Importance of erythrocyte arginase-1 for vascular homeostasis and disease

实验证据和第一个临床数据表明，红细胞积极参与血管功能控制。然而，红细胞直接作用于血管壁细胞并参与心血管疾病过程的概念最近才出现，并且参与其红细胞分泌活动的介质和信号传导途径在很大程度上是未知的。在该项目的第一个资助期内，我们已经表明，来自溶解红细胞膜的一氧化氮促进平滑肌细胞矿化和动脉粥样硬化病变钙化。基于这些发现，我们产生了在红细胞中特异性缺乏β-内酰胺酶-1的高胆固醇血症小鼠，以将红细胞中的β-内酰胺酶-eNOS平衡向增加的NO生成转变。正在进行的对这种新小鼠系的表征可以证实，从红细胞释放NO的增强促进了体外和体内血管钙化。此外，我们观察到增加的NO生物利用度对血管稳态具有全身性影响，表明红细胞在这些过程中起主要作用。因此，在第二个资助期，我们计划更详细地研究红细胞对血管稳态和疾病过程的贡献。具体来说，我们将表型小鼠与红细胞特异性的β-内酰胺酶-1删除和增加红细胞衍生的NO释放，无论是在C57 BL 6/J和高胆固醇血症载脂蛋白E缺陷的背景，包括血管功能调节（WP 1），新生内膜形成（WP 2）和动脉粥样硬化斑块的进展和稳定性（WP 3）。为了确定红细胞还原酶-1对全身NO生物利用度的贡献，将结果与内皮细胞特异性还原酶缺失的小鼠进行比较。基于NO可能通过诱导血管细胞中其他气体递质的合成起作用的观察结果和自己的初步数据，我们将研究内皮细胞和平滑肌细胞在药理学抑制或RNAi介导的H2S生成酶的遗传功能丧失后对小鼠和人红细胞的反应。为了将我们在小鼠和细胞中的发现转化为人类，将在来自风险患者或患有心血管疾病的患者的RBC中检查参与NO生物利用度和血管系统中信号转导的因子的表达和活性的差异。

Experimental evidence and first clinical data suggest that erythrocytes actively participate in vascular function control. However, the concept that erythrocytes directly act on cells of the vessel wall and contribute in cardiovascular disease processes has only recently emerged, and the mediators and signalling pathways involved in their erythrocrine activities are largely unknown. During the first funding period of this project we have shown that nitric oxide from lysed erythrocyte membranes promotes smooth muscle cell mineralisation and atherosclerotic lesion calcification. Based on these findings, we generated hypercholesterolaemic mice lacking arginase-1 specifically in red blood cells to shift the arginase-eNOS balance in erythrocytes towards increased NO generation. The ongoing characterisation of this new mouse line could confirm that the enhanced release of NO from erythrocytes promotes vascular calcification in vitro and in vivo. In addition, we observed that the increased NO bioavailability has systemic effects on vascular homeostasis suggesting a major role of erythrocytes in these processes. In the second funding period, we therefore plan to examine the contribution of erythrocytes to vascular homeostasis and disease processes in more detail. Specifically, we will phenotype mice with erythrocyte-specific arginase-1 deletion and increased erythrocyte-derived NO release, both on the C57BL6/J and the hypercholesterolaemic apolipoprotein E-deficient background, including vascular function regulation (WP1), neointima formation (WP2) and atherosclerotic plaque progression and stability (WP3). To determine the contribution of red blood cell arginase-1 to systemic NO bioavailability, findings will be compared to mice with endothelial cell-specific arginase deletion. Based on observations that NO may act via inducing the synthesis of other gasotransmitters in vascular cells and own preliminary data, we will examine the response of endothelial and smooth muscle cells after pharmacological inhibition or RNAi-mediated genetic loss-of-function of H2S generating enzymes to murine and human erythrocytes. To translate our findings in mice and cells to humans, differences in the expression and activity of factors involved in the bioavailability of NO and signal transduction in the vascular system will be examined in RBCs from patients at risk or with established cardiovascular disease.