Growth Factors and Cytokines Upregulate Gelatinase Expression in Bone Marrow CD34+ Cells and Their Transmigration Through Reconstituted Basement Membrane

Abstract

The mechanism(s) underlying the release of stem/progenitor cells from bone marrow into the circulation is poorly understood. We hypothesized that matrix metalloproteinases (MMPs), especially gelatinases, which are believed to participate in the proteolysis of basement membranes and in the migration of leukocytes, may facilitate this process. First, we investigated whether CD34+stem/progenitor cells express gelatinases A (MMP-2) and/or B (MMP-9) and whether growth factors and cytokines (granulocyte colony-stimulating factor [G-CSF], granulocyte-macrophage colony-stimulating factor [GM-CSF], stem cell factor [SCF], macrophage colony-stimulating factor [M-CSF], interleukin-3 [IL-3], IL-6, IL-8, and tumor necrosis factor- [TNF-]) are able to modulate their expression. Next, we examined the transmigration of these stem/progenitor cells through reconstituted basement membrane (Matrigel) and its modulation by growth factors and cytokines. CD34+ cells were obtained from steady-state bone marrow and peripheral blood (from leukapheresis products collected either in steady-state hematopoiesis or after mobilization with G-CSF plus chemotherapy or G-CSF alone). We found that peripheral blood CD34+ cells, regardless of whether they were mobilized or not, strongly expressed both gelatinases (MMP-2 and MMP-9) in contrast to steady-state bone marrow CD34+ cells, which did not. However, all the growth factors and cytokines tested could induce MMP-2 and MMP-9 secretion by the latter cells. Moreover, the stimulatory effects of G-CSF and SCF on both MMP-2 and MMP-9 secretion were found to be significantly higher in CD34+ cells isolated from bone marrow than in those from peripheral blood. In addition TNF-, GM-CSF, and IL-6 increased the secretion of a partially active form of MMP-2. Basal transmigration of bone marrow CD34+ cells through Matrigel was lower than that of peripheral blood CD34+ cells (P < .0001), but growth factors and cytokines increased it by 50% to 150%. Positive correlations were established between expression of gelatinases and CD34+cell migration (r > .9). The stimulatory effect of G-CSF was significantly greater on the migration of CD34+ cells from bone marrow than on those from peripheral blood (P = .004). Moreover, CD34+ cell migration was reduced to approximately 50% by antibodies to MMP-2 and MMP-9, tissue inhibitors of metalloproteinases (rhTIMP-1 and -2), ando-phenanthroline. TNF-–induced gelatinase secretion and migration of CD34+ cells and of clonogenic progenitors (colony-forming unit–granulocyte-macrophage [CFU-GM], burst-forming unit-erythroid [BFU-E], colony-forming unit granulocyte, erythroid, monocyte, megakaryocyte [CFU-GEMM], and colony-forming unit-megakaryocyte [CFU-MK]) were dose-dependent. Therefore, this study demonstrated that CD34+ cells that are circulating in peripheral blood express both MMP-2 and MMP-9 and transmigrate through Matrigel. In contrast, CD34+ cells from steady-state bone marrow acquire similar properties after exposure to growth factors and cytokines, which upregulate expression of gelatinases and transmigration of these cells when they enter the bloodstream. Hence, we suggest that growth factors and cytokines induce release of stem/progenitor cells from bone marrow into peripheral blood during mobilization, as well as during steady-state hematopoiesis, by signaling through gelatinase pathways.

Stromal‐derived factor 1 and matrix metalloproteinase 9 levels in bone marrow and peripheral blood of patients mobilized by granulocyte colony‐stimulating factor and chemotherapy. Relationship with mobilizing capacity of haematopoietic progenitor cells

“…Finally, in contrast to IL-8, the activity of which is mainly restricted to mature granulocytes, haematopoietic growth factors (HGF) and/or chemotherapy can induce a wider range of cells able to produce MMP-9, including marrow stromal and CD34 + progenitor cells (Janowska-Wieczorek et al, 1999;Heissig et al, 2002) and monocytes (Zhang et al, 1998). The production of MMP-9 induced by stromal cells may have a pivotal role in the control of stem cells via the haematopoietic niche.…”

The role of complement in the trafficking of hematopoietic stem/progenitor cells

“…[44][45][46][47] It soon became evident that proteolytic enzymes capable of degrading ECM components could play a role in the migration of HSPCs. 48 It is now recognized that homing is a complex multistep process that involves signaling through adhesion molecules; chemotactic molecules; and their receptors, proteases, and other factors. In addition, GM-CSF, IL-3, and SCF, which activate VLA-4 and VLA-5 and increase the adhesiveness of HSPC, as well as Flt3ligand, SCF, IL-3, IL-6, and HGF, which in addition to prostaglandin E2 (PGE2), up regulate the expression of CXCR4 on HSPC, also affect the homing of HSPC.…”

CD34+ cell responsiveness to stromal cell–derived factor‐1α underlies rate of engraftment after peripheral blood stem cell transplantation

“…Another aliquot of LP was used to isolate CD34+ cells using immunomagnetic beads (Miltenyi-Biotec, Auburn, CA) according to the manufacturer's instructions and as previously described by us. 19 Briefly, light-density mononuclear cells were separated by density gradient centrifugation over 60 percent Percoll (GE Healthcare, Quebec, Canada), labeled with a CD34+ antibody (QBEND 10) and passed through a positive selection column. The isolated CD34+ cells were further analyzed for surface CXCR4 expression and in vitro migration (as described below).…”