Pleiotrophin as a Swiss 3T3 cell-derived potent mitogen for adult rat hepatocytes

Rat parenchymal liver cells were cultured in the presence of lethally treated Swiss 3T3 cells. This co-culture allowed hepatocytes to produce colonies containing more than 300 cells in 30 days. Hepatocytes in colonies appeared morphologically normal and some of them were suggested to have bipotental differentiation capacity. The initial growth stimulatory activity of the feeder cells was replaceable with their conditioned medium (CM). Biochemical analysis of an active principle in the 3T3 cell-CM identified pleiotrophin. Pleiotrophin purified from the 3T3 cell-CM, recombinant human pleiotrophin, chemically synthesized human pleiotrophin, and midkine promoted the growth of hepatocytes as well. Reverse transcription-polymerase chain reaction clearly showed that the synthesis of mRNA of pleiotrophin was stimulated in the regenerating liver induced by either partial hepatectomy or the treatment with d-galactosamine, strongly suggesting a biological significance of pleiotrophin in the proliferation of hepatocytes in vivo. From these results we concluded that pleiotrophin is a new potent growth factor for adult parenchymal hepatocytes. This study indicates the importance of mesenchymal stimulation for the growth of adult rat hepatocytes.