Identification of the role and the signal transduction of different chemokines in the process of myeloma cell homing to the extravascular compartment of the bone marrow

Project Details

Description

Multiple Myeloma (MM) is a malignant proliferation of plasma cells in the bone marrow (BM). The tumor cells secrete a monoclonal immunoglobulin (paraprotein) and induce osteolytic lesions. Homing from the vascular to the extravacular compartment of the BM is necessary for an efficient distribution of the tumor cells over the different BM sites. Homing is a transendothelial migration of lymphocytes and consists out of several steps. Different groups of molecules are involved in the process. One group is that of the chemokines, small molecules that mediate the migration of a cell after binding to the correct receptor. We use the 5TMM experimental mouse model, which shows large homology with human MM, to study the chemokines. Our group has already identified several chemokines and chemokinereceptors on the 5TMM cells. MCP-1, secreted by the BM endothelial cells is a chemokine that induces chemotaxis of the MM cells via the CCR2 receptor. Laminin-1, an element of the basement membrane attracts the cells by binding to the 67kD laminin receptor. IGF-1, secreted by the BM fibroblasts binds to the IGF-1 receptor and mediates chemotaxis.
In this work we will try to identify new chemokine receptors on the MM cells (Rnase protection assay) and identify the functional ligands for these receptors. We will perform functional tests with purified chemokines and blocking tests with antibodies.
We will investigate the role of SDF-1, secreted by BM stromal cells, in the process of transendothelial migration as its receptor CXCR4 is present on the MM cells.
We will also study the signal transduction of the different chemokines. Migration and invasion are complex actions that depend on coordinated activation of different signalling pathways. We will start with IGF-1 and investigate whether the chemotaxis of the 5TMM cells is mediated by the PI3K or the MAPK pathway. For this we will test the effect of inhibitors for the different kinases on the migration of the cells. In a next step we will look at IGF-1 mediated MMP9 production. There is evidence that IGF-1 regulates motility and MMP9 production through the same pathway. PI3K and MAPK activity will be studied through phosphorylation tests. The transcriptionfactors involved, can be studied in a later stage.
In a next step we will determine the cytoskeletal changes in the MM cells after stimulation with the different chemokines. This will be correlated with quantifying migration through classical migration tests. To study cytoskeletal changes, F-actin will be labelled with phalloidin-FITC and quantified through flow cytometric analysis or labelled with phalloidin-rhodamin and studied through microscopy.
Other roles for the chemokines, for example upregulating MMP’s, will be investigated through functional tests.
It will also be important in the process of homing to determine whether the cells migrate via a trans or interendothelial way. For this, sequences of ultra thin slices will be studied in the transmission electron microscope.
If we are able to identify new chemokine receptors, we will determine whether they are differential expressed on CD45+ and CD45- MM cells. This is important as the CD45+ cells have a higher migratory capacity.
AcronymFWOTM250
StatusFinished
Effective start/end date1/10/0130/09/05

Keywords

  • multiple myeloma
  • chemokines
  • homing

Flemish discipline codes

  • Agriculture, forestry, fisheries and allied sciences
  • Basic sciences
  • Biological sciences