| Summary: | 碩士 === 國立成功大學 === 生物科技研究所碩博士班 === 95 === Studies of mesenchymal stem cells from various human tissues have brought new promises in regenerative medicine. However, most isolation methods result heterogeneous cell population. From recent studies on stem cells, expressions of ABCG2 (ATP-binding cassette, group 2) had been the key feature associated with a more primitive characteristic in most stem cells, while expression of CD90 was considered as common stem marker associated with stem cell residency in vivo. Therefore, by using human placenta as our tissue source, we isolated ABCG2+/CD90+ cells and characterized these cells by immunohistochemistry, flow cytometry and differentiation potentials.
In our study, flow cytometric analysis of fresh-isolated placental cells showed moderate shift for most MSC markers, except ABCG2, CD73, CD90 and CD105. Expression of ABCG2 was less than 2% while expression of CD90 was between 3-35% with strong individual variation. Interestingly, cells positive with ABCG2 also co-expressed CD90 in most cases. In addition, expression of hematopoietic cell and embryonic stem cell markers demonstrated the heterogeneous nature of placental cells.
When cells were cultured with a low seeding density of 1x104 cells/cm2, colony formation appeared approximately 10-14 days and exhibited a typical fibroblast-like morphology. Colony efficiency was around 4-40 per 10000 cells depending on placenta samples. These colonies uniformly expressed typical MSC markers, such as CD29, CD44, CD90, CD105 and HLA-ABC, and were devoid of hematopoietic cell marker. In addition, CD13, C49a, and CD63, were also weakly expressed in these cells. Furthermore, expressions of MSC markers and differentiation potentials were found to increase through culture.
When analyzing these colony-forming cells, two subpopulations (ABCG2+/CD90+ and ABCG2-/CD90+) were found. No significant difference of MSC marker expressions was found in both subpopulations. Similar results were obtained when expression of ESC markers was analyzed. Both subpopulations expressed Oct-4, Sox-2, E-cadherin, TRA-1-61 and TRA-1-80, but not SSEA-3 and SSEA-4. However, when differentiation potentials were analyzed between these subpopulations, individual variations were found in each colony with no correlation to ABCG2 expression.
In our study, we demonstrated that side population phenotype could also be found in placental chorionic villi similar to hematopoietic stem cells. However, further studies are still required in order to reveal the particular role of ABCG2 in mesenchymal stem cells.
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