Given that in Aab+ donors alpha cells become dysfunctional before donor’s blood glucose levels are impaired, it is important to differentiate whether a) alpha cells become dysfunctional first, which attracts and activates immune cells, or b) vice versa. In pre–T1D the first phase insulin secretion is dampened. We and others have shown that this phase is triggered by “first–responder” beta cells – a stable beta cell subpopulation (same cell or it’s neighbor remains 1st responder for 24–to–48h with frequent restimulation). We also found that majority of the 1st responders are alpha–cell–neighboring cells. Putting everything together, our question is whether alpha cells are targeted in T1D, which leads to the death/dysfunction of the neighboring beta cells (1st responders) first, and is manifested via loss of the first phase peak of GSIS in pre–T1D patients? This question is not currently pursued by the mainstream research direction, but if answered, could change our view of T1D pathogenesis and suggest novel targets for therapy.
We aim to study heterogeneity of alpha and beta cell function in live pancreatic tissue slices. For this we will be using advanced NADH and Ca²⁺ imaging techniques. We hypothesize that, in early–stage T1D, some alpha–and their neighboring beta cells will show impaired metabolism and Ca²⁺ response to nutrients, compared to non–diabetic (ND) tissue.
We will test how neoantigen–specific T–cell clones (isolated & provided by James Lab) interact with alpha– and neighboring beta cells in live tissue slices in recent–onset T1D donors. We expect that, compared to rested (naive) clones, the pre–activated T–cell clones will behave differently: with altered motility profiles, proliferation, and frequency of contact with alpha cells and their neighboring beta cells. Furthermore, we will manipulate alpha–T cell interactions using functional grade antibodies that block HLA presentation.
For additional insights into the mechanism (add–value experiment), we will develop a staining protocol to use fluorescently labeled HLA tetramers complexed with known immunogenic peptides that are expressed in alpha cells (designed & provided by James Lab) to tag T cells in T1D tissue. Using AI–assisted tools and 3D confocal microscopy, we will analyze alpha–T–cell immunologic synapse formation and networks to understand how T–cell specificity affects alpha–T cell interactions.
