Day Kenneth, Krishnegowda Naveen, Jagadeeswaran Pudur
Department of Cellular and Structural Biology, The University of Texas Health Science Center at San Antonio, San Antonio, TX 78229, USA.
Blood Cells Mol Dis. 2004 Jan-Feb;32(1):191-8. doi: 10.1016/j.bcmd.2003.09.004.
Thrombin is a serine protease generated from its zymogen, prothrombin, and plays a central role in the coagulation cascade. It is also important for mammalian development. The zebrafish has now been established as an excellent genetic model for studies on mammalian hemostasis and development. In this report, we used prothrombin-specific antisense morpholinos to knock down the levels of prothrombin to characterize the effects of prothrombin deficiency in the zebrafish embryo. Prothrombin morpholino-injected zebrafish embryos yielded an early phenotype exhibiting severe abnormalities that later showed occasional bleeding. In a second late phenotype, the embryos had no observable morphological abnormalities in early stages, but showed occasional bleeding at later stages. These phenotypes resembled characteristics shown by prothrombin knockout mice. Laser-induced vascular injury on some of the normal appearing phenotypic larvae showed a prolonged time to occlusion, and recombinant zebrafish prothrombin injected into these larvae restored a normal time to occlusion thus showing the specificity of the morpholino effect. The system developed here should be useful for investigation of the role of thrombin in vertebrate development.
凝血酶是一种由其酶原凝血素原产生的丝氨酸蛋白酶，在凝血级联反应中起核心作用。它对哺乳动物的发育也很重要。斑马鱼现已成为研究哺乳动物止血和发育的优秀遗传模型。在本报告中，我们使用凝血素原特异性反义吗啉代寡核苷酸来降低凝血素原水平，以表征斑马鱼胚胎中凝血素原缺乏的影响。注射了凝血素原吗啉代寡核苷酸的斑马鱼胚胎产生了一种早期表型，表现出严重异常，随后偶尔出现出血。在第二种晚期表型中，胚胎在早期没有明显的形态异常，但在后期偶尔出现出血。这些表型类似于凝血素原基因敲除小鼠所表现出的特征。对一些外观正常的表型幼虫进行激光诱导血管损伤，显示闭塞时间延长，而注射到这些幼虫体内的重组斑马鱼凝血素原恢复了正常的闭塞时间，从而显示了吗啉代寡核苷酸效应的特异性。这里开发的系统应该有助于研究凝血酶在脊椎动物发育中的作用。
