Immortalized Multipotential Mesenchymal Cells and the Hematopoietic Microenvironment

Abstract

In an attempt to analyze the cellular and molecular basis of the capacity of bone marrow stromal cells to support hematopoiesis in culture, we developed a series of murine stromal cell lines from a single long-term bone marrow culture (BMC). The cytokines produced by these cells were analyzed using immunohistochemical techniques, ribonuclease protection assays (RPA) and RT-PCR. We examined the capacity of these cloned cell lines to replace primary bone marrow-derived stromal cells in long-term bone marrow cultures (LT-BMC) and sought correlations between the capacity to support hematopoiesis in culture with the production of known cytokines. These immortalized lines replicate many of the functions of the hematopoietic microenvironment. They express cytokines known to play a role in hematopoiesis. All of the lines constitutively express mRNA for PBSF (SDF-1), macrophage colony-stimulating factor (M-CSF), stem cell factor (SCF), FLT-3, thrombopoietin (TPO), interleukin 7 (IL-7), leukemia inhibitory factor (LIF), tumor necrosis factor-beta (TNF-beta), and interferon-gamma (IFN-gamma). Most lines also express granulocyte-macrophage colony-stimulating factor (GM-CSF) and G-CSF. They vary in their expression of IL-6, tumor growth factor-beta1 (TGF-beta1), TGF-beta2, and TNF-alpha. Growing these lines in the presence of cytokines that influence hematopoiesis alters the levels of cytokine message. The most striking effects were produced by TNF-alpha. In addition to the cytokine mRNAs, the cell lines express factors associated with bone formation such as osteoblast-specific factor-2 (OSF-2) and bone morphogenetic protein-1 (BMP-1). They also express the neural cell-adhesion molecule neuropilin and neurotrophic factors including nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF). Several of the lines can maintain hematopoiesis in culture, as measured by the continuous production of myeloid colony-forming cells (CFU-c), for months. This capacity to support hematopoiesis does not correlate with any pattern of cytokine expression. Several of these lines also support the growth of human hematopoietic cells, and human CFU-c can be detected in the cultures in which CD34(+) bone marrow cells (BMC) are cultured on murine stromal cells. No correlation between the production of any of the known cytokines and the ability to support murine hematopoiesis was detected. In addition, there was no correlation between the capacity to support murine hematopoiesis and the capacity to maintain human HSC. Despite repeated cloning, the lines remain heterogeneous and are capable of producing cells with the properties of fibroblasts, osteoblasts, adipocytes, and myoblasts. In addition to the cytokine mRNAs, the cell lines express factors associated with bone formation such as OSF-2 and BMP-1. They also express the neural cell-adhesion molecule neuropilin and neurotrophic factors including NGF and BDNF.

Axonal sprouting into the denervated spinal cord and synaptic and postsynaptic protein expression in the spinal cord after transplantation of bone marrow stromal cell in stroke rats

“…Related studies have indicated that functional recovery in rodent ischemic stroke models is associated with collateral sprouting of the uninjured CST to the denervated side of spinal cord and is enhanced by treatment of inosine , bFGF (Kawamata et al, 1997), Nogo antibody (Papadopoulos et al, 2002) and Nogo receptor antagonist (Lee et al, 2004). Based on our and other studies, the BMSCs transplanted into the ischemic brain environment may provide their beneficial effects by secretion of neurotrophins and growth factors, including BDNF (Dormady et al, 2001;Li et al, 2002) and VEGF (Chen et al, 2003a). BMSC treatment can also reduce the gliosis and scar formation in the ischemic brain (Li et al, 2005).…”

Multipotent neural precursors express neural and hematopoietic factors, and enhance ex vivo expansion of cord blood CD34+ cells, colony forming units and NOD/SCID-repopulating cells in contact and noncontact cultures

“…They are polypeptides noted for their neurotrophic activity in the peripheral and central nervous system and might also be implicated in hematopoietic cells. 33,34 IGF-1 is a mediator of the action of growth hormone on normal tissues and it performs multiple functions, including cell proliferation and inhibition of apoptosis in hematopoietic cells. 35 The similarity in the expression profiles of AGM cells and C17.2 is in line with recent proposals on the close relationship between hematopoiesis and neuropoiesis during embryonic development and in the manifestation of 'adult stem cell plasticity'.…”

Bone marrow-derived mesenchymal stromal cells for the repair of central nervous system injury

“…7 MSCs contain a population of cells that are self-renewing, and are capable of differentiating into multiple mesodermal tissues, including bone, cartilage, fat and muscle. 8 They produce many neurotrophic factors and cytokines, 9 can assume other phenotypes, suppress inflammation and trigger production of reparative growth factors as demonstrated after transplantation into other organ systems such as the lung 10 and heart. 11 Potential advantages of MSCs over other types of transplanted cells are shown in Table 1 which include their ability to be harvested from autologous donors, their relatively rapid expansion ex vivo and the possibility that allogeneic MSCs are nonimmunogenic and, hence, readily available for clinical use, 12 although some studies have indicated that immune suppression improves transplant survival.…”