Effects of ACE2 gene therapy on Diabetes

描述(申请人提供)：糖尿病在世界各地都是一个日益严重的问题。I型和II型糖尿病的临床试验和动物模型表明，血管紧张素-II(Ang-II)信号通路的过度活跃参与了糖尿病和糖尿病并发症的发生。与临床相关的是，阻断肾素-血管紧张素系统(RAS)可预防新发糖尿病并降低糖尿病并发症的风险。血管紧张素转换酶(ACE)2是新近发现的一种单羧基肽酶，是ACE的第一个同源物。它被认为主要通过裂解Ang-II产生Ang-(1-7)来抑制Ang-II信号转导通路，Ang-(1-7)的作用与Ang-II相反，并由Mas受体介导。这种酶存在于各种组织和器官中，包括肾脏、肝脏、脂肪组织和胰腺。在糖尿病和糖尿病肾病早期，其在内分泌胰腺中的表达升高。胰岛同时表达RAS组分和NADPH氧化酶(NOx)组分，这两个组分是介导氧化应激的关键成分。在胰岛中，Ang-II和氧化应激都能降低胰岛素基因的表达和分泌。ACE2被认为与ACE/Ang-II/AT1受体轴相反，可能通过抑制NOx活性和Ang-II介导的胰岛素基因表达和分泌减少来保护胰岛β细胞功能。为了操纵ACE2的表达，我们产生了一种编码ACE2的新型腺病毒，并报告了ACE2在细胞和组织中的mRNA、蛋白质和活性增加。我们假设，ACE2在胰腺中的过度表达将减少氧化应激，改善β细胞功能，从而改善糖尿病小鼠的葡萄糖稳态。为了验证这一假设，我们将解决以下具体目标：1)确定ACE2表达和/或活动与糖尿病之间是否存在关系；2)评估ACE2过度表达在糖尿病小鼠中的后果；3)确定ACE2过度表达是否改善糖尿病患者的胰岛β细胞功能。为了实现这些目标，我们将首先使用ACE2基因敲除和db/db小鼠(2型糖尿病)模型来建立ACE2和糖尿病之间的关系。然后，我们将结合最先进的分子、药理学和生理学工具，对糖尿病前期和年轻糖尿病db/db小鼠进行体外(离体胰岛)和体内(胰腺)基因治疗。最后，我们将讨论ACE2可能通过哪些机制潜在地抵消糖尿病中过度活跃的RAS的有害影响。这项建议将展示ACE2过度表达对血糖正常化的有益影响的证据，并突出ACE2作为糖尿病治疗的新靶点。 公共卫生相关性：大约8%的美国人受到糖尿病的影响，糖尿病是众所周知的心血管疾病的危险因素，由于目前西方世界的肥胖症流行，预计这一比例还会增加。在2型糖尿病小鼠模型中使用基因疗法，这项应用将描述一种新的酶，ACE2，调节血糖水平的能力。如果得到证实，ACE2可能成为治疗2型糖尿病的新靶点。

DESCRIPTION (provided by applicant): Diabetes is a growing problem in all parts of the World. Clinical trials and animal models of type I and type II diabetes have shown that hyperactivity of angiotensin-II (Ang-II) signaling pathways contribute to the development of diabetes and diabetic complications. Of clinical relevance, blockade of the renin-angiotensin system (RAS) prevents new-onset diabetes and reduces the risk of diabetic complications. Angiotensin converting enzyme (ACE) 2 is a recently discovered mono- carboxypeptidase and the first homolog of ACE. It is thought to inhibit Ang-II signaling cascades mostly by cleaving Ang-II to generate Ang-(1-7), which effects oppose Ang-II and are mediated by the Mas receptor. The enzyme is present in various tissues and organs, including the kidney, liver, adipose tissue and pancreas. Its expression is elevated in the endocrine pancreas in diabetes and in the early phase during diabetic nephropathy. Pancreatic islets express both RAS components and NADPH oxidase (Nox) components, which are key elements in mediating oxidative stress. In the islet Ang-II and oxidative stress are both capable of decreasing insulin gene expression and secretion. ACE2 is hypothesized to oppose the ACE/Ang-II/AT1 receptor axis and may protect pancreatic beta- cell function by inhibiting both Nox activity and the Ang-II-mediated reduction of insulin gene expression and secretion. To manipulate ACE2 expression, we generated a novel adenovirus coding for ACE2 and reported increased ACE2 mRNA, protein and activity in cells and tissues. We hypothesize that ACE2 over-expression in the pancreas will reduce oxidative stress and ameliorate beta-cell function, thus leading to improved glucose homeostasis in diabetic mice. To test this hypothesis, we will address the following specific aims: 1) Determine the existence of a relationship between ACE2 expression and/or activity and diabetes; 2) Evaluate the consequences of ACE2 over-expression in diabetic mice; 3) Establish whether ACE2 over-expression improves pancreatic beta-cell function in diabetes. To achieve these goals, we will first use ACE2 knockout and db/db mouse (type 2 diabetes) models to establish the relationship between ACE2 and diabetes. Then we will combine state of the art molecular, pharmacological and physiological tools for in vitro (isolated islets) and in vivo (pancreas) gene therapy in pre-diabetic and young diabetic db/db mice. Finally, we will address the mechanisms by which ACE2 could potentially counterbalance the deleterious effects of the hyperactive RAS in diabetes. This proposal will show evidence of the beneficial effects of ACE2 over-expression on the normalization of blood glucose and highlight ACE2 as a new target for the treatment of diabetes. PUBLIC HEALTH RELEVANCE: Approximately 8% of Americans are affected by diabetes, a well known risk factor for cardiovascular diseases, and it is expected to grow due to the current obesity epidemic in the Western World. Using gene therapy in a mouse model of type 2 diabetes, this application will describe the ability of a new enzyme, ACE2, to regulate blood glucose levels. If confirmed, ACE2 could become a new target for the treatment of type 2 diabetes.