A microwell cell culture platform for the aggregation of pancreatic β-cells.
Cell-cell contact between pancreatic β-cells is important for maintaining survival and normal insulin secretion. Various techniques have been developed to promote cell-cell contact between β-cells, but a simple yet robust method that affords precise control over three-dimensional (3D) β-cell cluster size has not been demonstrated. To address this need, we developed a poly(ethylene glycol) (PEG) hydrogel microwell platform using photolithography. This microwell cell-culture platform promotes the formation of 3D β-cell aggregates of defined sizes from 25 to 210 μm in diameter. Using this platform, mouse insulinoma 6 (MIN6) β-cells formed aggregates with cell-cell adherin junctions. These naturally formed cell aggregates with controllable sizes can be removed from the microwells for macroencapsulation, implantation, or other biological assays. When removed and subsequently encapsulated in PEG hydrogels, the aggregated cell clusters demonstrated improved cellular viability (>90%) over 7 days in culture, while the β-cells encapsulated as single cells maintained only 20% viability. Aggregated MIN6 cells also exhibited more than fourfold higher insulin secretion in response to a glucose challenge compared with encapsulated single β-cells. Further, the cell aggregates stained positively for E-cadherin, indicative of the formation of cell junctions. Using this hydrogel microwell cell-culture method, viable and functional β-cell aggregates of specific sizes were created, providing a platform from which other biologically relevant questions may be answered.
Department of Chemical and Biological Engineering, University of Colorado, Boulder, CO 80309, USA.
Abigail B
AB
Bernard
Chien-Chi
CC
Lin
Kristi S
KS
Anseth
D000818:Animals; D015820:Cadherins / Q000378:metabolism; D002450:Cell Communication; D018929:Cell Culture Techniques / Q000379:methods*; D002470:Cell Survival; D003432:Cross-Linking Reagents / Q000737:chemistry; D020100:Hydrogels / Q000737:chemistry; D007328:Insulin / Q000378:metabolism; D050417:Insulin-Secreting Cells / Q000166:cytology*; D051379:Mice; D018613:Microscopy, Confocal / Q000379:methods; D008856:Microscopy, Fluorescence / Q000379:methods; D011092:Polyethylene Glycols / Q000737:chemistry; D011108:Polymers / Q000737:chemistry
D015820:Cadherins; D003432:Cross-Linking Reagents; D020100:Hydrogels; D007328:Insulin; D011108:Polymers; D011092:Polyethylene Glycols