ABSTRACT
Breast cancer cells (BCCs) show preference for the bone marrow (BM). An animal model showed 2 populations of BCCs in the BM with regard to their cycling states. An in vitro model of early BC entry into BM showed normal hematopoiesis. Here, we show a critical role for BCC-derived SDF-1alpha in hematopoietic regulation. The studies used a coculture of BM stroma and BCCs (cell lines and stage II BCCs). Northern blots and enzyme-linked immunosorbent assay (ELISA) showed gradual decreases in SDF-1alpha production in BCCs as they contact BM stroma, indicating partial microenvironmental effects caused by stroma on the BCCs. SDF-1 knock-down BCCs and increased exogenous SDF-1alpha prevented contact inhibition between BCCs and BM stroma. Contact inhibition was restored with low SDF-1alpha levels. Long-term culture-initiating assays with CD34(+)/CD38(-)/Lin(-) showed normal hematopoiesis provided that SDF-1alpha levels were reduced in BCCs. Gap junctions (connexin-43 [CX-43]) were formed between BCCs and BM stroma, with concomitant interaction between CD34(+)/CD38(-)/Lin(-) and BM stroma but not with the neighboring BCCs. In summary, SDF-1alpha levels are reduced in BCCs that contact BM stroma. The low levels of SDF-1alpha in BCCs regulate interactions between BM stroma and hematopoietic progenitors, consequently facilitating normal hematopoiesis.
Subject(s)
Breast Neoplasms/pathology , Cell Communication , Chemokines, CXC/physiology , Hematopoiesis , Stromal Cells/pathology , Aged , Animals , Bone Marrow Cells/pathology , Breast Neoplasms/chemistry , Chemokine CXCL12 , Chemokines, CXC/biosynthesis , Chemokines, CXC/genetics , Coculture Techniques , Contact Inhibition , Female , Gap Junctions , Humans , Mice , Middle Aged , Neoplasm Proteins , RNA, Small Interfering/pharmacologyABSTRACT
The bone marrow (BM) is home to at least two stem cells, hematopoietic (HSC) and mesenchymal. Hematopoiesis is partly regulated through neurokinin-1 (NK-1) and NK-2 belonging to the family of G-protein/7-transmembrane receptors. NK-1 and NK-2 show preference for the neurotransmitters, substance P (SP) and neurokinin-A (NK-A), respectively. Hematopoietic suppression mediated by NK-A could be partly explained through the production of TGF-beta1 and MIP-1alpha. This study further characterizes mechanisms by which NK-A inhibits progenitor cell proliferation. The study addresses the hypothesis that p53 is a mediator of NK-A activation and this occurs partly through p53-mediated expression of NK-2. The studies first analyzed two consensus sequences for p53 in supershift assays. Reporter gene assays with NK-2 gene constructs and p53 expressing wild-type and mutant vectors, combined with cell proliferation assays, show NK-A activating p53 to inhibit the proliferation of K562 progenitors. These effects were reversed by hematopoietic stimulators, GM-CSF and SP. Verification studies with human CD34+/CD38- and CD34+/CD38+ BM progenitors show similar mechanisms with the expression of p21. This study reports on p53 as central to NK-A-NK-2 interaction in cell cycle quiescence of hematopoietic progenitors. These effects are reversed by at least two hematopoietic stimulators, SP and GM-CSF, with concomitant downregulation of p53.
