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Tumor Biology |
Stimulates Human Glioblastoma Cell Growth through the Activation of Both Extracellular Signal-regulated Kinases 1/2 and Akt1
Pharmacology and Neuroscience, National Institute for Cancer Research c/o Advanced Biotechnology Center, Genoa, Italy [S. B., R. B., A. B., C. P., P. P., G. S.]; Section of Pharmacology, Department of Oncology, Biology and Genetic, University of Genoa, Genoa, Italy [S. B., R. B., A. B., C. P., T. F., G. S.]; Service of Pathology Hospital San Martino, Genoa, Italy [J. L. R.]; and Division of Neurosurgery, Department of Neurology and of the Vision Sciences, University of Genoa, Genoa, Italy [G. L. Z., R. S.]
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ABSTRACT |
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induces proliferation in a dose-dependent manner in both cell lines. Moreover, we observed that SDF-1-dependent proliferation is correlated with phosphorylation and activation of both extracellular signal-regulated kinases 1/2 and Akt and that these kinases are independently involved in glioblastoma cell proliferation. The role of CXCR4 stimulation in glioblastoma cell growth is further demonstrated by the ability of human monoclonal CXCR4 antibody (clone 12G5) to inhibit the SDF-1-induced proliferation as well as the proliferation induced by SDF-1-releasing treatments (lipopolysaccharide and 1% fetal bovine serum). These data support a role for SDF-1 in the regulation of glioblastoma growth in vitro, likely through an autocrine/paracrine mechanism. |
INTRODUCTION |
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TopABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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In this work, we studied the expression of the chemokine SDF-1, recently renamed CXCL12, and its receptor, CXCR4, in two glioblastoma cell lines and in human primary glioblastoma tissues and studied the possible role of these proteins in tumor progression. SDF-1 is a chemokine of the CXC subfamily originally characterized as a pre-B-cell stimulatory factor and cloned from bone marrow cell supernatants. SDF-1 exists in two alternative splicing variants, and ß, of which SDF-1 is the most abundant (2)
. SDF-1 nucleotide and amino acid sequences are highly conserved during evolution, suggesting that this molecule may play important biological roles. This chemokine is a chemotactic factor for T cells, monocytes, pre-B cells, dendritic cells, and hematopoietic progenitor cells and supports B-cell progenitor and CD34+ cell proliferation (3
, 4)
. However, its expression is not restricted to immune and hematic cells; SDF-1 has also been identified at the CNS level in neuronal, astroglial, microglial, and endothelial cells (3
, 5
, 6)
. SDF-1 exerts its activity by interacting with the CXCR4 receptor, a member of the G protein-coupled receptor superfamily. The interaction between CXCR4 and SDF-1 appears to be unique, whereas other chemokines may recognize multiple receptors. Disruption of the murine CXCR4 or SDF-1 genes displays a similar embryological lethal phenotype, characterized by deficient B-lymphopoiesis and myelopoiesis, abnormal cardiac and neuronal development, and defects in vasculogenesis (7
, 8)
. CXCR4, similar to SDF-1, is expressed in various tissues and also at brain level in different cell types, including endothelial cells, embryonic germinal neuroepithelium and mature neurons, glial cells, and microglia cells, and seems to be involved in different CNS pathologies (3
, 4
, 9)
. Recent data showed that CXCR4 and SDF-1 mRNAs are colocalized in glioblastomas and that their expression increases with tumor grade and is associated with regions of necrosis and angiogenesis (10)
. Other authors have demonstrated that SDF-1 and CXCR4 are involved in normal and malignant glial cell proliferation in vitro (11, 12, 13, 14)
.
In this study, we demonstrate that CXCR4 and SDF-1 are expressed in several human glioblastoma tumor tissues and cell lines. We also show that SDF-1 stimulates the proliferation of glioblastoma cells in vitro through the phosphorylation and activation of ERK1/2 and Akt. Moreover, we show that SDF-1 likely exerts its proliferative activity via the activation of an autocrine/paracrine pathway.
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MATERIALS AND METHODS |
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TopABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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was purchased from Pepro Tech EC Ltd. (London, United Kingdom).
