Genes and Development—A Workshop Report J. C. Cross a,* , P. M. Coan b , R. Fundele c , M. Hemberger a , M. Kibschull d and A. Ferguson-Smith b a Genes and Development Research Group, Department of Biochemistry and Molecular Biology, University of Calgary Faculty of Medicine, HSC Room 2279, 3330 Hospital Drive, N.W., Calgary, Alberta T2N 4N1, Canada; b Department of Anatomy, University of Cambridge, Cambridge, UK; c Department of Development and Genetics, Uppsala University, Uppsala, Sweden; d Institute of Anatomy, University of Duisburg-Essen, Essen, Germany INTRODUCTION The ‘Genes and Development’ workshop focused on recent advances in our understanding of the molecular control of placental development, focusing on the mouse model. Jay Cross gave an introduction to mouse placental development that outlined the major structures and cell types. He also reviewed the considerable progress that has been made in defining molecular pathways controlling development using mutant and transgenic mice as well as increasing knowledge of the biology underlying implantation and establishment of the mature placenta [1,2]. One of the important general findings to emerge in recent years is that development and function of the different sub-structures of the placenta can be interdependent and able to adapt to defects in other parts. This can complicate the analysis of a phenotype in that some types of changes can simply represent secondary changes reflecting attempts to compensate, and investigators need to take this into consider- ation before making conclusions. The remaining speakers were selected to discuss different aspects of placental development and highlight the use of new experimental approaches for studying mouse placental development and function. USE OF TROPHOBLAST STEM CELLS TO ANALYSE GENE FUNCTIONS Mark Kibschull from Elke Winterhager’s lab in Essen dis- cussed the use of primary trophoblast stem (TS) cell lines to analyse the function of connexin genes during placental devel- opment. TS cell lines can be generated from mouse post- implantation trophoblast cells. The breakthrough of this method published by Tanaka et al. [3] was the use of FGF4 and embryonic fibroblasts to keep the trophoblast cells prolif- erating and undifferentiated in vitro. In the presence of FGF4 and fibroblast conditioned medium (EMFI-CM) TS cells continue to divide in culture and express markers of early trophoblast. They can be generated from dissected extra- embryonic ectoderm or chorionic ectoderm but the easiest way is to derive them from blastocyst outgrowths. TS cells can only contribute to the trophoblast cell lineage in vivo as shown in chimeric embryo experiments and in vitro after induction of differentiation. Differentiation of TS cells can be induced by the removal of FGF4 or EMFI-CM from a proliferating TS cell culture, or by the addition of reagents like retinoic acid [3,4]. They differentiate along the trophoblast lineage into spongiotrophoblast and giant cells and probably also in syn- cytiotrophoblast. The advantage of using TS cell lines to study placental phenotypes is quite clear: TS cell lines can easily be generated from blastocysts of a knockout mouse resulting in a specific gene deficient trophoblast cell line which serves as a cell model for analysis. This is particularly significant if the placental phenotype results in embryonic lethality, as TS cell studies may be the only chance to investigate the underlying pathways in detail. Dr Kibschull then presented data showing the use of TS cell lines to analyse the role of connexins during placental development. Like in the early trophoblast lineage in vivo, undifferentiated wildtype TS cells express only Cx31 channels. Upon TS cell differentiation both Cx26 and Cx43 are induced each in different subpopulations of the culture. This connexin expression pattern is comparable to the placenta, where Cx26 channels connect the syncytiotrophoblast layers and Cx43 is expressed in giant cells. Null mutation of Cx31 results in a dysmorphogenesis of the placenta and as a consequence to embryonic death [5]. This phenotype is due to an impairment of the balance between proliferation and differentiation of trophoblast cells in favour of the giant cells pathway. Cx31- deficient TS cells showed accelerated differentiation compared to wildtype cells. This was indicated by an overall shift in expression of connexins and marker genes such as Mash2, Pl1 and Tpbpa to earlier differentiation stages. Furthermore, Cx31 mutant TS cells show a reduced proliferation but only when they start to differentiate. These results correspond to the enhanced differentiation and the reduced proliferation of the trophoblast cells observed in the Cx31 knockout placenta in vivo [5]. In addition, gene expression microarray analysis revealed that Cx31-deficiency in TS cells is correlated with a shift to giant cell differentiation. Taken together TS cells represent a powerful tool to study several aspects of tropho- blast lineage development. Furthermore, TS cell lines from knockout conceptuses make it possible to analyse the function of specific genes and their signalling pathways in trophoblast differentiation in vitro. * To whom correspondence should be addressed. Tel.: +1-403-220- 6876; Fax: +1-403-270-0737; E-mail: jcross@ucalgary.ca Placenta (2004), 25, Supplement A, Trophoblast Research, Vol. 18, S39–S41 doi:10.1016/j.placenta.2004.01.020 0143-4004/$–see front matter  2004 IFPA and Elsevier Ltd. All rights reserved.