Molecular Mechanisms of Impaired Angiogenesis in Aging Gastric Mucosa

老化胃粘膜血管生成受损的分子机制

• 批准号: 7929333
• 负责人: ANDRZEJ S TARNAWSKI
• 金额: --
• 依托单位: VETERANS HEALTH ADMINISTRATION
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-04-01 至 2014-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7929333
• 关键词: 5' -AMP-activated protein kinase; Acetylation; Acute; Age Reporting; Age-Years; Aging; Albumins; American; Angiogenesis Inhibition; Angiogenesis Inhibitors; Angiogenic Factor; Angiostatins; Antibodies; Apoptosis; Area; Arthritis; Asses; Binding; Biochemical; Blood Vessels; Blood capillaries; CREB1 gene; Cell Nucleus; Cell physiology; Cells; Characteristics; Chronic; Cicatrix; Clinical; Complementary DNA; Connective Tissue; DNA Binding; Dermal; Down-Regulation; Duodenal Ulcer; Economics; Endostatins; Endothelial Cells; Enzymes; Epithelial; Equilibrium; Ethanol; Exhibits; Extravasation; Fluorescein; Funding; Gastric mucosa; Gastric ulcer; Gastritis; Gelatinase B; Gene Activation; Gene Expression; Genes; Genetic Transcription; Goals; Granulation Tissue; HIF1A gene; Healed; Health; Heat shock proteins; Heat-Shock Proteins 90; Human; Hypoxia; Hypoxia Inducible Factor; Image; Imagery; Impaired wound healing; Impairment; Importins; In Vitro; Incidence; Indomethacin; Infarction; Injury; Investigation; Literature; Mediating; Molecular; Morbidity - disease rate; Myocardium; Natural regeneration; Non-Steroidal Anti-Inflammatory Agents; Nuclear; Nutrient; Organ; Oxygen; Pain; Patients; Peptic Ulcer; Pericytes; Permeability; Phosphorylation; Physiological; Plasmids; Population; Predisposition; Prevalence; Properdin; Proteins; Publications; Rattus; Relative (related person); Research; Research Design; Resolution; Role; STAT3 gene; Serum; Site; Small Interfering RNA; Stimulus; Stomach; Structure; Techniques; Technology; Testing; Therapeutic Intervention; Time; Tissues; Transcriptional Activation; Ulcer; Up-Regulation; Vascular Endothelial Growth Factor Receptor-2; Vascular Endothelial Growth Factors; Vascular Permeabilities; Veterans; Work; Wound Healing; age related; alcohol response; alpha Karyopherins; angiogenesis; annexin A5; autocrine; base; capillary; density; gene therapy; healing; high risk; hypoxia inducible factor 1; improved; in vivo; indexing; injured; instrument; mortality; mouse model; mutant; neutralizing antibody; novel; novel therapeutic intervention; novel therapeutics; overexpression; patient population; promoter; protein expression; protein kinase P; receptor; response; response to injury; stem; transcription factor; treatment strategy

DESCRIPTION (provided by applicant): ABSTRACT Background/Rationale: Gastric mucosa of aging humans and rats (aging gastric mucosa) exhibits increased susceptibility to injury and delayed healing. Healing of gastric injury requires re-establishment of mucosal blood vessels through the formation of new vessels (angiogenesis). Previous studies showed that healing of gastric injury in aging rats is delayed and impaired vs. young rats; but the underlying mechanisms are not explained, and angiogenesis in aging gastric mucosa has not been examined before. Our long-term objectives are: 1) to identify the molecular mechanisms of impaired angiogenesis in aging gastric mucosa and based on these findings 2) to develop new therapeutic strategies for treatment and reversing aging- related impaired angiogenesis and delayed healing. Our overall hypothesis is that an imbalance between angiogenic and anti-angiogenic factors suppresses angiogenesis and inhibits healing of injury in aging gastric mucosa. In this project, we will examine the mechanisms responsible for this imbalance and its consequences, ultimately demonstrating that manipulating this imbalance can improve angiogenesis and healing. We hypothesize that the mechanisms underlying