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Fibrinogen gamma-chain splice variant gamma' alters fibrin formation and structure
Amy V Cooper1, Kristina F Standeven, Robert A S Ariëns
1Academic Unit of Molecular Vascular Medicine, University of Leeds, Leeds General Infirmary, United Kingdom.
Abstract:
Fibrinogen gammaA/gamma' results from alternative splicing of mRNA. This variant, which constitutes approximately 8% to 15% of plasma fibrinogen, contains FXIII and thrombin binding sites. Our objective was to investigate whether gammaA/gamma' differs in fibrin formation and structure from the more common variant gammaA/gammaA. Both variants were separated and purified by anion-exchange chromatography. Fibrin formation and clot structure of the variants and unfractionated fibrinogen were investigated by turbidity and scanning electron microscopy (SEM). Thrombin cleavage of fibrinopeptides was analyzed by high-performance liquid chromatography (HPLC). Turbidity analysis showed significantly altered polymerization rates and overall fiber thickness in gammaA/gamma' clots compared with gammaA/gammaA and unfractionated fibrinogen. This finding was consistent with a range of thrombin concentrations. HPLC demonstrated reduced rates of fibrinopeptide B (FpB) release from gammaA/gamma' fibrinogen compared with gammaA/gammaA. Delayed FpB release was associated with delayed lateral aggregation of protofibrils and significant differences were found on SEM, with gammaA/gamma' clots consisting of smaller diameter fibers and increased numbers of branch points compared with both gammaA/gammaA and unfractionated fibrinogen. These results demonstrate that the gammaA/gamma' splice variant of fibrinogen directly alters fibrin formation and structure, which may help to explain the increased thrombotic risk associated with this variant.
Insights
The fibrinogen gammaA/gamma' splice variant alters fibrin clot formation and structure, leading to thinner fibers and more branch points. This structural change may contribute to the increased thrombotic risk associated with this fibrinogen variant.
Area of Science:
- Biochemistry
- Molecular Biology
- Hematology
Background:
- Fibrinogen gammaA/gamma' is an alternatively spliced mRNA variant, comprising 8-15% of plasma fibrinogen.
- This variant possesses FXIII and thrombin binding sites, suggesting functional differences.
Purpose of the Study:
- To investigate differences in fibrin formation and clot structure between fibrinogen gammaA/gamma' and gammaA/gammaA variants.
- To explore the potential link between gammaA/gamma' structure and thrombotic risk.
Main Methods:
- Purification of fibrinogen variants using anion-exchange chromatography.
- Analysis of fibrin formation and clot structure via turbidity and scanning electron microscopy (SEM).
- Assessment of thrombin cleavage of fibrinopeptides using high-performance liquid chromatography (HPLC).
Main Results:
- Turbidity assays revealed altered polymerization rates and fiber thickness in gammaA/gamma' clots.
- HPLC showed reduced fibrinopeptide B (FpB) release from gammaA/gamma' fibrinogen.
- SEM demonstrated that gammaA/gamma' clots have smaller diameter fibers and more branch points.
Conclusions:
- The gammaA/gamma' fibrinogen splice variant directly impacts fibrin formation and structure.
- Altered fibrin structure, characterized by thinner fibers and increased branching, may underlie the heightened thrombotic risk observed with this variant.