Primary Marrow-Derived Stromal Cells: Isolation and Manipulation
Marrow stromal cells (MSCs) are relatively rare cells difficult to visualize in marrow biopsies or detect in aspirated marrow. Under specific conditions MSC can be expanded in vitro and the population can give rise to several mesenchymal lineages. “MSC” also refers to mesenchymal stem cells which implies that all cells in the population are multipotent. It is generally agreed that while there may be a few multipotent stem cells in an MSC population the majority are not stem cells. In either case MSCs do not produce hematopoietic cells. Although MSCs have been isolated and characterized from several tissues, bone marrow is their most common source for research and clinical use. Primary MSC populations can be derived from bone marrow mononuclear cells with relative ease, but it is important to recognize the cellular heterogeneity within a culture and how this may vary from donor to donor. In this chapter, we describe methodology to derive primary MSCs from bone marrow screens, an otherwise discarded by-product of bone marrow harvests used for clinical transplantation. We also describe some useful techniques to characterize and manipulate MSCs—both primary and immortalized cell lines.
Key wordsMarrow stromal cells (MSCs) Bone marrow screen Reverse-transfection FACS AutoMACS CD146 siRNA miRNA Long-term culture (LTC)
This work was supported in part by NIH grants DK073701, DK082757, HL104070, DK082783, HL099993, and DK056465, Bethesda, MD, USA.
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