Effect of CTHRC1 on endothelial cell survival after acute ischemia

摘要 在以前的研究中，我们已经证明了CTHRC 1（胶原蛋白三重受阻重复包含1），一个因素， 在我们的实验室中发现的，只在大脑和骨骼中组成型表达，而不在其他组织中表达。 成年然而，CTHRC 1在组织的活化的成纤维细胞和间质细胞中高度表达 正在重塑和修复我们发现CTHRC 1是一种循环因子，但仅约 30%的健康人类受试者在血浆中具有可检测水平的CTHRC 1，浓度范围为 低pg/ml至近100 ng/ml。与大多数人相似，CTHRC 1的循环水平在小鼠中不可检测 并且在Pdgfrb启动子控制下CTHRC 1的强制转基因过表达也不会导致 在可检测的CTHRC 1血浆水平。因此，有两个CTHRC池1;一个是在本地生成的， 需要>48小时才能获得的正在进行修复的组织，以及循环CTHRC 1的第二池 在任何时候都可以得到，并且只在少数人类受试者中发现。循环CTHRC 1的意义 当我们从患有心血管疾病的患者中获得血浆样本时， 心脏骤停，死亡率约为50%。高CTHRC 1水平（≥0.75ng/ml）与 在经历心脏骤停的人中具有显著更高的存活率。CTHRC 1不表达 在成人心脏中，它在响应心肌梗塞而活化的成纤维细胞中高度诱导。测试 为了研究CTHRC 1在急性缺血性损伤中的作用，我们在Cthrc 1基因敲除小鼠中进行冠状动脉结扎， 在Cthrc 1空白背景下的转基因小鼠中， 在人体中发现的ng/ml范围。引人注目的是，70%的Cthrc 1基因敲除小鼠在缺血性损伤后3-4天死亡 而所有转基因小鼠都存活了下来。为了确定这一戏剧性发现的机制， 进行了体外研究，发现CTHRC 1促进多种细胞类型的细胞存活，包括 内皮细胞完整的代谢监测显示，Cthrc 1基因敲除小鼠的能量增加， 静息消耗和体外细胞代谢分析显示，在CTHRC 1存在下， 线粒体呼吸显著增加，而糖酵解减少，导致我们假设 CTHRC 1是代谢效率的调节者。总的来说，这一提议将检验以下假设： CTHRC 1在细胞应激条件下作为细胞存活的介导剂，通过增加代谢 效率，这反过来又保护急性缺血性损伤的致命后果。我们将确定是否 增加循环或CTHRC 1组织水平通过限制有害的 心肌缺血的后果。使用遗传小鼠模型和体外方法， 将确定作用机制，这将为新的治疗方法提供基础， 改善急性缺血性疾病预后。

Abstract In previous studies we have demonstrated that CTHRC1 (Collagen Triple Helix Repeat Containing 1), a factor discovered in our laboratory, is constitutively expressed only in brain and bone and not in other tissues during adulthood. However, CTHRC1 is highly expressed in activated fibroblasts and interstitial cells of tissues undergoing remodeling and repair. We discovered that CTHRC1 is a circulating factor but only approximately 30% of healthy human subjects have detectable levels of CTHRC1 in plasma, ranging in concentration from low pg/ml to almost 100ng/ml. Similar to most humans, circulating levels of CTHRC1 are not detectable in mice and rats and forced transgenic overexpression of CTHRC1 under Pdgfrb promoter control does also not result in detectable CTHRC1 plasma levels. Thus there are two pools of CTHRC1; one that is generated locally in tissues undergoing repair requiring >48 hours to be available, and a second pool of circulating CTHRC1 available at all times and found only in a minority of human subjects. The significance of circulating CTHRC1 for the cardiovascular system became apparent when we obtained plasma samples from patients experiencing cardiac arrest, a condition with approximately 50% mortality. High CTHRC1 levels (≥0.75ng/ml) are associated with substantially higher survival rates in humans experiencing cardiac arrest. While CTHRC1 is not expressed in the adult heart, it is highly induced in fibroblasts activated in response to myocardial infarction. To test the role of CTHRC1 in acute ischemic injury, we performed coronary artery ligation in Cthrc1 null mice and Cthrc1 transgenic mice on the Cthrc1 null background with physiologically relevant CTHRC1 plasma levels in the ng/ml range found in humans. Strikingly, 70% of Cthrc1 null mice died 3-4 days after the ischemic injury whereas all transgenic mice survived. With the goal of identifying the mechanism for this dramatic finding we performed in vitro studies and found that CTHRC1 promotes cell survival in variety of cell types including endothelial cells. Complete metabolic monitoring revealed that Cthrc1 null mice have increased energy expenditure at rest and analysis of cell metabolism in vitro revealed that in the presence of CTHRC1 mitochondrial respiration is significantly increased whereas glycolysis is reduced, leading us to hypothesize that CTHRC1 functions as a mediator of metabolic efficiency. Overall, this proposal will test the hypothesis that CTHRC1 functions as a mediator of cell survival under conditions of cell stress by increasing metabolic efficiency, which in turn protects from the deadly consequences of acute ischemic injury. We will determine if increasing circulating- or CTHRC1 tissue levels provides cardiovascular protection by limiting the deleterious consequences of myocardial ischemia. Using genetic mouse models and in vitro approaches the underlying mechanism of action will be identified, and this will provide the foundation for novel therapeutic approaches to improve outcomes of acute ischemic conditions.