Plastic adherent stromal cells from the bone marrow of commonly used strains of inbred mice: Variations in yield, growth, and differentiation

Abstract

Bone marrow stroma contains a unique cell population, referred to as marrow stromal cells (MSCs), capable of differentiating along multiple mesenchymal cell lineages. A standard liquid culture system has been developed to isolate MSCs from whole marrow by their adherence to plastic wherein the cells grow as clonal populations derived from a single precursor termed the colony-forming-unit fibroblast (CFU-F). Using this liquid culture system, we demonstrate that the relative abundance of MSCs in the bone marrow of five commonly used inbred strains of mice varies as much as 10-fold, and that the cells also exhibit markedly disparate levels of alkaline phosphatase expression, an early marker of osteoblast differentiation. For each strain examined, the method of isolating MSCs by plastic adherence yields a heterogeneous cell population. These plastic adherent cells also exhibit widely varying growth kinetics between the different strains. Importantly, of three inbred strains commonly used to prepare transgenic mice that we examined, only cells derived from FVB/N marrow readily expand in culture. Further analysis of cultures derived from FVB/N marrow showed that most plastic adherent cells express CD11b and CD45, epitopes of lymphohematopoietic cells. The later consists of both pre-B-cell progenitors, granulocytic and monocytic precursors, and macrophages. However, a subpopulation of the MSCs appear to represent bona fide mesenchymal progenitors, as cells can be induced to differentiate into osteoblasts and adipocytes after exposure to dexamethasone and into myoblasts after exposure to amphotericin B. Our results point to significant strain differences in the properties of MSCs and indicate that standard methods cannot be applied to murine bone marrow to isolate relatively pure populations of MSCs.

Critical role for Y1 receptors in mesenchymal progenitor cell differentiation and osteoblast activity

“…(17) ). However, it should be noted that BMSC cultures are heterogeneous and have been shown to include hematopoietic pre-B cells, granulocytic, and monocytic precursors, (18) as well as macrophages and endothelial cells. (19,20) It has been estimated that only 10% to 20% of the cells in a heterogeneous plasticadherent marrow cell culture are mesenchymal cells capable of differentiating into osteoblasts and adipocytes.…”

“…(19,20) It has been estimated that only 10% to 20% of the cells in a heterogeneous plasticadherent marrow cell culture are mesenchymal cells capable of differentiating into osteoblasts and adipocytes. (18) Several methods now have been developed to purify populations of MSCs from mouse bone marrow. Stem cell antigen 1 (Sca-1), a marker previously described on hematopoietic stem cells, also has been described on mesenchymal osteoprogenitor cells and is directly required for their self-renewal.…”

Comparative analysis of remyelinating potential of focal and intravenous administration of autologous bone marrow cells into the rat demyelinated spinal cord

“…Plastic-adherent cells from bone marrow are referred to as stromal cells (Friedenstein, 1976) and are thought to contain nonhematopoietic stem cells that can differentiate along multiple mesenchymal cell lineages (Prockop, 1997). The mononuclear cell fraction of bone marrow can be readily isolated by density gradient and contains stromal cells, hematopoietic and nonhematopoietic stem and precursor cells, and lymphocytes (Azizi et al, 1998;Phinney et al, 1999;Sasaki et al, 2001;Akiyama et al, 2002a). Transplantation of GFP-expressing mouse stromal cells by direct microinjection into the immunosuppressed rat indicates that these cells can form myelin in the demyelinated spinal cord (Akiyama et al, 2002b).…”

Effects of plating density and culture time on bone marrow stromal cell characteristics

“…Attempts have been made to classify these subpopulations by marker expression and by shape and growth kinetics, e.g., as spindle-shaped type I or flattened type II cells (19) or as rapidly self-renewing cells, or mature MSC (15,16,20). In addition to the observed heterogeneity in vitro, variations have been observed in MSC cultures obtained from different species as well as from different rodent strains or individual human donors (21)(22)(23). Some studies (23) found that variations even exist among different MSC donations obtained from the same human donor.…”