Tissue Samples
Nine human glioblastoma tumor tissues corresponding to grade IV gliomas according to the WHO classification were obtained from the Neurosurgery Division (University of Genoa, Genoa, Italy).
Cells and Culture Conditions
Human Glioblastoma Cell Lines.
U87- MG and DBTRG-05MG cell lines were purchased from the bank of biological material Interlab Cell Line Collection (Advanced Biotechnology Center, Genoa, Italy). Cells were cultured at 37°C in 5% CO2 in RPMI 1640 supplemented with 10% heat-inactivated FBS (Euroclone, West Yorkshire, United Kingdom), 2 mM glutamine, 100 IU/ml penicillin, 100 µg/ml streptomycin, and 1% nonessential amino acids.
Rat Cortical Type I Astrocytes.
Cultures of rat cortical type I astrocytes were performed as described previously (5)
.
RT-PCR
Total RNA was isolated from different human brain tumor tissues and human glioblastoma cells using acid phenol extraction (5)
. Before cDNA synthesis, RNA was treated with 40 units of RNase-free DNase I (Boehringer Mannheim, Indianapolis, IN) for 45 min. To control whether contaminating genomic DNA was present, RNA samples not reverse transcribed were subjected to PCR amplification. The gene-specific primers used for CXCR4 and SDF-1 amplification were as follows: CXCR4, 5'-ggccctcaagaccacagtca-3' (sense) and 5'-ttagctggagtgaaaacttgaag-3' (antisense); and SDF-1, 5'-atgaacgccaaggtcgtggtc-3' (sense) and 5'-ggtctgttgtgcttacttgttt-3' (antisense), which recognize both the and ß isoform. The primers for the ß-actin were 5'-tccggagacggggtca-3' (sense) and 5'-cctgcttgctgatcca-3' (antisense).
Western Blot
Human glioblastoma cells and astrocytes were serum starved for 48 h and then treated as described in "Results." Cells were lysed in 1% NP40, 20 mM Tris-HCl (pH 8), 137 mM NaCl, 10% glycerol, 2 mM EDTA, 1 mM phenylmethylsulfonyl fluoride, 1 µg/ml leupeptin, 1 mM sodium orthovanadate, and 10 mM NaF for 10 min at 4°C. Nuclei were removed by centrifugation in a minifuge at 5000 rpm for 10 min at 4°C, and cell lysates were assayed for protein content using the Bradford protein assay (Bio-Rad Laboratories, Hercules, CA). Proteins (5–10 µg) were resuspended in 2x reducing sample buffer [2% SDS, 62.5 mM Tris (pH 6.8), 0.01% bromphenol blue, 1.43 mM 2-mercaptoethanol, and 10% glycerol], separated by SDS-PAGE, transferred on polyvinylidene difluoride membrane (Bio-Rad), and blotted with polyclonal antibodies. The detection of immunocomplex was performed using the enhanced chemiluminescence kit (Amersham Pharmacia Biotech). Western blot of SDF-1 secretion in the culture supernatants of human glioblastoma cell lines was performed as described previously (5)
.
SDF-1 Quantification by ELISA
Human glioblastoma cells were serum deprived for 48 h and treated with LPS (10 µg/ml) or 1% FBS in the last 16 h. The culture supernatants were tested by solid-phase ELISA for SDF-1, according to the manufacturer’s recommendation (R&D Systems).
[3H]Thymidine Incorporation Assay
DNA synthesis activity was measured by means of [3H]thymidine uptake, as described previously (12)
. Cells were plated at 5 x 104 cells/well in 24-well plates and serum starved for 48 h before being treated with SDF-1 for 24 h; during the last 4 h, cells were pulsed with 1 µCi/ml [3H]thymidine (Amersham Pharmacia Biotech). When indicated, PD98059 (10 µM) was added to the cells for 10 min before SDF-1 stimulation and during the chemokine stimulation; LY294002 (10 µM) or 12G5 antibody (10 µg/ml) was added to the cells 30 min before chemokine stimulation and during chemokine stimulation.
Statistical Analysis
Experiments were performed in quadruplicate and repeated at least three times. Data are expressed as mean ± SE, and statistical significance was assessed by Student’s t test for independent groups. P 
0.05 was considered statistically significant.
