Cysteine-rich with EGF-like domains 2 – Exodosis and angiogenesis after myocardial infarction

急性心肌梗死对冠状动脉微循环造成巨大损伤，导致血管解体和毛细血管稀疏。几天内，梗死区域形成新的血管系统，有助于限制疤痕形成和左心室重塑的程度。内质网（ER）在许多自分泌和旁分泌血管生成生长因子的折叠、翻译后修饰和分泌中起着关键作用。然而，缺血再灌注损伤和心肌梗死后分泌活性的增加可能会破坏内质网稳态（内质网应激）。与KDEL受体的结合决定了蛋白质是否保留在内质网/高尔基体系统中或被分泌。因此，许多ER驻留伴侣具有经典的c端Lys-Asp-Glu-Leu (KDEL) ER保留信号。缺乏这个基序的生长因子被分泌。相反，具有弱内质网保留信号的蛋白质在基础条件下保留在内质网中，并在内质网胁迫下优先分泌（外源性）。我们提出，外逸提供了一种机制，刺激血管生成，特别是在梗死区域。我们在小鼠梗死区域进行了基于单个内皮细胞rna测序的生物信息学分泌组分析，以鉴定er应激内皮细胞产生的先前未表征的生长因子。因此，我们确定了富含egf样结构域2的半胱氨酸。CRELD2具有较弱的内质网保留信号（REDL），在内质网应激时被强烈诱导。根据我们的初步数据，我们假设内皮细胞在梗死区域通过外源性释放CRELD2，以自分泌的方式刺激血管生成和伤口愈合。这两种假设都将在我们的项目中进行研究。我们打算测量CRELD2在小鼠和人急性心肌梗死中的表达和分泌。我们想研究血管生成和心肌梗死后创面愈合在全局性或条条性creld2缺陷小鼠。我们的目的是确定CRELD2是否在梗死区域作为内质网驻留蛋白和/或分泌蛋白，并探索重组CRELD2在心肌梗死后的治疗潜力。最后，我们计划鉴定内皮细胞中假定的CRELD2细胞表面受体，并表征磷酸化蛋白组的下游变化。我们的项目将以CRELD2为例，确立外源性是心肌梗死后血管生成和创面愈合的重要刺激因素。

Acute myocardial infarction inflicts massive injury to the coronary microcirculation leading to vascular disintegration and capillary rarefication. Within days, a new vascular system develops in the infarct area and helps limiting the extent of scar formation and left ventricular remodelling. The endoplasmic reticulum (ER) plays a pivotal role in the folding, posttranslational modification, and secretion of many autocrine- and paracrine-acting angiogenic growth factors. However, ischemia-reperfusion injury and the increased secretory activity after myocardial infarction may overwhelm ER homeostasis (ER stress). Binding to KDEL receptors determines whether proteins are retained in the ER/Golgi system or are secreted. Accordingly, many ER-resident chaperones possess a classical, C-terminal Lys-Asp-Glu-Leu (KDEL) ER retention signal. Growth factors lacking this motif are secreted. Conversely, proteins with a weak ER retention signal are retained in the ER under basal conditions and are preferentially secreted under ER stress (exodosis). We propose that exodosis provides a mechanism to stimulate angiogenesis specifically in the infarct area. We have performed a single endothelial cell RNA-sequencing-based bioinformatic secretome analysis in the infarct area of mice to identify previously uncharacterised growth factors produced by ER-stressed endothelial cells. We thus identified cysteine-rich with EGF-like domains 2. CRELD2 has a weak ER retention signal (REDL) and is strongly induced during ER stress. Based on our preliminary data, we postulate that endothelial cells release CRELD2 in the infarct area via exodosis to stimulate angiogenesis and wound healing in an autocrine manner. Both hypotheses will be investigated in our project. We intend to measure the expression and secretion of CRELD2 in mice and humans with acute myocardial infarction. We want to study angiogenesis and wound healing after myocardial infarction in globally or conditionally Creld2-deficient mice. We aim to define if CRELD2 acts as an ER resident and/or secreted protein in the infarct area and to explore the therapeutic potential of recombinant CRELD2 after myocardial infarction. Finally, we plan to identify the putative CRELD2 cell surface receptor in endothelial cells and to characterise downstream alterations in the phosphoproteome. Using CRELD2 as an example, our project shall establish exodosis as an important stimulus of angiogenesis and wound healing after myocardial infarction.