Ex vivo expansion of adipose tissue‐derived stem cells in spinner flasks

Abstract

Recent reports indicate that adipose tissue is a novel source of multipotent stem cells that can be used in cell therapy and tissue engineering. However, using the traditional cultivation of adipose tissue-derived stem cells (ADSCs), it is hard to meet the needs of clinical applications. To obtain a large number of ADSCs while retaining their stemness, we seeded ADSCs in collagen/chitosan scaffolds and compared the proliferation of ADSCs in a 3-D static environment in dishes and a 3-D dynamic environment in spinner flask. The growth dynamic parameters of ADSCs were examined using a CCK-8 kit every other day, and the variations of glucose and lactic acid concentrations were analyzed every day. After 14 days, the cells were observed under a scanning electron microscope. The surface markers (CD29, CD34, CD44, CD45, CD73, CD105, CD166 and HLA-DR), the specific transcription factors (Nanog, Oct-4, Sox-2 and Rex-1) and the multi-differentiation potential (adipogenic, osteogenic and chondrogenic) were also assayed to identify the stemness of expanded cells. The results showed that the cells in scaffolds in spinner flask could be expanded by more than 26 times, and they presented better morphology and vitality and stronger differentiation ability than the cells cultivated in scaffolds statically. All the cells maintained stem cell characteristics after proliferation. Therefore, spinner flask cultivation is an easy-to-use, inexpensive system for expanding ADSCs in 3-D scaffolds.

The role of children's bone marrow mesenchymal stromal cells in the ex vivo expansion of autologous and allogeneic hematopoietic stem cells

“…Our results are in agreement to the one of Siegel et al () who recently reported the expression of Oct‐4, Nanog mRNA in BM‐MSCs to be much lower compared to pluripotent stem cells and Sox2 mRNA to be minimally detected in a number of BM‐MSC preparations. Similarly, Vicente López et al () and Zhu et al (), both working with human adipose‐derived stem cells (ASCs) though‐found: the former have no change on the expression of Nanog and Oct‐4 with low doses of bone morphogenetic protein (BMP) in their ex vivo expansion study and the latter additionally examined Sox‐2 and Rex‐1, which remained unaltered in a spinner flask cultivation.…”

The Necessity of a Systematic Approach for the Use of MSCs in the Clinical Setting

“…Therefore, due to the low number of cells and the invasive method required to isolate them, alternative sources of MSCs have been investigated. Cells with similar proprieties have been isolated from a broad range of tissues like skin [10, 11], peripheral blood [12], umbilical cord blood [13], muscle [14], adipose tissue [15], placenta [16, 17], dental pulp [18] or liver [19], and others. The differences in isolation protocols and tissues of origin lead to numerous definitions and use of various terms to refer to these cells such as mesenchymal stem cells, mesenchymal progenitor cells, or mesenchymal stromal cells.…”

Investigation of the Effective Action Distance Between Hematopoietic Stem/Progenitor Cells and Human Adipose-Derived Stem Cells During Their In Vitro Co-culture

“…Chondrogenic differentiation was induced by IMDM consisting of 0.1 μM dexamethasone, 50 μg/mL vitamin C, 100 μg/mL sodium pyruvate, 10 ng/mL transform growth factor (TFG-β3), 500 ng/mL bone morphogenetic protein (BMP-6), and 50 mg/mL ITS + , and detected by toluidine blue staining after 1, 2 and 3 weeks of induction, respectively. Adipogenic differentiation was induced by IMDM including 10% FBS, 0.1 mM 3-isobutyl-1-methyl xanthine, 1 μM dexamethasone, 0.2 μM indomethacin, and 10 μM insulin, and assayed with oil red staining after 1, 2 and 3 weeks of induction, respectively [15,16].…”