Comprehensive Phenotypic Characterization of Human Adipose-Derived Stromal/Stem Cells and Their Subsets by a High Throughput Technology

Abstract

The characterization of adipose-derived stromal/stem cells (ASCs) remains difficult due to the lack of a definitive and unique cellular marker. Therefore, a combination of markers is necessary to identify the cells. No comprehensive analysis of the immunophenotype of expanded plastic adherent ASCs has been published. Therefore, the aim of this study was to characterize the general phenotype of cultured ASCs and to further analyze cellular subsets. ASCs were isolated from lipoaspirates from patients undergoing cosmetic liposuction and cultured in standard cell culture. A comprehensive phenotype characterization was done with the BD Lyoplate™ Human Cell Surface Marker Screening Panel containing 242 antibodies and isotype controls. Cultured ASCs not only showed the characteristic expression profile of mesenchymal stem cells (MSCs), but also revealed donor-specific variability in the expression of 49 other markers. We further detected markers with a scattering in the fluorescence intensity, indicating subpopulations with different expression profiles. Therefore, a multi-color flow cytometric analysis was done after staining the cells with direct-labeled antibodies against CD73, CD90, CD105, and either CD34, CD140b, CD200, CD201, or CD36 to verify the selected subpopulations of ASCs. We detected no CD34-CD36 double-positive population, but CD34(+)-CD36(-) and CD34(-)CD36(+) subpopulations, both of which are positive for the 3 main MSC markers, CD73, CD90, and CD105. All other detected subpopulations also co-expressed the 3 main MSC markers, and therefore fulfill the minimal phenotypic criteria for the definition of cultured MSCs. Our study demonstrates the first comprehensive phenotypic characterization of ASCs and clearly highlights donor-specific variability in ASC preparations.

High-Throughput Screening of Surface Marker Expression on Undifferentiated and Differentiated Human Adipose-Derived Stromal Cells

“…9 A recent study by Baer et al comprehensively characterized the surface marker profile of cultured hASCs isolated from 13 different female donors. 10 Our study both confirms and builds on these results by comprehensively characterizing the surface marker profiles of undifferentiated hASCs as well as hASCs that have undergone either osteogenic or adipogenic differentiation using lyoplates that contain 242 purified monoclonal antibodies and corresponding isotype controls. These lyoplates allow for a high-throughput screening of hundreds of surface markers by flow cytometry analysis.…”

Human cytomegalovirus infection of human adipose-derived stromal/stem cells restricts differentiation along the adipogenic lineage

“…24,[33][34][35] Although very similar, ASCs and hMSCs do display differences in regards to surface receptor repertoire and differentiation potential. 36,37 Additionally, these surface antigens can change within a cell type as a function of passage number and/or time in culture. We demonstrate that ASCs express multiple putative HCMV receptors including PDGFR, CD61 (integrin b3), CD29 (integrin b1) and EGFR (Fig.…”

Clinical immunology Comparison of phenotype, chondrogenic and osteogenic potential of rheumatoid mesenchymal stem cells derived from articular and subcutaneous adipose tissue - the role of adipocytokines

“…CD34, a classical haematopoietic and endothelial cells marker, was expressed at about 10% of analyzed cells. The presence of CD34+ cells among ASCs has been already reported elswhere [10][11][12][13].…”