Understanding hepatocellular signaling occurring in biomaterial systems is important for successful hepatic tissue engineering. Toward this end, we employed synthetic glycopolymers, as artificial matrices, to examine integrin-mediated epidermal growth factor (EGF) signaling in primary hepatocyte cultures. We dispersed hepatocytes on a collagen matrix or on a synthetic glycopolymer matrix and subsequently stimulated them with EGF. Only hepatocytes cultured on collagen proliferated, and we observed significant expression of cyclin B1 in these cells. Pharmacological agents, LY294004 (a phosphatidylinositol [PI] 3-kinase inhibitor) and AG1478 (an EGF kinase receptor inhibitor), blocked hepatocyte proliferation and cyclin B1 expression. In addition, EGF-stimulated hepatocytes formed spheroids, exhibited membrane ruffling, and increased tryptophan 2,3-oxygenase (TO) expression when cultured on glycopolymer matrices. Interestingly, PI 3-kinase inhibition suppressed membrane ruffling, spheroid formation, and TO expression. Taken together, this data suggests PI 3-kinase plays an important role in mediating cross talk between integrin and the EGF signaling pathways in primary hepatocyte cultures.