Department of Genetic Medicine and Development, Faculty of Medicine, University of Geneva, Geneva, Switzerland; and.
Division of Pediatric Hematology/Oncology, Department of Pediatrics, School of Medicine, University of Michigan, Ann Arbor, MI.
Blood Adv. 2020 Nov 10;4(21):5480-5491. doi: 10.1182/bloodadvances.2020001472.
Plasma fibrinogen molecules comprise 2 copies of Aα, Bβ, and γ chains folded into a hexameric protein. A minor fibrinogen isoform with an extended Aα chain (AαE) is more abundant in newborn human blood than in adults. Larval zebrafish produce predominantly AαE-containing fibrinogen, but its functional significance is unclear. In 3-day-old zebrafish, when hemostasis is reliant on fibrinogen and erythrocyte-rich clotting but is largely thrombocyte-independent, we measured the time to occlusion (TTO) in a laser-induced venous thrombosis assay in 3 zebrafish strains (AB, TU, and AB × TL hybrids). AB larvae showed delayed TTO compared with the TU and AB × TL strains. Mating AB with TU or TL produced larvae with a TU-like TTO. In contrast to TU, AB larvae failed to produce fibrinogen AαE, due to a mutation in the AαE-specific coding region of fibrinogen α-chain gene (fga). We investigated whether the lack of AαE explained the delayed AB TTO. Transgenic expression of AαE, but not Aα, shortened the AB TTO to that of TU. AαE rescued venous occlusion in fibrinogen mutants or larvae with morpholino-targeted fibrinogen α-chain messenger RNA, but Aα was less effective. In 5-day-old larvae, circulating thrombocytes contribute to hemostasis, as visualized in Tg(itga2b:EGFP) transgenics. Laser-induced venous thrombocyte adhesion and aggregation is reduced in fibrinogen mutants, but transgenic expression of Aα or AαE restored similar thrombocyte accumulation at the injury site. Our data demonstrate a genetic modifier of venous thrombosis and a role for fibrinogen AαE in early developmental blood coagulation, and suggest a link between differentially expressed fibrinogen isoforms and the cell types available for clotting.
血浆纤维蛋白原分子由 2 个 Aα、Bβ 和 γ 链组成，折叠成六聚体蛋白。一种 Aα 链延长的较少纤维蛋白原同工型（AαE）在新生儿血液中的丰度高于成人。幼体斑马鱼主要产生含有 AαE 的纤维蛋白原，但它的功能意义尚不清楚。在 3 日龄的斑马鱼中，止血依赖于纤维蛋白原和富含红细胞的血栓形成，但很大程度上依赖于血小板，我们在 3 种斑马鱼品系（AB、TU 和 AB × TL 杂交种）的激光诱导静脉血栓形成试验中测量了闭塞时间（TTO）。与 TU 和 AB × TL 品系相比，AB 幼鱼的 TTO 延迟。将 AB 与 TU 或 TL 交配产生的幼鱼具有 TU 样的 TTO。与 TU 不同的是，AB 幼鱼由于纤维蛋白原α链基因（fga）中 AαE 特异性编码区的突变而不能产生纤维蛋白原 AαE。我们研究了缺乏 AαE 是否解释了 AB TTO 的延迟。AαE 的转基因表达而非 Aα 缩短了 AB TTO 至 TU 的时间。AαE 挽救了纤维蛋白原突变体或纤维蛋白原α链信使 RNA 靶向 morpholino 的幼鱼的静脉闭塞，但 Aα 的效果较差。在 5 日龄的幼鱼中，循环血小板有助于止血，如 Tg(itga2b:EGFP)转基因动物中所示。在纤维蛋白原突变体中，激光诱导的静脉血小板黏附和聚集减少，但 Aα 或 AαE 的转基因表达恢复了损伤部位类似的血小板聚集。我们的数据证明了静脉血栓形成的遗传修饰因子和纤维蛋白原 AαE 在早期发育性凝血中的作用，并提示差异表达的纤维蛋白原同工型与可用于血栓形成的细胞类型之间存在联系。
