MICROENCAPSULATION OF PANCREATIC ISLETS FOR CELL TRANSPLANTATION IN TYPE 1 DIABETES WITHOUT IMMUNOSUPPRESSION.

Type 1 Diabetes mellitus (T1D) is a chronic autoimmune disease caused by the attack of autoreactive T lymphocytes on pancreatic β-cells, leading to absolute insulin deficiency. For patients with T1D, exogenous insulin injections to control blood glucose are a lifesaving treatment. However, exogenous insulin administration does not prevent daily risk of hypoglycemic episodes, and does not guarantee a tight control of blood glucose. Pancreatic islet transplantation through the hepatic portal vein has recently emerged as one of the therapeutic approaches for improving blood glucose control in T1D patients with severe hypoglycemic unawareness. Despite promising results, most patients lose insulin-independence and graft function in variable times after transplantation. Among the causes is early graft loss due to immunological, anatomical, physiological and metabolic limitations of the transplant site. Also, islet transplantation requires life-long systemic immunosuppression (SI) to prevent graft rejection. SI causes lymphopenia and several side effects, included tumor development, frequent infections, and general toxicity. In addition, in order to overcome the shortage of allogeneic islets from cadaveric donors, islets from xenogeneic donors, such as pigs, represent an unlimited source. Islet encapsulation with biocompatible and non-degradable hydrogels may represent a valuable alternative to systemic immunosuppression as it provides a physical barrier that immunoisolates and protects the graft from the cytotoxic attacks of the host immune system. In the past thirty years, alginate encapsulation has been evaluated in pre-clinical and clinical settings. The successful use of alginate microcapsules has, however, been hampered by 1) the large diameters (600 and 1000μm), and 2) the mechanical instability, followed by the lack of immunoprotection. The large capsule size constitutes a diffusion barrier, impairing oxygen and nutrient exchanges, and limits the choice of transplant sites to areas that are not conceived for cell survival. Several alternative sites have been proposed to minimize early inflammatory reactions, promote vascularization and easy-accessibility, mimic physiological insulin release and protect from immune responses. Among all, the omental pouch, or the equivalent epididymal fat pad (EFP) in mice, is well vascularized and can accommodate large volumes. Conventional alginate microencapsulation has been optimized by minimizing capsule size (450-550μm in diameter), increasing the cell loading density (nearly 3%), and by using highly biocompatible Ultra-Pure medium viscosity sodium alginate (UP-MVG). This allowed for transplantation of microencapsulated islets in the EFP, engineered with a novel fibrin matrix to promote angiogenesis, decrease early graft loss, and improving islet engraftment. Under physiological conditions the capsules are also exposed to a combination of destabilizing forces, leading to swelling, increased pore size, dissolution, and capsule rupture. To protect the cells from the host immune system, the capsule must therefore be carefully designed, especially with respect to stability and porosity. Thus, novel alginate-based capsules have been designed with the goal to improve in vivo stability of alginate: 1) hybrid microcapsules (MicroMix) using an electrostatic droplet generator method by mixing UP-MVG with Polyethylene Glycol functionalized with Maleimide groups (Peg-Mal) 2) UP-MVG microcapsules double coated (Double) with Peg-Mal through an emulsification process. One of the main challenges of this part of the work has been to make alginate capsules stable under physiological conditions over extended periods of time. The hope is that the great, and still growing, knowledge about alginate-based capsule biocompatibility, mechanical properties and permselectivity will be useful for successful clinical transplantation.