Inducible Levels of Gelatinase B/Matrix Metalloproteinase-9 Gene Expression in Monocytes Are Associated with Marrow Cellularity in Myelodysplastic Syndrome (MDS).

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Evidence suggests that within the hematopoietic microenvironment (ME) stromal cell function can be modified by activities produced by monocytes/macrophages and that the reciprocal is also true; stroma can influence monocyte function. Critical regulatory molecules produced by stroma are often membrane bound until cleaved by metalloproteinases (MMP); cleavage can serve to either activate or inactivate their functions, making MMPs critical components of hematopoietic regulation. We report here that gene and protein expression of human matrix metalloproteinase-9 (MMP-9) is induced in monocytes in vitro by stromal cell conditioned media (CM). Briefly, flow sorted CD14+ peripheral blood monocytes were cultured for 5 days in the presence or absence of CM from HS-5 stromal cells, and MMP-9 gene expression was determined by real time PCR. Little or no MMP-9 gene expression was detected in CD14+ cells on day 0 prior to culture. In contrast, after 5 days of culture in control media MMP-9 gene expression was increased significantly (p=0.02). However culturing CD14+ cells in CM significantly increased expression another 3 fold (p<0.0001). MMP-9 protein secretion was also increased 12-fold after culture in CM. To identify which factors participate in the induction of MMP-9, the levels of MMP-9 mRNA were determined after CD14+ cells were cultured for 5 days in the presence or absence of 9 different recombinant factors. Factors chosen were those known to be present in CM and known to have their receptors expressed by monocytes. MMP-9 gene and protein expression increased 4 to 8-fold with 0.5ng/mL MCP-1/CCL2 or IL-1b, other factors tested had negligible effects. Immune cytochemical localization of MMP-9 protein in bone marrow biopsies from healthy donors and patients with myelodysplastic syndrome (MDS) indicated that mature myeloid cells including granulocytes and monocytes were stained strongly for MMP-9 protein, whereas stromal cells, fat cells, megakaryocytes, immature myeloid cells, blasts and cells of the erythroid lineage were negative. While levels of CM-induced MMP-9 gene expression among monocytes from 10 normal donors were relatively consistent, there was significant variation in inducible gene expression among CD14+ cells from 25 MDS patients (p= 0.02). Further analysis of MMP-9 gene expression in MDS mononuclear cells indicated that induced levels were negatively correlated with the degree of marrow cellularity (p=0.0002). Although marrow cellularity is a subjective estimate that can vary from one area of bone to another, the strong statistical correlation obtained suggests it may be related directly or indirectly to MMP-9 levels. In conclusion, monocytes can express and secrete MMP-9 in response to factors secreted by stromal cells. We hypothesize that the response of MDS monocytes to stromal signals can be abnormal resulting in unusually high or low levels of MMP-9, and that this may, directly or indirectly, influence marrow cellularity in MDS patients.