Abstract
Bone marrow stromal cell cultures contain multipotent cells that may have therapeutic utility for tissue restoration; however, the identity of the cell that maintains this function remains poorly characterized. We have utilized a unique model of murine bone marrow stroma in combination with liquid chromatography mass spectrometry to compare the nuclear, cytoplasmic and membrane associated proteomes of multipotent (MSC) (CD105+) and non-multipotent (CD105−) stromal cells. Among the 25 most reliably identified proteins, 10 were verified by both real-time PCR and Western Blot to be highly enriched, in CD105+ cells and were members of distinct biological pathways and functional networks. Five of these proteins were also identified as potentially expressed in human MSC derived from both standard and serum free human stromal cultures. The quantitative amount of each protein identified in human stromal cells was only minimally affected by media conditions but varied highly between bone marrow donors. This study provides further evidence of heterogeneity among cultured bone marrow stromal cells and identifies potential candidate proteins that may prove useful for identifying and quantifying both murine and human MSC in vitro.
Highlights
Bone marrow contains several different cell types including the precursors of blood, bone, fat and connective tissue
As mass spectrometry has proven a reliable method for analyzing cellular proteins [23,25,26], we utilized LC MS/MS to compare the proteome of Fluorescence Activated Cell Sorting (FACS) purified CD105+ and CD1052 stroma
Stromal cell populations were isolated to a purity of $95% and tested in differentiation culture assays to ensure that the multipotent potential of CD105+ and CD1052 fractions was maintained (Figure 1A)
Summary
Bone marrow contains several different cell types including the precursors of blood, bone, fat and connective tissue. It was shown that a monolayer of adherent fibroblast-like cells could be propagated in vitro by culturing whole bone marrow cells for several weeks in serum containing media [1]. These cells were found to support the growth of hematopoietic progenitors and could differentiate into fat (adipocytes), bone (osteocytes) and cartilage (chondrocytes) precursors both in vitro and in vivo [2,3]. MSC cultures have immense therapeutic potential and are currently being tested in clinical trials for the treatment of cartilage degeneration, myocardial infarction, diabetes, graft versus host disease and neurodegenerative disorders, among others [8,9,10,13,14,15,16,17]
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