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Upregulation of Adipogenesis and Chondrogenesis in MSC Serum-Free Culture.

ABSTRACT: Serum-free media have been shown to be effective in the expansion of mesenchymal stem cells (MSCs). However, the effects may go beyond cell expansion as the differentiation potentials of the cells may be modified, thus influencing their efficacy for downstream applications. The latter is poorly understood, and this has prompted an evaluation of the influence of a serum-free formulation on the chondrogenic, adipogenic, and osteogenic potential of MSCs. The media consisted of Knockout™ Serum Replacement (KSR) with a cocktail of growth factors coupled with either collagen or fibronectin coatings. Collagen coating was selected as it promoted consistent cellular attachment. When compared against fetal bovine serum (FBS) controls, cell proliferation in the serum-free media was enhanced at passage 1. Similar levels of surface markers were observed in the two groups with a slight reduction in CD90 and CD73 in the serum-free culture at passage 3. The cultures were screened under differentiation conditions and a better maintenance of the chondrogenic potential was noted in the serum-free media with higher expressions of glycoaminoglycans (GAGs) and collagen II. Chondrogenesis was deficient in the FBS group and this was attributed to the inherent inconsistency of animal serum. Adipogenesis was enhanced in the serum-free group with a higher PPARG expression and lipid accumulation. Similar levels of osteogenic mineralization was noted in the FBS and serum-free groups but collagen I gene expression was suppressed in the latter. This was initially observed during expansion. These observations were attributed to the signaling cascades triggered by the cytokines presented in the serum-free formulation and the interaction with the collagen substrate. The serum-free media helps to maintain and enhance the chondrogenic and adipogenic potentials of the MSCs, respectively. This advantage can be exploited for therapeutic applications in cartilage and adipose tissue engineering.

SUBMITTER: Ho ST

PROVIDER: S-EPMC4789315 | biostudies-literature | 2011

REPOSITORIES: biostudies-literature

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Upregulation of Adipogenesis and Chondrogenesis in MSC Serum-Free Culture.

Ho Saey Tuan Barnabas ST Tanavde Vivek Madhukar VM Hui James Hoi JH Lee Eng Hin EH

Cell medicine 20110601 1

Serum-free media have been shown to be effective in the expansion of mesenchymal stem cells (MSCs). However, the effects may go beyond cell expansion as the differentiation potentials of the cells may be modified, thus influencing their efficacy for downstream applications. The latter is poorly understood, and this has prompted an evaluation of the influence of a serum-free formulation on the chondrogenic, adipogenic, and osteogenic potential of MSCs. The media consisted of Knockout™ Serum Repla ...[more]

PMID: 26998400

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Project description:BackgroundSynovial fluid (SF) mesenchymal stem cells (MSCs) are derived from the synovial membrane and have cartilage repair potential. Their current use in clinical practice is largely exploratory. As their numbers tend to be small, therapeutic procedures using MSCs typically require culture expansion. Previous reports indicate that the stem cell-mobilizing device (STEM device) intraoperatively increases SF-MSCs.PurposeThis study evaluated the chondrogenic potential of non-culture expanded synovium-mobilized MSCs and SF-microfragments obtained after enrichment using the STEM device and ascertained if device-mediated synovial membrane manipulation facilitated ongoing MSC release.Study designControlled laboratory study.MethodsTwo samples of aspiration fluid were collected intraoperatively before and after STEM device utilization from patients (n = 16) undergoing diagnostic or therapeutic knee arthroscopy. Human knee synovium (n = 5) was collected during total knee replacement, and a suspended culture was performed to assess the effect of the STEM device on ongoing MSC release. Colony forming unit-fibroblastic assays were used to determine the number of MSCs. Additionally, cytometric characterization of stromal and immune cells and chondrogenesis differentiation assay were performed without culture expansion. Filtered platelet concentrates were prepared using the HemaTrate system.ResultsAfter STEM device use, a significant increase was evident in SF-MSCs (P = .03) and synovial fluid-resident synovial tissue microfragments (P = .03). In vitro-suspended synovium released significantly more MSCs following STEM device use than nonstimulated synovium (P = .01). The STEM device-released total cellular fraction produced greater in vitro chondrogenesis with significantly more glycosaminoglycans (GAGs; P < .0001) when compared with non-STEM device synovial fluid material. Nonexpanded SF-MSCs and SF-microfragments combined with autologous filtered platelet concentrate produced significantly more GAGs than the complete chondrogenic media (P < .0001). The STEM device-mobilized cells contained more M2 macrophage cells and fewer M1 cells.ConclusionNon-culture expanded SF-MSCs and SF-microfragments had the potential to undergo chondrogenesis without culture expansion, which can be augmented using the STEM device with increased MSC release from manipulated synovium for several days. Although preliminary, these findings offer proof of concept toward manipulation of the knee joint environment to facilitate endogenous repair responses.Clinical relevanceAlthough numbers were small, this study highlights 3 factors relevant to 1-stage joint repair using the STEM device: increased SF-MSCs and SF-microfragments and prolonged synovial release of MSCs. Joint repair strategies involving endogenous MSCs for cartilage repair without the need for culture expansion in a 1-stage procedure may be possible.

Dataset Information

Upregulation of Adipogenesis and Chondrogenesis in MSC Serum-Free Culture.

Publications

Upregulation of Adipogenesis and Chondrogenesis in MSC Serum-Free Culture.

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