Dept. of Internal Medicine, IRCCS Policlinico San Matteo Foundation, University of Pavia, Piazzale Golgi, 27100 Pavia, Italy.
Thromb Haemost. 2011 Oct;106(4):693-704. doi: 10.1160/TH11-02-0126. Epub 2011 Aug 11.
MYH9-related disease (MYH9-RD) is an autosomal-dominant thrombocytopenia caused by mutations in the gene for the heavy chain of non-muscle myosin-IIA (NMMHC-IIA). Recent in vitro studies led to the hypothesis that thrombocytopenia of MYH9-RD derives from an ectopic platelet release by megakaryocytes in the osteoblastic areas of bone marrow (BM), which are enriched in type I collagen, rather than in vascular spaces. SDF-1-driven migration of megakaryocytes within BM to reach the vascular spaces is a key mechanism for platelet biogenesis. Since myosin-IIA is implicated in polarised migration of different cell types, we hypothesised that MYH9 mutations could interfere with this mechanism. We therefore investigated the SDF-1-driven migration of a megakaryoblastic cell line, Dami cells, on type I collagen or fibrinogen by a modified transwell assay. Inhibition of myosin-IIA ATPase activity suppressed the SDF-1-driven migration of Dami cells, while over-expression of NMMHC-IIA increased the efficiency of chemotaxis, indicating a role for NMMHC-IIA in this mechanism. Transfection of cells with three MYH9 mutations frequently responsible for MYH9-RD (p.R702C, p.D1424H, or p.R1933X) resulted in a defective SDF-1-driven migration with respect to the wild-type counterpart and in increased cell spreading onto collagen. Analysis of differential localisation of wild-type and mutant proteins suggested that mutant NMMHC-IIAs had an impaired cytoplasmic re-organisation in functional cytoskeletal structures after cell adhesion to collagen. These findings support the hypothesis that a defect of SDF-1-driven migration of megakaryocytes induced by MYH9 mutations contributes to ectopic platelet release in the BM osteoblastic areas, resulting in ineffective platelet production.
肌球蛋白重链 9 相关疾病 (MYH9-RD) 是一种常染色体显性遗传性血小板减少症，由非肌肉肌球蛋白重链 IIA (NMMHC-IIA) 基因的突变引起。最近的体外研究提出假设，即 MYH9-RD 的血小板减少症源自骨髓 (BM) 成骨区巨核细胞的异位血小板释放，这些区域富含 I 型胶原，而不是血管空间。SDF-1 驱动的巨核细胞在 BM 内向血管空间的迁移是血小板生成的关键机制。由于肌球蛋白-IIA 参与不同细胞类型的极化迁移，我们假设 MYH9 突变可能干扰这一机制。因此，我们通过改良的 Transwell 测定法研究了巨核细胞系 Dami 细胞在 I 型胶原或纤维蛋白原上的 SDF-1 驱动迁移。肌球蛋白-IIA ATP 酶活性的抑制抑制了 Dami 细胞的 SDF-1 驱动迁移，而 NMMHC-IIA 的过表达增加了趋化作用的效率，表明 NMMHC-IIA 在该机制中起作用。常导致 MYH9-RD 的三种 MYH9 突变 (p.R702C、p.D1424H 或 p.R1933X) 的细胞转染导致 SDF-1 驱动的迁移相对于野生型对照存在缺陷，并增加了细胞在胶原上的扩散。对野生型和突变蛋白的差异定位分析表明，突变 NMMHC-IIA 在细胞粘附到胶原后，在功能性细胞骨架结构中细胞质再组织受损。这些发现支持这样一种假设，即 MYH9 突变引起的 SDF-1 驱动的巨核细胞迁移缺陷导致 BM 成骨区的异位血小板释放，导致无效的血小板生成。
