Dataset Information

Markers Are Shared Between Adipogenic and Osteogenic Differentiated Mesenchymal Stem Cells.

ABSTRACT: The stem cell differentiation paradigm is based on the progression of cells through generations of daughter cells that eventually become restricted and committed to one lineage resulting in fully differentiated cells. Herein, we report on the differentiation of adult human mesenchymal stem cells (hMSCs) towards adipogenic and osteogenic lineages using established protocols. Lineage specific geneswere evaluated by quantitative real-time PCR relative to two reference genes. The expression of osteoblast-associated genes (alkaline phosphatase, osteopontin, and osteocalcin)was detected in hMSCs that underwent adipogenesis. When normalized, the expression of adipocyte marker genes (adiponectin, fatty acid binding protein P4, and leptin) increasedin a time-dependent manner during adipogenic induction. Adiponectin and leptin were also detected in osteoblast-induced cells. Lipid vacuoles that represent the adipocyte phenotype were only present in the adipogenic induction group. Conforming to the heterogeneous nature of hMSCs and the known plasticity between osteogenic and adipogenic lineages, these data indicatea marker overlap between MSC-derived adipocytes and osteoblasts. Weproposea careful consideration of experimental conditions such as investigated timepoints, selected housekeeping genesand the evidence indicating lack of differentiation into other lineageswhen evaluating hMSC differentiation.

SUBMITTER: Kollmer M

PROVIDER: S-EPMC3765027 | biostudies-literature | 2013 May

REPOSITORIES: biostudies-literature

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Markers Are Shared Between Adipogenic and Osteogenic Differentiated Mesenchymal Stem Cells.

Köllmer Melanie M Buhrman Jason S JS Zhang Yu Y Gemeinhart Richard A RA

Journal of developmental biology and tissue engineering 20130501 2

The stem cell differentiation paradigm is based on the progression of cells through generations of daughter cells that eventually become restricted and committed to one lineage resulting in fully differentiated cells. Herein, we report on the differentiation of adult human mesenchymal stem cells (hMSCs) towards adipogenic and osteogenic lineages using established protocols. Lineage specific geneswere evaluated by quantitative real-time PCR relative to two reference genes. The expression of osteo ...[more]

PMID: 24013643

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Project description:Bone-marrow mesenchymal stem cells (BMSC) are considered the gold standard for use in tissue regeneration among mesenchymal stem cells (MSC). The abundance and ease of harvest make the adipose-derived stem cells (ASC) an attractive alternative to BMSC. The aim of the present study was to compare the transcriptome of ASC and BMSC, respectively isolated from subcutaneous adipose tissue and femur of 3 adult pigs, during in vitro osteogenic and adipogenic differentiation for up to four weeks. At 0, 2, 7, and 21 days of differentiation RNA was extracted for microarray analysis. A False Discovery Rate ?0.05 for overall interactions effect and P<0.001 between comparisons were used to determine differentially expressed genes (DEG). Ingenuity Pathway Analysis and DAVID performed the functional analysis of the DEG. Functional analysis of highest expressed genes in MSC and genes more expressed in MSC vs. fully differentiated tissues indicated low immunity and high angiogenic capacity. Only 64 genes were differentially expressed between ASC and BMSC before differentiation. The functional analysis uncovered a potential larger angiogenic, osteogenic, migration, and neurogenic capacity in BMSC and myogenic capacity in ASC. Less than 200 DEG were uncovered between ASC and BMSC during differentiation. Functional analysis also revealed an overall greater lipid metabolism in ASC, while BMSC had a greater cell growth and proliferation. The time course transcriptomic comparison between differentiation types uncovered <500 DEG necessary to determine cell fate. The functional analysis indicated that osteogenesis had a larger cell proliferation and cytoskeleton organization with a crucial role of G-proteins. Adipogenesis was driven by PPAR signaling and had greater angiogenesis, lipid metabolism, migration, and tumorigenesis capacity. Overall the data indicated that the transcriptome of the two MSC is relatively similar across the conditions studied. In addition, functional analysis data might indicate differences in therapeutic application.

