1736-P: Exploring Intrinsic Features of Human Pancreatic a- and ß-Cell Physiology Using Pseudoislets

Introduction: Mechanistic studies of pancreatic α- and β-cell physiology are challenging in whole islets due to interactions among different islet cell types. Reaggregation of sorted cell populations into pseudoislets offers the possibility to delineate the role of specific cell types, distinguish intrinsic cellular functions and paracrine interactions, and enhance accessibility for genetic interventions.

Methods: Building on methods of Walker et al, 2020, we generated >90% enriched human α- and β-cell aggregates and studied hormone secretion by perifusion, mitochondrial function by Seahorse, Ca²⁺ flux by fluorescence imaging, and electrophysiology by whole-cell patch-clamp. Native islets and pseudoislets of dissociated unsorted islet cells served as controls.

Results: The native islet kinetics of insulin and glucagon responses to glucose were preserved in β- and α-cell aggregates, indicating direct effects of glucose; however, rates of secretion, adjusted for IEQ and cell proportion, were markedly lower compared to native islets and reaggregated controls, suggesting that optimal α- and β-cell function depends on other islet cell types. The primary stimulus of oxygen consumption rate (OCR) and Ca²⁺ flux in β- and α-cell aggregates was glucose and amino acids, respectively. Baseline OCR (no substrate) was higher in α- than β-cell aggregates. Electrophysiologically, both native islet and α-cell aggregate action potentials (APs) were larger in size and faster in kinetics than β-cell APs; however, high glucose-induced inhibition of α-cell APs and subsequent hyperpolarization observed in native islets and control reaggregates was noticeably impaired in α-cell aggregates, suggesting a role for other cell types in glucose-mediated inhibition of α-cells.

Conclusion: Cardinal features of α- and β-cell physiology are largely preserved in α- and β-cell enriched pseudoislets, establishing its feasibility in the study of intrinsic α- and β-cell physiology.