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RESULTS |
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, Lanes 1–9, all nine tumor tissues express CXCR4 mRNA, whereas only three express SDF-1 mRNA. We also identified specific amplification products for CXCR4 and SDF-1 mRNAs in two different human glioblastoma cell lines, DBTRG-05MG and U87-MG (Fig. 1A
, Lanes 10 and 11). We also analyzed the ability of human glioblastoma cell lines to secrete SDF-1. DBTRG-05MG and U87-MG cells were cultured in serum-free medium in the presence or absence of LPS for 16 h, and the supernatants were analyzed by Western blot using an anti-SDF-1 polyclonal antibody. Our results showed that both tumor cell lines secreted a significant amount of SDF-1 under basal conditions and that this amount was even further increased after LPS stimulation (Fig. 1B)
.
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Induces DNA Synthesis in Human Glioblastoma Cell Lines in Vitro through ERK1/2 Activation. to induce cell proliferation in U87-MG and DBTRG-05MG cells by means of [3H]thymidine incorporation assay. Glioblastoma cells were serum starved for 48 h and treated with increasing concentrations of SDF-1 (3.12–50 nM) for 24 h. The results, shown in Fig. 2A
, indicate that in both cell lines, SDF-1 causes a dose-dependent induction of DNA synthesis with a maximum effect at 12.5 nM (90% and 80% for DBTRG-05MG and U87-MG cells, respectively). Using higher concentrations of SDF-1 (25 and 50 nM), a down-regulation of the response was observed. In fact, at the highest concentration tested (50 nM), the increase in [3H]thymidine incorporation was only 27% for DBTRG-05MG and 50% for U87-MG cells.
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treatment, we focused our attention on the activation of the MAPK ERK1/2, which converts extracellular stimuli to intracellular signals that control gene expression, cell proliferation, differentiation, and survival. By Western blot analysis, we analyzed SDF-1-induced ERK1/2 activation in DBTRG-05MG and U87-MG glioblastoma cell lines and cultured rat type I astrocytes, which were used as a model of normal glial cells. Cells were treated with SDF-1 (12.5 nM), and the cell lysates were analyzed for the presence of phosphorylated, and thus activated, forms of ERK1/2. In all of the cell types, a significant amount of phosphorylated ERK1/2 was detected under basal conditions, even after 48 h of serum deprivation (Fig. 2B)
. In both glioblastoma tumor cell lines, SDF-1 treatment induced ERK1/2 activation that was detectable after 15 min of treatment and lasted up to 30 min (Fig. 2B)
. Similarly, in normal astrocytes, SDF-1 induced activation of ERK1/2 that was clearly evident after 5 min of stimulation, further increased after 15 min, and lasted up to 30 min. The analysis of cell lysates for the total expression of MAPKs ensured the equal loading of proteins in the different lanes (data not shown).
To correlate the effects of SDF-1 on cell proliferation and ERK1/2 activation, we analyzed SDF-1-induced proliferation in U87-MG and DBTRG-05MG cells in the presence of PD98059, a pharmacological inhibitor of MAPK/ERK kinase (MEK). Cells were treated for 24 h with SDF-1 (12.5 nM) in the presence or absence of PD98059. SDF-1-dependent proliferation was completely blocked by pretreatment with PD98059 in both cell lines (Fig. 2C)
. Moreover, this compound inhibited basal proliferation significantly in DBTRG-05MG cells and to a lesser extent in U87-MG cells. In agreement with these results, PD98059 reduced both basal and SDF-1-stimulated ERK1/2 phosphorylation in both cell lines (Fig. 2D)
. Thus, these data show that ERK1/2 activation contributes to the U87-MG and DBTRG-05MG cell proliferation mediated by SDF-1.
Role of Akt Phosphorylation in SDF-1-dependent Cell Proliferation.
