GLYCEROPHOSPHORYLETHANOLAMINE (GPEA) IDENTIFIED AS AN HEPATOCYTE GROWTH STIMULATOR IN LIVER EXTRACTS

Extracts from weanling pig liver were found to act synergistically wit h growth factors such as hepatocyte growth factor and transforming gro wth factor-alpha to stimulate hepatocyte growth in serum-free cultures . In the absence of added growth factors, the extracts had no activity . The compound responsible for this activity was isolated by passing h eat-treated liver extract through anion-exchange and heparin columns f ollowed by gel filtration at neutral and low pH, reversed-phase HPLC, and a final gel filtration column at low pH. The activity was followed throughout the purification by its ability to increase substantially the incorporation of [H-3]thymidine into primary rat hepatocytes cultu red serum-free in the presence of hepatocyte growth factor. The active compound was identified by NMR and mass spectrometry as glycerophosph orylethanolamine (GPEA), a breakdown product of the phospholipid phosp hatidylethanolamine. The ethanolamine portion of the molecule was crit ical for the observed activity, whereas the glycerol phosphate portion was not necessary. In the absence of added growth factors, neither GP EA nor ethanolamine had any stimulatory effect on the cells. These res ults demonstrate that hepatocytes grown in culture, and especially tho se grown in serum-free media, require a supplement of ethanolamine and /or GPEA. In the absence of these compounds, their response to growth stimuli is greatly reduced. (C) 1996 Academic Press, Inc.