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  • buy AR-42 inhibitor br Experimental Procedures br Author Con

    2018-10-20


    Experimental Procedures
    Author Contributions
    Acknowledgments
    Introduction Bone marrow stromal cells (BMSCs, also called bone-marrow-derived mesenchymal stromal cells) are heterogeneous populations that likely contain varying levels of tripotent (osteogenic, adipogenic, and chondrogenic [OAC]) stem-cell-like cells; cells with restricted potency (bi-, uni-, and nullipotent), committed precursors, and other stromal cell types. Phenotypic variations probably reflect in vivo functional diversity and a biological requirement for distinct stromal subsets with specific roles in bone marrow maintenance. Single-cell-derived BMSC clone analysis has identified considerable variation in differentiation capacity, ranging from OAC tripotency to nullipotency in vitro (Muraglia et al., 2000; Okamoto et al., 2002; Russell et al., 2010, 2011) and in vivo (Kuznetsov et al., 1997), which may indicate the existence of BMSC subtypes with varied potencies and/or a hierarchical developmental progression. BMSCs also possess significant immunomodulatory characteristics and can influence all aspects of immune cell function via cell-cell interaction and immunoregulatory factor secretion (Nauta and Fibbe, 2007), although clear demarcation between the skeletogenic and immunomodulatory capacity of BMSCs has not been made. Identification of human BMSCs often relies on non-discriminatory epitope detection (such as CD29, CD44, CD73, CD90 [THY-1], CD105, CD106 [VCAM-1], and CD166) with lack of hematopoietic markers (such as CD34, CD14, and CD45) (Dominici et al., 2006). Additional BMSC surface buy AR-42 inhibitor have been described, including STRO-1 (Simmons and Torok-Storb, 1991), CD146 (MCAM) (Sacchetti et al., 2007), CD271 (LNGFR) (Quirici et al., 2002), Nestin (Méndez-Ferrer et al., 2010), platelet-derived growth factor receptor alpha (PDGFRα)/CD51 (Pinho et al., 2013), LNGFR+THY-1+VCAM-1hi+ (Mabuchi et al., 2013), and in mice, leptin receptor (LepR/CD295) (Zhou et al., 2014). However, cell-sorting experiments using these markers all show that they contain the colony-forming, differentiation-competent BMSC population, demonstrating that other, as yet undefined BMSC subtypes exist and further resolution of BMSC heterogeneity is required. In vitro studies of BMSC functionality are hindered by their limited lifespan as replicative senescence occurs during culture, thus limiting the number and depth of studies that can be performed. To address these issues, we immortalized human BMSCs using human telomerase reverse transcriptase (hTERT), followed by clonal isolation to generate a panel of BMSC lines (hTERT-BMSCs). This strategy enabled in-depth, multiparameter analysis of variation in human BMSC subpopulations with different behavioral traits that subsequently could be examined in heterogeneous primary BMSCs. We unveil a range of biophysico-chemical markers for identification of different BMSC subsets with specific immunomodulatory and differentiation competencies.
    Results
    Discussion
    Experimental Procedures
    Author Contributions
    Acknowledgments
    Introduction Human mesenchymal stromal cells (MSCs) are being used in cell therapy to support organ regeneration after injury, e.g., by injection into the heart after myocardial infarction (Behfar et al., 2014). However, the outcomes of MSC therapy have been variable and the reasons for success or failure are a matter of ongoing debate (Behfar et al., 2014; Bianco et al., 2013). First, the potential of MSC therapy to support organ regeneration depends on the intrinsic character of the transplanted cell population, which is often ill-defined (Bianco et al., 2013; Mishra et al., 2009; Prockop et al., 2014). Second, engraftment success, survival, phenotype, and activity of MSCs strongly depend on the microenvironment present at the site of delivery (Forbes and Rosenthal, 2014). This microenvironment often shares features of a healing wound, including inflammatory cells, neo-vasculature, and pro-fibrotic cytokines such as TGF-β1 (Forbes and Rosenthal, 2014). Tissue repair and tumor microenvironment can convert MSCs into contractile myofibroblasts (MFs) that de novo form α-smooth muscle actin (α-SMA)-containing stress fibers (Hinz, 2010a; Hinz et al., 2012). The most prominent examples are “cancer-associated fibroblasts” (CAFs) which originate at least in part from bone marrow-derived MSCs (Karnoub et al., 2007; Mishra et al., 2009; Öhlund et al., 2014; Quante et al., 2011).