Cytomimetic Engineering of Hepatocyte Morphogenesis and Function by Substrate-Based Presentation of Acellular E-Cadherin

Abstract

Although cadherin-mediated intercellular contacts can be integral to the maintenance of functionally competent hepatocytes in vitro, the ability to engineer hepatocellular differentiated function via acellular E-cadherin has yet to be thoroughly explored. To investigate the potential of substrate-presented, acellular E-cadherin to modulate hepatocellular self-assembly and functional fate, rat hepatocytes were cultured at sparse densities on surfaces designed to display recombinant E-cadherin/Fc chimeras. On these substrates, hepatocytes were observed to recognize microdisplayed E-cadherin/Fc and responded by modulating the spatial distribution of the intracellular cadherin-complexing protein beta-catenin. Substrate-presented E-cadherin/Fc was also found to markedly alter patterns of hepatocyte morphogenesis, as cellular spreading and two-dimensional reorganization were significantly inhibited under these conditions, leading to multicellular aggregates that were considerably more three-dimensional in nature. Increasing cadherin exposure was also associated with elevated levels of albumin and urea secretion, two markers of hepatocyte differentiation, over control cultures. This suggested that cell-substrate cadherin engagement established more functionally competent hepatocellular phenotypes, coinciding with the notion that E-cadherin is a differentiation-inducing ligand for these cells. The morphogenetic and function-promoting effects of substrate-bound E-cadherin/Fc were further enhanced under conditions in which protein A was utilized as an anchoring molecule to present cadherin molecules, suggesting that ligand mobility may play an important role in the effective establishment of cell-to-substrate cadherin interactions. Interestingly, the percent increase in function detected for conditions of high cadherin exposure versus control cultures was found to be substantially higher at extremely low cell densities. This observation indicated that hepatocytes respond to substrate-presented E-cadherin even in the absence of native intercellular interactions and associated juxtacrine signaling. The incorporation of acellular E-cadherin on biomaterial substrates may thus potentially present a means to prevent hepatocellular dedifferentiation by maintaining liver-specific function in otherwise severely functionally repressive culture conditions.