A comparative analysis of the adipogenic potential in human mesenchymal stromal cells from cord blood and other sources.

BACKGROUND AIMS: Mesenchymal stromal cells (MSCs) from umbilical cord blood (CB) attract attention by significantly impaired or absent adipogenic differentiation compared with MSCs derived from bone marrow (BM) and adipose tissue (AT). The diverging adipogenic propensity between the developmentally younger CB-MSCs and MSCs of the adult AT and BM resembles the age-dependent process in the BM, where adipose tissue increases with advancing age, accompanied by loss of bone stability. Thus, MSCs appeal as an attractive model to study the adipogenic process with respect to tissue sources and developmental ages. METHODS: We followed the expression of main adipogenic transcription factors, genes and protein markers in CB-, BM- and AT-MSCs under adipogenic induction, after silencing of preadipocyte factor 1 (Pref-1, PREF1) and after incubation with CB-plasma supplemented adipogenic media. RESULTS: An inverse relation in the expression of adipogenesis-associated markers and PREF1 in CB-MSCs suggested an inhibitory role of Pref-1 toward adipogenesis. However, Pref-1 protein was rarely detected in CB-MSCs, and siRNA silencing of Pref-1 failed to induce adipogenic differentiation in CB-MSCs. Thus, the impaired adipogenic differentiation of CB-MSCs in vitro was unrelated to endogenous Pref-1 protein expression. Nevertheless CB-plasma containing Pref-1 protein revealed an anti-adipogenic effect on AT-MSCs. CONCLUSIONS: Because Pref-1 is vastly abundant in CB-plasma and confers anti-adipogenic properties, Pref-1 in association with the ontogenic milieu probably induces long-lasting unresponsiveness toward adipogenic stimuli in CB-MSCs.

Altered gene expression in human adipose stem cells cultured with fetal bovine serum compared to human supplements.

Mesenchymal stromal cells (MSCs) are promising candidates for innovative cell therapeutic applications. For clinical scale manufacturing regulatory agencies recommend to replace fetal bovine serum (FBS) commonly used in MSC expansion media as soon as equivalent alternative supplements are available. We already demonstrated that pooled blood group AB human serum (HS) and thrombin-activated platelet releasate plasma (tPRP) support the expansion of multipotent adipose tissue-derived MSCs (ASCs). Slight differences in size, growth pattern and adhesion prompted us to investigate the level of equivalence by compiling the transcriptional profiles of ASCs cultivated in these supplements. A whole genome gene expression analysis was performed and data verified by polymerase chain reaction and protein analyses. Microarray-based screening of 34,039 genes revealed 102 genes differentially expressed in ASCs cultured with FBS compared to HS or tPRP supplements. A significantly higher expression in FBS cultures was found for 90 genes (fold change ≥2). Only 12 of the 102 genes showed a lower expression in FBS compared to HS or tPRP cultures (fold change ≤0.5). Differences between cells cultivated in HS and tPRP were hardly evident. Supporting previous observations of reduced adhesion of cells cultivated in the human alternatives we detected a number of adhesion and extracellular matrix-associated molecules expressed at lower levels in ASCs cultivated with human supplements. Confirmative assays analyzing transcript or protein expression with selected genes supported these results. Likewise a number of mesodermal differentiation-associated genes were higher expressed in cells grown in FBS. Quantifying adipogenic and osteogenic differentiation lacked to demonstrate a clear correlation to the supplement due to donor-specific variances. Our results emphasize the necessity of comparability studies as they indicate that FBS induces a culture adaptation exceeding that of ex vivo culture in human supplements and thus may contribute to the therapeutic potential.