Fibrinogen Philadelphia. A hereditary hypodysfibrinogenemia characterized by fibrinogen hypercatabolism. J Martinez, … , S S Shapiro, A J Erslev J Clin Invest. 1974;53(2):600-611. https://doi.org/10.1172/JCI107595. Research Article A new, autosomally inherited abnormal fibrinogen associated with hypofibrinogenemia has been described in several members of a family. Plasma fibrinogen measured either as thrombin-clottable protein or by immunodiffusion revealed a fibrinogen level ranging between 60 and 90 mg/100 ml. The thrombin time of plasma or purified fibrinogen was prolonged and only partially corrected by the addition of calcium. Purified fibrinogen prolonged the thrombin time of normal plasma. Fibrinopeptide release by thrombin was normal in rate and amount, but fibrin monomer aggregation was grossly disturbed, especially in a high ionic strength medium. We have designated this fibrinogen "fibrinogen Philadelphia." Acrylamide gel electrophoresis of mixtures of [121I]normal and [125I]abnormal fibrinogens revealed a slight increase in the anodal mobility of fibrinogen Philadelphia. Similarly, DEAE-cellulose chromatography showed slightly stronger binding of fibrinogen Philadelphia than normal. To elucidate the mechanism responsible for the low plasma fibrinogen concentration, simultaneous metabolic studies of autologous (patient) and homologous (normal) fibrinogen, labeled with 125I and 121I, respectively, were performed in two affected subjects. Autologous fibrinogen half-life was short and the fractional catabolic rate was markedly increased in both family members. In contrast, homologous fibrinogen half-life and fractional catabolic rate were normal. These metabolic studies demonstrate that rapid degradation of fibrinogen Philadelphia is largely responsible for the depressed levels of a plasma fibrinogen. This represents the first example of a […] Find the latest version: https://jci.me/107595/pdf Fibrinogen Philadelphia A HEREDITARY HYPODYSFIBRINOGENEMIA CHARACTERIZED BY FIBRINOGEN HYPERCATABOLISM JOSE MARTINEZ, RUTH R. HOLBURN, SANDOR S. SHAPIRO, and ALLAN J. ERSLEV From the Cardeza Foundation for Hematologic Research and the Departments of Medicine and Physiology, Jefferson Medical College of Thomas Jefferson University, Philadelphia, Pennsylvania 19107 A B STR A CT A new, autosomally inherited abnormal Philadelphia is largely responsible for the depressed fibrinogen associated with hypofibrinogenemia has been levels of a plasma fibrinogen. This represents the first described in several members of a family. Plasma fibri- example of a mutant plasma protein in which the mo- nogen measured either as thrombin-clottable protein or lecular defect is associated with an altered catabolism. by immunodiffusion revealed a fibrinogen level rang- ing between 60 and 90 mg/100 ml. The thrombin time INTRODUCTION of plasma or purified fibrinogen was prolonged and only partially corrected by the addition of calcium. Puri- Inherited functional abnormalities of fibrinogen have fied fibrinogen prolonged the thrombin time of normal been described in several families (1-21). Clinical ex- plasma. Fibrinopeptide release by thrombin was nor- pression of the defect is variable, ranging from no mal in rate and amount, but fibrin monomer aggrega- clinical manifestations (1-11), to bleeding (9, 12-20) tion was grossly disturbed, especially in a high ionic and even thrombotic episodes (14, 20, 21). Wound strength medium. We have designated this fibrinogen dehiscence also has been reported (1, 3). The most "fibrinogen Philadelphia." Acrylamide gel electro- frequent abnormalities of coagulation tests are pro- phoresis of mixtures of ["'I]normal and ['I]abnormal longed prothrombin and thrombin times and an inhibi- fibrinogens revealed a slight increase in the anodal tory effect of patient plasma or fibrinogen on the mobility of fibrinogen Philadelphia. Similarly, DEAE- thrombin time of normal plasma. The functional defect cellulose chromatography showed slightly stronger bind- in these dysfibrinogenemias has been related either ing of fibrinogen Philadelphia than normal. To eluci- to an abnormality in the proteolytic release of fibrino- date the mechanism responsible for the low plasma peptides from fibrinogen by thrombin or to an ab- fibrinogen concentration, simultaneous metabolic stud- normality in the subsequent aggregation of fibrin mono- ies of autologous (patient) and homologous (normal) mers. The plasma fibrinogen concentration, when mea- fibrinogen, labeled with 1'I and 181I, respectively, were sured by methods based on thrombin-clottable pro- performed in two affected subjects. Autologous fibrin- tein, has generally been low (22), but assay by chem- ogen half-life was short and the fractional catabolic ical or immunochemical