Implantation of Microencapsulated hPSC-Hep Spheroids

Microencapsulated hPSC-Hep spheroids were washed in PBS and briefly stored in CMRL-1066 serum-free medium prior to implantation. Implantation of alginate encapsulated cells and empty alginate beads was performed as previously described (King et al., 2001 (link); Bochenek et al., 2018 (link)). The intraperitoneal space was selected for the implantation site due to proximity to the liver and easy retrieval of the microbeads by saline lavage upon completion of the experiments. The implantation of microencapsulated hPSC-Hep spheroids was performed on 16 immunocompetent C57BL/6J male mice aged 16 weeks; empty alginate microbeads were transplanted into 8 mice as a control. The microbeads were retrieved from the peritoneal cavity after 10 (N = 12) and 20 (N = 12) days. The intervention (sulfated alginate) and control (unmodified alginate), and no-cell control groups were matched in size (n = 4 each). Images of the microbeads were collected using brightfield imaging by Operetta CLSTM High-Content Analysis System (Perkin Elmer), and the extent of PFO was evaluated manually in ImageJ.