Characterization of surface markers of human amnion mesenchym-derived side population cells

Abstract

Objective We aim to characterize the side population cells obtained by FACS from human amnion cells (hAMC) as stem cells. Methods hAMC were prepared by an enzymatic treatment from the amni-otic membrane separated mechanically from the placenta obtained by Caesarean section of 38-40 weeks after pregnancy. SP cells(hAMC-SP) were isolated by FACS from them and studied for stem cell antigens. Results hAMC-SP cells were about 0.3% among hAMCs. The SP cells at fourth to tenth passages after the culture start were positive for Nestin, Vimentin, integrin family antigens (CD49b、CD49c、CD49d、D49e) and other antigens such as CD9, CD13, CD19, CD29, CD44, CD46, CD51, CD59, CD166, and Oct-3/4. HLA-ABC,TRA-1-81 and SSEA-4 were weakly positive. In contrast, CD34, CD45, CD117, CD56, CD90, CD105, CD106, CD133, Flt-1, Musashi and HLA-DR were negative. Similarly, TRA-1-60 and SSEA-3 were negative. The in vitro transformation assay revealed that hAMC-SP cells did not form colonies in soft agarose cultures, whereas HepG2 cells, used as the positive controls of transformed cells, formed many colonies. Conclusion The cell surface marker studies suggest that hAMC-SP share characteristics with the stem cells obtained from other organs such as bone marrow mesenchymal cells and fat tissues. hAMC-SP cells rapidly and stably proliferate without transforming and they indeed differentiate into several cell types upon exposure to differentiation media. These results suggest that hAMC-SP, after proper depletion of HLA-ABC positive populations, can be used as ideal stem cells in a wide range of regenerative medicine over allogenicity because of their abundant presence. Key words: Amnion mesenchymal cells; Amnion stem cells; Surface marker; Regeneration medicine