Aims A novel heterozygous variant, FGA c.169_180+2 del (designated fibrinogen Shanghai), was identified in a patient with dysfibrinogenemia with antiphospholipid antibody syndrome (APS) and recurrent venous thrombosis, and in his asymptomatic father. We aimed to reveal the functional implication of structural change caused by this variant.
Methods Transcription analysis was performed with FGA minigene transfection assay to evaluate the impact of nucleosides deletion on mRNA editing. The fibrinogen isolated from propositus' plasma was used to characterise its functional defects. Fibrin polymerization and clot lysis experiments were performed by optical measurement of turbidity. Thrombin-catalysed fibrinopeptide release was analysed by the reversed-phase, high-performance liquid chromatography. The ultrastructures of fibrin clots were visualised by scanning electron microscopy.
Results FGA c.169_180+2 del led to an aberrant mRNA with exon 2 skipping and encoded an shortened Aα chain with 42 amino acids truncation at its N-terminal. The propositus' fibrinogen had an impaired release of fibrinopeptide A and abnormal polymerization with a significantly prolonged lag time, a slower maximum slope and reduced final turbidity. The fibrin clot formed with propositus' fibrinogen showed thicker fibres with looser network structure. Clot lysis was normal using the purified fibrinogen but was significantly impaired using the plasma sample from propositus, compared with that from his father.
Conclusions Fibrinogen Shanghai results in N-terminal truncation of Aα chain, which does not interfere with synthesis, assembly or secretion of fibrinogen, but compromises fibrin polymerization and clot formation. APS at least partially contributes to the development of thrombosis in the propositus.
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Handling editor Mary Frances McMullin
Contributors JZ designed the study, purified the fibrinogen from plasma, performed the functional fibrinogen assays, reviewed and interpreted the data, and wrote the manuscript. QD supervised research, helped to design the study and critically revised the manuscript. Wenman Wu wrote the manuscript and critically revised it. QO performed haemostatic assays and genetic analysis. YX analysed thrombin-catalysed fibrinopeptide release by HPLC. XW and QL performed transcription analysis with FGA minigene transfection assay to evaluate the impact of nucleosides deletion on mRNA editing. JD and YL reviewed and interpreted data. HW supervised research, helped to design the study. XW and YH supervised research, helped to design the study and critically revised the manuscript. All the authors approved the final version of the manuscript.
Funding This study was supported by Doctoral Innovation Fund Projects from Shanghai Jiaotong University School of Medicine (BXJ201412), the National Basic Research Program of China (2013CB966800), the General Program of National Natural Science Foundation of China (81300396 to J.D) and Novo Nordisk Haemophilia Research Fund in China.
Competing interests None declared.
Patient consent Obtained.
Ethics approval Ruijin Hospital affiliated to Shanghai Jiaotong University School of Medicine.
Provenance and peer review Not commissioned; externally peer reviewed.
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