Manufacturing of insulin-secreting spheroids with the RIN-5F cell line using a shaking culture method

Abstract

BACKGROUND: There have been many efforts to find methods to increase insulin production by islets or modified cells. Commercially available established cell lines can be a good source of artificial islets. We manufactured sphere-shaped cell clusters composed of insulin-secreting cells from the commercially available RIN-5F cell line.

METHODS: To generate artificial islets with insulin-secretion functions, we used the RIN-5F cell line. When cells cultured in RPMI-1640 medium containing 10% fetal bovine serum reached near confluency, they were trypsinized for suspension culture at high density, using a horizontal shaker. The cells were maintained for 5 days under 5% CO(2) with humidification. Next, the media from the RIN cell spheroid culture was collected over 5 consecutive days to test for insulin secretion.

RESULTS: Spheroids of artificial islets exhibited an oval shape with an approximate size of 94.13 ± 20.41 μm on day 5 during the shaking culture. Abnormal outgrowth of spheroids was not observed. Terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling-positive cells were not detected among the overall spheroids, including the core position. Insulin secretion, measured by enzyme-linked immunosorbent assay, was well maintained in the culture media over 5 days after spheroid formation.

CONCLUSION: This result suggested that a culture method with shaking can be applied to commercially available established cell lines to generate artificial islets, which might be used for a bioartificial pancreas