酪氨酸激酶BMX调控内皮细胞活化促进动脉粥样硬化斑块形成的作用及机制研究

Endothelial activation, manifesting as cell adhesion molecule upregulation and cytokine secretion, leading to monocyte adhesion is a critical event in atherosclerosis initiation. Bone marrow tyrosine kinase gene on chromosome X (BMX) has been suggested to be implicated in several pathological processes, including inflammatory response as well as angiogenesis. However, the role of BMX in atherosclerosis has not yet been assessed. Our preliminary experiment found that BMX levels were increased in atherosclerosis, and overexpression of BMX in ApoE-/- mice increased plaque burden and macrophage content, while silence of BMX in endothelial cells reduced TNF-α induced cell adhesion molecules expression. Thus, we hypothesized that BMX might play a role in promoting atherogenesis by facilitating endothelial activation and endothelial-monocyte adhesion, which is dependent on its tyrosine kinase activity. The effects of BMX and its kinase activity on development of atherosclerotic plaque will be verified in further experiment, and whether BMX could activate endothelial cells, promote monocyte adhesion and atherogenesis through phosphorylating related proteins in NF-κB signaling pathway will be investigated by means of several biological techniques. The purpose of this study is to explore the effect and cellular mechanism of BMX on atherogenesis and provide a novel therapeutic target for anti-atherosclerosis.

内皮细胞活化上调细胞粘附分子表达及炎症因子分泌，促进单核细胞粘附，是动脉粥样硬化（AS）启始的关键事件。位于X染色体上的骨髓酪氨酸激酶（BMX）参与炎症反应及血管新生等多种病理过程，但其在AS中的作用尚无报道。我们前期研究发现，AS斑块组织中BMX的表达上调，过表达BMX可增加ApoE-/-小鼠的斑块负荷及斑块内巨噬细胞含量，抑制BMX可减少TNF-α诱导的内皮细胞粘附分子的表达。据此，我们提出如下科学假说“BMX可能通过调控内皮细胞活化，增加内皮-单核细胞间粘附，促进AS斑块形成，且该作用依赖于其激酶活性”。本课题拟采用多种生物学实验方法，验证BMX及其激酶活性在AS斑块形成中的作用，探讨BMX是否通过磷酸化NF-κB信号通路上的相关蛋白，活化内皮细胞，促进单核细胞粘附，从而促进斑块形成。本研究旨在深入探讨BMX调控AS斑块形成的作用及分子机制，为AS的药物干预提供新靶点。

内皮细胞活化上调细胞粘附分子表达及炎症因子分泌，促进单核细胞粘附，是动脉粥样硬化（AS）启动的关键事件。以往研究发现，位于X染色体上的骨髓酪氨酸激酶（BMX）参与炎症反应及血管新生等多种病理过程，但其在AS中的作用尚未报道。本研究中我们发现，BMX主要表达在血管内皮细胞，在小鼠及人的斑块组织中均表达上调。以BMX-/-:apoE-/-双敲小鼠及BMX过表达腺病毒转染小鼠为研究对象，高脂喂养及颈动脉套管建立小鼠AS斑块模型，研究发现，敲除BMX能显著抑制小鼠AS斑块形成，抑制内皮细胞活化、炎症反应及斑块内血管新生，而过表达BMX能够显著促进AS斑块形成。体外脐静脉内皮细胞转染BMX小干扰序列能够显著抑制TNF-α诱导的细胞粘附分子ICAM-1,VCAM-1及E-selectin的蛋白表达，抑制内皮-单核细胞间粘附，内皮细胞迁移及体外成管。进一步机制研究发现，抑制BMX表达能够促进内皮细胞中KLF2的表达，而对NF-KB信号通路无关。蛋白质谱分析发现，TNF-α 刺激能够增加 BMX 与促进细胞增殖、迁移及血管新生相关蛋白 HSPG2、 NCOA5、 MATN2、 CTCF 等的结合。 总之，以上结果均表明BMX在AS形成中的重要作用，为AS的药物干预提供新靶点。