BBB seminar: Philippe Collas
Isolation and epigenetic characterization of mesenchymal stem cells from adipose tissue
Department of Biochemistry, Institute of Basic Medical Sciences, University of Oslo
Stem cells have been identified in several adult mesenchymal tissues and are believed to be responsible for tissue homeostasis. Mesenchymal stem cells undergo self-renewing divisions but also give rise to more committed progenitor cells which differentiate into specific cell types. Stem cells of stromal origin can be obtained from liposuction material. These cells display multilineage differentiation capacities in vitro and can promote repair of various tissues and functions in vivo.
We recently reported the purification and characterization of cells with a CD34+CD105+CD45-CD31- phenotype from the stromal vascular fraction of human adipose tissue, which exhibit mesenchymal stem cell properties. CD31- adipose stem cells (ASCs) are relatively quiescent but reenter the cell cycle in culture. They can be expanded clonally and differentiate into mesodermal lineages. Transcription profiling reveals expression of genes extending across all three germ layers. Further, single cell ASC clones display variations in their differentiation potential, raising the hypothesis of an epigenetic basis for this variation. To begin addressing this issue, we determined whether the DNA methylation status in undifferentiated ASCs would predict transcriptional activation potential upon differentiation.
Several features seem to epigenetically characterize ASCs. (i) Freshly isolated ASCs from three separate female donors display overall hypomethylated adipogenic loci, in contrast to myogenic or endothelial genes. (ii) ASCs display a mosaic CpG methylation profile, on the basis of heterogeneous methylation patterns between individual cells within a donor or within a clonal cell population, and from variations in the percentage of methylation of a given CpG between donors or between cell clones. (iii) DNA methylation profiles reflect neither the transcriptional status of any of the genes examined in undifferentiated cells, nor the potential for expression upon in vitro differentiation. (iv) Clonal culture of ASCs from single isolated cells does not affect global DNA methylation, but generates between-clone mosaicism at specific CpGs. (v) In vitro differentiation toward adipogenic, osteogenic or endothelial pathways maintains methylation patterns despite the induction of lineage-specific gene expression. (vi) Adipogenic genes are more methylated in primary differentiated cells unrelated to adipogenesis, arguing for ASC-specificity of the hypomethylated state of these loci. In conclusion adipogenic promoters are mosaically hypomethylated in ASCs; this hypomethylation may constitute a molecular signature of ASCs irrespective of gene expression. DNA hypomethylation of adipogenic genes may poise genes for transcriptional activation. DNA methylation, however, does not appear to be the sole determinant of differentiation potential of ASCs.
Host: Karl Tronstad, Department of Biomedicine
Philippe Collas received his PhD in 1991 from the University of Massachusetts Amherst, MA, USA. In 1995 he moved to Oslo where he has been active in the Department of Biochemistry at the Veterinary School and since 1999 at the University of Oslo, today as Professor of Biochemistry. Philippe Collas' research background is in transcriptional events in the nucleus and his current research focuses on dynamics of the nuclear envelope, on chromatin remodeling and on epigenetic reprogramming of nuclear function. His seminar will deal with isolation and epigenetic characterisation of mesenchymal stem cells from adipose tissue.
Boquest, A. C., Shahdadfar, A., Fronsdal, K., Sigurjonsson, O., Tunheim, S. H., Collas, P. and Brinchmann, J. E. (2005). Isolation and transcription profiling of purified uncultured human stromal stem cells: alteration of gene expression after in vitro cell culture. Mol Biol Cell 16, 1131-41.
Gaustad, K. G., Boquest, A. C., Anderson, B. E., Gerdes, A. M. and Collas, P. (2004). Differentiation of human adipose tissue stem cells using extracts of rat cardiomyocytes. Biochem Biophys Res Commun 314, 420-7.
Martins, S., Eikvar, S., Furukawa, K. and Collas, P. (2003). HA95 and LAP2 beta mediate a novel chromatin-nuclear envelope interaction implicated in initiation of DNA replication. J Cell Biol 160, 177-88.
Martins, S. B., Marstad, A. and Collas, P. (2003). In vitro modulation of the interaction between HA95 and LAP2beta by cAMP signaling. Biochemistry 42, 10456-61.
Collas, P. and Hakelien, A. M. (2003). Teaching cells new tricks. Trends Biotechnol 21, 354-61.