Investigating the Effects of 2-Aminoadipic Acid on Beta-Cell and Human Islet Function

• Author / Creator

  Hubert, Matthew

• 2-aminoadipic acid (2-AAA) has been identified as an accurate long-term biomarker for the future development of diabetes. An investigation of over 2000 patients found that patients in the upper quartile of initial blood 2-AAA levels showed a 2-4x higher incidence of diabetes development over the next 12-13 years. Mouse models and static mouse and human islet cultures both suggest that 2-AAA increases insulin release at low glucose. Additionally, previous 2-AAA studies indicate 2-AAA's toxicity towards glial cells is the result of inhibited glutathione production. This toxicity inducing inhibition could also be taking place in β-cells of the pancreatic islet, as the affected transporters and enzymes are present and required in both cell types. Decreases in glutathione production in β-cells could lead to increased intracellular reactive oxygen species (ROS) levels. This could increase hydrogen peroxide (H2O2) mediated insulin secretion, accounting for the increased basal insulin secretion observed in mice. Additionally, bacterial salt stress pathways are linked with lysine breakdown pathways which can produce 2-AAA. 2-AAA production in the body may decrease if the population of such bacteria in the gut decreases. To investigate if 2-AAA may be involved in the development of diabetes I performed the following experiments. The rat β-cell line INS-1 and human islets were treated with buffer containing 2-AAA along with H2O2 and/or 20mM glucose. Necrosis, H2O2 production, ROS and glutathione levels were measured in different experiments after treatment. Isolated human islets were cultured overnight, treated with 2.5mM glucose buffer containing 2-AAA for three hours, and then perifused at 2.5mM glucose and then 11.1mM glucose. Three-hours of static insulin secretion and one-hour of dynamic insulin secretion was then measured. Cystine and glutamate treatment was included in the buffer of many experiments to mirror previous 2-AAA studies in glial cells. Serum 2-AAA levels of germ-free mice were also measured and compared to wild-type mice. In summary, treating INS-1 cells with 2-AAA shows significant increases in necrosis, H2O2 production, and ROS levels, as well as a decrease in total glutathione levels. Treating human islets with 2-AAA shows significant decreases in total glutathione levels with no significant change in H2O2 production. Our data also suggests that 2-AAA does not modify insulin secretion from human islets at low 2.5mM or high 11.1mM glucose. A significant negative correlation was found between 2-AAA induced three-hour low glucose insulin secretion and the age of the male donors of those islets. No change in serum 2-AAA levels between germ-free and wild-type mice was observed. Similar toxic effects of 2-AAA as previously seen in glial cells were observed in the INS-1 β-cell line. Only some of 2-AAA’s toxic effects were observed in human islets. This may be due to the different cellular composition or oxidative resilience of human islets, or may be due to an insufficient treatment time of human islets due to limited culture viability of the explanted tissue. The lack of increased H2O2 production with 2-AAA treatment in human islets may account for a lack of insulin secretion increase with 2-AAA treatment. While germ-free mice did not have a lower 2-AAA level, it is still possible that modification of salt/lysine in diets may result in modifications of 2-AAA production from the gut microbiome. It is also possible that myeloperoxidase mediated 2-AAA production during inflammation may lead to modified blood 2-AAA levels. This research provides a more complete understanding of 2-AAA’s mechanism, suggesting a causative role for 2-AAA in the etiology of diabetes and provides support to utilize 2-AAA as a predictive biomarker to treat non-diabetic patients with high blood 2-AAA levels to prevent them from developing diabetes.

• Subjects / Keywords

  • 2-AAA
  • Diabetes
  • Islet

• Graduation date

  Fall 2020

• Type of Item

  Thesis

• Degree

  Master of Science

• DOI

  https://doi.org/10.7939/r3-x1e6-qx71

• License

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