Pathophysiology of Diabetic Gastroparesis

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.