Engineering hepatic and embryonic stem cell differentiation through the cell-cell adhesion molecule, E-cadherin.

摘要

The challenge in promoting hepatic differentiation lies in strategically engineering molecular signals that influence hepatic phenotype to ultimately drive functional response. This research investigated novel cell-biomaterial and cell-based signals to induce/maintain differentiated function in two model cells: primary adult rat hepatocytes and murine embryonic stem (ES) cells. The molecular signal was the cell-cell adhesion ligand, epithelial cadherin (E-cadherin), which is instrumental in liver tissue development.; Substrate-presented acellular E-cadherin fragments were engineered to mimic native cell-cell interactions. Our results revealed that hepatocytes successfully recognized the E-cadherin fragments which significantly altered cell morphology at increased E-cadherin concentration; moreover, albumin and urea secretion, liver-specific markers, were elevated. Thus, acellular E-cadherin incorporation in substrates may potentially prevent hepatocyte dedifferentiation by maintaining liver-specific function.; Next, the synergistic role of endogenous E-cadherins in murine ES cells and hepatotrophic growth factors were examined to elucidate their hepatic differentiating ability. Cadherin-expressing (CE-ES) cells were engineered to overexpress endogenous E-cadherins while cadherin-deficient (CD-ES) cells served as control cells (Dr. Larue, France). CE-ES cells intrinsically formed cell aggregates, whereas CD-ES cells were spread and elongated. By adding growth factors inspired from hepatic microenvironments (hepatocyte growth factor, dexamethasone, oncostatin M), CE-ES cells adopted more hepatic-like cuboidal appearance, whereas, CD-ES cells maintained elongated morphology. Furthermore, albumin and glucose-6-phosphatase (GO) levels, mid-late liver-specific markers, increased upon hepatotrophic stimulation in CE-ES cells. These trends suggested that E-cadherin expression enhanced CE-ES cell responsiveness to growth factor stimulation.; Lastly, ES cell integration/maturation was examined under organotypic conditions (co-cultivation with primary adult hepatocytes). Cadherin expression and priming predisposition on hepatodifferentiation was probed within microenvironments mimicking the liver. Direct cell contact with adult hepatocytes was requisite for enhanced ES cell commitment towards hepatodifferentiation. CE-ES cells acquired more hepatic-like phenotype with elevated levels of G6P expression. Hence, the heterotypic interface was significant in coordinating cellular responses. This will be significant for the future molecular optimization of transplantable ES cells.