The human constitutive androstane receptor promotes the differentiation and maturation of hepatic-like cells

Abstract

Expression of the constitutive androstane receptor (CAR, NR1I3) is enriched in the mature mammalian liver and increasingly recognized for its prominent role in regulating a myriad of processes including biotransformation, chemical transport, energy metabolism and lipid homeostasis. Previously, we demonstrated that CAR levels were markedly enhanced during the differentiation of hepatic-like cells derived from hESCs, prompting the hypothesis that CAR contributes a key functional role in directing human hepatogenesis. Here we demonstrate that over-expression of CAR in human embryonic stem cells (ESCs), transduced by a lentiviral vector, accelerates the maturation of hepatic-like cells, with CAR over-expressing cells exhibiting a 2.5-fold increase in albumin secretion by day 20 in culture differentiation, and significantly enhanced levels of mRNA expression of several liver-selective markers, including hepatic transcription factors, plasma proteins, biotransformation enzymes, and metabolic enzymes. CAR over-expressing cells also exhibited enhanced CITCO-inducible CYP3A7 enzymatic activity. Knockdown of CAR via siRNA attenuated the differentiation-dependent expression programs. In contrast, expression levels of the pregnane X receptor (PXR), a nuclear receptor most similar to CAR in primary sequence, were negligible in human fetal liver tissues or in the differentiating hESCs, and stable over-expression of PXR in hepatic-induced hESCs failed to enhance expression of hepatic phenotype markers. Together, these results define a novel role for human CAR in hepatic lineage commitment.