Akt is a known downstream effector of the PI3K-dependent signaling cascade. Recently, it was shown that besides the ERK1/2 pathway, chemokines stimulate PI3K, leading to activation of Akt (9)
. It has also been shown that the Akt pathway is strongly involved in the development of human glioblastomas (1)
. Thus, we examined the role of SDF-1 treatment in the activity of Akt by Western blot analysis. U87-MG and DBTRG-05MG cell lines were challenged with SDF-1 (12.5 nM), and their lysates were analyzed for Akt phosphorylation using an antibody that reacts specifically with the phosphorylated Ser473 located at the COOH terminus. Our results (shown in Fig. 3A
) demonstrate that, in contrast to normal astrocytes, in the two glioblastoma cell lines analyzed, a slight Akt activation had already occurred under basal conditions and was further increased after 15–30 min of SDF-1 stimulation. SDF-1 treatment also induces Akt activation in cultured type I astrocytes, although normal astrocytes do not show basal activation of Akt. The same lysates were also analyzed for total expression of Akt to ensure an equal loading of proteins in the different lanes (data not shown).
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, we studied the effect of LY294002, a specific PI3K inhibitor, on both SDF-1-dependent cell proliferation and Akt activation. As shown in Fig. 3B
, this compound strongly reduced U87-MG and DBTRG-05MG cell proliferation induced by SDF-1 treatment. LY294002 also reduced basal DNA synthesis in both cell lines. Moreover, both basal and SDF-1-dependent Akt activation were completely blocked by LY294002 in these cell lines (Fig. 3C)
. Furthermore, in U87-MG cells, LY294002 did not inhibit ERK1/2 phosphorylation, and PD98059 did not affect phosphorylation of Akt under both basal and SDF-1-stimulated conditions (Fig. 3D)
, indicating that ERK1/2 and PI3K/Akt pathways are independently involved in the proliferative signal mediated by SDF-1. Similar results were obtained in DBTRG-05MG cells (data not shown).
CXCR4/SDF-1 Interaction Is Responsible for Autocrine/Paracrine Regulation of Glioblastoma Growth in Vitro.
To demonstrate the involvement of CXCR4 in cell proliferation induced by SDF-1 and SDF-1-inducing agents, we used the monoclonal antihuman CXCR4 neutralizing antibody, clone 12G5. U87-MG and DBTRG-05MG cells were stimulated for 24 h with SDF-1 (12.5 nM) in the presence or absence of 12G5 (10 µg/ml). Fig. 4A
shows that 12G5 antibody reduced the SDF-1-dependent increase of [3H]thymidine incorporation by 50% in U87-MG cells and by 70% in DBTRG-05MG cells, whereas it did not significantly modulate basal proliferation of both cell lines.
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secretion in U87-MG cells is highly enhanced compared with the control cells (Fig. 4B)
. In light of these data, we evaluated the effect of 12G5 antibody on U87-MG cell proliferation in the presence or absence of LPS or 1% FBS. As shown in Fig. 4C
, the presence of LPS or FBS enhances the incorporation of [3H]thymidine by about 40% and 30%, respectively, over basal activity, and this increase is significantly blocked by 12G5 antibody. On the contrary, as shown above, this antibody does not influence the [3H]thymidine incorporation observed under basal conditions. These data indicate that SDF-1 produced and secreted by glioblastoma cells in the presence of LPS or serum induces cell proliferation via an autocrine/paracrine pathway in vitro. |
DISCUSSION |
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TopABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONREFERENCES |
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directly stimulates the proliferation of primary cultures of rat astrocytes (12)
, suggesting that SDF-1/CXCR4 molecules could be involved in the activation of astrocytes that occurs during astrogliosis and that abnormal CXCR4 signaling could play a role in the genesis of the aberrant proliferative behavior of glioblastoma tumors. It was recently reported that CXCR4 is implicated in the regulation of glioma proliferation because transfection of CXCR4 antisense or treatment with anti-CXCR4 antibody inhibited the proliferation of glioma cells in vitro (13)
. Similarly SDF-1 has also been involved in the progression of cancer of different histological derivation such as epithelial ovarian cancer, breast tumors, neuroblastomas, and pancreatic ductal adenocarcinomas (14, 15, 16)
. Rempel et al. (10)
have shown by immunohistochemical analysis of different glioblastoma tumors that CXCR4 and SDF-1 do not colocalize with regions highly expressing the proliferation marker MIB-1 but are present in tumoral regions characterized by necrosis and angiogenesis. Thus, these authors suggest that SDF-1 may promote neoangiogenesis to supply nutrients to sustain tumor growth. It was also proposed that SDF-1 induces up-regulation of several other chemokines in astroglioma cells, such as MCP-1 and interleukin 8, which appear to be involved in vascular endothelial cell proliferation and tumor neovascularization (9
, 17)
. All of this evidence indicates that CXCR4 and SDF-1 could play a primary role in brain tumor growth through diverse biological effects including cell proliferation, angiogenesis, and chemokine induction.
