Microencapsulation of embryonic stem cells for the generation of hepatocytes

The emergence of hepatocyte based clinical and pharmaceutical technologies, has been limited by the absence of a stable hepatocyte cell source. Embryonic stem cells may represent a potential solution to this cell source limitation problem since they are highly proliferative, renewable and pluripotent. Although many investigators have described techniques to effectively differentiate stem cells into a variety of mature cell lineages, their practicality is limited by: (1) low yields of fully differentiated cells, (2) absence of large scale processing considerations and (3) ineffective downstream enrichment protocols. Within this thesis we have sought to assess the feasibility of directing stem cell differentiation via microenvironment regulation, through the development of a murine embryonic stem cell (ES) alginate poly-L-lysine microencapsulation hepatocyte differentiation system. Our results indicate that the alginate microenvironment maintains cell viability, is conducive to ES cell differentiation, and maintains differentiated cellular function. In addition, we demonstrate that cellular aggregation is integral to the control of differentiation within the bead environment and this process is mediated by the E-cadherin protein. The temporal expression of surface E-cadherin and hepatocyte functional expression occur concomitantly and both cellular aggregation and albumin synthesis are blocked in the presence of anti E-cadherin immunoglobulin. Furthermore, by establishing a compartmental model of differentiation, which incorporates this aggregation phenomenon, we can optimize key encapsulation parameters, and predict hepatic function in various encapsulation regimes. As the final step in establishing a renewable cell source, we have also begun translational research initiatives, in order to move our resultant differentiated cell product "from bench to bedside".