Search

Research has provided evidence that tumor growth depends on the interaction of tumor cells with stromal cells, as already suggested in 1889 by Paget. Experimental and clinical studies have revealed that tumor stromal cells can be derived from bone marrow (BM)-derived progenitor cells, such as mesenchymal stern cells (MSCs), which can be mobilized into the circulation and incorporate into tumor micro-environments. Many observations indicate that, in the tumor micro-environment, MSCs have several tumor growth promoting functions, including expression of growth factors, promotion of tumor vessel formation and creation of tumor stem cell niches. This review will discuss the currently known tumor growth promoting BM-derived cells and focus on the role of MSCs in modulating tumor micro-environments. In addition, we will discuss the potential of inhibiting BM-derived cells and their utilization as cellular vehicles for selective delivery of cancer therapeutics as additional strategies in the treatment of cancer. (C) 2008 Elsevier Ireland Ltd. All rights reserved.

Stem cell factor (SCF) has a potent synergistic effect during megalkaryopoiesis when administered in combination with the major megalkaryocytic cytokine, thrombopoietin (TPO). In this study we analyzed the underlying mechanisms with regard to STAT5 activity. TPO stimulation of MO7e cells resulted in STAT5 transactivation, which could be enhanced 1.6-fold by costimulation with SCF, whereas SCF alone did not induce STAT5 transcriptional activity. This costimulatory effect of SCF was reflected in an increase in TPO-induced STAT5 DNA binding and increased and prolonged STAT5 tyrosine phosphorylation in both MO7e cells and primary human megakaryocyte progenitors. In contrast, serine phosphorylation of STAT5 was constitutive and associated with an inhibitory effect on STAT5 transactivation. Signal transduction pathways that might synergize in TPO-mediated STAT5 transactivation were analyzed using specific pharmacological inhibitors and indicated an essential role for Janus-activated kinase 2 (JAK2) and a partial role for Src-family kinases. Costimulation with SCF was found to increase and prolong tyrosine phosphorylation of JAK2 and the TPO receptor c-mpl. In addition, the Src kinase inhibitor SU6656 partially downregulated the additional effect of SCF costimulation on STAT5 tyrosine phosphorylation. SCF-induced enhancement of JAK2 phosphorylation was not affected by inhibition of Src kinase, suggesting that both JAK2 and Src kinase mediate STAT5 tyrosine phosphorylation. Synergistic activation of JAK2 and Src kinase may thus contribute to the enhanced STAT5 signaling in the presence of TPO and SCF.

Purpose: To test magnetic resonance (MR) contrast media of different molecular weights (MWs) for their potential to characterize noninvasively microvascular changes in an experimental tumor treatment model.Materials and Methods: MD-MBA-435, a poorly differentiated human breast cancer cell line, was implanted into 31 female homozygous athymic rats. Animals were assigned randomly to a control (saline) or drug treatment (monoclonal antibody vascular endothelial growth factor (MabVEGF) antibody) group. In both groups, dynamic MR imaging (MRI) was performed in each animal using up to three different contrast media on sequential days at baseline and follow-up examination. The MWs of the contrast media used ranged from 557 Da to 92 kDa. Using a bidirectional kinetic model, tumor microvessel characteristics, including the fractional plasma volume (fPV) and transendothelial permeability K-PS, were estimated for each contrast medium. These microvascular characteristics were compared between drug and control groups and between contrast media of different MWs.Results: Tumors grew significantly slower (P Conclusion: Therapeutic effects of a Mab-VEGF antibody on tumor microvessel characteristics can be monitored by dynamic MRI. Intermediate-size agents, such as Gadomer-17, offer a substantial dynamic range and are less limited by imaging precision and therefore should be considered a practical alternative to monitor antiangiogenesis treatment effects in a clinical setting.

