In vitro recapitulation of the urea cycle using murine embryonic stem cell-derived in vitro liver model

Ammonia, a toxic metabolite, is converted to urea in hepatocytes via the urea cycle, a process necessary for cell/organismal survival. In liver, hepatocytes, polygonal and multipolar structures, have a few sides which face hepatic sinusoids and adjacent hepatocytes to form intercellular bile canaliculi connecting to the ductules. The critical nature of this three-dimensional environment should be related to the maintenance of hepatocyte function such as urea synthesis. Recently, we established an in vitro liver model derived from murine embryonic stem cells, IVLmES, which included the hepatocyte layer and a surrounding sinusoid vascular-like network. The IVLmES culture, where the hepatocyte is polarized in a similar fashion to its in vivo counterpart, could successfully recapitulate in vivo results. l-Ornithine is an intermediate of the urea cycle, but supplemental l-ornithine does not activate the urea cycle in the apolar primary hepatocyte of monolayer culture. In the IVLmES, supplemental l-ornithine could activate the urea cycle, and also protect against ammonium/alcohol-induced hepatocyte death. While the IVLmES displays architectural and functional properties similar to the liver, primary hepatocyte of monolayer culture fail to model critical functional aspects of liver physiology. We propose that the IVLmES will represent a useful, humane alternative to animal studies for drug toxicity and mechanistic studies of liver injury.