Final Activity Report Summary - HSC (Migration of HSCs from the AGM region to other haemopoietic organs during mouse ontogeny) The haemopoietic stem cells (HSCs) are the foundation of the adult haemopoietic system and give rise to all mature blood cells. In mammals distinct sites of haemopoietic activity have been identified during ontogeny. Quantitative temporal measurement of various haemopoietic activities present in the yolk sac, AGM and foetal liver suggests that the foetal liver activity is a consequence of colonisation by cells emigrating from the AGM and yolk sac (Kumaravelu, 2002). However, there is only limited experimental data where this migration was shown (Emambokus, 2003; Gothert, 2004; Samokhvalov 2007). Therefore, to investigate whether HSCs generated in the AGM are able to migrate and seed other haemopoietic organs, such as foetal liver and bone marrow we followed a different approach using a more tightly regulated and inducible TetON system and Cre-lox technology. We attempted to mark HSCs with the fluorescent protein YFP at the time of their generation in the AGM and track marked cells through development. For the specific cell lineage expression of Cre recombinase the Sca-1 expression cassette, generated by the host laboratory, was used (Miles,1997; Ma, 2001). Sca-1 (stem cell antigen-1) is a widely used HSC marker (Spangrude,1988). One part of the TetON system (rtTAM2) was expressed under the control of the Sca-1 locus and several transgenic mouse lines were established. Subsequently these mice were supposed to be bred against two other transgenic mouse lines, TetO-Cre mice and ROSAYFP reporter mice. In the presence of doxycycline, rtTAM2 should activate the TetO-Cre promoter allowing the expression of Cre-recombinase. Active Cre-recombinase would then delete a stop cassette, flanked by loxP sites, and lead to the generation of the fluorescent protein YFP as a permanent mark. Unfortunately, a careful characterisation of all established transgenic rtTAM2 mouse lines revealed that none of the lines was expressing the rtTAM2 protein in HSCs.
