Bioprocess development for the cell-based treatment of diabetes

• Main Author: Hoesli, Corinne
• Language: English
• Published: University of British Columbia 2010
• Online Access: http://hdl.handle.net/2429/30255

The widespread cellular treatment of type 1 diabetes by islet transplantation is limited by tissue shortage and graft rejection. This work describes two novel bioprocesses to immobilize pancreatic cells in alginate using: (1) hollow fiber bioreactors or (2) alginate bead generation by an adapted emulsion and internal gelation process. After optimization, live cell recovery rates and growth rates were not significantly different between these more scalable processes and conventional methods of alginate immobilization. After 10 days of alginate-immobilized culture, the insulin content of neonatal porcine cells cultured in a hollow fiber bioreactor increased by 4 fold, while the insulin expression of human isletdepleted tissue cultured in emulsion beads increased by 67 ± 32 fold, matching previous reports that used small-scale cultures. Solutions with >100 Pa·s viscosity could be used with the emulsion process to generate beads with higher concentration and greater antibody exclusion than has been so far permitted by nozzle-based encapsulators. The 5% alginate beads generated by the emulsion process led to blood glucose normalization of allogeneic β- cells transplanted into diabetic mice within 2 weeks, while mice transplanted with 1.5% alginate beads generated by a conventional encapsulator remained hyperglycemic after 20 days. The improved result with the 5% alginate emulsion beads was associated with lower graft-specific antibody plasma levels. These results suggest that the 5% alginate beads provided improved immune isolation of the graft. If human pancreatic progenitors are to be used for the large-scale generation of insulin+ cells in alginate, their expansion in serum-free medium will be a prerequisite. Pancreatic duct-like cells are expected to have more potential to generate insulin+ cells than the fibroblast-like cells that overgrow unsorted cultures of islet-depleted human pancreatic tissue. The last part of this thesis describes the magneticactivated depletion of CD90-expressing cells, which reduced the fraction of CD90+ fibroblast-like cells from 34 ± 20% to 1.3 ± 0.6%. This allowed the expansion of the duct-like cell population in an optimized serum-free medium. These novel pancreatic cell culture methods could be used to generate and/or offer immune protection to insulin+ cells for the clinical-scale cellular treatment of diabetes.