impaired angiogenesis in aging gastric mucosa are: 1) Aging-related alterations in gastric microvascular endothelial cell (EC) function - reduced transcriptional activation of the VEGF gene due to: A) downregulation of importin and B) downregulation of the transcriptional factors P-CREB and P-STAT3, 2) Angiogenic imbalance due to reduced pro-angiogenic VEGF and concomitant increased anti-angiogenic endostatin in aging gastric mucosa leads to inhibition of angiogenesis, and 3) Therapeutic interventions aimed at the mechanisms: A) local VEGF gene therapy, B) upregulation and activation of importin with AICAR treatment, or C) specific neutralizing antibody against endostatin will significantly reverse impaired angiogenesis, and will improve microvascular regeneration and healing of chronic gastric ulcers in aging rats. The rationale for these hypotheses stems from our preliminary work that demonstrated in gastric mucosa of aging (vs. young) rats significantly decreased VEGF, increased endostatin, reduced angiogenesis in response to injury, and distorted and impaired mucosal healing. Furthermore, we showed that EC isolated from the gastric mucosa of aging rats exhibit reduced VEGF gene activation, impaired in vitro angiogenesis and reduced angiogenic response to hypoxia. We also demonstrated that importin is an essential requirement for VEGF gene activation and for angiogenesis since its silencing with specific siRNA significantly reduced angiogenesis in young EC. Conversely, activation of importin by treatment with AICAR significantly reversed impaired angiogenesis in aging EC. Research Design: We will examine the above hypotheses in EC isolated from gastric mucosa of young and aging rats and in vivo in gastric mucosa of aging vs. young rats following injury and ulceration. In vitro studies will include analysis of: expression of proteins involved in angiogenesis - VEGF, VEGFR2, P-VEGFR2 as well as HSP90, HIF1, HIF1, importin , AMPK, P-CREB and P-STAT3; their interactions and binding of HIF1, P-CREB and P- STAT3 to the VEGF gene. For in vivo studies, we will examine in gastric mucosa of rats, injury (erosion and ulcer) and will quantitatively asses angiogenesis during gastric ulcer healing, and determine expression of endostatin, MMP9 & others listed proteins. Significance: In 2006, ~39 million Americans (12% of population) were 65 years of age or older, and this number is projected reach ~70 million by 2030. This population is at high risk of tissue and organ injuries. Impaired angiogenesis is a key factor in delayed healing, morbidity and mortality. The central topic of our project is angiogenesis, a critical requirement for "wound healing", which is one of the VA priority areas. PUBLIC HEALTH RELEVANCE: Relevance to Veterans Health. In 2006 nearly 39 million Americans (12% of population) were 65 years of age or older, and this number is projected to grow to about 70 million by 2030. The veteran patient population is aging and patients 70 years or older constitute a significant proportion. This population is at high risk of tissue and organ injuries. Impaired angiogenesis is a key factor in delayed tissue injury healing, morbidity, and mortality. Impaired healing of wounds, both internal, such as gastric and duodenal ulcers or external, such as non-healing dermal ulcers constitute important clinical and economic problems. Gastric injury - gastritis and ulcers are frequently encountered in VA patients often as a result of chronic treatment NSAIDs for arthritis or pain. Lifetime prevalence of gastroduodenal ulcers in the US population is 3-6% but their incidence among the VA population is 2.5-fold higher. The central topic of our proposed investigations is directly related to angiogenesis that is a critical requirement for "wound healing", which is one of the VA priority areas.

