Blood Research Institute, BloodCenter of Wisconsin, Milwaukee, WI;
Blood Research Institute, BloodCenter of Wisconsin, Milwaukee, WI; Medical College of Wisconsin, Milwaukee, WI; and Children's Research Institute, Children's Hospital of Wisconsin, Milwaukee, WI.
Blood. 2014 Jun 12;123(24):3706-13. doi: 10.1182/blood-2014-02-555151. Epub 2014 Apr 4.
The cellular source of coagulation factor VIII (FVIII) remains controversial. Like many coagulation proteins, FVIII is produced in the liver, and FVIII synthesis has long been associated with hepatocytes. But extrahepatic synthesis also occurs, and mounting evidence suggests that hepatocytes are not responsible for FVIII production. To determine the tissue that synthesizes FVIII, we developed a Cre/lox-dependent conditional knockout (KO) model in which exons 17 and 18 of the murine factor VIII gene (F8) are flanked by loxP sites, or floxed (F8(F)). In cells expressing Cre-recombinase, the floxed sequence is deleted, resulting in F8(F→KO) gene inactivation. When F8(F) mice were crossed with various tissue-specific Cre strains, we found that hepatocyte-specific F8-KO mice are indistinguishable from controls, whereas efficient endothelial-KO models display a severe hemophilic phenotype with no detectable plasma FVIII activity. A hematopoietic Cre model was more equivocal, so experimental bone marrow transplantation was used to examine hematopoietic FVIII synthesis. FVIII(null) mice that received bone marrow transplants from wild-type donors were still devoid of plasma FVIII activity after hematopoietic donor cell engraftment. Our results indicate that endothelial cells are the predominant, and possibly exclusive, source of plasma FVIII.
凝血因子 VIII（FVIII）的细胞来源仍存在争议。像许多凝血蛋白一样，FVIII 在肝脏中产生，FVIII 的合成长期以来一直与肝细胞有关。但也存在肝外合成，越来越多的证据表明肝细胞不负责 FVIII 的产生。为了确定合成 FVIII 的组织，我们开发了一种 Cre/lox 依赖性条件敲除（KO）模型，其中小鼠 FVIII 基因（F8）的外显子 17 和 18 被loxP 位点侧翼，或 floxed（F8(F)）。在表达 Cre 重组酶的细胞中，floxed 序列被删除，导致 F8(F→KO)基因失活。当 F8(F)小鼠与各种组织特异性 Cre 株系杂交时，我们发现肝细胞特异性 F8-KO 小鼠与对照小鼠没有区别，而有效的内皮细胞-KO 模型则表现出严重的血友病表型，血浆 FVIII 活性检测不到。造血细胞 Cre 模型则更为模棱两可，因此进行了实验性骨髓移植以检查造血细胞 FVIII 的合成。接受来自野生型供体骨髓移植的 FVIII(null)小鼠在造血供体细胞移植后仍然缺乏血浆 FVIII 活性。我们的结果表明，内皮细胞是血浆 FVIII 的主要来源，可能是唯一来源。
