Amelioration of the macrothrombocytopenia associated with the murine Bernard-Soulier syndrome
Amelioration of the macrothrombocytopenia associated with the murine Bernard-Soulier syndrome
复制标题
DOI：
10.1182/blood-2002-03-0997
复制
发表时间：
2002-09-15
期刊：
影响因子：
20.3
通讯作者：
Ware, J
中科院分区：
文献类型：
--
作者：
Kanaji, T;Russell, S;Ware, J
血小板糖蛋白（GP）Ib-IX受体缺失导致Bernard-Soulier综合征，其特征为严重出血和实验室表现为巨血小板减少症。虽然巨血小板减少症的表型与GP Ib-IX复合物的缺失直接相关，但产生巨血小板减少症的破坏分子机制尚不清楚。我们利用Bernard-Soulier综合征的小鼠模型来设计表达GP Ib α-亚基（GP Ibalpha）的血小板，其中大部分胞质外序列已被人白细胞介素-4受体（IL-4 R α）α-亚基的分离结构域取代。IL-4 R α/GP Ib α融合蛋白在中国仓鼠卵巢（CHO）细胞中膜表达，人GP IX和GP Ib β亚基的存在促进了其表达。产生表达嵌合受体的转基因动物，并将其培育成缺乏小鼠GP I α但表达IL-4 R α/GP I α融合序列的产生鼠Bernard-Soulier综合征的动物。这些小鼠的表征显示，与来自Bernard-Soulier综合征小鼠模型的血小板相比，循环血小板计数增加2倍，血小板大小减少50%。免疫沉淀证实，IL-4 R α/GP Ibalpha亚基与细丝蛋白-1和14-3- 3 ζ相互作用，这是已知的GP Ibalpha细胞质tall的结合蛋白。表达嵌合受体的小鼠保留严重出血表型，证实GP Ibalpha胞质外结构域在止血中的关键作用。这些结果提供了在体内的GP Ibalpha亚基，有助于正常的巨核细胞成熟和血小板生成的结构要素的见解。
An absent platelet glycoprotein (GP) Ib-IX receptor results in the Bernard-Soulier syndrome and is characterized by severe bleeding and the laboratory presentation of macrothrombocytopenia. Although the macrothrombocytopenic phenotype is directly linked to an absent GP Ib-IX complex, the disrupted molecular mechanisms that produce the macrothrombocytopenia are unknown. We have utilized a mouse model of the Bernard-Soulier syndrome to engineer platelets expressing an alpha-subunit of GP Ib (GP Ibalpha) in which most of the extracytoplasmic sequence has been replaced by an isolated domain of the alpha-subunit of the human interleukin-4 receptor (IL-4Ralpha). The IL-4Ralpha/GP Ibalpha fusion is membrane expressed in Chinese hamster ovary (CHO) cells, and its expression is facilitated by the presence of human GP IX and the beta-subunit of GP Ib. Transgenic animals expressing a chimeric receptor were generated and bred into the murine Bernard-Soulier syndrome-producing animals devoid of mouse GP Ibalpha but expressing the IL-4Ralpha/GP Ibalpha fusion sequence. The characterization of these mice revealed a 2-fold increase in circulating platelet count and a 50% reduction in platelet size when compared with platelets from the mouse model of the Bernard-Soulier syndrome. Immunoprecipitation confirmed that the IL-4Ralpha/GP Ibalpha subunit interacts with filamin-1 and 14-3-3zeta, known binding proteins to the GP Ibalpha cytoplasmic tall. Mice expressing the chimeric receptor retain a severe bleeding phenotype, confirming a critical role for the GP Ibalpha extracytoplasmic domain in hemostasis. These results provide in vivo insights into the structural elements of the GP Ibalpha subunit that contribute to normal megakaryocyte maturation and thrombopoiesis.