Pancreatic islet cell reaggregation systems: Efficiency of cell reassociation and endocrine cell topography of rat islet-like aggregates

Single cells isolated from rat islets of Langerhans were cultured under conditions that support reassociation into islet-like aggregates. Comparisons were made of enzymatic methods of islet dissociation, rotational or static culture conditions, and culture at basal or stimulatory glucose concentrations. Over a period of 4 days the aggregates progressed through three stages of organization: Cell coalescence to cellular chains, rearrangement of chains into small spheroids, and growth of spheroids. The numerical yield of aggregates was optimum after islets were dissociated with dispase. Culture under rotation resulted in the production of more aggregates of significantly larger diameter than under static conditions. Medium glucose concentrations of 4 and 11 mM supported cell reassociation under rotator culture, but no aggregation occurred under static culture at the basal (4 mM) glucose level. Aggregates resulting from 4-day rotator culture exhibited endocrine cell distributions similar to intact islets. Islet aggregates released insulin in response to glucose, but nonaggregated cells, maintained in culture, did not. The present comparisons reveal significant variability in the cellular composition, rate of formation, and yield of aggregates, and suggest that the methodology for producing aggregates should be carefully considered in experimental design.