Masahiro Yoda1, Takahiro Kaido1, Tomu Kamijo2,3, Chiaki Taira1, Yumiko Higuchi1, Shinpei Arai4, Nobuo Okumura1,5. 1. Department of Clinical Laboratory Investigation, Graduate School of Medicine, Shinshu University, 3-1-1 Asahi, Matsumoto, 390-8621, Japan. 2. Department of Medical Sciences, Graduate School of Medicine, Science and Technology, Shinshu University, Matsumoto, Japan. 3. Department of Laboratory Medicine, Shinshu University Hospital, Matsumoto, Japan. 4. Department of Clinical Laboratory Investigation, Graduate School of Medicine, Shinshu University, 3-1-1 Asahi, Matsumoto, 390-8621, Japan. arais@shinshu-u.ac.jp. 5. Department of Clinical Laboratory Sciences, School of Health Sciences, Shinshu University, Matsumoto, Japan.
Abstract
INTRODUCTION: We identified a patient with a novel heterozygous variant fibrinogen, γp.C352R (Niigata II; N-II), who had a bleeding episode and failed infertility treatment and was suspected to have hypodysfibrinogenemia based on low and discordant fibrinogen levels (functional assay 0.33 g/L, immunological assay 0.91 g/L). We analyzed the mechanism of this rare phenotype of a congenital fibrinogen disorder. MATERIALS AND METHODS: Patient plasma fibrinogen was purified and protein characterization and thrombin-catalyzed fibrin polymerization performed. Recombinant fibrinogen-producing Chinese hamster ovary (CHO) cells were established and the assembly and secretion of variant fibrinogen analyzed by ELISA and western blotting. RESULTS: Purified N-II plasma fibrinogen had a small lower molecular weight band below the normal γ-chain and slightly reduced fibrin polymerization. A limited proportion of p.C352R fibrinogen was secreted into the culture medium of established CHO cell lines, but the γ-chain of p.C352R was synthesized and variant fibrinogen was assembled inside the cells. CONCLUSION: We demonstrated that fibrinogen N-II, γp.C352R was associated with markedly reduced secretion of variant fibrinogen from CHO cells, that fibrin polymerization of purified plasma fibrinogen was only slightly affected, and that fibrinogen N-II produces hypodysfibrinogenemia in plasma.
INTRODUCTION: We identified a patient with a novel heterozygous variant fibrinogen, γp.C352R (Niigata II; N-II), who had a bleeding episode and failed infertility treatment and was suspected to have hypodysfibrinogenemia based on low and discordant fibrinogen levels (functional assay 0.33 g/L, immunological assay 0.91 g/L). We analyzed the mechanism of this rare phenotype of a congenital fibrinogen disorder. MATERIALS AND METHODS:Patient plasma fibrinogen was purified and protein characterization and thrombin-catalyzed fibrin polymerization performed. Recombinant fibrinogen-producing Chinese hamster ovary (CHO) cells were established and the assembly and secretion of variant fibrinogen analyzed by ELISA and western blotting. RESULTS: Purified N-II plasma fibrinogen had a small lower molecular weight band below the normal γ-chain and slightly reduced fibrin polymerization. A limited proportion of p.C352Rfibrinogen was secreted into the culture medium of established CHO cell lines, but the γ-chain of p.C352R was synthesized and variant fibrinogen was assembled inside the cells. CONCLUSION: We demonstrated that fibrinogen N-II, γp.C352R was associated with markedly reduced secretion of variant fibrinogen from CHO cells, that fibrin polymerization of purified plasma fibrinogen was only slightly affected, and that fibrinogen N-II produces hypodysfibrinogenemia in plasma.
Authors: Kristina B Kruse; Amy Dear; Erin R Kaltenbrun; Brandan E Crum; Peter M George; Stephen O Brennan; Ardythe A McCracken Journal: Am J Pathol Date: 2006-04 Impact factor: 4.307
Authors: A Casini; T Brungs; C Lavenu-Bombled; R Vilar; M Neerman-Arbez; P de Moerloose Journal: J Thromb Haemost Date: 2017-03-06 Impact factor: 5.824