Regulation of cell morphology and cytochrome P450 expression in human hepatocytes by extracellular matrix and cell-cell interactions

The influence of extracellular matrix conditions and plating density on cel l cytoarchitecture and the constitutive and chemically induced expression o f cytochrome P450 3A4 (CYP3A4) was examined in primary cultures of human he patocytes. Constitutive and drug-induced microsomal CYP3A4 expression occur red equally well in human hepatocyte cultures maintained on either a comple x or simple substratum (Matrigel vs collagen, type I), or in a sandwich con figuration (i.e., between two layers of extracellular matrix), despite the markedly different morphological properties exhibited by each condition. Ho wever, a density-dependent decrease in both the constitutive and induced le vels of CYP3A4 was observed in hepatocytes maintained on a simple collagen substratum as plating density was reduced from 100% to 25%. Marked alterati ons in cell shape and cytoarchitecture were noted concomitant with decrease s in the expression and localization of intercellular gap junctions and E-c adherin-mediated cell adhesions. In addition, the intracellular distributio n of microtubules and microfilaments was altered substantially and the expr ession of immunoreactive actin and beta -tubulin increased as cell density was decreased. These effects were reversed to some extent by overlaying mon olayers with extracellular matrix or by co-culturing with another cell type . Efforts to maintain normal cell shape and cytoskeletal distribution in he patocytes at low cell density with a Matrigel substratum failed to restore normal basal levels of CYP3A4 expression or responsiveness to rifampicin (R IF). Likewise, E-cadherin and Cx-32 expression was again reduced, even thou gh the distribution and expression of cytoskeletal elements returned to nor mal levels. These results suggest that cell-cell contacts, but not the extr acellular matrix configuration or composition, play a critical role in dete rmining normal responsiveness to chemical modulators in human hepatocytes.