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, IVL^mES, which included the hepatocyte layer and a surrounding sinusoid vascular-like network. The IVL^mES 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 IVL^mES, supplemental l-ornithine could activate the urea cycle, and also protect against ammonium/alcohol-induced hepatocyte death. While the IVL^mES 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 IVL^mES will represent a useful, humane alternative to animal studies for drug toxicity and mechanistic studies of liver injury.