Incorporation of fibrin molecules containing fibrinopeptide A alters clot ultrastructure and decreases permeability

Previous studies have shown that a heterozygous mutation in the fibrinogen Aalpha chain gene, which results in an Aalpha R16C substitution, causes fibrinolytic resistance in the fibrin clot. This mutation prevents thrombin cleavage of fibrinopeptide A from mutant Aalpha R16C chains, but not from wild-type Aalpha chains. However, the mechanism underlying the fibrinolytic resistance is unclear. Therefore, this study investigated the biophysical properties of the mutant fibrin that contribute to fibrinolytic resistance. Fibrin clots made from the mutant fibrinogen incorporated molecules containing fibrinopeptide A into the polymerised clot, which resulted in a 'spiky' clot ultrastructure with barbed fibrin strands. The clots were less stiff than normal fibrin and were cross-linked slower by activated FXIII, but had an increased average fiber diameter, were more dense, had smaller pores and were less permeable. Protein sequencing showed that unclottable fibrinogen remaining in the supernatant consisted entirely of homodimeric Aalpha R16C fibrinogen, whereas both cleaved wild-type alpha chains and uncleaved Aalpha R16C chains were in the fibrin clot. Therefore, fibrinolytic resistance of the mutant clots is probably a result of altered clot ultrastructure caused by the incorporation of fibrin molecules containing fibrinopeptide A, resulting in larger diameter fibers and decreased permeability to fibrinolytic enzymes.     

Studies on the ultrastructure of fibrin lacking fibrinopeptide B (beta- fibrin)

Release of fibrinopeptide B from fibrinogen by copperhead venom procoagulant enzyme results in a form of fibrin (beta-fibrin) with weaker self-aggregation characteristics than the normal product (alpha beta-fibrin) produced by release of fibrinopeptides A (FPA) and B (FPB) by thrombin. We investigated the ultrastructure of these two types of fibrin as well as that of beta-fibrin prepared from fibrinogen Metz (A alpha 16 Arg----Cys), a homozygous dysfibrinogenemic mutant that does not release FPA. At 14 degrees C and physiologic solvent conditions (0.15 mol/L of NaCl, 0.015 mol/L of Tris buffer pH 7.4), the turbidity (350 nm) of rapidly polymerizing alpha beta-fibrin (thrombin 1 to 2 U/mL) plateaued in less than 6 min and formed a "coarse" matrix consisting of anastomosing fiber bundles (mean diameter 92 nm). More slowly polymerizing alpha beta-fibrin (thrombin 0.01 and 0.001 U/mL) surpassed this turbidity after greater than or equal to 60 minutes and concomitantly developed a network of thicker fiber bundles (mean diameters 118 and 186 nm, respectively). Such matrices also contained networks of highly branched, twisting, "fine" fibrils (fiber diameters 7 to 30 nm) that are usually characteristic of matrices formed at high ionic strength and pH. Slowly polymerizing beta-fibrin, like slowly polymerizing alpha beta-fibrin, displayed considerable quantities of fine matrix in addition to an underlying thick cable network (mean fiber diameter 135 nm), whereas rapidly polymerizing beta-fibrin monomer was comprised almost exclusively of wide, poorly anastomosed, striated cables (mean diameter 212 nm). Metz beta-fibrin clots were more fragile than those of normal beta-fibrin and were comprised almost entirely of a fine network. Metz fibrin could be induced, however, to form thick fiber bundles (mean diameter 76 nm) in the presence of albumin at a concentration (500 mumol/L) in the physiologic range and resembled a Metz plasma fibrin clot in that regard. The diminished capacity of Metz beta-fibrin to form thick fiber bundles may be due to impaired use or occupancy of a polymerization site exposed by FPB release. Our results indicate that twisting fibrils are an inherent structural feature of all forms of assembling fibrin, and suggest that mature beta-fibrin or alpha beta-fibrin clots develop from networks of thin fibrils that have the ability to coalesce to form thicker fiber bundles.     

Comparison of polymerization of ancrod and thrombin fibrin monomers.

The polymerization of thrombin and ancrod fibrin monomers was studied with a standardized technique that evaluated turbidity changes and protein incorporation into the clot. Ancrod fibrin monomers were found to polymerize more slowly and form less turbid clots (at identical protein concentrations). Changes in ionic strength and pH influences ancrod fibrin monomer polymerization to a greater extent than thrombin fibrin monomer polymerization. Benzyltriethylammonium chloride was shown to be a potent inhibitor of fibrin monomer polymerization, with a greater inhibitory effect on ancrod fibrin monomers than on thrombin fibrin monomers. The differences between ancrod and thrombin fibrin may play a role in the infrequent thrombotic complications reported with ancrod therapy.     

