Concerted Regulation of CD34 and CD105 Accompanies Mesenchymal Stromal Cell Derivation from Human Adventitial Stromal Cell

Mesenchymal stromal cells (MSC) have been intensively studied for innovative therapeutic applications. MSC in vitro are characterized by plastic-adherent proliferation, their specific immunophenotype and multipotency, whereas MSC progenitors in vivo are described as perivascular cells. Whether MSC progenitors acquire in vitro MSC characteristics upon in vitro culture is still unclear. This question can be experimentally accessed by analyzing changes in cellular properties that occur during the early in vitro culture phase, the MSC derivation phase. Here, we examined dynamics in morphology, proliferation, and expression of surface markers used for MSC characterization (such as CD34, CD105, CD146, and CD271) in tight kinetics during the MSC derivation phase of adipose tissue-derived MSC (AT-MSC). Using multiparametric flow cytometry, we identified 3 major ex vivo stromal vascular cell subsets: CD34(+)CD146(-)CD271(+/-) adventitial stromal cell-like cells (AdSC), CD34(-)CD146(+)CD271(+/-) pericyte-like cells (PC), and CD34(+)CD31(+)CD146(+) endothelial cells. Of these subsets, only AdSC, but not PC gave rise to MSC under MSC culture conditions. At day 4 of culture, AdSC became fibroblastoid and upregulated CD105, CD146, and CD271. Following this phenotypic transition, AdSC commenced proliferation and downregulated CD34. In our study, we demonstrate that AdSC are more clonogenic AT-MSC progenitors than PC. Moreover, we, for the first time have dissected the phenotypic transitions from MSC progenitors to in vitro MSC during the MSC derivation phase using multiparametric flow cytometry. Hence, we propose a model describing how de novo acquisition of the typical MSC morphology by AdSC is accompanied by concerted regulation of surface marker expression upon in vitro culture.