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Defining alpha-cell proglucagon processing for type 2 diabetes treatment

定义 2 型糖尿病治疗的 α 细胞胰高血糖素原加工过程

基本信息

批准号：
10331361
负责人：
Bethany Paige Cummings
金额：
$ 38.72万
依托单位：
UNIVERSITY OF CALIFORNIA AT DAVIS
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-09-15 至 2022-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10331361
关键词：
Defining alpha cell proglucagon processing

项目摘要

PROJECT SUMMARY: We propose to use state-of-the-art mouse modeling, genetically modified human islets and a unique single cell RNA-seq technique to generate new insights into the regulation of pancreatic islet function that will serve as the basis for developing new therapeutics for type 2 diabetes mellitus (T2DM). The proglucagon-derived peptides, glucagon and glucagon-like peptide-1 (GLP-1), potentiate glucose-stimulated insulin secretion (GSIS). However, unlike GLP-1, glucagon promotes hepatic glucose production and is a weaker inducer of GSIS. We identified a pathway that shifts the alpha cell from producing glucagon to producing GLP-1. We propose to define this pathway to establish a new model describing GLP-1 function and to identify drug targets that increase alpha cell GLP-1 production. Proglucagon is expressed in gut L cells and islet alpha cells and is differentially cleaved depending on the prohormone convertase (PC) present. Canonically, PC1/3 (Pcsk1) is expressed in L cells to produce GLP-1 and PC2 (Pcsk2) is expressed in alpha cells to produce glucagon. However, our data reveal that beta cell GLP-1 receptor (GLP-1R) signaling increases alpha cell GLP-1 and Pcsk1 expression. We hypothesize that beta cell GLP-1R signaling increases alpha cell GLP-1 production to augment GSIS in a paracrine positive feedback loop. We will pursue three aims to define the alpha cell to beta cell (aims 1 and 2) and the beta cell to alpha cell (aim 3) cross-talk involved in our model of beta cell GLP-1R function to identify novel targets that raise alpha cell GLP-1:glucagon levels for T2DM treatment. In aim 1, we will define the contribution of alpha cells to beta cell GLP-1R function in response to endogenous GLP-1 in high fat diet-fed mice. To this end, we will assess glucose regulation and GSIS in low fat diet-fed or high fat diet-fed beta cell GLP-1R wild-type and knockout mice, with or without alpha cell ablation, with or without L cell-derived GLP-1 stimulation. In aim 2, we will define the contribution of alpha cells to beta cell GLP-1R function in response to exogenous GLP-1. To this end, we will assess glucose regulation and GSIS in beta cell GLP-1R wild-type and knockout mice, with or without alpha cell ablation, with or without treatment with a GLP-1R agonist. In aim 3, we will determine the impact of beta cell GLP-1R signaling on alpha cell fate. We show that beta cell GLP-1R signaling increases the expression of Pcsk1 and other beta cell-specific genes in human alpha cells and extend upon previous work showing that alpha cells that express GLP-1 are immature. Thus, we hypothesize that beta cell GLP-1R signaling regulates alpha cell transcriptional programs to promote conversion of alpha cells into beta-like cells. To test this hypothesis and identify putative mediators we will perform IHC, lineage tracing and conditioned media studies in mice. In parallel, we will use a highly sensitive single cell RNA-seq platform that we have developed to assess markers of islet cell identity and transcriptome on mouse and human islets. These data will enable the development of therapies that increase alpha cell GLP-1 production for T2DM treatment.

项目摘要：我们建议使用最先进的小鼠模型，转基因人类胰岛和一种独特的单细胞rna-seq技术，以产生对胰岛调节的新见解。这将成为开发治疗2型糖尿病(T2 DM)的新疗法的基础。这个胰高血糖素原衍生的多肽、胰升糖素和胰升糖素样肽-1(GLP-1)增强葡萄糖刺激胰岛素分泌(GSIS)。然而，与GLP-1不同的是，胰升糖素促进肝脏葡萄糖的产生，是一种 GSIS的诱导剂较弱。我们发现了一条将阿尔法细胞从产生胰高血糖素转移到生产GLP-1。我们建议定义这一途径以建立一个新的模型来描述GLP-1的功能和以确定增加阿尔法细胞GLP-1产量的药物靶点。胰高血糖素原在肠道L细胞和胰岛α细胞，根据前激素转换酶(PC)的存在而不同地被切割。规范地，PC1/3(Pcsk1)在L细胞中表达以产生GLP-1，而PC2(Pcsk2)在α中表达细胞产生胰高血糖素。然而，我们的数据显示，β细胞GLP-1受体(GLP-1R)信号增加阿尔法细胞GLP-1和Pcsk1的表达。我们假设β细胞GLP-1R信号增加在旁分泌正反馈环中，α细胞GLP-1的产生增强了GSIS。我们将追求三个目标定义阿尔法细胞到贝塔细胞(目标1和2)和贝塔细胞到阿尔法细胞(目标3)的串扰我们的β细胞GLP-1R功能模型用于识别提高α细胞GLP-1水平的新靶点： T2 DM治疗。在目标1中，我们将定义α细胞对β细胞GLP-1R功能的贡献高脂饮食小鼠对内源性GLP-1的反应。为此，我们将评估血糖调节和低脂饮食或高脂饮食喂养的GLP-1R野生型和基因敲除小鼠中的GSIS 细胞消融，加或不加L细胞来源的GLP-1刺激。在目标2中，我们将定义阿尔法的贡献细胞对外源GLP-1的反应使细胞向β细胞GLP-1R转化。为此，我们将评估葡萄糖 β细胞GLP-1R野生型和基因敲除小鼠的调节和GSIS，有或没有α细胞消融，或不使用GLP-1R激动剂治疗。在目标3中，我们将确定β细胞GLP-1R的影响阿尔法细胞命运的信号。我们发现，β细胞GLP-1R信号增加了Pcsk1和Pcsk1的表达人类阿尔法细胞中的其他β细胞特异性基因，并在先前工作的基础上延伸，表明阿尔法细胞表达GLP-1的人是不成熟的。因此，我们假设β细胞GLP-1R信号调节α细胞促进阿尔法细胞转化为类贝塔细胞的转录程序。为了检验这一假说，确定可能的介质我们将在小鼠身上进行IHC、谱系追踪和条件培养研究。在……里面同时，我们将使用我们开发的高度敏感的单细胞rna-seq平台来评估标记。小鼠和人类胰岛细胞特性和转录组的研究。这些数据将使开发为治疗2型糖尿病而增加α细胞GLP-1产生的疗法。