Here we studied the direct role of SDF-1-CXCR4 interaction in glioblastoma proliferation in vitro using two different human glioblastoma cell lines, U87-MG and DBTRG-05MG. We demonstrate that these cell lines express both CXCR4 and SDF-1 mRNAs and are also able to secrete SDF-1 under basal conditions or in response to LPS or FBS stimulation. Moreover, we showed that in these glioblastoma cell lines, stimulation with exogenous SDF-1 induces a significant dose-dependent proliferation, starting at concentrations as low as 3.15 nM and reaching maximal effect at 12.5 nM. At the higher concentration tested (25 and 50 nM), we observed a progressive decrease of SDF-1-dependent proliferation, which is probably due to CXCR4 down-regulation. In a recent publication, Geminder et al. (18)
demonstrated that in neuroblastoma cells, which constitutively express both CXCR4 and SDF-1, exposure to different doses of exogenous SDF-1 induces a dose-dependent reduction of CXCR4 expression. In a previous study, we also proposed that in cortical type I astrocytes, the SDF-1 secretion induced after LPS stimulation may cause CXCR4 down-regulation (5)
.
In normal and tumor glial cells, the transduction of proliferative signals involves a family of ERKs, whose enzymatic activity increases in response to SDF-1 (9)
. We directly studied the role of SDF-1 on ERK1/2 activation in glioblastoma cells. Our data show that in both tumor cell lines analyzed, basal ERK1/2 activation is increased after SDF-1 stimulation, as observed in cortical type I astrocytes. We also demonstrate that PD98059, a MEK inhibitor, reduced both SDF-1-induced cell proliferation and ERK1/2 activation, indicating that ERK1 and ERK2 are involved in the proliferative signal of SDF-1. Similarly, basal cell proliferation and ERK1/2 activation observed under serum-free conditions were reduced by PD98059 treatment, suggesting that ERK1/2 may also be an important component of basal glioblastoma cell proliferation.
Another important signal transduction pathway in glioblastoma is the PI3K/Akt pathway. Akt is a serine-threonine kinase whose phosphorylation and activation by specific kinases are dependent on PI3K activation in response to mitogens. However, besides its well-documented role in cell survival, Akt can regulate a variety of cellular functions including growth, differentiation, cell cycle progression, transcription, translation, and cellular metabolism. Constitutive activation of the PI3K/Akt pathway has been observed in several human cancers, and up to 80% of glioblastoma express elevated levels of Akt (1)
. Moreover, in a recent work (19)
, it was proposed that gliomas, in which the Akt pathway is impaired, show limited ability to proliferate. In our study, we demonstrate that both the glioblastoma cell lines analyzed show constitutive activation of Akt, which, on the contrary, is not observed in normal astrocytes. Moreover, SDF-1 treatment is able to increase this basal Akt activation. We also show that LY294002, a specific PI3K inhibitor, completely blocked both basal and SDF-1-induced cell proliferation and Akt activation observed in U87-MG and DBTRG-05MG cells. However, although both PI3K/Akt and ERK1/2 pathways are involved in glioblastoma cell proliferation, these pathways appear to operate independently: the activation of ERK1/2 does not require PI3K/Akt; and the activation of PI3K/Akt does not require ERK1/2. In cultures of cortical type I astrocytes, SDF-1 also induces cell proliferation through phosphorylation of ERK1/2 and activation of the PI3K pathway, but at odds with tumor cells, in normal astrocytes wortmannin, a PI3K inhibitor, reduced both SDF-1-dependent proliferation and ERK1/2 activation, indicating that ERK1/2 and PI3K pathways are intermingled (12)
. Thus, the loss of inter-regulation between ERK1/2 and PI3K/Akt pathways in CXCR4 signaling, and consequently of their multiple downstream targets, may contribute to glioblastoma tumor proliferation.
