The overarching goal of this project is to determine the role of angiotensin converting enzyme 2 (ACE2) in the diabetic gut, how it impacts hyperglycemia and glycemic variability, and thus contributes to the pathogenesis of diabetic retinopathy (DR). The protective arm of the renin angiotensin system (RAS) consists of ACE2, which converts angiotensin II (Ang-II) to angiotensin 1-7 (Ang-1-7). Ang-1-7 opposes the effects of Ang-II by virtue of its actions on the MAS receptor. While the systemic (endocrine) RAS works with local (tissue) RAS such as that in the eye and gut to achieve homeostasis in health, in diabetes loss of key components of the protective RAS can lead to widespread pathology. The literature and our preliminary data support that diabetes results in loss of expression of ACE2 in the gut, bone marrow, and retina. Glycemic variability is implicated in DR pathogenesis. Intestinal ACE2 can regulate glucose homeostasis by modulating tryptophan absorption and incretin release and by generating Ang 1- 7 from luminal Ang II. Ang 1-7 by binding to Mas receptor can block glucose transport in the gut similar to what has been described in the pancreas. Based on this, we hypothesis: In T2D, loss of enterocyte ACE2 decreases: i) tryptophan uptake and incretin secretion leading to hyperglycemia; ii) MAS receptor activation increasing gut glucose absorption; and iii) gut barrier integrity resulting in leakage of gut microbial peptides into the circulation. All three mechanisms increase retinal permeability and activating immune cells promoting DR pathology. Aim 1 will test if dysregulation of ACE2 in the gut epithelium results in i) interruption of tryptophan transport by B0AT1 decreasing incretin secretion and ii) reduced MAS receptor activation leading to increased glucose absorption in the gut. Aim 2 will test if in db/db mice, loss of intestinal ACE2 will result in increasing circulating levels of gut microbial peptides that will activate TLRs on retinal endothelial cells and lead to increased retinal leukostasis and blood retinal barrier dysfunction. Aim 3 will examine if nutraceuticals or probiotics can restore the balance of the intestinal RAS (ACE2/Ang-1-7/MAS) in db/db mice to prevent development of DR. Impact: We propose a novel mechanism for deterioration of glucose homeostasis and increased glucose variability in diabetes- the loss of function of the ACE2:B0AT1 oligomer form of ACE2 (unique to the intestinal epithelium) and reduced levels of intestinal Ang 1-7 resulting in less intestinal MAS receptor activation. The dysregulated intestinal RAS can lead to serious retinal pathology promoting DR.