Abstract Number: PB0415
Meeting: ISTH 2020 Congress
Background: Mice lacking tissue factor (TF) are embryonically lethal (E10.5). The lethality has been attributed to defects in vascular development and circulatory failure, which suggests additional roles for TF in embryonic development beyond coagulation. Zebrafish has two TF paralogs (f3a and f3b) with unknown functions. Knockdown of f3b gene was shown to reduce coagulation and angiogenesis. Both f3a and f3b are necessary for normal hemostasis, albeit with differential contributions in the arterial vs. venous beds. The role of f3a in vascular development remains unknown.
Aims: To characterize the in vivo role of zebrafish f3a in vascular development.
Methods: Transgenic line of zebrafish with endothelial-specific reporter gene expression (flk1:egfp-NLS/kdrl:mCherry-CAAX) was used to image vascular development in vivo. The role of f3a during embryonic development was investigated by mRNA knockdown using Morpholino antisense oligonucleotides (MO). The f3a morphants were examined at 52 hours after fertilization (hpf) for defects in morphological appearance, bleeding, and vascular patterning.
Results: Phylogenetic and sequence homology analyses suggested that f3a exhibits greater similarity (amino acid and nucleotide sequence) to its human ortholog than f3b. f3a expression was seen in all developmental stages (64 cells stage to 72 hpf), indicating that f3a transcripts are maternally deposited, as well as zygotically expressed. MOs targeting the exon 3-intron 3 junction (e3-i3) dose-dependently reduced the expression of f3a, but not that of f3b, documenting target specificity. F3aMO-injected embryos exhibited a shortened body axis (11.45%), a short/crooked tail (27.5%) as well as bleeding in trunk (5.44%) and head (9.52%) regions. The caudal vein plexus (CVP), which forms through a process of sprouting immediately following the onset of circulation, showed a 3-fold decrease in the number of CVP loops and sprouting events, relative to controls.
Conclusions: This study reveals that zebrafish f3a, in addition to its function in coagulation, has an essential and non-redundant role in developmental angiogenesis.
To cite this abstract in AMA style:Subramaniam S, Eisa-Beygi S, Ramachandran R, Fletcher C, Weiler H. Coagulation Factor IIIa (f3a) Knockdown in Zebrafish Leads to Defective Angiogenesis and Bleeding Phenotype [abstract]. Res Pract Thromb Haemost. 2020; 4 (Suppl 1). https://abstracts.isth.org/abstract/coagulation-factor-iiia-f3a-knockdown-in-zebrafish-leads-to-defective-angiogenesis-and-bleeding-phenotype/. Accessed January 27, 2022.
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