Intraperitoneal heparin in peritoneal dialysis and its effect on fibrinopeptide A in plasma and dialysate

In 6 patients on continuous ambulatory peritoneal dialysis we investigated the inhibition of intraperitoneal fibrin formation by heparin. A continuous addition of 500 U of heparin per liter dialysate was used for 52 h. In plasma no heparin activity could be detected, even 52 h after intraperitoneal administration of heparin. The fibrin formation was determined by fibrinopeptide A, a thrombin-induced split product of fibrinogen. In patients under regular continuous ambulatory peritoneal dialysis we determined the fibrinopeptide A concentrations in plasma. The values were comparable with the fibrinopeptide A concentrations measured in disseminated intravascular coagulopathy. They decreased during intraperitoneal administration of heparin from 63.2 +/- 11.8 to 4.9 +/- 1.7 ng/ml. The fibrinopeptide A concentration in the 4-hour intraperitoneal dialysate (155.8 +/- 15.7 ng/ml) decreased after heparin administration to 8.5 +/- 2.0 ng/ml and was always higher than in plasma. We conclude that 500 U heparin per liter dialysate prevents the intraperitoneal fibrin formation. The low antithrombin III concentration (0.44 +/- 0.13 mg/dl) in protein-poor dialysate seems to be sufficient to inhibit the thrombin activity after acceleration by heparin.     

Autoantibody to plasma fibrinopeptide A in a patient with a severe acquired haemorrhagic syndrome.

We describe a 50-year-old man with a severe acquired haemorrhagic syndrome. He had slightly prolonged clotting times using bovine thrombin, human thrombin and reptilase. His plasma contained a polyclonal IgG which interfered with the generation of fibrin monomers without inhibiting the aggregation of preformed monomers. The inhibitor delayed thrombin-induced fibrinopeptide A release. The IgG bound to insolubilized synthetic fibrinopeptide A (one binding site per molecule) and, with higher affinity, to fibrinogen (two binding sites per molecule). It did not bind to insolubilized fibrin monomers. The IgG did not impair the catalytic activity of thrombin toward a small synthetic substrate but inhibited the binding of thrombin to fibrinogen without binding to thrombin. The binding of the anti-fibrinopeptide A autoantibody to fibrinogen might have impaired thrombin-induced fibrinogen to fibrin conversion in vivo. This may have favoured the reported haemorrhagic syndrome which was associated with severe chronic renal insufficiency.     

Determination of human fibrinopeptide A by radioimmunoassay in purified systems and in the blood.

The formation of fibrin clots or circulating soluble fibrin is accompanied by the appearance of fibrinopeptides. Measurement of the fibrinopeptide concentration in plasma can provide important information on the rate of conversion of fibrinogen to fibrin by thrombin. This rate varies under different physiologic and pathologic conditions. Fibrinopeptide A is a better molecular marker of the conversion than fibrinopeptide B since it is the first peptide to be cleaved by thrombin. A radioimmunoassay technique has been developed for the quantitative determination of human fibrinopeptide A. The procedure detects human fibrinopeptide A at a concentration of approximately 0.05 ng/ml. The variation of fibrinopeptide A content in normal persons may reflect its rapid formation and catabolism. A significantly increased concentration of this peptide was found in a patient during defibrination therapy with a purified enzyme from the venom of Agkistrodon rhodostoma and in patients suffering from retinal vascular occlusions.     

Fibrin clots obtained from plasma containing heparin show a higher sensitivity to t-PA-induced lysis.

