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Mesenchymal stem cells (MSCs) exhibit immune-suppressive properties, follow a pattern of multilineage differentiation, and exhibit transdifferentiation potential. Ease in expansion from adult bone marrow, as well as its separation from ethical issues, makes MSCs appealing for clinical application. MSCs treated with retinoic acid resulted in synaptic transmission, based on immunostaining of synaptophysin and electrophysiological studies. In situ hybridization indicated that the neurotransmitter gene preprotachykinin-I was expressed in these cells. However, translation of this gene only occurred after stimulation with interleukin (IL)-1α. This effect was blunted by costimulation with IL-1 receptor antagonist. This study reports on the ability of MSCs to be transdifferentiated into neurons with functional synapses with the potential to become polarized towards producing specific neurotransmitters. Stem Cells 2005;23:383-391

BackgroundAn understanding of BC cell (BCC) entry into bone marrow (BM) at low tumor burden is limited when compared to highly metastatic events during heavy tumor burden. BCCs can achieve quiescence, without interfering with hematopoiesis. This occurs partly through the generation of gap junctions with BM stroma, located close to the endosteum. These events are partly mediated by the evolutionary conserved gene, Tac1.Methodogy/Principal FindingsThis study focuses on the role of mesenchymal stem cells (MSCs), Tac1, SDF-1 and CXCR4 in BCC entry into BM. The model is established in studies with low numbers of tumor cells, and focuses on cancer cells with low metastatic and invasion potential. This allowed us to recapitulate early event, and to study cancer cells with low invasive potential, even when they are part of larger numbers of highly metastatic cells. A novel migration assay showed a facilitating role of MSCs in BCC migration across BM endothelial cells. siRNA and ectopic expression studies showed a central role for Tac1 and secondary roles for SDF-1α and CXCR4. We also observed differences in the mechanisms between low invasive and highly metastatic cells. The in vitro studies were verified in xenogeneic mouse models that showed a preference for low invasive BCCs to BM, but comparable movement to lung and BM by highly metastatic BCCs. The expressions of Tac1 and production of SDF-1α were verified in primary BCCs from paired samples of BM aspirates and peripheral blood.Conclusions/SignificanceMSC facilitate BCC entry into BM, partly through Tac1-mediated regulation of SDF-1α and CXCR4. We propose a particular population of BCC with preference for BM could be isolated for characterization. This population might be the subset that enter BM at an early time period, and could be responsible for cancer resurgence and resistance to current therapies.

The generation of dopamine (DA) neurons from stem cells holds great promise in the treatment of Parkinson’s disease and other neural disease associated with dysfunction of DA neurons. Mesenchymal stem cells (MSCs) derived from the adult bone marrow show plasticity with regards to generating cells of other germ layers. In addition to reduced ethical concerns, MSCs could be transplanted across allogeneic barriers, making them desirable stem cells for clinical applications. We have reported on the generation of DA cells from human MSCs using sonic hedgehog (SHH), fibroblast growth factor 8 and basic fibroblast growth factor. Despite the secretion of DA, the cells did not show evidence of functional neurons, and were therefore designated DA progenitors. Here, we report on the role of brain‐derived neurotrophic factor (BDNF) in the maturation of the MSC‐derived DA progenitors. 9‐day induced MSCs show significant tropomyosin‐receptor‐kinase B expression, which correlate with its ligand, BDNF, being able to induce functional maturation. The latter was based on Ca2+ imaging analyses and electrophysiology. BDNF‐treated cells showed the following: increases in intracellular Ca2+ upon depolarization and after stimulation with the neurotransmitters acetylcholine and GABA and, post‐synaptic currents by electrophysiological analyses. In addition, BDNF induced increased DA release upon depolarization. Taken together, these results demonstrate the crucial role for BDNF in the functional maturation of MSC‐derived DA progenitors.

