Anat Embryol (2004) 208:359–366 DOI 10.1007/s00429-004-0403-4 ORIGINAL ARTICLE J. Dudas · M. Papoutsi · M. Hecht · A. Elmaouhoub · B. Saile · B. Christ · S. I. Tomarev · C. S. von Kaisenberg · L. Schweigerer · G. Ramadori · J. Wilting The homeobox transcription factor Prox1 is highly conserved in embryonic hepatoblasts and in adult and transformed hepatocytes, but is absent from bile duct epithelium Accepted: 30 April 2004 / Published online: 30 June 2004  Springer-Verlag 2004 Abstract Prox1 is a transcription factor with two highly conserved domains, a homeobox and a prospero domain. It has been shown that Prox1 knock-out mice die during early embryonic stages and display a rudimentary liver. We have studied the expression of Prox1 at RNA and protein levels in chick, rat, mouse and human liver and in transformed and non-transformed hepatic cell lines. Prox1 is expressed in early embryonic hepatoblasts and is still expressed in adult hepatocytes. Prox1 protein is located in the nuclei of hepatoblasts, which grow into the neigh- boring embryonic mesenchyme. The expression pattern in chick, mouse, rat and human embryos is highly con- served. Besides albumin and a-fetal protein, Prox1 be- longs to the earliest markers of the developing liver. In adult liver, Prox1 is expressed in hepatocytes but is absent from bile duct epithelial and non-parenchymal cells (Kupffer cells, hepatic stellate cells, sinusoidal endothe- lial cells and myofibroblasts). Isolated primary hepato- cytes and hepatoma cell lines (HepG2, Hep3B) are Prox1 positive, whereas the immortalized murine liver cell-line MMH, which constitutively expresses the receptor c-met, is Prox1 negative. Transfection of MMH with Prox1 cDNA increases the expression level significantly as com- pared to control transfectants. In HepG2 and Hep3B, the Prox1 levels are even up to 100 times higher. Our studies show that Prox1 is a highly conserved transcription factor, expressed in hepatocytes from the earliest stages of de- velopment into adulthood and over-expressed in hepato- ma cell lines. Its absence from bile duct epithelial cells suggests a function for the specification of hepatoblasts into hepatocytes. The genes controlled by Prox1 need to be studied in the future. Keywords Liver development · Chick · Rat · Human · MMH · Hepatoma cells Introduction The parenchyma of the liver is of endodermal origin. Development of the endoderm is controlled by tran- scription factors such as HNF3b and GATA-4 (Ang and Rossant 1994; Kuo et al. 1997). The endoderm then forms a tube, which can be subdivided into fore-, mid- and hindgut. Regional expression of HNF3a,-b and -g sug- gests that these transcription factors are involved in specification of endodermal compartments (Ang et al. 1993). The liver starts developing at embryonic day 8 (ED) in the mouse and ED 2.5 in the chick at the tran- sition of fore- and midgut. The first molecular evidence for liver development is the expression of albumin and a- fetal protein, which can be observed in the mouse around ED 9 (Gualdi et al. 1996). Liver development is regulated by paracrine interac- tions of the cardiogenic and septum transversum meso- J. Dudas · A. Elmaouhoub · B. Saile · G. Ramadori Department of Gastroenterology and Endocrinology, Georg August University Göttingen, Robert Koch Strasse 40, 37075 Göttingen, Germany M. Papoutsi · M. Hecht · L. Schweigerer · J. Wilting ( ) ) Pediatrics I, Children’s Hospital, Georg August University Göttingen, Robert Koch Strasse 40, 37075 Göttingen, Germany e-mail: joerg.wilting@med.uni-goettingen.de Tel.: +49-551-396270 Fax: +49-551-396231 B. Christ Institute of Anatomy and Cell Biology, Albert Ludwigs University Freiburg, Albertstrasse 17, 79104 Freiburg, Germany S. I. Tomarev Laboratory of Molecular and Developmental Biology, National Eye Institute, National Institutes of Health, Bethesda, MD 20892-2730, USA C. S. von Kaisenberg Universitätsfrauenklinik, Michaelisstrasse 16, 24105 Kiel, Germany