Although heparin is currently used in concomitance with thrombolytic agents to improve their efficacy, its effect on fibrinolysis is controversial. We have evaluated the sensitivity to t-PA-induced lysis of clots prepared from plasma preincubated in vitro with therapeutic concentrations of heparin. The extent of t-PA-induced lysis was significantly increased by preincubation of plasma with 0.5 and 1.0 U/ml heparin. The concentration of t-PA required to give similar lysis rates were reduced by up to five times after adding 1.0 U/ml heparin to plasma prior to clot formation. Heparin added to the t-PA-containing medium after clot formation did not exert any significant effect. The effect of heparin was not mediated by the inhibition of thrombin as preincubation of plasma with hirudin did not modify clot sensitivity to t-PA. We also found that heparin significantly modified fibrin assembly and clot structure as assessed by a turbidimetric assay. Pre-incubation of fibrinogen with heparin caused an increase in the speed of fibrin fibre polymerization and in the turbidity of the final fibrin gel; changes known to be associated with the formation of thicker fibrin fibres. Thus the effect of heparin on clot sensitivity to lysis appears to be due to an increased permeability of these clots to fibrinolytic components. This may contribute to the antithrombotic activity and to the haemorrhagic risk of heparin. These findings could be particularly important for clinical thrombolysis.(ABSTRACT TRUNCATED AT 250 WORDS)     

Early cross-linked fibrin in human plasma contains alpha-polymers with intact fibrinopeptide A.

alpha-polymer formation, as opposed to gamma-chain dimerization has been considered a relatively late event in factor XIII-induced fibrin stabilization. Recently it has been shown, however, that plasma from healthy individuals and from patients with fibrinaemia contains small amounts of soluble fibrin/fibrinogen oligomers interlinked through dimerized gamma-chains as well as cross-linked alpha-chains. The present work was carried out to see if these early alpha-chain polymers also arise during coagulation of plasma in vitro. Plasma samples from healthy individuals, prepared by immediate centrifugation of blood collected without anticoagulant, were allowed to clot spontaneously for varying periods. The plasma clots were solubilized in SDS-urea-mercaptoethanol and samples were subjected to SDS-PAGE and Western blotting using polyclonal antibodies to human fibrinogen, or monoclonal antibodies specific either for A alpha/alpha-chains, for fibrinopeptide A-containing chains, for the N-terminus of the fibrin beta-chain or for the gamma-chains. Fibrin/fibrinogen oligomers were seen to form long before visible gelation of plasma. These oligomers were cross-linked through gamma-chain dimerization, but also through A alpha- or alpha-chain polymerization. The number and amount of alpha-polymers containing A alpha-chains increased immediately after clot formation, but these disappeared about 20 min later, due to complete removal of fibrinopeptide A (FPA) by thrombin. It is concluded that alpha-polymer formation is a very early event during plasma coagulation in vitro, and that both A alpha- and alpha-chains are involved.     

Disintegration and reorganization of fibrin networks during tissue-type plasminogen activator-induced clot lysis.

In this study, we investigated tissue-type plasminogen activator (tPA)-induced lysis of glutamic acid (glu)-plasminogen-containing or lysine (lys)-plasminogen-containing thrombin-induced fibrin clots. We measured clot development and plasmin-mediated clot disintegration by thromboelastography, and used scanning electron microscopy (SEM) to document the structural changes taking place during clot formation and lysis. These events occurred in three overlapping stages, which were initiated by the addition of thrombin, resulting first in fibrin polymerization and clot network organization (Stage I). Autolytic plasmin cleavage of glu-plasminogen at lys-77 generates lys-plasminogen, exposing lysine binding sites in its kringle domains. The presence of lys-plasminogen within the thrombin-induced fibrin clot enhanced network reorganization to form thicker fibers as well as globular complexes containing fibrin and lys-plasminogen having a greater level of turbidity and a higher elastic modulus (G) than occurred with thrombin alone. Lys-plasminogen or glu-plasminogen that had been incorporated into the fibrin clot was activated to plasmin by tPA admixed with the thrombin, and led directly to clot disintegration (Stage II) concomitant with fibrin network reorganization. The onset of Stage III (clot dissolution) was signaled by a sustained secondary rise in turbidity that was due to the combined effects of lys-plasminogen presence or its conversion from glu-plasminogen, plus clot network reorganization. SEM images documented dynamic structural changes in the lysing fibrin network and showed that the secondary turbidity rise was due to extensive reorganization of severed fibrils and fibers to form wide, occasionally branched fibers. These degraded structures contributed little, if anything, to the structural integrity of the residual clot, and eventually collapsed completely during the course of progressive clot dissolution. These results provide new perspectives on the major structural events that occur in the fibrin clot matrix during fibrinolysis.     

Fibrinogen Baltimore II: congenital hypodysfibrinogenemia with delayed release of fibrinopeptide B and decreased rate of fibrinogen synthesis.

