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Different culture conditions modulate the immunological properties of adipose stem cells.

ABSTRACT: The potential of human adipose stem cells (ASCs) for regenerative medicine has received recognition owing to their ease of isolation and their multilineage differentiation capacity. Additionally, low immunogenicity and immunosuppressive properties make them a relevant cell source when considering immunomodulation therapies and allogeneic stem cell treatments. In the current study, immunogenicity and immunosuppression of ASCs were determined through mixed lymphocyte reactions. The immunogenic response was analyzed after cell isolation and expansion in fetal bovine serum (FBS), human serum (HS)-supplemented medium, and xeno-free and serum-free (XF/SF) conditions. Additionally, the immunophenotype and the secretion of CXC chemokine ligand 8 (CXCL8), CXCL9, CXCL10, C-C chemokine ligand 2 (CCL2), CCL5, interleukin 2 (IL-2), IL-4, IL-6, IL-10, IL-17A, tumor necrosis factor-?, interferon-?, transforming growth factor-?1, indoleamine 2,3-deoxygenase, Galectin-1, and Galectin-3 were analyzed. The results showed that ASCs were weakly immunogenic when expanded in any of the three conditions. The significantly strongest suppression was observed with cells expanded in FBS conditions, whereas higher ASC numbers were required to display suppression in HS or XF/SF conditions. In addition, statistically significant differences in protein secretion were observed between direct versus indirect cocultures and between different culture conditions. The characteristic immunophenotype of ASCs was maintained in all conditions. However, in XF/SF conditions, a significantly lower expression of CD54 (intercellular adhesion molecule 1) and a higher expression of CD45 (lymphocyte common antigen) was observed at a low passage number. Although culture conditions have an effect on the immunogenicity, immunosuppression, and protein secretion profile of ASCs, our findings demonstrated that ASCs have low immunogenicity and promising immunosuppressive potential whether cultured in FBS, HS, or XF/SF conditions.

SUBMITTER: Patrikoski M

PROVIDER: S-EPMC4181391 | biostudies-literature | 2014 Oct

REPOSITORIES: biostudies-literature

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Different culture conditions modulate the immunological properties of adipose stem cells.

Patrikoski Mimmi M Sivula Jyrki J Huhtala Heini H Helminen Mika M Salo Fanny F Mannerström Bettina B Miettinen Susanna S

Stem cells translational medicine 20140813 10

The potential of human adipose stem cells (ASCs) for regenerative medicine has received recognition owing to their ease of isolation and their multilineage differentiation capacity. Additionally, low immunogenicity and immunosuppressive properties make them a relevant cell source when considering immunomodulation therapies and allogeneic stem cell treatments. In the current study, immunogenicity and immunosuppression of ASCs were determined through mixed lymphocyte reactions. The immunogenic res ...[more]

PMID: 25122689

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Project description:INTRODUCTION: Currently, human adipose stem cells (hASCs) are differentiated towards osteogenic lineages using culture medium supplemented with L-ascorbic acid 2-phosphate (AsA2-P), dexamethasone (Dex) and beta-glycerophosphate (?-GP). Because this osteogenic medium (OM1) was initially generated for the differentiation of bone marrow-derived mesenchymal stem cells, the component concentrations may not be optimal for the differentiation of hASCs. After preliminary screening, two efficient osteogenic media (OM2 and OM3) were chosen to be compared with the commonly used osteogenic medium (OM1). To further develop the culture conditions towards clinical usage, the osteo-inductive efficiencies of OM1, OM2 and OM3 were compared using human serum (HS)-based medium and a defined, xeno-free medium (RegES), with fetal bovine serum (FBS)-based medium serving as a control. METHODS: To compare the osteo-inductive efficiency of OM1, OM2 and OM3 in FBS-, HS- and RegES-based medium, the osteogenic differentiation was assessed by alkaline phosphatase (ALP) activity, mineralization, and expression of osteogenic marker genes (runx2A, DLX5, collagen type I, osteocalcin, and ALP). RESULTS: In HS-based medium, the ALP activity increased significantly by OM3, and mineralization was enhanced by both OM2 and OM3, which have high AsA2-P and low Dex concentrations. ALP activity and mineralization of hASCs was the weakest in FBS-based medium, with no significant differences between the OM compositions due to donor variation. However, the qRT-PCR data demonstrated significant upregulation of runx2A mRNA under osteogenic differentiation in FBS- and HS-based medium, particularly by OM3 under FBS conditions. Further, the expression of DLX5 was greatly stimulated by OM1 to 3 on day 7 when compared to control. The regulation of collagen type I, ALP, and osteocalcin mRNA was modest under induction by OM1 to 3. The RegES medium was found to support the proliferation and osteogenic differentiation of hASCs, but the composition of the RegES medium hindered the comparison of OM1, OM2 and OM3. CONCLUSIONS: Serum conditions affect hASC proliferation and differentiation significantly. The ALP activity and mineralization was the weakest in FBS-based medium, although osteogenic markers were upregulated on mRNA level. When comparing the OM composition, the commonly used OM1 was least effective. Accordingly, higher concentration of AsA2-P and lower concentration of Dex, as in OM2 and OM3, should be used for the osteogenic differentiation of hASCs in vitro.

Project description:Tendon resident cells (TCs) are a mixed population made of terminally differentiated tenocytes and tendon stem/progenitor cells (TSPCs). Since the enrichment of progenitors proportion could enhance the effectiveness of treatments based on these cell populations, the interest on the effect of culture conditions on the TSPCs is growing. In this study the clonal selection and the culture in presence or absence of basic fibroblast growth factor (bFGF) were used to assess their influences on the stemness properties and phenotype specific features of tendon cells.Cells cultured with the different methods were analyzed in terms of clonogenic and differentiation abilities, stem and tendon specific genes expression and immunophenotype at passage 2 and passage 4.The clonal selection allowed to isolate cells with a higher multi-differentiation potential, but at the same time a lower proliferation rate in comparison to the whole population. Moreover, the clones express a higher amounts of stemness marker OCT4 and tendon specific transcription factor Scleraxis (SCX) mRNA, but a lower level of decorin (DCN). On the other hand, the number of cells obtained by clonal selection was extremely low and most of the clones were unable to reach a high number of passages in cultures. The presence of bFGF influences TCs morphology, enhance their proliferation rate and reduce their clonogenic ability. Interestingly, the expression of CD54, a known mesenchymal stem cell marker, is reduced in presence of bFGF at early passages. Nevertheless, bFGF does not affect the chondrogenic and osteogenic potential of TCs and the expression of tendon specific markers, while it was able to downregulate the OCT4 expression.This study showed that clonal selection enhance progenitors content in TCs populations, but the extremely low number of cells produced with this method could represent an insurmountable obstacle to its application in clinical approaches. We observed that the addition of bFGF to the culture medium promotes the maintenance of a higher number of differentiated cells, reducing the proportion of progenitors within the whole population. Overall our findings demonstrated the importance of the use of specific culture protocols to obtain tendon cells for possible clinical applications.

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Different culture conditions modulate the immunological properties of adipose stem cells.

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Different culture conditions modulate the immunological properties of adipose stem cells.

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