LEPTIN REGULATION OF BETA CELL PROLIFERATION

瘦素对 β 细胞增殖的调节

• 批准号: 10625939
• 负责人: RICHARD A COX
• 金额: $ 20万
• 依托单位: BAYLOR COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-07-19 至 2023-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10625939
• 关键词: Affect; Alpha Cell; Beta Cell; Biology; Blood Glucose; Calcium; Cell Proliferation; Cell physiology; Chronic Disease; Collaborations; D Cells; Data; Diabetes Mellitus; Endocrine; Equilibrium; Exhibits; Exocytosis; Exposure to; Feedback; Future; Gene Expression; Gene Targeting; Genetic Transcription; Glucagon; Goals; Hormone secretion; Hormones; Human; Insulin; Islet Cell; Knockout Mice; Knowledge; Lead; Learning; Leptin; Manuscripts; Mediating; Membrane; Metabolic; Modeling; Molecular; Mus; Neurons; Obesity; Outcome; Pancreatic delta Cell; Paracrine Communication; Pathologic; Patients; Persons; Physiological; Play; Positioning Attribute; Protein Tyrosine Kinase; Regulation; Role; STAT3 gene; Signal Transduction; Small Interfering RNA; Somatostatin; Testing; Therapeutic; Time; Universities; Visit; blood glucose regulation; calcium indicator; energy balance; fluorescence imaging; in vivo Model; inhibitor; innovation; insight; insulin secretion; islet; leptin receptor; loss of function; paracrine; response; single-cell RNA sequencing; tool; transcription factor; translational potential

Future Efforts to Strengthen 1R01DK128098-01A1 My goal for the next year is to define molecular and transcriptional mechanisms that govern leptin regulation of islet delta cell functions. The delta cell has emerged as an important regulator of islet function through paracrine actions of somatostatin (SST) that inhibit alpha and beta cells. However, we know little about endocrine signals that perform gene expression and metabolic outcomes in delta cells. Leptin suppresses insulin secretion; however, the leptin receptor (LepR) is exclusively expressed on delta cells of human islets, suggesting leptin may act indirectly on beta cells. Our preliminary data demonstrate for the first time that leptin stimulates SST secretion from human islets which, in turn, decreases insulin secretion from the beta cell. Thus, we propose a model whereby leptin serves as a negative feedback regulator of beta cell insulin secretion by stimulating SST release from delta cells. We will build out our model by achieving the following goals: Cement our central hypothesis that leptin stimulates delta cell SST secretion to inhibit beta cell function. Our preliminary data demonstrate that leptin stimulates SST secretion from human and mouse islets, which suppresses insulin secretion under static conditions. We will employ islet perifusion studies to test dynamic changes in hormone secretion and paracrine signaling that, ultimately, better model in vivo islet responses. - I will visit Duke University to learn islet perifusion from Drs. Jonathan Campbell and David D’Alessio. These efforts will empower me to define dynamic islet responses and paracrine signals following exposure to leptin. - I will leverage new expertise in islet perifusion to define the delta cell specific effects of leptin (loss of function: SST-Cre;LepR-lp/lp mice (previously generated) or siRNA-mediated LepR deletion in human islets) and consequent paracrine effects of SST on insulin and glucagon secretion. These data will firmly establish the premise for my new R01 application that leptin suppresses beta cell function through stimulation of delta cell SST secretion. Define molecular mechanisms of leptin action in delta cells. Islet hormone release involves membrane depolarization and an elevation in intracellular calcium to mediate hormone exocytosis. Although leptin signaling through LepR involves tyrosine kinase activity and JAK-STAT signaling, there is evidence in neurons that leptin increases intracellular calcium (PMID: 30304668). Here, we will test whether leptin increases calcium in delta cells to mediate SST secretion. - I set up a collaboration with Dr. Mark Huising to determine if leptin stimulated SST secretion involves classical exocytotic signals, most notably increased intracellular calcium. Islets expressing the calcium indicator GCaMP6 in delta cells (SST-Cre;Rosa26-lsl-GCaMP; PMID: 28380380, PMID: 27408771) ) will be treated with leptin and GCaMP6 activation will be read out by fluorescence imaging. Revealing leptin effects on calcium levels in delta cells will position future studies to define the signals downstream of LepR that lead to SST secretion. Define transcriptional mechanisms of leptin action in delta cells. Our preliminary data suggest delta cell expression of Stat3 is required for leptin-induced SST secretion. Islets from SST-Cre;Stat3-fl/fl (dStat3 KO) mice exhibit a significantly blunted secretion of SST when treated with leptin compared to control islets. - We will block STAT3 during islet perifusion in the presence of leptin in human (STAT3 inhibitor, TTI-101; Tvardi Therapeutics) or mouse (dStat3 KO) islets to assess the impacts on hormone secretion. - Human islets treated with leptin +/- STAT3 inhibitor (TTI-101) will be subjected to scRNA-seq analysis (expertise previously established as part of a separate manuscript in prep) to determine the delta cell specific gene targets of leptin-STAT3 signaling that define delta cell activities. We will also uncover other transcription factors and close key knowledge gaps that define the regulation of delta cells. Significance. Our proposed studies will reveal leptin stimulates JAK-STAT signaling in delta cells to ready SST secretion, which suppresses beta cell insulin secretion. Leptin is a critical regulator of energy balance and glucose homeostasis and therefore, these findings will provide key insights into leptin action during physiologic (post-prandial) and pathologic (obesity, diabetes) states. We will also determine the potential translational benefits of STAT3 inhibition. Our revised premise focused on islet function combined with new preliminary data using innovative tools will significantly strengthen our new R01 application for late 2022 or early 2023. Our proposed studies for the next year will reveal new biology about delta cells and define molecular and transcriptional mechanisms through which leptin regulates islet function.