Project description:BackgroundMesenchymal stem cells (MSCs) are promising targets for therapeutic use in regenerative medicine and tissue engineering. In the previous study, we have found that MSCs could be reverted to a primitive stem cell population after in vitro induction of osteogenic and de-osteogenic differentiation (de-osteogenic differentiated MSCs, De-Os-MSCs). De-Os-MSCs showed improved cell survival and osteogenic potential. However, the underlying mechanism and its potential effect on fracture healing has not been explored.MethodsMSCs were isolated from the rat bone marrow. MicroRNAs were cloned into lentiviral vectors and transduced into MSCs to observe the effects on osteogenesis. The expression levels of marker genes were evaluated by quantitative RT-PCR. Ectopic bone formation model was used to evaluate the bone regeneration ability of mir-92b transduced MSCs in vivo. An open femur fracture model was established, and MSCs or De-Os-MSCs were administrated to the fracture sites. Histological, biomechanical and microCT analysis were used to evaluate the quality of bone.ResultsIn the present study, we found that mir-92b was significantly increased in the secretions of De-Os-MSCs. And mir-92b could promote the osteogenic differentiation potential of MSCs by activating pERK and JNK signaling pathways. The ectopic bone formation assay showed that MSCs overexpressing mir-92b formed more bone like tissues in vivo. Most importantly, we found local administration of De-Os-MSCs could accelerate fracture healing using an open femur fracture model in rats. The quality of bone property was much better as shown by microCT and biomechanical testing.ConclusionTaken together, our study demonstrated that mir-92b promoted osteogenesis of MSCs, which was partially accounted for the enhanced osteogenic differentiation potential of De-Os-MSCs. And De-Os-MSCs had shown better regenerative capacity in accelerating fracture healing when they were locally given.The translational potential of this articleDe-Os-MSCs could be used to accelerate fracture healing, and reduce the occurrence of delayed unions and non-unions.

Project description:Understanding the molecular basis underlying the formation of bone-forming osteocytes and lipid-storing adipocytes will help provide insights into the cause of disorders originating in stem/progenitor cells and develop therapeutic treatments for bone- or adipose-related diseases. In this study, the role of RGS2 and RGS4, two members of the regulators of G protein signaling (RGS) family, was investigated during adipogenenic and osteogenenic differentiation of human mesenchymal stem cells (hMSCs).Expression of RGS2 and RGS4 were found to be inversely regulated during adipogenesis induced by dexamethasone (DEX) and 3-isobutyl-methylxanthine, regardless if insulin was present, with RGS2 up-regulated and RGS4 down-regulated in response to adipogenic induction. RGS2 expression was also up-regulated during osteogenesis at a level similar to that induced by treatment of DEX alone, a shared component of adipogenic and osteogenic differentiation inducing media, but significantly lower than the level induced by adipogenic inducing media. RGS4 expression was down-regulated during the first 48 h of osteogenesis but up-regulated afterwards, in both cases at levels similar to that induced by DEX alone. Expression knock-down using small interfering RNA against RGS2 resulted in decreased differentiation efficiency during both adipogenesis and osteogenesis. On the other hand, expression knock-down of RGS4 also resulted in decreased adipogenic differentiation but increased osteogenic differentiation.RGS2 and RGS4 are differentially regulated during adipogenic and osteogenic differentiation of hMSCs. In addition, both RGS2 and RGS4 play positive roles during adipogenesis but opposing roles during osteogenesis, with RGS2 as a positive regulator and RGS4 as a negative regulator. These results imply that members of RGS proteins may play multifaceted roles during human adipogenesis and osteogenesis to balance or counterbalance each other's function during those processes.

Dataset Information

Markers Are Shared Between Adipogenic and Osteogenic Differentiated Mesenchymal Stem Cells.

Publications

Markers Are Shared Between Adipogenic and Osteogenic Differentiated Mesenchymal Stem Cells.

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