Elucidation of the mechanisms of giant platelet production in MYH9 disorders

MYH 9疾病（如May-Hegglin异常）的特征是由MYH 9（非肌肉肌球蛋白重链IIA（NMMHC-IIA）基因）突变引起的巨血小板减少症和胞浆粒细胞包涵体。我们检测了MYH 9 5770 delG和5818 delG突变患者外周血细胞中突变型NMMHC-IIA多肽的表达。针对5818 delG产生的异常羧基端残基，制备了针对突变型NMMHC-IIA（NT 629）的特异性抗体。NT 629与重组5818 delG NMMHC-IIA反应，但不与野生型NMMHC-IIA反应，并且不识别正常外周血细胞的任何细胞组分。免疫荧光和免疫印迹显示，突变NMMHC-IIA是目前和隔离仅在包涵体内的中性粒细胞，弥漫性分布在整个淋巴细胞的细胞质中，稀疏地定位在单核细胞的弥漫性细胞质背景，并均匀地分布在减少的水平，只有在大血小板。突变型NMMHC-IIA在表面活化的血小板中不移位至板状伪足。野生型NMMHC-IIA均匀分布于患者外周血CD 34+细胞来源的巨核细胞中，但粗突变型NMMHC-IIA不均匀分散，细胞质中无异常聚集。我们显示了突变型NMMHC-IIA的差异表达，并推测MYH 9疾病中细胞特异性调节机制起作用。

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.