Adipose-derived and bone marrow mesenchymal stem cells: a donor-matched comparison

Abstract

Background: Adipose-derived stem cells (ASCs) have been introduced as an alternative to bone marrow mesenchymal stem cells (BMSCs) for cell-based therapy. However, different studies comparing ASCs and BMSCs have shown conflicting results. In fact, harvesting ASCs and BMSCs from different individuals might influence the results, making comparison difficult. Therefore, this study aimed to characterize donor-matched ASCs and BMSCs in order to investigate proliferation, differentiation potential and possible effects of donor variation on these mesenchymal stem cells (MSCs).

Methods: Human bone marrow and adipose tissue samples were obtained from nine donors aged 8-14. ASCs and BMSCs were isolated and characterized based on expression of surface markers using flow cytometry. The proliferation up to 21 days was investigated. Multi-lineage differentiation was induced using osteogenic, chondrogenic and adipogenic differentiation media. Alkaline phosphatase (ALP) activity was monitored and collagen type I formation was evaluated by immunofluorescence staining. In vitro multi-potency was studied using tissue-specific stains and lineage-specific gene expression. In addition, the osteogenic lineage was evaluated at protein level.

Results: Isolated ASCs and BMSCs from all donors demonstrated morphologic and immunophenotypic characteristics of MSCs, with expression of MSCs markers and negative expression of hematopoietic markers. Unlike BMSCs, ASCs showed high expression of CD49d and low expression of Stro-1. In general, ASCs showed significantly higher proliferation and adipogenic capacity with more lipid vesicle formation and expression of the adipogenesis-related genes than BMSCs. In contrast, BMSCs showed significantly higher osteogenic and chondrogenic capacity compared to ASCs. BMSCs had earlier and higher ALP activity, calcium deposition, and expression of the osteogenesis- and chondrogenesis-related genes and the osteogenesis-related protein osteopontin. Proliferation and differentiation capacity of ASCs and BMSCs varied significantly among the donors.

Conclusions: ASCs and BMSCs showed tissue-specific differentiation abilities, but with significant variation between donors. The similarities and differences in the properties of ASCs and BMSCs should be taken into consideration when planning stem cell-based therapy.

Keywords: Adipose-derived stem cells; Bone marrow mesenchymal stem cells; Characterization; Differentiation; Proliferation.

Fig. 1

Morphology and immunophenotype characteristics of BMSCs and ASCs. a Representative microscopic illustrations of BMSCs and ASCs. Scale bar 100 μm. b Representative histograms from the flow cytometry analysis showing surface marker expression on BMSCs and ASCs, monoclonal antibody control (blue) and the stained cells (red). c and d Percentage of surface markers expression, average of nine donors (c) and in each donor (d). Each symbol represents one donor. *p < 0.05, **p < 0.01, ***p < 0.001. ASCs adipose-derived stem cells, BMSCs bone marrow-derived stem cells

Fig. 2

Proliferation of BMSCs and ASCs. a-c MTT assay at day 3, 7, 14 and 21, average of nine donors (a) and in each donor (b and c). d and e Relative gene expression of PCNA in BMSCs and ASCs at day 7, 14 and 21, average of nine donors (d) and in each donor (e). Each symbol represents one donor. **p < 0.01, ***p < 0.001. ASCs adipose-derived stem cells, BMSCs bone marrow-derived stem cells, MTT 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide, PCNA proliferating cell nuclear antigen

Fig. 3

ALP activity and collagen type I formation in BMSCs and ASCs. a Representative images of ALP staining of BMSCs and ASCs at day 3, 7 and 14. Scale bar 100 μm. b and c ALP activity assay of BMSCs and ASCs at day 14, average of nine donors (b) and in each donor (c). d Representative images of IF staining of collagen type I in BMSCs and ASCs at day 14. Scale bar 100 μm. Each symbol represents one donor. ALP alkaline phosphatase, ASCs adipose-derived stem cells, BMSCs bone marrow-derived stem cells, DAPI 4′,6-diamidino-2-phenylindole

Fig. 4

Osteogenic gene and protein expression, and calcium deposition in BMSCs and ASCs. a and b Relative gene expression of the osteogenic genes Runx2, collagen type I, ALP and osteopontin in BMSCs and ASCs at day 7, 14 and 21, average of nine donors (a) and in each donor (b). c Western blot analysis of osteopontin in BMSCs and ASCs at day 21. d Representative images after Alizarin red S staining of BMSCs and ASCs at day 21 and 28. Scale bar 100 μm. e and f Quantification of Alizarin red S staining of BMSCs and ASCs at day 21 and 28, average of nine donors (e) and in each donor (f). Each symbol represents one donor. **p < 0.01, ***p < 0.001. ALP alkaline phosphatase, ASCs adipose-derived stem cells, BMSCs bone marrow-derived stem cells, Runx2 runt-related transcription factor 2

Fig. 5

Chondrogenic gene expression and cartilaginous matrix formation in BMSCs and ASCs. a and b Relative gene expression of the chondrogenic gene aggrecan in BMSCs and ASCs at day 28, average of nine donors (a) and in each donor (b). c Representative images of Alcian blue-stained sections of BMSC and ASC pellets at day 28. Scale bar 50 μm. Each symbol represents one donor. ***p < 0.001. ASCs adipose-derived stem cells, BMSCs bone marrow-derived stem cells

Fig. 6

Adipogenic gene expression and lipid vesicle formation in BMSCs and ASCs. a and b Relative gene expression of the adipogenic genes LPL and PPARG in BMSCs and ASCs at day 14, average of nine donors (a) and in each donor (b). c Representative images of Oil red O stained BMSCs and ASCs at day 14. Scale bar 100 μm. d and e Quantification of Oil red O-stained BMSCs and ASCs at day 14, average of nine donors (d) and in each donor (e). Each symbol represents one donor. ***p < 0.001. ASCs adipose-derived stem cells, BMSCs bone marrow-derived stem cells, LPL lipoprotein lipase, PPARG peroxisome proliferator activated receptor gamma