Studying the early pathophysiology of pancreatic islet dysfunction is key to preventing or reversing type 2 diabetes (T2D). Long before T2D is diagnosed, islets become hyperactive, causing systemic hyperinsulinemia. Pulsatile insulin secretion also becomes disordered, leading to the loss of efficient insulin secretion from the islets and reduced insulin signaling and clearance at the liver. A constantly high demand for insulin can eventually lead to exhaustion and failure of the islets. Preventing these early maladaptations to nutrient excess in islets may prevent the progression of obesity to T2D.
To study islet and hepatocyte dysfunction in the context of T2D, innovative in vitro technologies are needed to overcome limitations in current models. Therefore, we created a series of microfluidic chips to investigate the role of pulsatile insulin secretion in islet and hepatocyte function. A syringe pump system was used to force islets to oscillate with alternating stimulatory and inhibitory solutions. Murine islets were treated chronically with high glucose to cause dysfunction followed by forced overnight oscillation treatment or a constant reduction in glycolytic activity with the glucokinase inhibitor mannoheptulose. Glucose-stimulated calcium recordings revealed that islets forced to oscillate had a greater recovery of function compared to islets with constantly reduced glycolytic activity, indicating a role for oscillations in maintaining beta-cell function. Using chips designed to hold hepatocytes or both islets and hepatocytes, we determined that pulsatile insulin delivery to hepatocytes improves nutrient storage compared to continuous insulin delivery.
Another new in vitro model has emerged from the advancements in stem-cell-derived islets (SC-islets) that facilitate the study of beta cells that are relevant to human disorders. However, SC-islets are still functionally immature and can benefit from new techniques that augment their insulin secretion. Using tissue culture plates containing 200 µM by 200 µM square microwells, we created SC-islet microaggregates that secreted significantly more insulin than standard sized aggregates. Automated imaging allowed us to normalize insulin secretion to aggregate size, improving the throughput of functional assays. Combining the enhanced functionality of microaggregates with the improved normalization procedure can expand the utility of SC-islets as a research model.
The earliest detectible changes in islets during the progression of T2D are an increase in insulin secretion and an impairment in beta-cell metabolism. Reducing islet activity back to a normal level in the face of hyperglycemia can potentially prevent these adaptations from progressing to the point of beta-cell failure. We determined that pancreatic islet adaptations to hyperglycemia are prevented by glycolytic inhibition and amplified by inhibiting membrane depolarization. Mathematical modeling of beta-cell activity revealed that reducing intracellular calcium levels causes a buildup of glycolytic metabolites that may initiate a signaling pathway responsible for islet adaptation to hyperglycemia. Taken together, the outlined in vitro tools and studies move toward a better understanding of the islet dysfunction that leads to T2D.