Efficient Derivation and Concise Gene Expression Profiling of Human Embryonic Stem Cell-Derived Mesenchymal Progenitors (EMPs)

Department of Primary Care Medicine & Department of Obstetrics/Gynecology, National Taiwan University Hospital & College of Medicine, National Taiwan University, Taipei, Taiwan
Cell Transplantation (Impact Factor: 4.42). 09/2011; 20(10):1529-1545. DOI: 10.3727/096368910X564067

ABSTRACT New potential sources of stem cells for clinical application include bone marrow mesenchymal stem cells (BMMSCs), human embryonic stem cells (hESCs), and induced pluripotent stem cells (iPS). However, each source is not without its own concerns. While research continues in an effort to overcome these problems, the generation of mesenchymal progenitors from existing hESC lines may circumvent many of these issues. We report here a simple and efficient method of generating hESC-derived mesenchymal progenitors (EMPs) and transcriptome profiling using a concise, custom-designed, oligomnucleotide gene expression microarray. Characterization of EMPs shows that these cells are similar to BMMSCs in terms of differentiation capacity as well as cell surface marker expression. In addition, EMPs express several ESC markers and HLA-G, a nonclassical MHC class I molecule with immunomodulatory properties. Morevoer, EMPs possess significantly enhanced proliferative ability over BMMSCs during which karyotypic stability was maintained. Although derived from hESCs, EMPs do not form any tumors in immunocompromised mice. To efficiently profile gene expression in multiple samples, we designed an oligoarray to probe just over 11,000 genes highly expressed in stem cells. We found that the transcriptome of EMPs is more similar to BMMSCs than hESCs. Both cell types highly express genes involved in processes related to the cytoskeleton, extracellular matrix, and cell adhesion, but EMPs show higher expression of genes involved in cell proliferation whereas BMMSCs showed higher expression of immune-related genes. Based on our data, EMPs may be an accessible source of mesenchymal progenitor for therapeutic use.

1 Bookmark
  • [Show abstract] [Hide abstract]
    ABSTRACT: Mesenchymal stem cells (MSCs) have a high potential for therapeutic efficacy in treating diverse musculoskeletal injuries and cardiovascular diseases, and ameliorating the severity of graft-versus-host and autoimmune diseases. While most of these clinical applications require substantial cell quantities, the number of MSCs that can be obtained initially from a single donor is limited. Reports on the derivation of MSC-like cells from pluripotent stem cells (PSCs) are thus of interest, since the infinite proliferative capacity of PSCs opens the possibility to generate large amounts of uniform batches of MSCs. However, characterization of such MSC-like cells is currently inadequate, especially concerning the question of whether these cells are equivalent or identical to MSCs. In this study, we have derived MSC-like cells (iMPCs) using four different methodologies from a newly established induced PSC line reprogrammed from human bone marrow stromal cells (BMSCs), and compared the iMPCs directly to the originating parental BMSCs. The iMPCs exhibited typical MSC/fibroblastic morphology and MSC-typical surface marker profile, and were capable of differentiation in vitro along the osteogenic, chondrogenic, and adipogenic lineages. However, compared to the parental BMSCs, iMPCs displayed a unique expression pattern of mesenchymal and pluripotency genes and were less responsive to traditional BMSC differentiation protocols. We therefore conclude that iMPCs generated from PSCs via spontaneous differentiation represent a distinct population of cells that comprise MSC-like characteristics.
    Stem cells and development 03/2014; · 4.15 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Skeletal muscle (SkM) comprise approximately 40% of human body weight. Injury or damage to this important tissue can result in physical disability, and in severe cases is difficult for its endogenous stem cell-the satellite cell-to reverse effectively. Mesenchymal stem cells (MSC) are post-natal progenitor/stem cells that possess multilineage mesodermal differentiation capacity, including towards SkM. Adult bone marrow (BM) is the best-studied source of MSCs; however, aging also decreases BMMSC numbers and can adversely affect differentiation capacity. Therefore, we asked whether human sources of developmentally early-stage mesenchymal stem cells (hDE-MSCs) isolated from embryonic stem cells, fetal bone, and term placenta could be cellular sources for SkM repair. Under standard muscle-inducing conditions, hDE-MPCs differentiate towards a SkM lineage rather than cardiomyocytic or smooth muscle lineages, as evidenced by increased expression of SkM-associated markers and in vitro myotube formation. In vivo transplantation revealed that SkM-differentiated hDE-MSCs can incorporate into host SkM tissue efficiently in a mouse model of SkM injury. In contrast, adult BMMSCs do not express SkM-associated genes after in vitro SkM differentiation nor engraft in vivo. Further investigation of possible factors responsible for this difference in SkM differentiation potential revealed that, compared to adult BMMSCs, hDE-MSCs expressed higher levels of serum response factor (SRF), a transcription factor critical for SkM lineage commitment. Moreover, knockdown of SRF in hDE-MSCs resulted in decreased expression of SkM-related genes after in vitro differentiation and decreased in vivo engraftment. Our results implicate SRF as a key factor in age-related SkM differentiation capacity of MSCs, and demonstrate that hDE-MSCs are possible candidates for SkM repair.
    Stem cells and development 02/2014; · 4.15 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Embryonic stem cells (ESCs) and mesenchymal stem cells (MSCs) have been studied for years as primary cell sources for regenerative biology and medicine. MSCs have been derived from cell and tissue sources, such as bone marrow (BM), and more recently from ESCs. This study investigated MSCs derived from BM, H1- and H9-ESC lines in terms of morphology, surface marker and growth factor receptor expression, proliferative capability, modulation of immune cell growth and multipotency, in order to evaluate ESC-MSCs as a cell source for potential regenerative applications. The results showed that ESC-MSCs exhibited spindle-shaped morphology similar to BM-MSCs but of various sizes, and flow cytometric immunophenotyping revealed expression of characteristic MSC surface markers on all tested cell lines except H9-derived MSCs. Differences in growth factor receptor expression were also shown between cell lines. In addition, ESC-MSCs showed greater capabilities for cell proliferation, and suppression of leukocyte growth compared to BM-MSCs. Using standard protocols, induction of ESC-MSC differentiation along the adipogenic, osteogenic, or chondrogenic lineages was less effective compared to that of BM-MSCs. By adding bone morphogenetic protein 7 (BMP7) into transforming growth factor beta 1 (TGFβ1)-supplemented induction medium, chondrogenesis of ESC-MSCs was significantly enhanced. Our findings suggest that ESC-MSCs and BM-MSCs show differences in their surface marker profiles and the capacities of proliferation, immunomodulation, and most importantly multi-lineage differentiation. Using modified chondrogenic medium with BMP7 and TGFβ1, H1-MSCs can be effectively induced as BM-MSCs for chondrogenesis.
    Cell and Tissue Research 06/2014; · 3.68 Impact Factor