Role of Pertussis Toxin-sensitive G Protein-coupled Receptor Signaling in the Proliferation of Bone Marrow Mesenchymal Stem Cells

Background: Bone marrow (BM) mesenchymal stem cells (MSCs) can be expanded over 20～30 cell doublings in vitro even in the absence of any growth factors. However, the mechanisms that govern MSC proliferation are not well understood. Methods: We investigated the role of signaling of the pertussis toxin (PTX)-sensitive G protein-coupled receptor in the proliferation of BM MSCs. Results: PTX inhibited the proliferation of human BM MSCs and murine BM stromal MS-5 cells in a dose-dependent manner. Among the chemokines produced by the BM stromal cells, stromal cell-derived factor-1 (SDF-1) enhanced the proliferation of BM MSCs, while MIP-1α, MCP-3 or RANTES did not. PTX also inhibited the proliferation of some fibroblasts, such as MRC-5 and NIH-3T3, but did not affect the proliferation of HeLa and HSF cells. HSF cells did not express CXCR4 mRNA, but did produce SDF-1. In contrast, HeLa cells expressed CXCR4 strongly on the cell surface, but did not produce SDF-1. BM MSCs, MS-5, MRC-5, and NIH-3T3 cells all expressed CXCR4 minimally on the cell surface. These cells, however, had abundant CXCR4 protein in their cytoplasm, which was demonstrated by flow cytometric analysis performed after permeabilization of the cells. In addition, an ELISA performed on the culture supernatants of the cells revealed that these cells constitutively produce and secrete SDF-1. Conclusion: These results indicate that the signaling through the PTX-sensitive G protein-coupled receptor, which is induced by autocrine factors, plays an important role in the proliferation of BM MSCs and in some fibroblasts, and that SDF-1 is the most probable candidate for the autocrine growth factor.