We directly demonstrated the role of CXCR4 on SDF-1-induced proliferation in U87-MG and DBTRG-05MG cells using the monoclonal antihuman CXCR4 neutralizing antibody, clone 12G5, which partially blocks the CXCR4 activity. This antibody inhibits SDF-1-dependent DNA synthesis by 50% and 70% in U87-MG and DBTRG-05MG cells, respectively. No significant effect on DNA synthesis was observed in untreated cells, indicating that under basal conditions, the amount of SDF-1 secretion by these cells is not sufficient to promote cell proliferation. Using an ELISA test specific for SDF-1, we demonstrate that both LPS and low FBS concentration significantly increased SDF-1 secretion in U87-MG cells. Similarly, by RT-PCR, we identified an increase in SDF-1 mRNA expression in U87-MG cells cultured in the presence of FBS compared with serum-deprived cells (data not shown). Moreover, we show that 12G5 antibody significantly reduces the increase of [3H]thymidine incorporation observed in U87-MG cells performed in the presence of LPS or 1% FBS. Thus, these data suggest that the amount of SDF-1 produced and secreted by U87-MG cells in the presence of LPS or FBS may lead to an autocrine/paracrine regulation of cell growth. Interestingly, a different responsiveness with regard to this autocrine loop was observed in primary cultures of rat astrocytes. Indeed, whereas a significant basal SDF-1 secretion was observed in the tumor cell lines studied, normal astrocytes did not show any constitutive secretion (5)
. This observation suggests that only in pathological conditions, such as AIDS dementia complex during HIV infection, this autocrine loop may take place in normal cells, likely due to the infection/inflammation process. Conversely, the constitutive activation of this pathway may occur during tumor transformation and progression. Furthermore, an autocrine/paracrine proliferative loop in glioma and nonglioma tumors was also demonstrated for other chemokines, such as interleukin 8 (20)
. This CXC chemokine is constitutively expressed in several human cancer tissues and cell lines and can act as an autocrine growth factor in different tumors such as human melanoma, gliomas, pancreatic cancer, colon cancer, and malignant mesothelioma (9
, 20)
. Because our studies demonstrated that although CXCR4 mRNA was present in all GBMs analyzed, SDF-1 mRNA was expressed by only one-third of tumor tissues analyzed, we can hypothesize that other cell types present in the tumor microenvironment may also contribute to the regulation of glioblastoma tumor growth and diffusion through the production of SDF-1. GBMs are the most vascularized tumors and are often infiltrated with numerous lymphoid/myeloid cells that are attracted by the cytokines and chemokines secreted by the tumor cells (1)
. Thus, the activation of macrophages and T lymphocytes results in the secretion of additional chemokines including SDF-1, which may contribute to tumor expansion by interacting with the CXCR4 present on the tumor extracellular membranes. In addition, we cannot exclude the role of endothelial cells, neurons, and microglial and/or reactive astrocytes, which are known to secrete SDF-1 (9)
, in sustaining tumor development.
In summary, we demonstrate that SDF-1 and CXCR4 are expressed in both human glioblastoma tissues and cell lines. Our experiments also indicate that stimulation with exogenous SDF-1 induces cell growth, likely activating ERK1/2 and Akt pathways that are independently involved in glioblastoma proliferation. Moreover, we show that glioblastoma cells are able to secrete SDF-1, which is responsible for an autocrine/paracrine proliferative loop in vitro.
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FOOTNOTES |
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1 Supported by Associazione Italiana Ricerca sul Cancro 2001-2002, Progetti Finalizzati MISAN 2001, and a grant from the San Paolo Foundation (Torino, Italy) to G. S., S. B. was supported by a fellowship from Federazione Italiana Ricerca Cancro. 
2 To whom requests for reprints should be addressed, at Service of Pharmacology and Neuroscience, National Institute for Cancer Research c/o Advanced Biotechnology Center, Largo Rosanna Benzi, 10, 16132 Genoa, Italy. Phone: 39-010-5737254; Fax: 39-010-5737257; E-mail: schettini{at}cba.unige.it
3 The abbreviations used are: CNS, central nervous system; ERK, extracellular signal-regulated kinase; FBS, fetal bovine serum; GBM, glioblastoma multiforme; LPS, lipopolysaccharide; MAPK, mitogen-activated protein kinase; MEK, MAPK/ERK kinase; PI3K, phosphatidylinositol 3'-kinase; SDF, stromal cell-derived factor; RT-PCR, reverse transcription-PCR; CXCR4, CXC chemokine receptor 4.
