努南综合征（Noonan Syndrome，NS）患者存在反复不明原因性出血及术后出血量增加，现有针对血小板以及凝血因子方面的疗法效果不佳。血管内皮屏障功能障碍是影响出血的另一关键因素。编码的酪氨酸磷酸酶Shp2的PTPN11基因突变是NS患者的主要遗传改变，突变产生活化的Shp2。预实验结果显示：NS相关Shp2活化突变（Shp2A72G）通过激活肌球蛋白轻链（MLC），增加骨架张力，破坏内皮屏障。因此我们提出假说：NS相关Shp2活化突变通过破坏内皮屏障参与NS导致的出血。为此本课题引入Shp2活化突变基因修饰小鼠，结合原代内皮细胞，明确Shp2活化突变对屏障的作用。机制上通过蛋白相互作用，尤其是Shp2作用底物的磷酸化修饰进行阐述。项目成果有望明确Shp2激活突变引起血管内皮屏障破坏的作用及机制，并为NS不明原因性出血的临床干预提供理论基础与实验依据。

Noonan syndrome (NS) patients present serious unexplained bleeding repeatedly or prolonged bleeding following surgery. Furthermore, conventional treatment including platelets and clotting factors do not appear to be efficient in NS. Vascular endothelial barrier dysfunction has been reported as another key point recently. The missense mutations in the PTPN11 gene are the dominant genetic changes in NS, resulting in a gain-of-function of the tyrosine phosphatase Shp2. Our preliminary data showed that constitutively active mutants of Shp2 (Shp2A72G) resulted in endothelial barrier dysfunction by activating the MLC and increasing cytoskeleton tension. Collectively, these data support the hypothesis that NS-associated active mutants of Shp2 induced bleeding disorders in NS patients is mediated by the disruption of endothelial barrier function. Therefore we would make the function clear with endothelial specific SHP2 knockout mouse line and primary cell line HUVECs. For the molecular mechanism, protein - protein interaction, especially the phosphorylation modification of substrate of Shp2 would be used to explain the phenotype. This project is expected to demonstrate the function of Shp2 in endothelium and provides theoretical and experimental clues for novel intervention strategy in unexplained bleeding.