From MRC Clinical Sciences Centre (L.D., S.P., A.S., Z.W., O.B., M.D., L.C., E.K., J.T., J.L.) and National Heart and Lung Institute Vascular Science Unit (N.D., G.B., A.M.R.), Faculty of Medicine, Imperial College London, UK.
Circulation. 2015 Jun 23;131(25):2217-25. doi: 10.1161/CIRCULATIONAHA.114.015064. Epub 2015 Apr 24.
Asymmetrical dimethylarginine (ADMA) is an endogenous inhibitor of nitric oxide synthesis and a risk factor for cardiovascular disease. Dimethylarginine dimethylaminohydrolase (DDAH) enzymes are responsible for ADMA breakdown. It has been reported that endothelial DDAH1 accounts for the majority of ADMA metabolism. However, we and others have shown strong DDAH1 expression in a range of nonendothelial cell types, suggesting that the endothelium is not the only site of metabolism. We have developed a new endothelium-specific DDAH1 knockout mouse (DDAH1(En-/-)) to investigate the significance of endothelial ADMA in cardiovascular homeostasis.
DDAH1 deletion in the DDAH1(En-/-) mouse was mediated by Tie-2 driven Cre expression. DDAH1 deletion was confirmed through immunocytochemistry, whereas Western blotting showed that DDAH1 remained in the kidney and liver, confirming expression in nonendothelial cells. Plasma ADMA was unchanged in DDAH1(En-/-) mice, and cultured aortas released amounts of ADMA to similar to controls. Consistent with these observations, vasoreactivity ex vivo and hemodynamics in vivo were unaltered in DDAH1(En-/-) mice. In contrast, we observed significantly impaired angiogenic responses both ex vivo and in vivo.
We demonstrate that endothelial DDAH1 is not a critical determinant of plasma ADMA, vascular reactivity, or hemodynamic homeostasis. DDAH1 is widely expressed in a range of vascular and nonvascular cell types; therefore, the additive effect of DDAH1 expression in multiple organ systems determines plasma ADMA concentrations. Endothelial deletion of DDAH1 profoundly impairs the angiogenic capacity of endothelial cells, indicating that intracellular ADMA is a critical determinant of endothelial cell response.
不对称二甲基精氨酸（ADMA）是一氧化氮合成的内源性抑制剂，也是心血管疾病的危险因素。二甲基精氨酸二甲氨基水解酶（DDAH）酶负责 ADMA 的分解。据报道，内皮细胞 DDAH1 负责大部分 ADMA 的代谢。然而，我们和其他人已经在一系列非内皮细胞类型中发现了强烈的 DDAH1 表达，这表明内皮细胞并不是唯一的代谢部位。我们已经开发了一种新的内皮细胞特异性 DDAH1 敲除小鼠（DDAH1(En-/-)），以研究内皮 ADMA 在心血管稳态中的重要性。
DDAH1(En-/-)小鼠中的 DDAH1 缺失是由 Tie-2 驱动的 Cre 表达介导的。通过免疫细胞化学证实了 DDAH1 的缺失，而 Western blot 显示 DDAH1 仍存在于肾脏和肝脏中，证实了其在非内皮细胞中的表达。DDAH1(En-/-)小鼠的血浆 ADMA 没有变化，培养的主动脉释放的 ADMA 量与对照组相似。与这些观察结果一致，DDAH1(En-/-)小鼠的血管反应性和体内血液动力学在体外和体内均未改变。相比之下，我们观察到体外和体内的血管生成反应都明显受损。
我们证明内皮细胞 DDAH1 不是血浆 ADMA、血管反应性或血液动力学稳态的关键决定因素。DDAH1 在多种血管和非血管细胞类型中广泛表达；因此，多个器官系统中 DDAH1 表达的加性效应决定了血浆 ADMA 浓度。内皮细胞 DDAH1 的缺失严重损害了内皮细胞的血管生成能力，表明细胞内 ADMA 是内皮细胞反应的关键决定因素。
