Hepatocytes in primary cultures de-differentiate and re-differentiate following addition of Engelbreth-Holm-Swarm mouse sarcoma (matrigel) to the cultures. The Wnt/beta-catenin pathway has been shown to be important in liver growth and development. Here, we investigate changes in beta-catenin and its mechanism, during matrigel-induced hepatocyte differentiation. Primary rat hepatocytes were cultured for 8 days, and matrigel was added to half of the cultures. Total and nuclear protein and total RNA were extracted at different days of culture and examined for beta-catenin and other Wnt pathway components. A significant increase in total beta-catenin protein was observed upon matrigel addition, during hepatocyte differentiation, despite a decrease in beta-catenin and frizzled-1 (Wnt receptor) expression. A concurrent decrease in the glycogen synthase kinase-3beta (GSK3beta), axin, and ser45/thr41-phosphorylated beta-catenin proteins was observed in matrigel-treated cultures, implying decreased degradation of beta-catenin. Interestingly, a decrease in nuclear beta-catenin and total active beta-catenin was observed in the presence of matrigel. Matrigel also induced an increased association of beta-catenin with Met (hepatocyte growth factor receptor), whereas association with E-cadherin remained unchanged. This coexisted with decreased beta-catenin tyrosine phosphorylation. Thus, beta-catenin undergoes multifactorial regulation during matrigel-induced hepatocyte differentiation and maturation; this induces its stabilization and membrane translocation, possibly contributing to hepatocyte differentiation.