Mesenchymal stem cells (MSCs) are multipotent, can be easily isolated from bone marrow and other tissues and expanded ex vivo also for a potential clinical use. So far, MSCs are characterized by their adherent properties, immunophenotype, and their differentiation potential in vitro. Because of a lack of distinctive markers for MSCs, the current understanding of their endogenous localization and functions in vivo remain elusive. However, the identification of the MSC niche is important in order to better understand the cell biological characteristics of these cells and their physiological and pathophysiological role.

We therefore aimed to generate a transgenic mouse model in which MSCs are labeled by a live reporter gene. CD73, an established membrane surface molecule on MSCs, appeared to be a suitable marker with only moderate distribution in other cell types. We confirmed high expression of CD73 in various MSC populations by FACS, immunostainings and PCR. A reporter vector expressing EGFP under control of the CD73 promotor was generated from bacterial artificial chromosomes (BACs). Transfection of the vector into established MSC lines gave rise to EGFP positive cells with the ability to differentiate into the osteogenic and adipogenic lineage, proving expression of the construct in multipotent MSCs. Next, we generated transgenic embryonic stem cell (ESC) lines using the CD73-BAC-EGFP reporter vector. EGFP positive cells could be detected starting from day eight of ESC differentiation. Then, transgenic mice were generated from ESC clones by diploid aggregation. EGFP expressing cells could be found in different tissues of embryonic and adult CD73-BAC-EGFP mice. Primary cultures from bone marrow, white fat, uterus and kidney gave rise to adherent growing EGFP positive cells that were able to differentiate into mesenchymal lineages, underlining the residence of MSCs in various organs. In late stage embryos EGFP positive cells were visible in developing bones at the sites of endochondral ossification, demonstrating osteogenic precursor cells.

We are currently further analyzing the transgenic mouse model to characterize the EGFP positive cells in regard to the specificity of the labeling for MSCs and to investigate their in vivo distribution.