A congenital hypodysfibrinogenemia, fibrinogen Baltimore II, was found in a young asymptomatic Caucasian female. Prothrombin, partial thromboplastin, and euglobulin lysis times were normal, as were platelet function and coagulation factor assays. Subnormal plasma fibrinogen levels were found using chronometric, rate-independent, and immunologic assay methods. Kinetic analysis of fibrinopeptide release revealed a delay in the thrombin-catalyzed release of fibrinopeptide B from the abnormal protein. Proteolysis of fibrinopeptide A by thrombin or Arvin, fibrin monomer polymerization, and fibrin polymer ligation occurred at normal rates. Catabolism of radiolabeled autologous and homologous fibrinogen was also normal, but the fibrinogen synthetic rate was less than half the normal value. Comparison of the coagulation characteristics of fibrinogen Baltimore II with those of other abnormal fibrinogens indicates that it represents a unique example of hypodysfibrinogenemia.     

Fibrinogen Milano. VI: A heterozygous dysfibrinogenemia (A alpha 16 Arg----His) with bleeding tendency

Congenital heterozygous dysfibrinogenemia was diagnosed in a young woman with bleeding tendency. 3 other asymptomatic members of her family (mother and the 2 sisters) had abnormal fibrinogen. The proposita's plasma exhibited prolonged thrombin and reptilase times. Plasma fibrinogen concentration determined by functional assay was 0.3 g/l, whereas immunologic assay revealed normal fibrinogen levels. Turbidity curves, representing the rate of thrombin-induced fibrin formation, were markedly delayed both in the presence and absence of Ca2+. Isoelectric focusing and SDS electrophoresis of reduced fibrinogen showed normal charge and size of the subunit chains. Release of fibrinopeptide B by thrombin was normal, whereas HPLC elution diagrams of fibrinopeptide A showed an abnormal peak A* with a slightly shorter retention time than the normal fibrinopeptide A. The amino acid analysis showed that the arginine in peak A* is replaced by histidine (A alpha 16 Arg----His).     

A novel mutation (deletion of Aalpha-Asn 80) in an abnormal fibrinogen: fibrinogen Caracas VI. Consequences of disruption of the coiled coil for the polymerization of fibrin: peculiar clot structure and diminished stiffness of the clot.

An abnormal fibrinogen was identified in a 10-year-old male with a mild bleeding tendency; several years later, the patient developed a thrombotic event. Fibrin polymerization of plasma from the propositus and his mother, as measured by turbidity, was impaired. Plasmin digestion of fibrinogen and thrombin bound to the clot were both normal. The structure of clots from both plasma and purified fibrinogen was characterized by permeability, scanning electron microscopy and rheological measurements. Permeability of patients' clots was abnormal, although some measurements were not reliable because the clots were not mechanically stable. Consistent with these results, the stiffness of patients' clots was decreased approximately two-fold. Electron microscopy revealed that the patients' clots were very heterogeneous in structure. DNA sequencing of the propositus and his mother revealed a new unique point mutation that gives rise to a fibrinogen molecule with a missing amino acid residue at Aalpha-Asn 80. This new mutation, which would disrupt the alpha-helical coiled-coil structure, emphasizes the importance of this part of the molecule for fibrin polymerization and clot structure. This abnormal fibrinogen has been named fibrinogen Caracas VI.     

Polyphosphate enhances fibrin clot structure

Polyphosphate, a linear polymer of inorganic phosphate, is present in platelet dense granules and is secreted on platelet activation. We recently reported that polyphosphate is a potent hemostatic regulator, serving to activate the contact pathway of blood clotting and accelerate factor V activation. Because polyphosphate did not alter thrombin clotting times, it appeared to exert all its procoagulant actions upstream of thrombin. We now report that polyphosphate enhances fibrin clot structure in a calcium-dependent manner. Fibrin clots formed in the presence of polyphosphate had up to 3-fold higher turbidity, had higher mass-length ratios, and exhibited thicker fibers in scanning electron micrographs. The ability of polyphosphate to enhance fibrin clot turbidity was independent of factor XIIIa activity. When plasmin or a combination of plasminogen and tissue plasminogen activators were included in clotting reactions, fibrin clots formed in the presence of polyphosphate exhibited prolonged clot lysis times. Release of polyphosphate from activated platelets or infectious microorganisms may play an important role in modulating fibrin clot structure and increasing its resistance to fibrinolysis. Polyphosphate may also be useful in enhancing the structure of surgical fibrin sealants.     