Received 8/ 6/02. Accepted 2/19/03.
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D. Zheng, S.-h. Oh, Y. Jung, and B. E. Petersen Oval Cell Response in 2-Acetylaminofluorene/Partial Hepatectomy Rat Is Attenuated by Short Interfering RNA Targeted to Stromal Cell-Derived Factor-1 Am. J. Pathol., December 1, 2006; 169(6): 2066 - 2074. [Abstract] [Full Text] [PDF]
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 M. Aghi, K. S. Cohen, R. J. Klein, D. T. Scadden, and E. A. Chiocca Tumor Stromal-Derived Factor-1 Recruits Vascular Progenitors to Mitotic Neovasculature, where Microenvironment Influences Their Differentiated Phenotypes. Cancer Res., September 15, 2006; 66(18): 9054 - 9064. [Abstract] [Full Text] [PDF]
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 S. Brule, N. Charnaux, A. Sutton, D. Ledoux, T. Chaigneau, L. Saffar, and L. Gattegno The shedding of syndecan-4 and syndecan-1 from HeLa cells and human primary macrophages is accelerated by SDF-1/CXCL12 and mediated by the matrix metalloproteinase-9 Glycobiology, June 1, 2006; 16(6): 488 - 501. [Abstract] [Full Text] [PDF]
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A. Ottaiano, R. Franco, A. Aiello Talamanca, G. Liguori, F. Tatangelo, P. Delrio, G. Nasti, E. Barletta, G. Facchini, B. Daniele, et al. Overexpression of Both CXC Chemokine Receptor 4 and Vascular Endothelial Growth Factor Proteins Predicts Early Distant Relapse in Stage II-III Colorectal Cancer Patients. Clin. Cancer Res., May 1, 2006; 12(9): 2795 - 2803. [Abstract] [Full Text] [PDF]
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 T. Kakinuma and S. T. Hwang Chemokines, chemokine receptors, and cancer metastasis J. Leukoc. Biol., April 1, 2006; 79(4): 639 - 651. [Abstract] [Full Text] [PDF]
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 F. Vianello, N. Papeta, T. Chen, P. Kraft, N. White, W. K. Hart, M. F. Kircher, E. Swart, S. Rhee, G. Palu, et al. Murine B16 Melanomas Expressing High Levels of the Chemokine Stromal-Derived Factor-1/CXCL12 Induce Tumor-Specific T Cell Chemorepulsion and Escape from Immune Control. J. Immunol., March 1, 2006; 176(5): 2902 - 2914. [Abstract] [Full Text] [PDF]
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T. Florio, S. Casagrande, F. Diana, A. Bajetto, C. Porcile, G. Zona, S. Thellung, S. Arena, A. Pattarozzi, A. Corsaro, et al. Chemokine Stromal Cell-Derived Factor 1{alpha} Induces Proliferation and Growth Hormone Release in GH4C1 Rat Pituitary Adenoma Cell Line through Multiple Intracellular Signals Mol. Pharmacol., February 1, 2006; 69(2): 539 - 546. [Abstract] [Full Text] [PDF]
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J. Zhang, S. Sarkar, and V.W. Yong The chemokine stromal cell derived factor-1 (CXCL12) promotes glioma invasiveness through MT2-matrix metalloproteinase Carcinogenesis, December 1, 2005; 26(12): 2069 - 2077. [Abstract] [Full Text] [PDF]
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H. Kishimoto, Z. Wang, P. Bhat-Nakshatri, D. Chang, R. Clarke, and H. Nakshatri The p160 family coactivators regulate breast cancer cell proliferation and invasion through autocrine/paracrine activity of SDF-1{alpha}/CXCL12 Carcinogenesis, October 1, 2005; 26(10): 1706 - 1715. [Abstract] [Full Text] [PDF]
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H. Nakagawa, K. Yoshioka, E. Miyahara, Y. Fukushima, M. Tamura, and K. Itoh Intrathecal Administration of Y-27632, a Specific Rho-Associated Kinase Inhibitor, for Rat Neoplastic Meningitis Mol. Cancer Res., August 1, 2005; 3(8): 425 - 433. [Abstract] [Full Text] [PDF]
