Broudy V C, Lin N L, Sabath D F, Papayannopoulou T, Kaushansky K
Department of Medicine, University of Washington, Seattle 98195, USA.
Blood. 1997 Mar 15;89(6):1896-904.
Thrombopoietin (Tpo) is a major regulator of megakaryopoiesis both in vivo and in vitro. Tpo initiates its biologic effects by binding to the Mpl receptor, which is a member of the hematopoietin receptor family. To define the Tpo binding characteristics of the Mpl receptor, we iodinated purified 70-kD recombinant human Tpo using the Bolton-Hunter reagent. Autoradiographic analysis of (125)I-Tpo binding to normal human marrow mononuclear cells showed many grains specifically associated with megakaryocytes; there were no grains specifically associated with myeloblasts or erythroblasts. Equilibrium binding experiments with (125)I-Tpo and normal human platelets showed a single class of high-affinity receptors (kd, 190 pmol/L) with approximately 30 Mpl receptors per platelet. Affinity cross-linking with (125)I-Tpo showed that the Mpl receptor on platelets is of molecular weight approximately 98 kD. Despite their sequence similarity, erythropoietin and Tpo did not cross-compete for binding to BaF3 cells engineered to coexpress Mpl receptor and erythropoietin receptor. Progeny of normal human burst-forming units-erythroid (BFU-E) contained Mpl receptor mRNA, and flow cytometric analysis showed the presence of Mpl receptor protein on the surface of these cells. These data indicate that display of the Mpl receptor is not limited to the megakaryocytic lineage, but also includes progeny of BFU-E. Like receptors for other hematopoietic cytokines, the binding affinity of the Mpl receptor for Tpo is high, with relatively few receptors displayed per cell. These results suggest that the effects of Tpo to speed red blood cell recovery after myelosuppressive therapy in vivo and to enhance colony-forming unit-erythroid generation in vitro may be mediated by direct interaction of Tpo and erythroid progenitor cells.
血小板生成素（Tpo）是体内和体外巨核细胞生成的主要调节因子。Tpo通过与Mpl受体结合来启动其生物学效应，Mpl受体是造血受体家族的成员。为了确定Mpl受体的Tpo结合特性，我们使用博尔顿-亨特试剂对纯化的70-kD重组人Tpo进行碘化。对(125)I-Tpo与正常人骨髓单个核细胞结合的放射自显影分析显示，许多颗粒与巨核细胞特异性相关；没有颗粒与成髓细胞或成红细胞特异性相关。用(125)I-Tpo和正常人血小板进行的平衡结合实验显示有一类单一的高亲和力受体（kd，190 pmol/L），每个血小板约有30个Mpl受体。用(125)I-Tpo进行的亲和交联显示血小板上的Mpl受体分子量约为98 kD。尽管促红细胞生成素和Tpo序列相似，但它们不会相互竞争与经基因工程改造共表达Mpl受体和促红细胞生成素受体的BaF3细胞的结合。正常人红系爆式集落形成单位（BFU-E）的子代含有Mpl受体mRNA，流式细胞术分析显示这些细胞表面存在Mpl受体蛋白。这些数据表明，Mpl受体的表达不仅限于巨核细胞系，还包括BFU-E的子代。与其他造血细胞因子的受体一样，Mpl受体对Tpo的结合亲和力很高，每个细胞表达的受体相对较少。这些结果表明，Tpo在体内骨髓抑制治疗后加速红细胞恢复以及在体外增强红系集落形成单位生成的作用可能是由Tpo与红系祖细胞的直接相互作用介导的。
