Membrane Type 1-Matrix Metalloproteinase Is Directly Involved in G-CSF Induced Human Hematopoietic Stem and Progenitor Cell Mobilization.

Abstract

Abstract G-CSF induced hematopoietic stem/progenitor cell (HSC/HPC) mobilization is regulated by a complex interplay between extracellular matrix (ECM), cytokines/chemokines, adhesion molecules, and proteases, which affect stem cell retention and proliferation in the bone marrow microenvironment. Several studies suggest that membrane type 1-matrix metalloproteinase (MT1-MMP), expressed on the surface of various cell types, is a key enzyme for normal cell motility and tumor cell migration and invasion. We found that enriched human CD34+ cells express various surface MT1-MMP levels, depending on the cell source and G-CSF treatment. CD34+ cells obtained from BM of healthy donors treated with G-CSF were found to have the highest mean fluorescence intensity (>900 arbitrary units), while level of expression was lower in CD34+ cells derived from G-CSF mobilized peripheral blood from healthy donors (159±40), human steady-state (SS) BM (80±19), and human cord blood (41±4). Following 48 hr incubation of human SS-BM CD34+ cells with G-CSF, the expression of MT1-MMP increased 2-fold compared to untreated cells, whereas treatment with other cytokines, such as SCF, SDF-1 and IL-6, had only a minimal impact. Immunocytochemical analysis of human cord blood CD34+ enriched cells plated on fibronectin or hyaluronate-coated cover slips revealed that in response to SDF-1, MT1-MMP changes its localization in the polarized and spreading cells, suggesting a role in the process of HPCs directional migration. Indeed, incubation with neutralizing antihuman MT1-MMP Ab, targeting its catalytic site, significantly reduced human progenitor migration towards a gradient of SDF-1 in transwells. Interestingly, low concentrations of tissue inhibitor of the metalloproteinase-2 (TIMP-2) enhanced SDF-1 induced transwell migration, while higher concentrations hampered this process. More importantly, 5 daily injections of G-CSF to NOD/SCID mice previously engrafted with human cells, up-regulated MT1-MMP expression on CD45+ and CD34+ human cells in the BM and peripheral blood of the mobilized chimeric mice compared to untreated, control chimeras. Treatment of chimera mice with antihuman MT1-MMP Ab on days 3–5 of G-CSF induced mobilization, significantly reduced the number of CD45+ and CD34+ human cells in peripheral blood. In summary, based on our data we suggest that following G-CSF treatment, increased levels of MT1-MMP on the surface of human progenitors in the BM facilitates their mobilization most probably due to pericellular ECM degradation and/or activation of other regulatory molecules, pointing to the essential role of MT1-MMP in G-CSF induced mobilization.