The precise outgrowth and arborization of dendrites is crucial for their function as integrators of signals relayed from axons and, hence, the functioning of the brain. Proper dendritic differentiation is particularly resonant for Purkinje cells as the intrinsic activity of this cell-type is governed by functionally distinct regions of its dendritic tree. Activity-dependent mechanisms, driven by electrical signaling and trophic factors, account for the most active period of dendritogenesis. An as yet unexplored trophic modulator of Purkinje cell dendritic development is corticotropin-releasing factor (CRF) and family member, urocortin, both of which are localized in climbing fibers. Here, we use rat organotypic cerebellar slice cultures to investigate the roles of CRF and urocortin on Purkinje cell dendritic development. Intermittent exposure (12 h per day for 10 days in vitro) of CRF and urocortin induced significantly more dendritic outgrowth (45% and 70%, respectively) and elongation (25% and 15%, respectively) compared with untreated cells. Conversely, constant exposure to CRF and urocortin significantly inhibited dendritic outgrowth. The trophic effects of CRF and urocortin are mediated by the protein kinase A and mitogen-activating protein kinase pathways. The study shows unequivocally that CRF and urocortin are potent regulators of dendritic development. However, their stimulatory or inhibitory effects are dependent upon the degree of expression of these peptides. Furthermore, the effects of CRF and urocortin on neuronal differentiation and re-modeling may provide a cellular basis for pathologies such as major depression, which show perturbations in the expression of these stress peptides.

This is the first study that comprehensively describes the effects of the platelet-derived growth factor (PDGF) isoforms C and D during in vitro expansion of human mesenchymal stem cells (hMSCs). Our results show that PDGFs can enhance proliferation of hMSCs without affecting their multipotency. It is of great value to culture and expand hMSCs in a safe and effective manner without losing their multipotency for manipulation and further development of cell-based therapies. Moreover, differential effects of PDGF isoforms have been observed on lineage-specific differentiation induced by BMP2 and Vitamin D3. Based on label-free LC-based quantitative proteomics approach we have furthermore identified specific pathways induced by PDGFs during the proliferation process, showing the importance of bioinformatics tools to study cell function.

In order to study the development and functional properties of single, isolated, rat mesencephalic trigeminal neurones, a cell-culture procedure was developed for these specific primary sensory neurones. Mesencephalic trigeminal neurones were isolated from the brainstem of 16-day-old rat embryos. Various factors thought to promote the survival and growth of these neurones in vitro were examined. Outgrowth and maintenance of mesencephalic trigeminal neurones in vitro appeared to be stimulated by a muscle-derived factor, present in muscle-conditioned medium or in muscle extract. Of the neurotrophic factors examined, brain-derived neurotrophic factor and neurotrophin-3, but not nerve-growth factor, promoted the survival of rat mesencephalic trigeminal neurones. Optimal survival of these neurones was found to occur on a monolayer of astrocytes, an effect mediated through direct cell-to-cell interactions.

Intracerebroventricularly (icv) administered corticotropin-releasing hormone (CRH) produces a dose-dependent increase in heart rate in association with behavioral activation. The present study was designed to investigate whether these CRH-induced responses are dependent on adrenal function. The effects of adrenalectomy (ADX) and subsequent corticosterone replacement were studied. Administration icv of 300 ng of CRH failed to produce behavioral activation and tachycardia in ADX rats. Corticosterone replacement restored the CRH-induced behavioral response to preoperative levels, whereas the CRH-induced tachycardia was partially restored. This latter result may be related to the fact that the baseline heart rate of ADX animals appeared to be significantly higher than that of corticosterone-treated ADX animals. It is concluded that circulating adrenal corticosterone in ADX rats is involved in the expression of the behavioral and cardiac effect of central CRH.

Research has provided evidence that tumor growth depends on the interaction of tumor cells with stromal cells, as already suggested in 1889 by Paget. Experimental and clinical studies have revealed that tumor stromal cells can be derived from bone marrow (BM)-derived progenitor cells, such as mesenchymal stern cells (MSCs), which can be mobilized into the circulation and incorporate into tumor micro-environments. Many observations indicate that, in the tumor micro-environment, MSCs have several tumor growth promoting functions, including expression of growth factors, promotion of tumor vessel formation and creation of tumor stem cell niches. This review will discuss the currently known tumor growth promoting BM-derived cells and focus on the role of MSCs in modulating tumor micro-environments. In addition, we will discuss the potential of inhibiting BM-derived cells and their utilization as cellular vehicles for selective delivery of cancer therapeutics as additional strategies in the treatment of cancer. (C) 2008 Elsevier Ireland Ltd. All rights reserved.

