Two Cases of Dysfibrinogenemia in Childhood

Introduction

Hemostasis in childhood differs from that of adults; and the hemostatic system is still developing during childhood. These differences offer a protective advantage to children with hemorrhagic and thrombotic complications. Plasma levels of coagulation factors (except for fibrinogen, factor V and factor VIII), as well as plasma levels of protein C, protein S and antithrombin are reduced. Hereditary dysfibrinogenemia is a rare disorder wherein an inherited abnormality in fibrinogen structure may result in defective fibrin function and/or structure. Clinical symptoms may vary from asymptomatic to life-threatening bleeding complications. Venous or arterial thrombotic complications occur extremely rarely during childhood. Fibrinogen, a key component in hemostasis, is a 340-kDa glycoprotein. The molecule consists of three different pairs of polypeptide chains (Aα, Bβ, and γ) each encoded by a distinct gene (FGA, FGB, and FGG). N-terminal parts of the Aα chain - fibrinopeptides A and the Bβ chain - fibrinopeptides B are situated in the central part of the molecule and block polymerization of the molecules. Conversion of fibrinogen to fibrin occurs after the cleavage of N-terminal fibrinopeptides by the serine protease thrombin. Correct conformation of fibrinopeptides is important for the cleavage by thrombin.

Methods

Routine coagulation tests were performed with citrated plasma samples on a STA-R coagulation analyzer. The functional fibrinogen level was measured by the Clauss method. Total fibrinogen level was determined by an immunoturbidimetric assay performed on a UV-2401PC spectrophotometer. Fibrin polymerization induced by either thrombin or reptilase and fibrinolysis experiments were obtained by the turbidimetrical method. Fibrinopeptide release was measured as a function of time; and the fibrinopeptides were determined by the reversed-phase, high-performance liquid chromatography (RP-HPLC) method. The purified genomic DNA was amplified by polymerase chain reaction, using specific primers; and dideoxysequencing was performed with Dye Terminator Cycle Sequencing with a Quick Start kit and a CEQ 8000 genetic analysis system).

Results

We have examined two unrelated boys aged 4 (boy 1) and 14 (boy 2) for susp. dysfibrinogenemia. Routine coagulation tests revealed prolonged thrombin and reptilase time in both boys and also showed decreased functional fibrinogen levels in both kids (Tab. 1). Boy 1 presented with bleeding manifestation – easy bruising, epistaxis, and bleeding after tooth extraction. His 26-year-old mother presented with similar coagulation findings and with mild bleeding complications. Boy 2 was asymptomatic. Fibrin polymerization experiments carried out on plasma samples showed prolonged lag time and significantly reduced final turbidity in both cases. Measurement of kinetics of fibrinopeptide release showed a decreased amount of the released fibrinopeptide A in both patients. DNA sequencing of boy 1 revealed a point mutation in exon 2 of the FGA gene at the position 3456 G/A, which causes the substitution of Aα 16 Arg to His (fibrinogen Praha V). The boy was found to be heterozygous for the mutation as well as his mother. DNA analysis of boy 2 revealed the same point mutation in the FGAgene, causing the same substitution of Aα 16 Arg to His (fibrinogen Kralupy nad Vltavou).

Conclusion

In this study we report two cases of congenital defects in the fibrinogen Aα Arg16-Gly17 bond found during childhood. It has been described earlier that the replacement of Aα 16 arginine by histidine decelerates thrombin catalyzed fibrinopeptide A release. Although both kids presented with the same genetic defect and with similar coagulation results, they have different clinical manifestation of the disease.

Acknowledgment

This work was supported by the project of the Ministry of Health of the Czech Republic for conceptual development of the research organization 00023736, by Grant from the Academy of Sciences, Czech Republic (P205/12/G118), and by ERDF OPPK CZ.2.16/3.1.00/28007.