Substitution of the gamma-chain Asn308 disturbs the D:D interface affecting fibrin polymerization, fibrinopeptide B release, and FXIIIa-catalyzed cross-linking

Crystallographic structures indicate that gamma-chain residue Asn308 participates in D:D interactions and indeed substitutions of gammaAsn308 with lysine or isoleucine have been identified in dysfibrinogens with impaired polymerization. To probe the role of Asn308 in polymerization, we synthesized 3 variant fibrinogens: gammaAsn308 changed to lysine (gammaN308K), isoleucine (gammaN308I), and alanine (gammaN308A). We measured thrombin-catalyzed polymerization by turbidity, fibrinopeptide release by high-performance liquid chromatography, and factor XIIIa-catalyzed cross-linking by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. In the absence of added calcium, polymerization was clearly impaired with all 3 variants. In contrast, at 0.1 mM calcium, only polymerization of gammaN308K remained markedly abnormal. The release of thrombin-catalyzed fibrinopeptide B (FpB) was delayed in the absence of calcium, whereas at 1 mM calcium FpB release was delayed only with gammaN308K. Factor XIIIa-catalyzed gamma-gamma dimer formation was delayed with fibrinogen (in absence of thrombin), whereas with fibrin (in presence of thrombin) gamma-gamma dimer formation of only gammaN308K was delayed. These data corroborate the recognized link between FpB release and polymerization. They show fibrin cross-link formation likely depends on the structure of protofibrils. Together, our results show substitution of Asn308 with a hydrophobic residue altered neither polymer formation nor polymer structure at physiologic calcium concentrations, whereas substitution with lysine altered both.     

Hereditary dysfibrinogenemia in a patient with thrombotic disease

A new case of congenital dysfibrinogenemia, in which the patient has severe thrombotic disease, is reported. The abnormal fibrinogen molecules are characterized by normal fibrinopeptide release with thrombin and defective polymerization in the formation of fibrin. Clotting times with ancrod and reptilase are significantly prolonged. All other coagulation tests (except those for fibrinogen function) are normal, and the patient has no other underlying disease. The apparent paradox of defective fibrinogen, which clots abnormally and is yet associated with thrombotic disease, can be explained by further analysis of the patient's fibrinogen. The two important functional properties of this fibrinogen are: (1) it forms fibrin gels that are extremely rigid, and (2) the fibrin is highly resistant to lysis by plasmin. Thus, although the abnormal fibrinogen forms defective clots, the fibrin that is formed cannot be removed by the fibrinolytic system. These results provide a molecular explanation for the thrombotic disease in this patient. This abnormal fibrinogen appears to have unique characteristics and has been designated as fibrinogen Chapel Hill Ill.     

An acquired inhibitor that produced a delay of fibrinopeptide B release in an asymptomatic patient

An asymptomatic, 29-year-old woman was referred to our hospital before surgery because in the basic study of hemostasis she showed a prolonged thrombin time (TT) and a normal reptilase time (RT). She had not received any anticoagulants so, to account for these abnormal results the presence of an inhibitor or a dysfibrinogenemia was suspected. A 1:1 mixture of the patient's plasma with control plasma did not correct the TT. Dysfibrinogenemia was excluded because the defibrinated plasma retained the inhibitory activity when mixed with normal plasma. When 0.02 mg/ml of Protamine Sulphate (a concentration that neutralizes 1 U/mL of heparin in normal plasma) was added to the patient's plasma, the inhibitory activity did not disappear. IgG from the patient and from normal serum was isolated. The patient's IgG was able to prolong the TT of a normal plasma and of a purified fibrinogen. The patient IgG did not impair the catalytic activity of thrombin, because no difference was observed in the hydrolysis of S-2238 by 1 U NIH human thrombin with normal or patient IgG. The time course of the thrombin-mediated fibrinopeptide-release from normal fibrinogen with the patient's IgG, showed a delay in the fibrinopeptide B (FPB) release without affecting the fibrinopeptide A (FPA) release. This patient has an IgG antibody that delays fibrinopeptide B release of fibrinogen.     