Purpose: We hypothesized that downstream effects of endogenous vascular endothelial growth factor (VEGF)/VEGF receptor signaling on acute myelogenous leukemia (AML) cell survival resulted in increased in vitro cellular drug resistance and a longer time to kill most leukemic cells in vivo upon drug exposure. Experimental Design: In primary AML cells from pediatric patients, VEGFA and VEGFC mRNA expression and in vitro cellular resistance to nine cytotoxic drugs were studied. As in vivo equivalents for in vitro drug resistance, in vivo AML blast reduction upon drug exposure, measured as blast cell reduction on day 15 in the bone marrow and as time in days from diagnosis to complete remission (CR) were used. Results: Increased endogenous VEGFC levels significantly correlated with increased in vitro resistance for six typical AML drugs in primary AML cells from pediatric patients. Patients with >5% blasts on day 15 showed a 12.9-fold increase in the median VEGFC level compared with patients with ≤5% blasts (P = 0.002). Time to reach CR was studied using linear regression analysis with VEGFC, age at diagnosis, sex, treatment protocol, FAB type, cytogenetic risk profile, and WBC counts as variables. There was a significant positive independent association between VEGFC levels and time to CR (b = 6.02, SE = 1.58, P ≤ 0.0001, n = 72). Conclusions: These results suggest for the first time that higher endogenous VEGFC levels of AML cells are related to decreased in vitro and in vivo drug responsiveness.

Corticotropin releasing factor receptor 2 (CRF-R2) is strongly expressed in the cerebellum and plays an important role in the development of the cerebellar circuitry, particularly in the development of the dendritic trees and afferent input to Purkinje cells. However, the mechanisms responsible for the distribution and stabilization of CRF-R2 in the cerebellum are not well understood. Here, we provide the first detailed analysis of the cellular localization of the full-length form of CRF-R2 in rat cerebellum during early postnatal development. We document unique and developmentally regulated subcellular distributions of CRF-R2 in cerebellar cell types, e.g. granule cells after postnatal day 15. The presence of one or both receptor isoforms in the same cell may provide a molecular basis for distinct developmental processes. The full-length form of CRF-R2 may be involved in the regulation of the first stage of dendritic growth and at later stages in the controlling of the structural arrangement of immature cerebellar circuits and in the autoregulatory pathway of the cerebellum.

Objective. To define whether the observed synergistic effects of erythropoietin (EPO) and stem cell factor (SCF) on erythroid cells can, in part, be mediated by the signal transducer and activator of transcription 5 (STAT5).Methods. STAT5 activation was examined in erythroid cell lines by analyzing the effects of EPO and SCF on STAT5 tyrosine phosphorylation, serine phosphorylation, DNA binding, and STAT5-mediated gene transactivation.Results. EPO induced a 5.0-fold +/- 0.4-fold increase in STAT5 transactivation, which could be further enhanced by SCF. SCF pretreatment followed by EPO stimulation resulted in a 9.0-fold +/- 0.9-fold increase in STAT5 transactivation, while SCF alone did not increase STAT5 transactivation. This costimulatory effect of SCF was not mediated by increased STAT5 tyrosine or serine phosphorylation or increased STAT5 DNA binding. In addition, enhanced STAT5 transactivation was independent of the phosphatidyl inositol 3-kinase and MAPK(p42/p44) pathways. Instead, the protein kinase A (PKA) inhibitor protein PKI and the PKA inhibitor H89 prevented the costimulatory SCF effect. Furthermore, the PKA target CREB showed a strongly increased and prolonged serine-133 phosphorylation after costimulation with SCF + EPO. The involvement of CREB in STAT5 transactivation was demonstrated by overexpression of serine-133-mutated CREB, which completely blocked the SCF effect. In addition, the CREB-binding protein CBP/p300 was shown to be essential for EPO- and SCF-mediated STAT5 transactivation.Conclusion. SCF enhances the EPO-mediated STAT5 transactivation by triggering a PKA/CREB-dependent pathway. (C) 2003 International Society for Experimental Hematology. Published by Elsevier Inc.

In both larval and adult urodele amphibians, limb blastema formation requires the presence of an adequate nerve supply. In previous research, we demonstrated that the hindlimb of early Xenopus laevis larvae formed a regeneration blastema even when denervated, while the denervated limb of late larvae did not. We hypothesized that the nerve-independence was due to the autonomous synthesis of a mitogenic neurotrophic-like factor by undifferentiated limb bud cells. In this paper, we demonstrate that fgf-2 mRNA is present in larval limb tissues and that its level is correlated to the extent of mesenchymal cells populating the limb: in early limbs, fgf-2 mRNA is present at high levels all over the limb, while, in late limbs, the fgf-2 expression is low and detectable only in the distal autopodium. After denervation, fgf-2 mRNA synthesis increases in amputated early limbs but not in amputated late limbs. The implantation of anti-FGF-2 beads into amputated early limbs hardly lowers the mitotic activity of blastema cells. However, FGF-2 beads implanted into the blastema of late limbs prevent the denervation-induced inhibition of mitosis and oppose blastema regression. Our data indicate that FGF-2 is a good candidate for the endogenous mitogenic factor responsible for blastema formation and growth in amputated and denervated early limbs. However, in amputated late limbs, the very limited fgf-2 expression is not sufficient to promote blastema formation in the absence of nerves. (C) 2001 Academic Press. ispartof: Developmental biology vol:231 issue:2 pages:436-446 ispartof: location:United States status: published

