In contrast to most of the other research groups at IPK our group is using mammalian cells to study early differentiation processes based on stem cells.

The main focus is the analysis of differentiation mechanisms of mouse embryonic stem (ES) cells in vitro, but also of cord blood-derived stem/progenitor cells. Moreover, the identification and characterization of progenitor cells generated during ES cells differentiation, and potentially found in adult tissues, is investigated. Signalling mechanisms and effects of growth/differentiation factors and extra-cellular matrix proteins are further analysed during differentiation of ES cells into endodermal (pancreatic, hepatic), mesodermal (cardiac) and ectodermal (neuronal) cell types. The following research fields are of interest to our group:

Differentiation of ES cells into pancreatic insulin-producing cells: The pancreatic differentiation of wild type ES cells and of ES cells constitutively expressing Pax4 (Pax4+) was analysed. By using a new in vitro differentiation model, we show that ES cells differentiate into insulin-producing islet-like clusters. At an intermediary stage, the intermediate filament proteins nestin and cytokeratin 19, the pancreatic transcription factor Isl-1 and the proinsulin marker protein C-peptide are transiently expressed. The differentiated cells show functional properties of beta cells, including glucose-dependent insulin release, pancreas-specific ion channel activity and normalization of blood glucose level after transplantation into diabetic mice. Of special interest is the analysis of signalling factors involved in the regulation of differentiation of ES cells into pancreatic versus neuronal cells. To this aim, we performed transcriptional profiling of wild type and transgenic Pax4+ cells and studied pathways of early lineage differentiation. In addition, we analyse the regulation of ES cells into endoderm (pancreatic/hepatic) versus ectoderm (neural) lineage by using exogenous differentiation factors (i.e. Activin A).

Analysis of cardiac differentiation: During the last years, we were successful in showing the differentiation of ES cells into functional cardiomyocytes. On the basis of new Affymetrix data, we are studying the role of neuroectodermal cells on early cardiac commitment and differentiation. In addition, the effects of small molecules on cardiac differentiation, especially into pacemaker cells, are investigated.

Isolation and characterization of nestin-positive progenitor cells from mouse intestinal epithelium (intestinal epithelial-derived nestin-positive progenitors, INPs): We generated nestin-expressing cells called INPs by co-culture of intestinal epithelial cells on mouse embryonic fibroblasts via WNT-dependent mechanisms. The INPs showed high, but limited proliferation capacity. Following differentiation of INPs, cells expressed tissue-specific transcripts and proteins. However, the functional capacity of the cells was limited as demonstrated by functional assays, such as ELISA (pancreatic, hepatic cells), integration into hippocampal brain slices (neuronal cells) and transplantation into mouse blastocysts.

Ongoing experiments concentrate on the characterization of ES cells constitutively expressing nestin. We are interested to study the effects of the neural stem cell marker nestin on the development of progenitor cells, which eventually differentiate into multiple lineage.

Characterization of Prominin-1 as common marker of progenitor cells expressed in ES cells: Prominin-1 (the orthologue of the human CD133 antigen), previously described as marker of neuroepithelial and hematopoietic stem cells, was detected on committed and early progenitor cells derived from mouse ES cells. The co-expression with nestin during early differentiation in multi-lineage progenitors suggests a role for prominin-1 as a new marker defining early ES-derived progenitor cells. The potential role of prominin-1 in regulating symmetric versus asymmetric cell division is of special interest.

Analysis of the effects of electromagnetic fields (EMF) on ES cells and neural progenitor cells: ES cells were exposed to low (ELF) and high (RF) frequency electromagnetic fields. EMF exposure to neural progenitor cells generated from ES cells during neuronal differentiation resulted in a modification of transcript levels of genes involved in apoptosis (bax, bcl-2, GADD45) suggesting that apoptotic mechanisms may be involved in the immediate response to EMFs. The data have been verified by several cell biological assays, including COMET and FACS analysis.

The group has ongoing collaborations with the Samuel Lunenfeld Research Institute, Mount Sinai Hospital, University of Toronto, Canada (Prof.  A. Nagy), Laboratory of Cardiovascular Science, GRC, National Institute on Aging (NIA), NIH, Baltimore, USA (Prof. K. Boheler), Laboratory of Molecular Medicine, IZKF, University of Leipzig (Dr. M. Cross), Institute of Reconstructive Neurobiology, University of Bonn (Prof. O. Brüstle), Institute for Anatomy and Cell Biology, University of Halle (Dr. A. Navarrete-Santos), Department of Pharmacology and Toxicology, Technical University Dresden (Prof. U. Ravens), Biocenter of the TU Dresden (Dr. D. Corbeil), Institute of Biological Interfaces (IBG), Research Centre Karlsruhe (Dr. A. Rolletschek), Fraunhofer Institute IGB, Stuttgart (Prof. Heike Mertsching).