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Biomed Rep. 2017 Mar;6(3):300-306. doi: 10.3892/br.2017.845. Epub 2017 Jan 19.

Cell culture density affects the stemness gene expression of adipose tissue-derived mesenchymal stem cells.

Kim DS1, Lee MW1, Lee TH2, Sung KW1, Koo HH1,3,4, Yoo KH1,3,5.

Author information

1
Department of Pediatrics, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Republic of Korea.
2
Department of Laboratory of Cancer and Stem Cell Biology, Plant Engineering Institute, Sejong University, Seoul, Republic of Korea.
3
Stem cell and Regenerative Medicine Center, Research Institute for Future Medicine, Samsung Medical Center, Sungkyunkwan University, Seoul, Republic of Korea.
4
Department of Health Sciences and Technology, SAIHST, Sungkyunkwan University, Seoul, Republic of Korea.
5
Department of Medical Device Management and Research, SAIHST, Sungkyunkwan University, Seoul, Republic of Korea.

Abstract

The results of clinical trials using mesenchymal stem cells (MSCs) are controversial due to the heterogeneity of human MSCs and differences in culture conditions. In this regard, it is important to identify gene expression patterns according to culture conditions, and to determine how the cells are expanded and when they should be clinically used. In the current study, stemness gene expression was investigated in adipose tissue-derived MSCs (AT-MSCs) harvested following culture at different densities. AT-MSCs were plated at a density of 200 or 5,000 cells/cm2. After 7 days of culture, stemness gene expression was examined by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) analysis. The proliferation rate of AT-MSCs harvested at a low density (~50% confluent) was higher than that of AT-MSCs harvested at a high density (~90% confluent). Although there were differences in the expression levels of stemness gene, such as octamer-binding transcription factor 4, nanog homeobox (Nanog), SRY-box 2, Kruppel like factor 4, v-myc avian myelocytomatosis viral oncogene homolog (c-Myc), and lin-28 homolog A, in the AT-MSCs obtained from different donors, RT-qPCR analysis demonstrated differential gene expression patterns according to the cell culture density. Expression levels of stemness genes, particularly Nanog and c-Myc, were upregulated in AT-MSCs harvested at a low density (~50% confluent) in comparison to AT-MSCs from the same donor harvested at a high density (~90% confluent). These results imply that culture conditions, such as the cell density at harvesting, modulate the stemness gene expression and proliferation of MSCs.

KEYWORDS:

cell density; mesenchymal stem cell; stemness

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