Mice deficient in α-sarcoglycan (Sgca-null mice) develop progressive muscular dystrophy and serve as a model for human limb girdle muscular dystrophy type 2D. Sgca-null mice suffer a more severe myopathy than that of mdx mice, the model for Duchenne muscular dystrophy. This is the opposite of what is observed in humans and the reason for this is unknown. In an attempt to understand the cellular basis of this severe muscular dystrophy, we isolated clonal populations of myogenic progenitor cells (MPCs), the resident postnatal muscle progenitors of dystrophic and wild-type mice. MPCs from Sgca-null mice generated much smaller clones than MPCs from wild-type or mdx dystrophic mice. Impaired proliferation of Sgca-null myogenic precursors was confirmed by single fiber analysis and this difference correlated with Sgca expression during MPC proliferation. In the absence of dystrophin and associated proteins, which are only expressed after differentiation, SGCA complexes with and stabilizes FGFR1. Deficiency of Sgca leads to an absence of FGFR1 expression at the membrane and impaired MPC proliferation in response to bFGF. The low proliferation rate of Sgca-null MPCs was rescued by transduction with Sgca-expressing lentiviral vectors. When transplanted into dystrophic muscle, Sgca-null MPCs exhibited reduced engraftment. The reduced proliferative ability of Sgca-null MPCs explains, at least in part, the severity of this muscular dystrophy and also why wild-type donor progenitor cells engraft efficiently and consequently ameliorate disease. Mice deficient in alpha-sarcoglycan (Sgca-null mice) develop progressive muscular dystrophy and serve as a model for human limb girdle muscular dystrophy type 2D. Sgca-null mice suffer a more severe myopathy than that of mdx mice, the model for Duchenne muscular dystrophy. This is the opposite of what is observed in humans and the reason for this is unknown. In an attempt to understand the cellular basis of this severe muscular dystrophy, we isolated clonal populations of myogenic progenitor cells (MPCs), the resident postnatal muscle progenitors of dystrophic and wild-type mice. MPCs from Sgca-null mice generated much smaller clones than MPCs from wild-type or mdx dystrophic mice. Impaired proliferation of Sgca-null myogenic precursors was confirmed by single fiber analysis and this difference correlated with Sgca expression during MPC proliferation. In the absence of dystrophin and associated proteins, which are only expressed after differentiation, SGCA complexes with and stabilizes FGFR1. Deficiency of Sgca leads to an absence of FGFR1 expression at the membrane and impaired MPC proliferation in response to bFGF. The low proliferation rate of Sgca-null MPCs was rescued by transduction with Sgca-expressing lentiviral vectors. When transplanted into dystrophic muscle, Sgca-null MPCs exhibited reduced engraftment. The reduced proliferative ability of Sgca-null MPCs explains, at least in part, the severity of this muscular dystrophy and also why wild-type donor progenitor cells engraft efficiently and consequently ameliorate disease. Funding This work was supported in part by FWO-Odysseus Program Grant No. G.0907.08; Research Council of the University of Leuven, Grant No. OT/09/053 and EME-C2161-GOA/11/012; Wicka Charity Funds, USA, Grant No. zkb8720; the Italian Ministry of University and Scientific Research Grant No. 2005067555_003, PRIN 2006-08, CARIPLO 2007.5639 and 2005.2008; EC FP7 CARE-MI No. 242038; and Optistem No. 223098. Acknowledgements We thank Gianpaolo Papaccio for critical discussion of the manuscript; K. P. Campbell for providing Sgca-null mice; Laura Perani, Rossana Tonlorenzi and Rudi Micheletti for technical assistance; Christina Vochten for professional secretarial service; Paolo Luban for a kind donation; Dr Shea Carter for manuscript editing; and Dr Maurizio Moggio (Bank of DNA, Cell Lines and Nerve-Muscle-Cardiac Tissues/Telethon Genetic BioBank Network, Ospedale Maggiore Policlinico Mangiagalli e Regina Elena, Milan, Italy) for providing the LGMD2D sample