Longitudinal single cell responses in human islet organoids to beta cell regenerative drugs - Summary All forms of diabetes result from insufficient numbers of insulin-producing beta cells. Thus, beta cell regenerative drug therapies could provide a scalable and affordable approach for millions of people with diabetes. The most advanced beta cell regenerative therapies include inhibitors of the kinase DYRK1A, exemplified by harmine, alone, or with GLP1 receptor agonists (GLPRA’s). However, the complete mechanisms of action for DYRK1A inhibitors and GLP1RA’s are incompletely understood. Therefore, to further optimize their efficacy, and to more deeply understand the mechanisms of action of beta cell regenerative therapies, a deeper cellular and molecular analysis of regenerative drug-treated human islets is needed. My Preliminary Data from single studies on regenerative drug-treated human islets suggest that “cycling alpha cells” serve as a beta cell progenitor reservoir in response to DYRK1A inhibitors such as harmine. However, most of our studies and reports from others have focused on beta cell proliferation at one single time point. As it is clear that islet cell subpopulations may progress through different phenotypes and proliferation phases over time following drug exposure, the mechanisms through which beta cell expansion occurs over time with regenerative drug treatment remain largely unexplored. Therefore, in this application I will address this knowledge gap by utilizing a human islet-derived organoid (islet MT) system that allows for long-term culturing and drug treatment. This will allow accurate study of lineage trajectory dynamics through collection of data at multiple time points. More specifically, in Aim 1 of this project, I will perform single cell and single nucleus RNAseq as well as single nuclear ATACseq, to explore temporal evolution of cellular mechanisms through which harmine enhances human beta cell regeneration in vitro. These experiments will also provide deeper insights into the gene regulatory networks that regulate transcriptional as well as epigenetic control. Since MT organoids are not identical to normal human islets, it is critical to compare data from Aim 1 with my existing single cell transcriptomic profiles of drug-treated human islets. Therefore, In Aim 2, I will integrate the single cell transcriptomic and accessible chromatin profiles of the human islet organoids from Aim 1 and compare and contrast these data with the single cell transcriptomic profile of human islets. Collectively, these analyses will be instrumental to pinpoint the temporal control of the fate decisions of cycling alpha cells and elucidate other possible islet cell types with the potential to transdifferentiate into beta cell.