The factor XIII V34L polymorphism accelerates thrombin activation of factor XIII and affects cross-linked fibrin structure

Factor XIII on activation by thrombin cross-links fibrin. A common polymorphism Val to Leu at position 34 in the FXIII A subunit is under investigation as a risk determinant of thrombosis. Because Val34Leu is close to the thrombin cleavage site, the hypothesis that it would alter the function of FXIII was tested. Analysis of FXIII subunit proteolysis by thrombin using sodium dodecyl sulfate-polyacrylamide gel electrophoresis and high-performance liquid chromatography showed that FXIII 34Leu was cleaved by thrombin more rapidly and by lower doses than 34Val. Mass spectrometry of isolated activation peptides confirmed the predicted single methyl group difference and demonstrated that the thrombin cleavage site is unaltered by Val34Leu. Kinetic analysis of activation peptide release demonstrated that the catalytic efficiency (k(cat)/K(m)) of thrombin was 0.5 for FXIII 34Leu and 0.2 (micromol/L)(-1) x sec(-1) for 34Val. Presence of fibrin increased the catalytic efficiency to 4.8 and 2.2 (micromol/L)(-1) x sec(-1), respectively. Although the 34Leu peptide was released at a similar rate as fibrinopeptide A, the 34Val peptide was released more slowly than fibrinopeptide A but more quickly than fibrinopeptide B generation. Cross-linking of gamma- and alpha-chains appeared earlier when fibrin was incubated with FXIII 34Leu than with 34Val. Fully activated 34Leu and 34Val FXIII showed similar cross-linking activity. Analysis of fibrin clots prepared using plasma from FXIII 34Leu subjects by turbidity and permeability measurements showed reduced fiber mass/length ratio and porosity compared to 34Val. The structural differences were confirmed by electron microscopy. These results demonstrate that Val34Leu accelerates activation of FXIII by thrombin and consequently affects the structure of the cross-linked fibrin clot.     

Zinc inhibits FPA release and increases fibrin turbidity

Physiologic concentrations of Zn(II) (4-40 microM) can increase the rate of thrombin-induced fibrin clot formation (decreased clotting time, CT) and increase the turbidity of the fibrin gel. Both the initial and ultimate turbidity (AbS 600 nm) of fibrin gels are increased in the presence of Zn(II). Two techniques were used to elaborate the mechanisms of Zn+2 procoagulant effect. Analytical ultracentrifugation indicates that Zn(II) does not induce the formation of fibrinogen multimers. Radioimmunoassay for FPA indicates that thrombin activation of fibrinogen is decreased by Zn(II), with 50% inhibition of FPA release observed at 35 microM Zn(II). These experiments indicate that the critical feature of Zn(II) procoagulant effect is not due to the induction of fibrinogen proteolysis by thrombin, which is actually decreased. Rather, it appears that Zn(II) accelerates the polymerization step of fibrin assembly and concomitantly modifies fibrin gel structure.     

Functional characterization of fibrinogen Bicêtre II: a gamma 308 Asn-->Lys mutation located near the fibrin D:D interaction sites.

The effects of the gamma-308 Asn-->Lys substitution of fibrinogen Bicêtre II on clot formation, structure and properties were determined to elucidate the role of this part of the molecule in fibrin polymerization. This process was followed by measurement of turbidity, and the structure and biophysical characteristics of the clots were studied by permeation, scanning electron microscopy, and rheological techniques. Turbidity studies revealed an increased lag period and greater final turbidity for fibrin BII clots, indicating impaired oligomer formation. By permeation it was found that BII clots had greater network porosity, four times more than that of the control. The clot architecture visualized by scanning electron microscopy was similar to that of control clots with pore size and fiber diameter slightly increased. BII clots had a stiffness decreased by more than half, and an increased loss tangent, a measure of the inelastic deformation of the clot. All these results suggest a disruption of the proper alignment of fibrin monomers during oligomer formation. Consistent with these results, fibrin cross-linking by adding the physiological concentration of factor XIII to the purified protein showed that gamma and alpha chain cross-linking was impaired in BII clots. This amino acid substitution defines distinctive effects on the surface of the D:D interaction sites that are reflected in the clot structure and functional properties.     

Abnormal fibrin ultrastructure, polymerization, and clot retraction in multiple myeloma

We have investigated two patients with multiple myeloma who had a prolonged thrombin clotting time and absent clot retraction but no haemorrhagic diathesis. Coagulation and platelet function studies, as well as electron microscopy of the fibrin clot, indicate abnormal fibrin polymerization; adherence of myeloma protein to very thin fibrin strands results in a rigid translucent clot which fails to retract. This abnormality is not necessarily associated with a haemorrhagic diathesis.