Mechanisms of Dietary Restriction mediated by Creb-binding Protein in Mice

描述（由申请人提供）：在动物模型中，饮食限制是一种减少热量摄入并随后延长寿命和预防与年龄相关疾病的操作。这种操作的保护范围包括癌症、糖尿病、心血管疾病和阿尔茨海默病等流行疾病。因此，有必要了解这种范式的潜在代谢机制。CBP是一种转录调节因子，与饮食限制的保护和延长年龄特性有关。重要的是，有证据表明CBP主要通过作用于下丘脑（代谢状态的感觉中心）来发挥这些作用。这些研究的目的是研究CBP的作用和饮食限制的代谢特征，重点是下丘脑的反应。为了进行这些研究，我们将培养出生后缺乏CBP的小鼠。然后，这些小鼠将受到急性（8周）或慢性（18个月）饮食限制。在此之后，将收集相关组织并分析其代谢谱。具体来说，我们将1)使用定制的qPCR板监测代谢基因表达的变化；2)利用染色质免疫沉淀法和RT-PCR法观察代谢基因靶向CBP的变化；3)用液相色谱法和质谱法定量代谢物的使用。这些检测将在慢性和急性组中进行。此外，我们将使用衰老遗传标记来评估慢性饮食限制小鼠，这将有助于辨别保护作用是否归因于下丘脑中的CBP表达。其他神经内分泌反应将监测所有组进一步表征这种饮食操作的生理学。该项目完成后，预计将建立更清晰的饮食限制的代谢和转录谱，使我们离临床应用的发展更近一步。

DESCRIPTION (provided by applicant): Dietary Restriction is a manipulation that reduces caloric intake and subsequently extends lifespan and protects against age-related diseases in animal models. The protective spectrum of this manipulation includes prevalent diseases such as cancer, diabetes, cardiovascular disease, and Alzheimer's disease. It is therefore imperative to understand the underlying metabolic mechanisms of this paradigm. CBP is a transcriptional regulator that has been implicated in both the protective and age-extending properties of dietary restriction. Of importance, evidence suggests CBP exerts these effects by mainly acting in the hypothalamus, a sensory center of metabolic state. The purpose of these studies is to study the role of CBP and the metabolic profile of dietary restriction with an emphasis in hypothalamic responses. To carry these studies we will develop mice that lack CBP post-natally. These mice will then be subjected to either acute (8 weeks) or chronic (18 month) dietary restriction. Following this period, relevant tissues will be collected and analyzed for metabolic profiles. Specifically, we will 1) monitor metabolic gene expression changes using custom qPCR plates; 2) observe metabolic gene targeting changes of CBP using Chromatin immunoprecipitation followed by RT-PCR; 3) quantify metabolite usage by liquid chromatography followed by mass spectrometry. These assays will be performed in both chronic and acute groups. Additionally, we will assess the chronically dietary restricted mice using genetic markers of senescence, which will help discern if the protective effects are attributable to CBP expression in the hypothalamus. Other neuroendocrine responses will be monitored for all groups further characterizing the physiology of this dietary manipulation. Upon completion of this project it is expected that a clearer metabolic and transcriptional profile of dietary restriction will be established, taking us one step closer to the development of clinical applications.