Biopolymers for Bioartificial Pancreas

The concept of the bioartificial pancreas has developed substantially in the last decade, after the first successful islet transplantation in 2000. Islets are clusters of endocrine cells found in the pancreas. Amongst these cells, the insulin secreting beta cells are targeted by the immune system in Type 1 Diabetes. One of the major challenges islet transplantation has faced is the rapid loss of islets, post-transplantation. To mitigate this, a bioartificial pancreas that provides support and immuno-isolation for islets before transplantation has been investigated.
Polyhydroxyalkanoates (PHAs) are intracellular energy storage polymers synthesized by Gram-positive and Gram-negative bacteria. Because of their high degree of biocompatibility and structural properties, PHAs have been chosen as the scaffold material for the bioartificial pancreas. Alginate is a naturally occurring polysaccharide made up of mannuoronate and guluronate units. Because of its ability to mimic the extracellular matrix, it has been widely investigated for its use as a hydrogel and is being investigated for its use in islet transplantation.
This project aimed to produce a bioartificial pancreas that contains pancreatic cells encapsulated in an alginate hydrogel environment (mimicking islets) and a PHA scaffold providing support and an additional layer of immuno-isolation.
To identify a scaffold material, two PHAs were produced- P(3HB), a stiff, brittle polymer by Bacillus subtilis OK2 fermentation and a novel

elastomeric P(3HO-co-HD) by Pseudomonas mendocina CH50. Several studies were performed on these produced polymers. Initially, they were characterised chemically, structurally, mechanically and thermally and compared to PLLA, a FDA approved polymer. It was observed that when these polymers were compared in terms of viability and insulin release, cells seeded in P(3HO-co-HD) performed best. Porosity was introduced into the P(3HO-co-HD) scaffold to mimic the mechanical properties of the native pancreas and facilitate exchange of nutrients and waste. The effect of the type of porogen, porogen size and concentration was also investigated. It was observed that the scaffold obtained using 100μm NaCl, at a concentration of 15% w/v was the best scaffold for the BRIN BD11cells. Next, the polymers were fabricated into 2D & 3D structures and evaluated for function. No statistical difference was observed when the mechanical properties of both 2D & 3D structures were compared. The same trend was observed for the viabilities and insulin release of BRIN BD11 cells seeded in these structures.
In addition, three P(3HO-co-HD) dominated P(3HO-co-HD)/P(3HB) blends: 95:5, 90:10 & 80:20 were made in order to improve the handling of P(3HO-co-HD). When the handling of the scaffolds, the viability and insulin release of BRIN BD11 cells seeded in these blends were considered, the 95:5 P(3HO-co-HD)/P(3HB) blend was selected as the best combination. Further, the 95:5 P(3HO-co-HD)/P(3HB) blend was processed into 2D & 3D structures, evaluated and compared to 2D & 3D P(3HO-co-HD) structures. No significant difference was observed when the

95:5 P(3HO-co-HD)/P(3HB) blend and P(3HO-co-HD) structures were compared based on cell viability and insulin release.
Finally, two kinds of alginate hydrogel structures were made- alginate microbeads and 3D printed block hydrogels. The effect of cell densities (1X105cells/ml & 5X105cells/ml) and alginate concentration (2%, 4% & 5% w/v) on cell viability was evaluated. The two structures were then compared based on cell viability and insulin release. Of all the conditions, 5% w/v alginate 3D printed block hydrogels containing 5x105cells/ml performed best based on cell viability and insulin release of BRIN BD11 cells seeded.
In conclusion, in this work, two PHAs including a novel PHA have been successfully produced. In addition, the best blend structure for scaffold creation has been identified and evaluated. Further, alginate hydrogels with optimum cell densities and structures have been selected and used for the creation of the inner hydrogel environment. These represent the framework for the successful development of bioartificial pancreas in the future.