Elucidation of the mechanisms of giant platelet production in MYH9 disorders

Myh9疾病，如May-Hegglin异常，其特征是大血小板减少和细胞质粒细胞包涵体，这是由Myh9突变引起的，Myh9是非肌肉肌球蛋白重链IIA(NMMHC-IIA)基因的突变。我们检测了Myh9 5770delG和5818delG突变患者外周血细胞中突变型NMMHC-IIA多肽的表达。针对5818delG产生的异常羧基末端残基，提出了针对突变型NMMHC-IIA(NT629)的特异性抗体。NT629能与重组5818delG NMMHC-IIA反应，但不能与野生型NMMHC-IIA反应，不能识别正常外周血细胞的任何细胞成分。免疫荧光和免疫印迹显示，突变型NMMHC-IIA仅存在于中性粒细胞内的包涵体中，弥漫分布于淋巴细胞胞浆中，在单核细胞中稀疏分布于弥漫的胞浆背景中，仅在大血小板中均匀分布。突变型NMMHC-IIA在表面活化的血小板中没有移位到片状脂膜。野生型NMMHC-IIA在患者外周血CD34+细胞来源的巨核细胞中分布均匀，而粗突变型NMMHC-IIA分布不均，胞浆内无异常聚集。我们显示了突变的NMMHC-IIA的差异表达，并假设细胞特异性调控机制在Myh9疾病中发挥作用。

MYH9 disorders such as May-Hegglin anomaly are characterized by macrothrombocytopenia and cytoplasmic granulocyte inclusion bodies that result from mutations in MYH9, the gene for nonmuscle myosin heavy chain-IIA(NMMHC-IIA). We examined the expression of mutant NMMHC-IIA polypeptide in peripheral blood cells from patients with MYH9 5770delG and 5818delG mutations. A specific antibody to mutant NMMHC-IIA(NT629) was raised against the abnormal carboxyl-terminal residues generated by 5818delG. NT629 reacted to recombinant 5818delG NMMHC-IIA but not to wild-type NMMHC-IIA, and did not recognize any cellular components of normal peripheral blood cells. Immunofluorescence and immunoblotting revealed that mutant NMMHC-IIA was present and sequestrated only in inclusion bodies within neutrophils, diffusely distributed throughout lymphocyte cytoplasm, sparsely localized on a diffuse cytoplasmic background in monocytes, and uniformly distributed at diminished levels only in large platelets. Mutant NMMHC-IIA did not translocate to lamellipodia in surface activated platelets. Wild-type NMMHC-IIA was homogeneously distributed among megakaryocytes derived from the peripheral blood CD34+ cells of patients, but coarse mutant NMMHC-IIA was heterogeneously scattered without abnormal aggregates in the cytoplasm. We show the differential expression of mutant NMMHC-IIA and postulatethat cell-specific regulation mechanisms function in MYH9 disorders.