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Spheroid Fabrication Using Concave Microwells Enhances the Differentiation Efficacy and Function of Insulin-Producing Cells via Cytoskeletal Changes.

ABSTRACT: Pancreatic islet transplantation is the fundamental treatment for insulin-dependent diabetes; however, donor shortage is a major hurdle in its use as a standard treatment. Accordingly, differentiated insulin-producing cells (DIPCs) are being developed as a new islet source. Differentiation efficiency could be enhanced if the spheroid structure of the natural islets could be recapitulated. Here, we fabricated DIPC spheroids using concave microwells, which enabled large-scale production of spheroids of the desired size. We prepared DIPCs from human liver cells by trans-differentiation using transcription factor gene transduction. Islet-related gene expression and insulin secretion levels were higher in spheroids compared to those in single-cell DIPCs, whereas actin-myosin interactions significantly decreased. We verified actin-myosin-dependent insulin expression in single-cell DIPCs by using actin-myosin interaction inhibitors. Upon transplanting cells into the kidney capsule of diabetic mouse, blood glucose levels decreased to 200 mg/dL in spheroid-transplanted mice but not in single cell-transplanted mice. Spheroid-transplanted mice showed high engraftment efficiency in in vivo fluorescence imaging. These results demonstrated that spheroids fabricated using concave microwells enhanced the engraftment and functions of DIPCs via actin-myosin-mediated cytoskeletal changes. Our strategy potentially extends the clinical application of DIPCs for improved differentiation, glycemic control, and transplantation efficiency of islets.

SUBMITTER: Lee YN

PROVIDER: S-EPMC7768489 | biostudies-literature | 2020 Nov

REPOSITORIES: biostudies-literature

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Spheroid Fabrication Using Concave Microwells Enhances the Differentiation Efficacy and Function of Insulin-Producing Cells via Cytoskeletal Changes.

Lee Yu Na YN Yi Hye Jin HJ Goh Hanse H Park Ji Yoon JY Ferber Sarah S Shim In Kyong IK Kim Song Cheol SC

Cells 20201127 12

Pancreatic islet transplantation is the fundamental treatment for insulin-dependent diabetes; however, donor shortage is a major hurdle in its use as a standard treatment. Accordingly, differentiated insulin-producing cells (DIPCs) are being developed as a new islet source. Differentiation efficiency could be enhanced if the spheroid structure of the natural islets could be recapitulated. Here, we fabricated DIPC spheroids using concave microwells, which enabled large-scale production of spheroi ...[more]

PMID: 33261076

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Project description:Cells with various structures and proteins naturally come together to cooperate in vivo. This study used cell spheroids cultured in agarose micro-wells as a 3D model to study the movement of cells or spheroids toward other spheroids. The formation dynamics of tumor spheroids and the interactions of two batches of cells in the agarose micro-wells were studied. The results showed that a concave bottom micro-well (diameter: 2 mm, depth: 2 mm) prepared from 3% agarose could be used to study the interaction of two batches of cells. The initial tumor cell numbers from 5 × 103 cells/well to 6 × 104 cells/well all could form 3D spheroids after 3 days of incubation. Adding the second batch of DU 145 cells to the existing DU 145 spheroid resulted in the formation of satellite cell spheroids around the existing parental tumor spheroid. Complete fusion of two generation cell spheroids was observed when the parental spheroids were formed from 1 × 104 and 2 × 104 cells, and the second batch of cells was 5 × 103 per well. A higher amount of the second batch of cells (1 × 104 cell/well) led to the formation of independent satellite spheroids after 48 h of co-culture, suggesting the behavior of the second batch of cells towards existing parental spheroids depended on various factors, such as the volume of the parental spheroids and the number of the second batch cells. The interactions between the tumor spheroids and Human Umbilical Vein Endothelial Cells (HUVECs) were modeled on concave agarose micro-wells. The HUVECs (3 × 103 cell/well) were observed to gather around the parental tumor spheroids formed from 1 × 104, 2 × 104, and 3 × 104 cells per well rather than aggregate on their own to form HUVEC spheroids. This study highlights the importance of analyzing the biological properties of cells before designing experimental procedures for the sequential fusion of cell spheroids. The study further emphasizes the significant roles that cell density and the volume of the spheroids play in determining the location and movement of cells.

Dataset Information

Spheroid Fabrication Using Concave Microwells Enhances the Differentiation Efficacy and Function of Insulin-Producing Cells via Cytoskeletal Changes.

Publications

Spheroid Fabrication Using Concave Microwells Enhances the Differentiation Efficacy and Function of Insulin-Producing Cells via Cytoskeletal Changes.

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