thrombocytopenia and low Factor V levels and 13 had reduced Factor II. The mean Factor VIII level was 31% (Fig. 1); only one patient had a level within the normal range. The euglobulin lysis time was abnormally short in one patient, whereas 83% had abnormal amounts of FSP's in their sera. Three children with acute leukemia and fatal hemorrhagic chickenpox showed evidence of hepatitis at necropsy. Two had been in remission and all were taking various cytotoxic agents. The data (Table 3) show that all had thrombocytopenia, reduced Factors II, V, and fibrinogen, and elevated FSP's. Cases 2 and 3 had low Factor VIII. Case 1 had 80% reduction in Factor VIII as the varicella was <4.0.0 progressing. All patients received plasma and platelet concentrates. In Case 2 the patient was also given heparin (100 units per kg. body weight every 4 hours intravenously) and cytosine arabinoside (50 mg. per kg. intravenously). Discussion In this investigation patients were selected on the basis of having fibrinogen concentrations below 150 mg. per 100 ml. Two main groups were studied, those with severe hepatocellular disease and those with DIC. The data show that plasma coagulation Factors II and V could not be used to differentiate between the two groups. Although the platelet counts and serum FSP's tended to be normal in hepatic disease, and abnormal in DIC, 26% of the patients who had hepatic disease had thrombocytopenia and 23% had elevated FSP's. The reasons for this are probably related to the presence of hypersplenism in five cases, systemic hyperfibrinolysis as seen in 20%, and other factors as reported by others Factor VIII levels were clearly different in the two groups, being elevated in hepatic disease and reduced in DIC. The data suggest that when hypofibrinogenemia is due to underproduction, Factor VIII is normal to elevated, and conversely, if it is due to excessive utilization, Factor VIII is re-

5 December 1973 FACTOR VIII A HYPOFIBRINOGENEMIA 901 duced. Occasionally, one may find both mechanisms operative, as was demonstrated in the three cases of hemorrhagic varicella in this study. To define the mechanism, better turnover rates for fibrinogen could be performed. 4 However, at present these technics have not been refined, adequate controls are not completely defined, and availability is not universal, making this procedure impractical. Other investigators have suggested that DIC may be operative in hepatic disease, basing this presumption mainly on the response of patients to heparin. 19 Although heparin was not employed in the hepatic-disease cases in this study, the data do not support DIC as the mechanism for hypofibrinogenemia. Since heparin has protean therapeutic actions, its effect in this setting must be interpreted with caution. The liver is known to be the site of synthesis of many of the plasma proteins, and it could be anticipated that it is also the site of Factor VIII production. 26 It has been reported that Factor VIII levels are normal to increased in adult patients with chronic hepatocellular disease 20 and elevated in fatal hepatic necrosis. 213 In the present study the two children who died of acute viral hepatitis and had no identifiable parenchymal cells at necropsy and the one child with a 90% hepatectomy had elevated Factor VIII levels. In addition, normal levels have been noted in orthotopic transplantation of the human liver. 11 The data suggest that if the liver is the predominant site of Factor VIII production, it is not accomplished solely by way of the parenchymal cells. The reticuloendothelial part of the liver and other extrahepatic sites need to be studied further to elucidate this mechanism in man. 26 Since Factor VIII is measured as activity and not concentration, an alternative explanation could be that the Factor VIII molecule is made more reactive somehow and thus would assay as normal or elevated. 16 However, the use of two-stage systems of assay has not supported this view Nevertheless, in patients with acquired hypofibrinogenemia, Factor VIII levels appear to be a valuable guide to differentiate severe hepatocellular disease from DIC. References 1. Astrup T, Mullertz S: The fibrin plate method for estimating fibrinolytic activity. Arch Biochem 40: , Bohmig HJ, Abouna GM, Diez-Pardo JA: Coagulation studies in acute hepatic necrosis. Thromb Diath Haemorrh 26: , Brecher G, Cronkite EP: Estimation of the number of platelets by phase microscopy, Blood Coagulation, Hemorrhage and Thrombosis: Methods of Study. Second edition. Edited by LM Tocantin, LA Kazal. New York, Grune andstratton, 1964, p Brodsky I, Siegel NH, Kahn SB, et al: Simultaneous fibrinogen and platelet survival with 75-Se selenomethionine in man. Studies in diseases with normal coagulation and in hepatocellular disease with abnormal coagulation. Br J Haematol 18: , Buckell M: The effect of citrate on euglobulin methods of estimating fibrinolytic activity. J Clin Pathol 11: , Corrigan JJ Jr: Value of factor VIII levels in acquired hypofibrinogenemic states (abstr). Clin Res XX:263, Corrigan JJ Jr, Jordan CM: Heparin therapy in septicemia with disseminated intravascular coagulation. N Engl J Med 283: , Finbiner RB, McGovern JJ, Goldstein R, et al: Coagulation defects in liver disease and response to transfusions. Am J Med 26: , Fletcher AP, Bierderman O, Moore D, et al: Abnormal plasminogen-plasmin system activity (fibrinolysis) in patients with hepatic cirrhosis. Its causes and consequences. J Clin Invest 43: , GallusAS, Lucas CR, HirshJ: Coagulation studies in patients with acute infectious hepatitis. Br J Haematol 22: , Groth CG, Pechet L, Starzl TE: Coagulation during and after orthotopic transplantation of the human liver. Arch Surg 98:31-34, Hardisty RM, Ingram GIC: Bleeding Disorders: Investigation and Management. Oxford, Blackwell Scientific Publications, 1965, p Meili FO, Straub PW: Elevation of factor VIII in acute fatal liver necrosis. Thromb Diath Haemorrh 24: , Merskey C, Johnson AJ, Kleiner GJ, et al: The defibrination syndrome: Clinical features and laboratory diagnosis. Br J Haematol 13: , Merskey C, Lalezari P, Johnson AJ: A rapid, simple, sensitive method for measuring fibrinolytic split products in human serum. Proc Soc Exp Biol Med 131: , 1969

6 902 CORRIGAN r^l. A.J.C.P. Vol Niemetz J, Nossel HL: Activated coagulation factors: In-vivo and in-vitro studies. Br J Haematol 16: , Owren PA, Aas K: The control of dicumarol therapy and the quantitative determination of prothrombin and proconvertin. Scand J Lab Invest 3: , Quick AJ: Hemorrhagic Diseases. Philadelphia, Lea and Febiger, 1957, p Rake MO, Shilkin KB, Winch J, et al: Early and intensive therapy of intravascular coagulation in acute liver failure. Lancet II: , Rapaport SI, Ames SB, Mikkelsen S, et al: Plasma clotting factors in chronic hepatocellular disease. N Erigl J Med 263: , Ratnoff OD, Menzie C: New method for the determination of fibrinogen in small samples of plasma. J Lab Clin Med 37: , Shanberge JN, Matsuoka T, Fukui H: A simple method for preparation of a substrate for one-stage factor V assay. Am J Clin Path 47: , Simone JV, Vanderheiden J, Abildgaard CF: A semiautomatic one-stage factor VIII assay with commercially prepared standard. J Lab Clin Med 69: , Spector I, Corn M: Laboratory tests of hemostasis. The relation to hemorrhage in liver disease. Arch Intern Med 119: , Spector I, Corn M, Ticktin HE: Effect of plasma transfusions on the prothrombin time and clotting factors in liver disease. N Engl J Med 275: , Webster WP, Zukoski CF, Hutchin P, et al: Plasma factor VIII synthesis and control as revealed by canine organ transplantation. Am J Physiol 220: , 1971