methods has resulted in normal *rate was markedly increased in both family members. values in all cases but one (12). In contrast, homologous fibrinogen half-life and frac- We have recently had the opportunity to study a tional catabolic rate were normal. These metabolic family with an abnormal fibrinogen characterized by studies demonstrate that rapid degradation of fibrinogen disturbed fibrin monomer aggregation and a low cir- culating plasma fibrinogen concentration determined by This work wvas presented in part at the 64th Annual both biological and immunochemical methods. Radio- Meeting of the American Society for Clinical Investigation, isotopic studies have demonstrated a much more rapid Atlantic City, N. J., April 29. 1972 and published in abstract form (J. Clin. Invest. 51: 62a, 1972). catabolism of this mutant fibrinogen that the normal Received for publication 18 June 1973 and in rcvised species, apparently accounting for the depressed fibri- form 15 October 1973. nogen levels found in affected members. These fea- 600 The Journal of Clinical Investigation Volume 53 February 1974- 600-L-611 tures differentiate this abnormal fibrinogen from pre- thrombin time of normal plasma was tested by mixing nor- viously described dysfibrinogenemias, and we propose mal plasma with various amounts of normal or propositus "fibrinogen Philadelphia." plasma or normal or propositus purified fibrinogen. The that it be designated purified fibrinogen solutions had equivalent concentrations (1.3 mg/ml) so that the total amount of fibrinogen was METHODS constant in the test system. Reptilase times were performed with venom from Bothrops atroxrs diluted with distilled Blood was collected with plastic syringes into 1/9 vol 0.1 water. The system was identical to the one described for M trisodium citrate and transferred to plastic tubes. After the thrombin time method, except that 0.1 ml (20 jug/ml) centrifugation for 15 min at 4VC and 2,500 g, the platelet- of Reptilase was substituted for thrombin. poor plasma was removed and either tested immediately or Fibrinogen concentration was determined by thrombin quick-frozen in acetone-dry ice and stored at -850C. clottability with the method of Ellis and Stransky (36), Fibrinogen purification. Normal fibrinogen was purified ammonium sulphate precipitation (37), and quantitative im- from ACD plasma obtained from healthy Australia antigen munodiffusion, by using commercial antibody-containing negative donors by the method of Kazal, Amsel, Miller, agar plates.' and Tocantins (23). The abnormal fibrinogen was purified Fibrinopeptide release during coagulation was investi- by the same method from ACD plasma obtained from the gated by a method similar to that described by Gralnick, propositus (J. S.) and her son (D. S.). Normal and D. S. Givelber, Shainoff, and Finlayson (17). Lyophilized fibrino- fibrinogens were lyophilized and stored at -850C; J. S. gen, containing sodium citrate, was dissolved in distilled fibrinogen was quick-frozen and stored at the same tem- water to a concentration of 3.5 mg/ml fibrinogen and 0.055 perature. The amount of fibrinogen present in the purified M citrate. 0.5-ml aliquots of this fibrinogen solution (1.75 materials was measured spectrophotometrically at 280 nm, mg) were added to a series of tubes containing 0.05 ml of pH 7.1, by assuming an A'801% of 13.9, the figure obtained bovine thrombin (2.5 U). At intervals 0.5 ml of 4% acetic by Kazal et al. (23). When normal fibrinogen is prepared acid was added to each tube. After the clot had dissolved, by this technique, the protein is consistently more than the mixture was treated with 0.05 ml 100% trichloroacetic 94% clottable. acid (TCA).5 30 min later the samples were centrifuged, Coagulation studies. Platelets were counted by the and the protein present in the supernate was measured by method of Brecher and Cronkite (24). The bleeding time the Folin-Lowry method (38), as modified by Gralnick was determined by the method of Ivy, Nelson, and Bucher et al. (17). Fibrinopeptides were also measured as the (25). Partial thromboplastin time and prothrombin time amount of TCA-soluble arginine present in the supernate were done by standard methods (26). Plasma prothrombin (17). was assayed by the two-stage technique of Ware and Fibrin monomer aggregation studies were done by several Seegers (27) slightly modified (28). Factor V was mea- methods. One was similar to the procedure described by sured by the correction of the prolonged prothrombin time von Felten, Frick, and Straub (39). 100 ml of plasma, con- of aged oxalated plasma (29). Factors VII and X were taining 0.5% Na EDTA and 25,000 U Trasylol 6 was diluted each measured by the correction of the one-stage prothrom- 1: 5 with 0.15 M NaCl and clotted with 100 U