Abstract
BackgroundComplex immunophenotypic repertoires defining discrete adipose-derived stem cell (ASC) subpopulations may hold a key toward identifying predictors of clinical utility. To this end, we sorted out of the freshly established ASCs four subpopulations (SPs) according to a specific pattern of co-expression of six surface markers, the CD34, CD73, CD90, CD105, CD146, and CD271, using polychromatic flow cytometry.MethodUsing flow cytometry-associated cell sorting and analysis, gating parameters were set to select for a CD73+CD90+CD105+ phenotype plus one of the four following combinations, CD34−CD146−CD271− (SP1), CD34−CD146+CD271− (SP2), CD34+CD146+CD271− (SP3), and CD34−CD146+CD271+ (SP4). The SPs were expanded 700- to 1000-fold, and their surface repertoire, trilineage differentiation, and clonogenic potential, and the capacity to support wound healing were assayed.ResultsUpon culturing, the co-expression of major epitopes, the CD73, CD90, and CD105 was maintained, while regarding the minor markers, all SPs reverted to resemble the pre-sorted population with CD34−CD146−CD271− and CD34−CD146+CD271− representing the most prevalent combinations, followed by less frequent CD34+CD146−CD271− and CD34+CD146+CD271− variants. There was no difference in the efficiency of adipo-, osteo-, or chondrogenesis by cytochemistry and real-time RT-PCR or the CFU capacity between the individual SPs, however, the SP2CD73+90+105+34-146+271- outperformed others in terms of wound healing.ConclusionsOur study shows that ASCs upon culturing inherently maintain a stable distribution of immunophenotype variants, which may potentially disguise specific functional properties of particular downstream lines. Furthermore, the outlined approach suggests a paradigm whereby discrete subpopulations could be identified to provide for a therapeutically most relevant cell product.Electronic supplementary materialThe online version of this article (doi:10.1186/s13287-016-0435-8) contains supplementary material, which is available to authorized users.
Highlights
Complex immunophenotypic repertoires defining discrete adipose-derived stem cell (ASC) subpopulations may hold a key toward identifying predictors of clinical utility
Evolution of the subpopulation phenotype The cells expanded during P0 were sorted into four subpopulations (SP1–4) based on the co-expression of three major epitopes CD73, CD90, and CD105, and four different combinations of minor markers, including CD34 −CD146−CD271−, CD34−CD146+CD271−, CD34+ CD146 +CD271−, and CD34−CD146+CD271+ (Fig. 3)
Downstream lines from ASCs selected on the basis of specific surface repertoires have a tendency to assume upon culturing a phenotypical distribution reminiscent of the original population
Summary
Complex immunophenotypic repertoires defining discrete adipose-derived stem cell (ASC) subpopulations may hold a key toward identifying predictors of clinical utility. To this end, we sorted out of the freshly established ASCs four subpopulations (SPs) according to a specific pattern of co-expression of six surface markers, the CD34, CD73, CD90, CD105, CD146, and CD271, using polychromatic flow cytometry. Apart from the intracellular molecules, which are less suitable for prospective analysis of live subpopulations, such as Sox2 [17], Nanog [18], a whole array of surface markers, including CD117 [19], CD24, CD34, CD146, CD271 [20,21,22,23], CD133, CD200, CD362, Stro-1 [24], and SSEA4 [25] have been reported.
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