Breast cancer remains the most prevalent cancer among women in the United States. Substance P, a peptide derived from the TAC1 gene, mediates oncogenic properties in breast and other cancers. TAC1 expression facilitates the entry of breast cancer cells into bone marrow. The transcriptional repressor element 1-silencing transcription factor (REST) has been implicated in both oncogenic and tumor-suppressor functions. REST binds to the 5 untranslated region of the TAC1 promoter and suppresses its expression. This study investigated a role for REST in TAC1 induction in breast cancer. Western blots and real-time PCR indicated that REST expression in breast cancer cells was inversely proportional to the cells' aggressiveness, for both cell lines and primary breast cancer cells. REST knockdown in low-metastatic T47D cells and nontumorigenic MCF12A cells resulted in increases in TAC1 induction, proliferation, and migration. These parameters were negatively affected by ectopic expression of REST in highly aggressive MDA-MB-231 cells. Together, these findings show a central role for REST in the oncogenic function of TAC1 and suggest a tumor-suppressor role for REST in breast cancer.REST ͉ substance P ͉ tachykinin D espite advances in diagnostic technologies and treatments, breast cancer (BC) continues to be among the leading causes of cancer-related deaths among women (1). The TAC1 gene has been implicated in the development of breast and other cancers (2-4). TAC1 (preprotachykinin-A) is a single-copy gene with 7 exons (5). Through alternative splicing and posttranslational modification, TAC1 produces peptides belonging to the tachykinin family (5-7). Substance P (SP) is the predominant peptide produced from TAC1 (8, 9).BC cells (BCCs) produce high levels of SP (6). SP binds with varying affinities to 3 G protein-coupled, 7-transmembrane receptors: neurokinin (NK)1, NK2, and NK3 (6, 10). NK1 and NK2 have been reported on BCCs. SP mediates BCC proliferation, imparts radiation resistance, protects from apoptosis, induces growth-and angiogenic-promoting factors, and facilitates BC metastasis to bone marrow (2,4,11,12). The truncated NK1 has also been linked to oncogenesis, partly through the induction of TAC1 (2).Stromal-derived factor-1␣, which is implicated in BC biology, regulates TAC1 expression (4, 13). Other cytokines also induce TAC1 expression in BCCs via autocrine stimulation (2, 14). RE-1 silencer of transcription (REST) represses TAC1 expression in mesenchymal stem cells (15). The link between REST and TAC1 could be relevant to disorders such as tumorigenesis and hippocampal seizure (16).REST, also known as neural restrictive silencing factor, is a zinc finger transcriptional repressor of neuronal genes in nonneuronal and immature neuronal cells (17). REST facilitates chromatin remodeling and inhibits gene expression by assembling a repressor complex (18), Sin3A/histone deacetylase, and co-REST at the N and C termini, respectively (18). REST has been implicated in both oncogenic and tumor-suppressor roles (19,20)....

The generation of dopamine (DA) neurons from stem cells holds great promise for future biomedical research and in the clinical treatment of neurodegenerative diseases, such as Parkinson's disease. Mesenchymal stem cells (MSCs) derived from the adult human bone marrow (BM) can be easily isolated and expanded in culture while maintaining their immense plasticity. Here, we describe a protocol to generate DA-producing cells from adult human MSCs using a cocktail that includes sonic hedgehog (SHH), fibroblast growth factor 8 (FGF8), and basic fibroblast growth factor (bFGF). Electrophysiological functional DA neurons could be achieved by further treatment with brain-derived neurotrophic factor (BDNF). In summary, a protocol is described for the induction of primary BM-derived human MSCs to specific transdifferentiation; in this case, functional DA neurons. The MSC-derived DA cells express DA-specific markers, synthesize, and secrete dopamine. The described method could be used to generate DA cells for various model systems in which DA-producing cells are implicated in pathophysiological conditions.

Mesenchymal stem cells (MSCs) are located in postnatal bone marrow, show plasticity, are linked to various bone marrow disorders, exhibit phagocytosis, exert Ag-presenting properties (APC), and are immune suppressive. Unlike professional APCs, MSCs respond bimodally to IFN-γ in MHC-II expression, with expression at 10 U/ml and baseline, and down-regulation at 100 U/ml. The effects at high IFN-γ could not be explained by down-regulation of its receptor, IFN-γRI. In this study, we report on the mechanisms by which IFN-γ regulates MHC-II expression in MSCs. Gel shift assay and Western blot analyses showed dose-dependent increases in activated STAT-1, indicating responsiveness by IFN-γRI. Western blots showed decreased intracellular MHC-II, which could not be explained by decreased transcription of the master regulator CIITA, based on RT-PCR and in situ immunofluorescence. Reporter gene assays with PIII and PIV CIITA promoters indicate constitutive expression of PIII in MSCs and a switch to PIV by IFN-γ, indicating the presence of factors for effect promoter responses. We explained decreased MHC-II at the level of transcription because CIITA protein was observed in the cytosol and not in nuclei at high IFN-γ level. The proline/serine/threonine region of CIITA showed significant decrease in phosphorylation at high IFN-γ levels. An understanding of the bimodal effects could provide insights on bone marrow homeostasis, which could be extrapolated to MSC dysfunction in hematological disorders.

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