增强1R01DK128098-01A1的未来努力 我明年的目标是定义控制胰岛Delta细胞功能的瘦素调节的分子和转录机制。三角细胞通过抑制α和β细胞的生长抑素（SST）的旁分泌作用成为胰岛功能的重要调节剂。但是，我们对在三角洲细胞中执行基因表达和代谢结果的内分泌信号知之甚少。瘦素抑制胰岛素分泌；但是，瘦素接收器（LEPR）仅在人类胰岛的三角细胞上表达，这表明瘦素可能间接作用于β细胞。我们的初步数据首次证明了瘦素刺激人类胰岛的SST分泌，从而减少了β细胞中胰岛素的分泌。因此，我们提出了一个模型，瘦素通过刺激从三角细胞释放SST释放SST来作为β细胞胰岛素分泌的负反馈调节剂。我们将通过实现以下目标来建立我们的模型： 巩固了我们的中心假设，即瘦素刺激三角细胞SST分泌以抑制β细胞功能。我们的初步数据表明，瘦素刺激人类和小鼠胰岛的SST分泌，这在静态条件下抑制了胰岛素分泌。我们将利用胰岛渗出研究来测试同型分泌和旁分泌信号的动态变化，最终在体内胰岛响应中更好地模型。 - 我将访问杜克大学，从博士那里学习胰岛的渗透。乔纳森·坎贝尔和大卫·阿莱西奥。这些努力将使我能够在暴露于瘦素后定义动态胰岛反应和旁分泌信号。 - 我将利用胰岛渗透方面的新专业知识来定义瘦素的Delta细胞特异性作用（功能丧失：SST-CRE； LEPR-LP/LP小鼠（以前产生的）或siRNA介导的LEPR缺失在人类胰岛中）以及随之而来的SST对SST对胰岛素和肝甲分泌的旁分泌效应。 这些数据将首先建立我的新R01应用程序的前提，该应用程序通过刺激Delta细胞SST分泌来抑制β细胞功能。 定义三角细胞中瘦素作用的分子机制。胰岛酮释放涉及膜去极化和细胞内钙的升高，以介导雌激素胞吐作用。尽管通过LEPR信号传导涉及酪氨酸激酶活性和JAK-STAT信号传导，但有证据表明瘦素会增加细胞内钙（PMID：30304668）。在这里，我们将测试三角细胞中钙的瘦素是否增加以介导SST分泌。 - 我与Mark Huising博士建立了合作，以确定瘦素刺激的SST分泌是否涉及经典的胞吐信号，最著名的是增加细胞内钙。在三角洲细胞中表达钙指示剂GCAMP6的胰岛（SST-CRE; ROSA26-LSL-GCAMP; PMID：28380380，PMID：27408771）将通过leptin和gcamp6激活处理。 揭示瘦素对三角洲细胞中钙水平的影响将定位未来的研究，以定义LEPR下游的信号，从而导致SST分泌。 定义三角细胞中瘦素作用的转录机制。我们的初步数据表明STAT3的Delta细胞表达是瘦素诱导的SST分泌所必需的。与对照胰岛相比，SST-CRE; STST-CRE; STAT3-FL/FL（DSTAT3 KO）小鼠SST的分泌显着钝化。 - 在人（Stat3抑制剂，TTI-101； TVARDI THERAPETICS）或小鼠（DSTAT3 KO）胰岛中，我们将在胰岛渗透期间阻止STAT3渗透期间的STAT3，以评估对Horsene分泌的影响。 - 用瘦素+/- STAT3抑制剂（TTI-101）处理的人类胰岛将进行SCRNA-SEQ分析（先前作为PREP中单独手稿的一部分建立的专业知识），以确定lepta细胞特异性基因的特定基因靶标，以定义定义Delta细胞活性。我们还将发现其他转录因子和关键知识差距，以定义三角洲细胞的调节。 意义。我们提出的研究将揭示瘦素刺激三角细胞中的jak-stat信号传导，以抑制β细胞胰岛素分泌。瘦素是能量平衡和葡萄糖稳态的关键调节剂，因此，这些发现将在生理（后态）和病理（肥胖症，糖尿病）状态期间对瘦素作用的关键见解。我们还将确定STAT3抑制的潜在转化益处。我们修订的专注于胰岛功能的前提以及使用创新工具结合新的初步数据将显着增强我们在2022年末或2023年初的新R01应用。我们对明年的拟议研究将揭示有关Delta细胞的新生物学，并定义有关分子和转录机制，从而通过莱特蛋白调节液体调节小岛的功能。