Adipogenic Differentiation of Human Adult Stem Cells From Bone Marrow Stroma (MSCs)

Abstract

We assayed gene expressions during adipogenesis of human MSCs. Microarray assays demonstrated time-dependent increases in expression of 67 genes, including 2 genes for transcription factors that were not previously shown to be expressed during adipogenesis.Introduction: Increased numbers of bone marrow adipocytes have been observed in patients with osteoporosis and aplastic anemia, but the pathological mechanisms remain unknown. Recently, microarray assays for mRNAs were used to follow adipogenic differentiation of the preadipocytic cell line, 3T3-L1, but adipogenic differentiation has not been examined in primary cells from bone marrow. Here we defined the sequence of gene expression during the adipogenesis ex vivo of human cells from bone marrow referred to as either mesenchymal stem cells or marrow stromal cells (MSCs). Materials and Methods: MSCs were plated at extremely low densities to generate single-cell derived colonies, and adipogenic differentiation of the colonies assayed by accumulation of fat vacuoles, time-lapse photomicroscopy, microarrays, and reverse transcriptase-polymerase chain reaction (RT-PCR) assays. Results and Conclusions: About 30% of the colonies differentiated to adipocytes in 14 days and about 60% in 21 days. Cell proliferation was inhibited by approximately 50% in adipogenic medium. The differentiation occurred primarily at the center of the colonies, and a few adipocytes that formed near the periphery migrated toward the centers. RT-PCR assays demonstrated that the differentiation was accompanied by increases in a series of genes previously shown to increase with adipogenic differentiation: peroxisome proliferator activated receptor ␥, CCAAT enhancer-binding protein ␣, acylCoA synthetase, lipoprotein lipase, and fatty acid binding protein 4. We also followed differentiation with microarray assays. Sixty-seven genes increased more than 10-fold at day 1 and 20-fold at day 7, 14, or 21. Many of the genes identified were previously shown to be expressed during adipocytic differentiation. However, others, such as zinc finger E-box binding protein and zinc finger protein 145, were not. This study should serve as a basis for future study to clarify the mechanisms of adipocyte differentiation of MSCs.

Functional expression of glucose-dependent insulinotropic polypeptide receptors is coupled to differentiation in a human adipocyte model

“…Differentiation studies in 3T3-L1 cells have reported that the expression of mRNA for FABP4 is absent in undifferentiated cells, is detectable by day 3 of differentiation and then rapidly increases towards a steady state around day 8 (several 100-fold increase in gene expression), for up to 28 days. 20,22,27 Similar profiles of FABP4 gene expression have been observed during the differentiation of human marrow and adipose tissue stromal cells into adipocytes, 28,29 as well as during the differentiation of SGBS cells. 16 We specifically analysed the expression profile of FABP4 during the differentiation of SGBS cells to relate GIPR expression to a well-established molecular differentiation marker of adipogenesis.…”

Osteogenic lineage commitment of mesenchymal stem cells from patients with ossification of the posterior longitudinal ligament

“…Cells from passage 2 were seeded at densities of 10,000 cells/cm 2 . Upon reaching the appropriate subconfluent level, cells were incubated in adipogenic induction medium (complete culture medium supplemented with 100 nM dexamethasone, 0.5 mM isobutylmethylxanthine and 100 μM indomethacin (Sigma-Aldrich)) for 21 days, as previously reported [14]. To evaluate lipid vesicle formation, cells were fixed with 4% formaldehyde and stained with 0.3% Oil Red O for 20 min.…”

Chondrogenic differentiation of bovine bone marrow mesenchymal stem cells (MSCs) in different hydrogels: Influence of collagen type II extracellular matrix on MSC chondrogenesis

“…Bone marrow mesenchymal stem cells (MSCs) are currently among the best characterized adult stem cells (Majumdar et al, 1998;Pittenger et al, 1999) isolated from various tissue sources such as fat, muscle, and bone (Jiang et al, 2002b;Sottile et al, 2002;Zuk et al, 2002). These cells are able to differentiate into bone, fat, cartilage, muscle tissue, and neurons (Mackay et al, 1998;Reyes et al, 2001;Sekiya et al, 2004;Woodbury et al, 2000) and were ultimately reported in almost all cell lineages (Jiang et al, 2002;Krause et al, 2001). For these reasons, the isolation and manipulation of adult stem cells represents a promising tool for understanding tissue development and regeneration, as well as for studying the engineered repair of tissues and organs (Orlic et al, 2001;Pittenger and Martin, 2004;Wakitani et al, 2002).…”