Nerve growth factor – The key factor in endothelial aging, angiogenesis and gastric ulcer healing

 描述(由申请人提供)： 背景/原理：血管生成--新的毛细血管的形成是组织损伤修复的基本过程，如胃糜烂(GE)和溃疡(GU)。衰老的人和大鼠的胃粘膜(衰老的胃粘膜)表现出对损伤的易感性，血管生成受损和愈合延迟。[我们的总体假设是，神经生长因子(NGF)对胃内皮细胞(EC)的活性、功能、血管生成和GU愈合至关重要，而衰老ECs中NGF缺陷是老年人胃血管生成和GU愈合的主要机制。]这一假说的理论基础来源于我们的初步研究，结果表明：1)衰老的胃内皮细胞(AGECs)NGF表达减少，NGF蛋白和/或NGF基因治疗在体外恢复了衰老ECs的血管生成，并逆转了NSAIDs对血管生成的抑制；[2)NGF局部治疗老年大鼠GU显著增加NGF和VEGF的局部表达，促进血管生成，加速溃疡愈合，促进粘膜再生。]我们的具体目标是确定是否以及在多大程度上：1)NGF及其信号转导对胃内皮细胞的活性、功能和血管生成至关重要，并确定 2)NGF缺乏是内皮细胞衰老和衰老相关的内皮细胞增殖和血管生成障碍的关键机制，以及(B)NGF治疗或NGF基因治疗是否能在体外恢复衰老内皮细胞的血管生成；[3)使用NGF局部治疗GUS将促进血管再生和GU愈合，以及这些事件是否可以通过激光共聚焦内窥镜(CLE)在体内和实时显示。]我们的长期目标是揭示调节胃血管生成及其在衰老过程中的损伤的新机制，为NGF治疗用于促进老年人GU的血管生成和愈合提供基础，并利用CLE在体内评估GU的愈合和血管再生，以及识别可能是溃疡复发的基础的GU瘢痕异常。设计：由于内皮细胞是血管生成的重要靶点和效应细胞，在体外分离的幼年和老年大鼠胃粘膜内皮细胞的研究中，我们将确定增龄性内皮细胞功能损害的机制，NGF及其受体、PI3K/Akt和MAPK信号通路的表达，以及NGF在EC存活、迁移、增殖和血管生成中的作用。接下来，我们将在体内检测老年和年轻大鼠GU愈合和血管再生的一系列事件，并将确定局部注射NGF是否会促进老年大鼠GU愈合并防止非类固醇抗炎药诱导的GU复发。我们还将使用来自年轻人和老年人的人类GUS样本来确定我们的实验结果与人类的相关性。意义：我们项目的中心主题是新生血管、损伤愈合和衰老的影响。2006年，约有3900万美国人(占总人口的12%)年龄在65岁或以上，预计到2030年，这一数字将达到约7000万。这一人群是组织和器官损伤以及胃溃疡并发症的高危人群。随着年龄的增长，血管生成受损是延迟愈合、发病率和死亡率的关键因素。[最近的研究表明，非H。幽门螺杆菌，非NSAIDs溃疡(“特发性”溃疡)正在增加，它们占所有溃疡和患者高达30% 有“特发性”出血性溃疡病史的患者复发性溃疡出血风险高，死亡率高达83-92%。因此，胃溃疡的复发及其并发症仍然是重要的临床问题，尤其是在高龄和重症患者中。这项拟议的研究将深入了解NGF和血管生成在GU愈合过程中的新机制和调节作用，以及它们与这些过程的衰老相关损害的关系。这一新的信息可以适用于其他领域的粘膜愈合，包括炎症性肠病，食道侵蚀和溃疡，以及癌症相关的新生血管。

 DESCRIPTION (provided by applicant): Background/Rationale: Angiogenesis - formation of new capillary blood vessels is a fundamental process essential for healing of tissue injury, e.g. gastric erosions (GE) and ulcers (GU). Gastric mucosa of aging humans and rats (aging gastric mucosa) exhibits increased susceptibility to injury, impaired angiogenesis and delayed healing. [Our overall hypothesis is that nerve growth factor (NGF) is critical for gastric endothelial cell (EC) viability, function, angiogenesis and GU healing, and that NGF deficiency in aging ECs is the major mechanism of aging-related impairment of gastric angiogenesis and GU healing in aging individuals.] The rationale for this hypothesis stems from our preliminary studies showing that: 1) aging gastric ECs (AGECs) have reduced expression of NGF, and NGF protein and/or NGF gene therapy restores in vitro angiogenesis in aging ECs cells and reverses NSAIDs-induced inhibition of angiogenesis, and [2) local treatment of GU in aging rats with NGF significantly increases local expression of NGF and VEGF, angiogenesis, accelerates ulcer healing, and improves mucosal regeneration.] Our specific objectives are to determine whether and to what extent: 1) NGF and its signaling are critical for gastric ECs viability, function and angiogenesis and to identify the underlying molecular mechanisms; 2) NGF deficiency is: (a) crucial mechanism for ECs' aging and for aging-related impairment of EC proliferation and angiogenesis and (b) whether NGF treatment or NGF gene therapy restores in vitro angiogenesis in aging ECs; [3) Local treatment of GUs in aging rats with NGF will improve vascular regeneration and GU healing and whether these events can be visualized in vivo and in real time using confocal laser endomicroscopy (CLE).] Our long-term objective is to uncover novel mechanisms that regulate gastric angiogenesis and its impairment in aging, to provide a basis for the therapeutic use of NGF to improve angiogenesis and healing of GU in aging individuals and to use CLE for in vivo assessment of GU healing, vascular regeneration and for identifying GU scar abnormalities that may be the basis for the ulcer recurrence. Design: Since ECs are critical targets and effectors of angiogenesis, in in vitro studies in ECs isolated from gastric mucosa of young and aging rats we will determine the mechanisms underlying aging related impairment of EC functions, expression of NGF, VEGF, their receptors, PI3K/Akt and MAPK signaling, and the role of NGF in EC survival, migration, proliferation and angiogenesis. Next we will examine in vivo in aging and young rats sequential events of GU healing and vascular regeneration and will determine whether locally injected NGF will improve GU healing in aging rats and prevent NSAIDs-induced GU recurrence. We will also determine human relevance of our experimental findings using human specimens of GUs from young and aging individuals. Significance: The central topic of our project is neovascularization, injury healing and impact of aging. In 2006, ~39 million Americans (12% of population) were 65 years of age or older, and this number is projected to reach ~70 million by 2030. This population is at high risk of tissue and organ injuries, and GU complications. Impaired angiogenesis in aging is a key factor in delayed healing, morbidity and mortality. [Recent studies indicate that the incidence of non-H. pylori, non-NSAIDs ulcers ("idiopathic" ulcers) is increasing and they constitute up to 30% of all ulcers and patients with a history of "idiopathic" bleeding ulcers have a high risk of recurrent ulcer bleeding and a high mortality 83 - 92%. Therefore the GU recurrence and its complications still remain important clinical issue, especially in aging and severely ill patients.] The proposed studies will provide insight into novel mechanisms and the regulatory role of NGF and angiogenesis in GU healing and their relation to aging-related impairment of these processes. This novel information can be applicable to other areas of mucosal healing including inflammatory bowel disease, esophageal erosions and ulcers, as well as to cancer-related neovascularization.