Fibronectin and Collagen IV Microcontact Printing Improves Insulin Secretion by INS1E Cells

Extracellular matrix (ECM) molecules play significant roles in regulating -cell function and viability within pancreatic islets by providing mechanical and biological support, stimulating cell survival, proliferation, and their endocrine function. During clinical islet transplantation, the -cell's ECM environment is degraded by enzymatic digestion. Literature suggests that interactions between islet cells and ECM molecules, such as fibronectin (FN), collagen type IV (Col4), and laminin (LN), are essential for maintaining, or stimulation of islet function and survival, and can effect differentiation and proliferation of the endocrine cells. It is also thought that three-dimensional (3D) culture of -cells can improve glucose responsiveness by providing a specific niche, in which cells can interact with each other in a more natural manner. Conventional suspension cultures with -cells results generally in a heterogeneous population with small and large aggregates, in which cells experience different nutrient diffusion limitations, negatively affecting their physiology and function. In this study, we have explored the effect of FN, Col4, and LN111 on INS1E insulinoma cells by using microcontact printing (CP) to investigate whether a controlled environment and aggregate dimensions would improve their endocrine function. Using this method, we produced a pattern of well-defined circular spots of FN, Col4, and LN111 on polydimethylsiloxane with high spatial resolution. Cell seeding of the INS1E cells on these ECM protein spots resulted in the formation of 3D -cell aggregates. We show that these INS1E aggregates have very reproducible dimensions, and that the cell culture method can be easily adjusted, leading to a highly accurate way of forming 3D -cell aggregates on an ECM-functionalized substrate. In addition, we show that ECM molecules can act as anchoring points for -cells on an otherwise non-cell-adherent material, and this can improve both the endocrine function and viability. We found a significant increase in the secretion of insulin by INS1E cells cultured on CP FN and Col4 substrates, in comparison to cells that were growing in monolayers on substrates without ECM molecules. Moreover, INS1E cells growing on circular ECM spots in a 3D manner showed improved endocrine function in comparison to their two-dimensional counterparts.

Impact Statement This research deals with finding a proper bioengineering strategy for the creation of improved -cell replacement therapy in type 1 diabetes. It specifically deals with the microenvironment of -cells and its relationship to their endocrine function.