Engineering the Hepatocyte Differentiation–Proliferation Balance by Acellular Cadherin Micropresentation

Abstract

The successful development of bioartificial and cell-based liver support systems relies on the identification of molecular mechanisms controlling the balance between hepatocellular proliferation and differentiation. Although a definitive function-inductive role for the cell-cell adhesion molecule, E-cadherin, was established through lateral cadherin-cadherin engagement in hepatocyte cocultures (Brevia, T.A., and Moghe, P.V. Biotechnol. Bioeng. 76, 295, 2001), the roles of other modes of cadherin presentation are not well understood. Further, alternative cadherin display configurations promoting cell growth/proliferative pathways, a major requisite for sustainable engineered tissues, remain to be identified. In this report, we employed protein A-functionalized polymeric microsphere substrates that specifically bound self-dimerizing cadherin-IgG/Fc fusion chimeras via their Fc regions, thereby orienting them outward for active adhesion, and presented the E-cadherin chimeras basally to cultured rat hepatocytes to study the effects of cadherin display on cell proliferative potential and differentiated function. In contrast to the previously documented function-inductive roles of laterally expressed cadherin, basal acellular cadherin presentation resulted in an increase in hepatocyte DNA synthesis and cell divisions, accompanied by a decrease in the expression of a key marker of liver-specific function, albumin message levels. Next, we probed the relative effect of basal exogenous display of acellular cadherins on the inductive phase of differentiation within hepatocyte cocultures with cadherin-expressing L929 cells. When acellular cadherins were applied to hepatocyte cocultures involving chaperone cell-mediated cadherin presentation, the previously reported function-inductive effects of cadherins on hepatocyte function were reversed, resulting in lower levels of albumin and urea secretion indicating a dominance of acellular cadherin effects. Our results demonstrate that cadherins are important regulators of the balance between hepatocyte differentiation and proliferation, and furthermore that the direction of balance shift may be dependent on the method of cadherin presentation. Thus, the geometric display of cadherin could be a potential parameter to switch hepatocyte functional-proliferative balance and may aid in customizing scaffolds for regulating hepatic tissue dynamics.

Control of hepatic differentiation via cellular aggregation in an alginate microenvironment

“…As a final point, E-cadherin expression has also been documented in other tissue engineering based work, namely the increase in E-cadherin expression of hepatocytes cultured in alginate/galactosylated chitosan/ heparin scaffolds (Seo et al, 2006). In addition, E-cadherin presented to mature hepatocytes in the form of modified microspheres, modulated the functional state of the hepatocytes, existing either as a proliferating cell population or a differentially functioning cell population (Brieva and Moghe, 2004). E-selectin too, has been described for modulating cellular aggregation, though it has not been shown to be involved in hepatic differentiation.…”

“…However we do see E-cadherin production later on in the differentiation process, Figure 2A. The E-cadherin protein itself has been identified within liver tissue samples, (Figarella-Branger et al, 1995) and has been reported to play a role in hepatic differentiation, (Brieva and Moghe, 2004;Dasgupta et al, 2005). Furthermore, the presence of tight junctions facilitated by the presence of E-cadherin, are necessary for normal liver function.…”

Downregulation of KLF6 is an early event in hepatocarcinogenesis, and stimulates proliferation while reducing differentiation

“…E-cadherin, along with the adenomatous polyposis coli (APC) gene, is frequently hypermethylated in HCC, associated with poorer survival [34,35]. Cadherins can regulate the balance between hepatocyte differentiation and proliferation [21] and Ecadherin is upregulated in HCC cell lines by agents that induce differentiation, including sodium butyrate and interferon alpha [36].…”

“…Therefore, we next explored whether inhibition of hepatocellular growth by KLF6 was associated with increased differentiation, as assessed by analysis of several growth-related (cyclin D1 and beta catenin) and differentiation-related transcripts (albumin, transthyretin (TTR)), as well as alpha fetoprotein (AFP), a marker of the undifferentiated, oncofetal state of hepatocytes. In addition, Ecadherin was assessed as a marker for differentiation, as well as growth inhibition [21]. The effect of KLF6 overexpression on these genes is summarized in Table 1.…”

Characterization of porous poly(D,L‐lactic‐co‐glycolic acid) sponges fabricated by supercritical CO2 gas‐foaming method as a scaffold for three‐dimensional growth of Hep3B cells

“…A possible mechanism to explain the low cell proliferation and increased function of hepatocytes could involve a Zhu et al: Supercritical CO 2 Gas-Foaming Method reciprocate relationship between cell growth and differentiation (Brieva and Moghe, 2004). Hepatocytes with high proliferation were believed to have been stimulated into the S phase with a concomitant down-regulation of differentiated functions (Zhu et al, 2007b).…”