Magnetic Bead-Guided Assembly of 3D Primary Human Islet Cells in Decellularized Pancreatic Scaffolds

Background: Three-dimensional (3D) cell cultures are increasingly recognized as effective models for studying diseases and developing cell therapies. In the endocrine pancreas field, organoids/spheroids derived from human islet cells enable advances in diabetes research, drug screening, and tissue engineering. While various 3D culture methods exist, approaches such as magnetic bead-assisted aggregation remain underexplored for endocrine pancreatic cells. Additionally, the use of biological scaffolds, especially those derived from decellularized pancreatic extracellular matrix, provides a biomimetic environment that promotes adhesion, proliferation, and functionality of pancreatic cells. This study presents a protocol for magnetic bead-guided 3D culture of human islet cells within decellularized pancreatic scaffolds. Methods: Human pancreas from adult brain-dead donors was harvested for both islets' isolation processing and decellularization to generate an acellular pancreatic bioscaffold. Primary human pancreatic islets were first grown in two-dimensional adherent cultures, then enzymatically harvested from the surface and reassembled into three-dimensional clusters using different initial cell amounts (small clusters 0.5 × 10⁴-1 × 10⁴ and larger clusters 2.5 × 10⁴-5 × 10⁴ cells) and then placed within acellular pancreatic slices of different thickness, namely 50 and 90 μm. Optic microscopic examination, scanning electron microscopy analysis, and assessment of insulin and lactate dehydrogenase (LDH) levels were used to evaluate these 3D islet-like cluster cultures. Results: We report the establishment of 3D cultures derived from primary pancreatic islet cells using a magnetic approach in a remarkable 18 h period for the complete formation of 3D clusters. The small clusters (0.5 × 10⁴-1 × 10⁴ cells) exhibited a faster attachment to the acellular matrix, with cells visibly spreading outside the cluster interacting with the bioscaffold slice, when compared to the larger clusters (2.5 × 10⁴-5 × 10⁴ cells). These cells continued to produce insulin, and no statistically significant differences in LDH levels were found under these different conditions. Conclusions: Here, we demonstrate that a magnetic bead-based protocol can be successfully applied to endocrine pancreatic cells, enabling the rapid formation of compact, viable, and functional 3D structures. Despite limitations such as higher cost and prolonged retention of magnetic particles, the approach supports size-dependent interactions with decellularized pancreatic scaffolds. These findings are valuable for researchers designing experiments tailored to specific objectives and underscore the potential of this platform for advancing diabetes research and pancreatic tissue engineering.