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L. J. Bendall, R. Baraz, J. Juarez, W. Shen, and K. F. Bradstock Defective p38 Mitogen-Activated Protein Kinase Signaling Impairs Chemotaxic but not Proliferative Responses to Stromal-Derived Factor-1{alpha} in Acute Lymphoblastic Leukemia Cancer Res., April 15, 2005; 65(8): 3290 - 3298. [Abstract] [Full Text] [PDF]
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A. Katayama, T. Ogino, N. Bandoh, S. Nonaka, and Y. Harabuchi Expression of CXCR4 and Its Down-Regulation by IFN-{gamma} in Head and Neck Squamous Cell Carcinoma Clin. Cancer Res., April 15, 2005; 11(8): 2937 - 2946. [Abstract] [Full Text] [PDF]
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S.-B. Peng, V. Peek, Y. Zhai, D. C. Paul, Q. Lou, X. Xia, T. Eessalu, W. Kohn, and S. Tang Akt Activation, but not Extracellular Signal-Regulated Kinase Activation, Is Required for SDF-1{alpha}/CXCR4-Mediated Migration of Epitheloid Carcinoma Cells Mol. Cancer Res., April 1, 2005; 3(4): 227 - 236. [Abstract] [Full Text] [PDF]
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C. C. Schimanski, S. Schwald, N. Simiantonaki, C. Jayasinghe, U. Gonner, V. Wilsberg, T. Junginger, M. R. Berger, P. R. Galle, and M. Moehler Effect of Chemokine Receptors CXCR4 and CCR7 on the Metastatic Behavior of Human Colorectal Cancer Clin. Cancer Res., March 1, 2005; 11(5): 1743 - 1750. [Abstract] [Full Text] [PDF]
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S. Scala, A. Ottaiano, P. A. Ascierto, M. Cavalli, E. Simeone, P. Giuliano, M. Napolitano, R. Franco, G. Botti, and G. Castello Expression of CXCR4 Predicts Poor Prognosis in Patients with Malignant Melanoma Clin. Cancer Res., March 1, 2005; 11(5): 1835 - 1841. [Abstract] [Full Text] [PDF]
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 I. Tegeder and G. Geisslinger Opioids As Modulators of Cell Death and Survival--Unraveling Mechanisms and Revealing New Indications Pharmacol. Rev., September 1, 2004; 56(3): 351 - 369. [Abstract] [Full Text] [PDF]
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G. G. Vaday, S.-B. Hua, D. M. Peehl, M. H. Pauling, Y.-H. Lin, L. Zhu, D. M. Lawrence, H. D. Foda, and S. Zucker CXCR4 and CXCL12 (SDF-1) in Prostate Cancer: Inhibitory Effects of Human Single Chain Fv Antibodies Clin. Cancer Res., August 15, 2004; 10(16): 5630 - 5639. [Abstract] [Full Text] [PDF]
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K. Kawada, M. Sonoshita, H. Sakashita, A. Takabayashi, Y. Yamaoka, T. Manabe, K. Inaba, N. Minato, M. Oshima, and M. M. Taketo Pivotal Role of CXCR3 in Melanoma Cell Metastasis to Lymph Nodes Cancer Res., June 1, 2004; 64(11): 4010 - 4017. [Abstract] [Full Text] [PDF]
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 X. Wu, D.-J. Li, M.-M. Yuan, Y. Zhu, and M.-Y. Wang The Expression of CXCR4/CXCL12 in First-Trimester Human Trophoblast Cells Biol Reprod, June 1, 2004; 70(6): 1877 - 1885. [Abstract] [Full Text] [PDF]
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 J. B. Rubin, A. L. Kung, R. S. Klein, J. A. Chan, Y. Sun, K. Schmidt, M. W. Kieran, A. D. Luster, and R. A. Segal A small-molecule antagonist of CXCR4 inhibits intracranial growth of primary brain tumors PNAS, November 11, 2003; 100(23): 13513 - 13518. [Abstract] [Full Text] [PDF]
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