A method and
system are disclosed for differentiating non-human
primate induced pluripotent stem cells (NHP iPSCs) into
hepatocyte-like cells through a defined, multi-stage protocol. The method includes sequentially exposing NHP iPSCs to six distinct
cell differentiation media over an approximate eighteen-day period. Each medium comprises defined combinations of growth factors, small molecules, and inhibitors that direct progressive lineage specification through anterior primitive streak,
definitive endoderm, posterior foregut, liver
bud progenitors, hepatoblasts, and
hepatocyte-like cells. Concentration ranges and
exposure durations are provided for each stage, ensuring reproducibility and reducing heterogeneity across
cell populations. In some embodiments, treatment with a
transforming growth factor beta (TGF-β )
receptor I inhibitor during early hepatoblast specification enhances lineage fidelity. The resulting
hepatocyte-like cells are validated by
immunostaining and western blotting for hepatic markers. Applications include preclinical
drug metabolism, hepatotoxicity assessment, and
in vitro disease modeling using non-human
primate–derived systems.