描述（由申请人提供）： 摘要背景/基本原理：衰老人类和大鼠的胃粘膜（老化胃粘膜）表现出对损伤和愈合延迟的敏感性增加。胃损伤的愈合需要通过新血管的形成（血管生成）来重建粘膜血管。先前的研究表明，与年轻大鼠相比，老年大鼠胃损伤的愈合被延迟和受损。但其潜在机制尚未得到解释，并且以前也没有研究过老化胃粘膜的血管生成。我们的长期目标是：1）确定老化胃粘膜血管生成受损的分子机制，并基于这些发现2）开发新的治疗策略来治疗和逆转与衰老相关的血管生成受损和延迟愈合。我们的总体假设是，血管生成因子和抗血管生成因子之间的不平衡会抑制血管生成并抑制老化胃粘膜损伤的愈合。在这个项目中，我们将研究造成这种不平衡及其后果的机制，最终证明操纵这种不平衡可以改善血管生成和愈合。我们假设老化胃粘膜血管生成受损的机制是：1) 胃微血管内皮细胞 (EC) 功能的老化相关改变 - VEGF 基因转录激活减少，原因是：A) importin 下调和 B)转录因子 P-CREB ​​和 P-STAT3，2) 由于老化胃粘膜中促血管生成 VEGF 减少和随之而来的抗血管生成内皮抑素增加导致血管生成失衡，导致抑制血管生成，以及 3) 针对以下机制的治疗干预：A) 局部 VEGF 基因治疗，B) 使用 AICAR 治疗上调和激活 importin，或 C) 针对内皮抑素的特异性中和抗体将显着逆转受损的血管生成，并改善微血管衰老大鼠慢性胃溃疡的再生和愈合。这些假设的基本原理源于我们的初步工作，即在衰老（与年轻）大鼠的胃粘膜中，VEGF 显着减少，内皮抑素增加，响应损伤的血管生成减少，以及粘膜愈合扭曲和受损。此外，我们发现从衰老大鼠胃粘膜中分离出的 EC 表现出 VEGF 基因激活降低、体外血管生成受损以及对缺氧的血管生成反应降低。我们还证明了 importin 是 VEGF 基因激活和血管生成的必要条件，因为用特异性 siRNA 沉默它会显着减少年轻 EC 中的血管生成。相反，通过 AICAR 治疗激活 importin 可以显着逆转老化 EC 中受损的血管生成。研究设计：我们将在从年轻和衰老大鼠的胃粘膜中分离出的 EC 以及在损伤和溃疡后的衰老大鼠与年轻大鼠的胃粘膜体内检验上述假设。体外研究将包括分析： 参与血管生成的蛋白质的表达 - VEGF、VEGFR2、P-VEGFR2 以及 HSP90、HIF1、HIF1、importin、AMPK、P-CREB ​​和 P-STAT3； HIF1、P-CREB ​​和 P-STAT3 与 VEGF 基因的相互作用以及结合。对于体内研究，我们将检查大鼠胃粘膜的损伤（糜烂和溃疡），定量评估胃溃疡愈合过程中的血管生成，并测定内皮抑素、MMP9 和其他列出的蛋白质的表达。意义：2006 年，约 3900 万美国人（占人口的 12%）年龄在 65 岁或以上，预计到 2030 年这一数字将达到约 7000 万。这一人群面临组织和器官损伤的高风险。血管生成受损是延迟愈合、发病和死亡的关键因素。我们项目的中心主题是血管生成，这是“伤口愈合”的关键要求，也是 VA 的优先领域之一。 公共卫生相关性： 与退伍军人健康的相关性。 2006年，近3900万美国人（占人口的12%）年龄在65岁或以上，预计到2030年这一数字将增长到约7000万。退伍军人患者群体正在老龄化，70岁或以上的患者占很大比例。该人群是组织和器官损伤的高风险人群。血管生成受损是组织损伤愈合延迟、发病率和死亡率的关键因素。伤口愈合受损，无论是内部伤口，例如胃和十二指肠溃疡，还是外部伤口，例如不愈合的皮肤溃疡，都构成重要的临床和经济问题。胃损伤 - VA 患者经常会出现胃炎和溃疡，通常是由于长期使用非甾体抗炎药治疗关节炎或疼痛而导致的。美国人口中胃十二指肠溃疡的终生患病率为 3-6%，但退伍军人群体中的发病率高出 2.5 倍。我们提出的研究的中心主题与血管生成直接相关，血管生成是“伤口愈合”的关键要求，也是 VA 的优先领域之一。