Pathophysiology of Diabetic Gastroparesis

项目总结/摘要 糖尿病与多种并发症相关，包括糖尿病性胃轻瘫。全身 糖尿病的影响改变了体内的所有细胞，但与其他糖尿病并发症，胃轻瘫， 并不是所有的病人都有。在没有梗阻的情况下，胃排空延迟（GE）定义为 胃轻瘫，并且也发生在一些但不是所有的糖尿病小鼠中。这些观察表明，与 由于遗传学的原因，全身变化与局部因素的融合是糖尿病发生迟发性GE的必要条件。 我们以前已经发现，延迟GE与Cajal间质细胞（ICC）的损失有关， 反过来与更少的CD 206+固有肌层巨噬细胞（MPM）相关。MPM在以下方面发挥着核心作用： 改变健康和疾病中GI固有肌层的细胞内容物和生物环境。 胃MPM的来源和糖尿病激活MPM中损伤分子的机制是 不知道。这一提议的总体假设是，延迟的潜在中心决定因素是 糖尿病的胃排空是由胃粘膜中诱发的程序性激活反应的组合， 单核细胞和胃肌层中新的单核细胞源性MPM的组织环境 propria。这将在三个具体目标（SA）中进行测试。SA 1将确定糖尿病对循环系统的影响。 单核细胞，使其在胃固有肌层中浸润和活化。SA 2将确定 在糖尿病和SA 3发病后，聚集在胃固有肌层的激活的MPM的起源将 确定新建立和激活的MPM如何导致糖尿病胃细胞损伤。的SAs 有大量的初步数据支持。1)我们发现全身暴露于糖尿病会改变 循环单核细胞的表观基因组景观，使得单核细胞准备表达限制性的 当它们进入组织并成为MPM时，信号通路的补充; 2）糖尿病患者中的MPM 3）糖尿病导致存在新的、不同的、离散的和不连续的细胞， 4）谱系标记的、长寿的MPM不被糖尿病改变; 5)糖尿病引起的胃固有肌层细胞代谢紊乱激活单核细胞源性 MPM与ICC紧密贴壁，产生有害的细胞因子，导致ICC损伤和中断 胃功能为了解决总体假设，我们开发了分离已识别MPM的技术 和单核细胞，应用高度复杂的技术通过单细胞RNASeq，ATAC-Seq， 和蛋白质，并确定了先天免疫缺陷病毒中表观基因组景观的变化。 基于单细胞研究中鉴定的表型的免疫细胞。初步数据和建议 实验使我们能够确定决定先天免疫系统对糖尿病反应的关键步骤， 导致延迟GE发展的细胞。所获得的新知识对于直接瞄准 单核巨噬细胞系统对糖尿病胃轻瘫的预测、预防和治疗。

PROJECT SUMMARY/ABSTRACT Diabetes mellitus is associated with multiple complications including diabetic gastroparesis. The systemic effects of diabetes modify all cells in the body but in common with other diabetic complications, gastroparesis does not develop in all patients. Delayed gastric emptying (GE) in the absence of obstruction defines gastroparesis and also occurs in some but not all diabetic mice. These observations suggest that together with genetics, convergence of systemic changes with local factors is necessary to develop delayed GE in diabetes. We previously have found that delayed GE that is associated with loss of interstitial cells of Cajal (ICC), which in turn is associated with fewer CD206+ muscularis propria macrophages (MPMs). MPMs play a central role in modifying the cellular content and biological environment of the GI muscularis propria in health and disease. The source of gastric MPMs and the mechanisms for activation of injurious molecules in MPMs by diabetes are not known. The overall hypothesis of this proposal is that the underlying central determinant of delayed gastric emptying in diabetes is the combination of a programmed activation response evoked in monocytes and the tissue environment of new monocyte-derived MPMs in the gastric muscularis propria. This will be tested in three specific aims (SA). SA1 will determine the effect of diabetes on circulating monocytes that primes them for infiltration and activation in gastric muscularis propria. SA2 will determine the origin of activated MPMs that populate the gastric muscularis propria following onset of diabetes and SA3 will determine how newly established and activated MPMs cause cellular injury in the diabetic stomach. The SAs are supported by extensive preliminary data. 1) We have found that systemic exposure to diabetes modifies the epigenomic landscape of circulating monocytes so that monocytes are poised to express a restricted complement of signaling pathways when they enter the tissue and become MPMs; 2) MPMs in the diabetic stomach are derived from infiltrating monocytes; 3) Diabetes results in the presence of new, distinct, discrete types of MPMs that are unique to the tissue; 4) Lineage-tagged, long-lived MPMs are not changed by diabetes; 5) Disruption of cellular metabolism in the gastric muscularis propria by diabetes activates monocyte-derived MPMs in close apposition to ICC to produce injurious cytokines, which cause damage to ICC and disrupted gastric function. To address the overall hypothesis we have developed techniques to isolate identified MPMs and monocytes, applied highly sophisticated technology for profiling genes by single cell RNASeq, ATAC-Seq, and proteins by mass cytometry (CyTOF), and determined changes in the epigenomic landscape in innate immune cells based on phenotypes identified in single cell studies. The preliminary data and the proposed experiments position us to identify the key steps that determine the responses to diabetes in innate immune cells that result in development of delayed GE. The new knowledge gained will be crucial to direct targeting of the mononuclear phagocyte system for prediction, prevention and treatment of diabetic gastroparesis.