Role of the hormone LEAP2 in eating, body weight, blood glucose and survival

项目总结 激素酰基-生长激素是饮食、体重、血糖和生存的关键调节因素。 与其受体GHSR结合。在过去的项目期间，我们研究了Ghrelin分泌的调节。一位少校 研究发现，在模拟饥饿的严重热量限制协议下，Ghrelin细胞表达的1- 肾上腺素能受体(1AR)介导Ghrelin的分泌，而Ghrelin反过来又能防御显著的低血糖 和死亡率。此外，随着最近发现富含肝脏的抗菌肽2(Leap2)是一种 内源性GHSR拮抗剂，我们将研究扩展到在细胞水平上表征Leap2的功能 并测定受试者和小鼠因各种代谢紊乱引起的血浆Leap2变化。 我们发现Leap2既能超极化，又能阻止酰基-Ghrelin激活弓状NPY神经元， 提示Leap2既是一种GHSR反向激动剂，又是一种拮抗剂。此外，我们还发现，血浆 Leap2受代谢状态调节：其水平随着肥胖和血糖升高而增加，然后降低 禁食和减肥。这些变化大多与酰基-Ghrelin相反。总的来说，这导致了 美国提出Leap2功能的以下模型：1)在肥胖状态下，Leap2升高，酰基-Ghrelin下降， 使血浆Leap2/acyl-ghrelin摩尔比升高，从而限制acyl-ghrelin恶化的能力 通过增加食物摄入量、体重和血糖来减少肥胖和糖耐量。2)在营养方面 缺乏状态，例如由严重的卡路里限制引起的状态，Leap2的下降在 哪种升高的酰基胃促生长素能最有效地预防危及生命的低血糖。在当前 R01提案，我们在3个AIMS上测试了这个模型，使用了一系列最新的工具来删除、击倒、 和/或在肥胖和严重热量限制的情况下中和Leap2。在目标1中，我们测试删除 或阻断Leap2会加剧肥胖环境中的肥胖和糖耐量异常，我们确定 Leap2和Ghrelin进入不同中枢神经系统区域的程度。在目标2中，我们测试在 严重卡路里限制协议建模饥饿，删除Leap2将进一步增强 激活生长激素受体以提高血糖和存活率。在目标3中，我们选择性地从肝脏或 肠道-Leap2的两个主要来源-并评估血浆Leap2和代谢的变化 对长期高脂肪饮食暴露和严格的热量限制的反应。我们将使用一个包含四个元素的集合 新的、未发表的重组小鼠品系，允许我们删除或选择性删除Leap2和/或 Ghrelin，与诱导Leap2表达下调的新型病毒载体一起，以及Leap2 中和性单抗。我们的研究将提供对功能意义的基本见解 最近鉴定的GHSR拮抗剂和反向激动剂Leap2以及相关的GHSR激动剂acyl- Ghrelin在营养过剩和糖耐量过剩环境下肥胖和糖耐量异常的发生 在严重热量限制的情况下发生危及生命的低血糖。

PROJECT SUMMARY The hormone acyl-ghrelin serves as a key regulator of eating, body weight, blood glucose, and survival upon binding to its receptor, GHSR. In the past project period, we studied regulation of ghrelin secretion. A major finding was that under a severe caloric restriction protocol modeling starvation, ghrelin cell-expressed 1- adrenergic receptors (1ARs) mediate ghrelin secretion, which in turn defends against marked hypoglycemia and mortality. Also, following the recent identification of liver-enriched antimicrobial peptide 2 (LEAP2) as an endogenous GHSR antagonist, we extended our studies to characterize LEAP2 function at the cellular level and to determine plasma LEAP2 changes due to various metabolic perturbations in human subjects and mice. We found that LEAP2 both hyperpolarizes and prevents acyl-ghrelin from activating arcuate NPY neurons, suggesting that LEAP2 serves as both a GHSR inverse agonist and antagonist. Also, we found that plasma LEAP2 is regulated by metabolic status: its levels increase with obesity and rising blood glucose and decrease with fasting and weight loss. These changes were mostly opposite of those of acyl-ghrelin. Collectively, this led us to propose the following model of LEAP2 function: 1) In obese states, LEAP2 rises and acyl-ghrelin falls, shifting the plasma LEAP2/acyl-ghrelin molar ratio higher and thus limiting acyl-ghrelin’s capacity to worsen obesity and glucose intolerance by raising food intake, body weight and blood glucose. 2) In nutritionally deficient states, such as that induced by severe caloric restriction, a fall in LEAP2 creates an environment in which elevated acyl-ghrelin can most effectively act to prevent life-threatening hypoglycemia. In the current R01 proposal, we test this model in 3 aims by using a combination of mostly novel tools to delete, knockdown, and/or neutralize LEAP2 in settings of obesity and severe caloric restriction. In Aim 1, we test whether deleting or blocking LEAP2 will exacerbate obesity and glucose intolerance in obesogenic settings, and we determine the extent to which LEAP2 and ghrelin gain access to different CNS regions. In Aim 2, we test whether under a severe caloric restriction protocol modeling starvation, deleting LEAP2 will further enhance the capacity of activated GHSRs to boost blood glucose and survival. In Aim 3, we delete LEAP2 selectively from liver or intestine – the two predominant sources of LEAP2 – and assess changes in plasma LEAP2 and metabolism in response to long-term high fat diet exposure and severe caloric restriction. We will use a collection of four new, unpublished recombinant mouse lines that allow us to delete or site-selectively delete LEAP2 and/or ghrelin, together with a novel viral vector that induces knockdown of LEAP2 expression, and a LEAP2 neutralizing monoclonal antibody. Our studies will provide fundamental insight into the functional significance of the recently characterized GHSR antagonist and inverse agonist LEAP2 and the related GHSR agonist acyl- ghrelin in the development of obesity and glucose intolerance under settings of nutritional overabundance and in the development of life-threatening hypoglycemia under settings of severe caloric restriction.